PREPUBLICATION COPY UNCORRECTED PROOFS

MARIJUANA AND MEDICINE: ASSESSING THE SCIENCE BASE

[Image]

INSTITUTE OF MEDICINE ---------------------------------------------------------------------------

MARIJUANA AND MEDICINE: ASSESSING THE SCIENCE BASE

Janet E. Joy, Stanley J. Watson, Jr., and John A. Benson, Jr., Editors

Division of Neuroscience and Behavioral Health

INSTITUTE OF MEDICINE

NATIONAL ACADEMY PRESS Washington, D.C. 1999 ---------------------------------------------------------------------------

NATIONAL ACADEMY PRESS · 2101 Constitution Avenue, N.W.*Washington, D.C. 20418

NOTICE: The project that is the subject of this report was approved by the Governing Board of the National Research Council, whose members are drawn from the councils of the National Academy of Sciences, the National Academy of Engineering, and the Institute of Medicine. The Principal Investigators responsible for the report were chosen for their special competences and with regard for appropriate balance.

The Institute of Medicine was chartered in 1970 by the National Academy of Sciences to enlist distinguished members of the appropriate professions in the examination of policy matters pertaining to the health of the public. In this, the Institute acts under both the Academy's 1863 congressional charter responsibility to be an adviser to the federal government and its own initiative in identifying issues of medical care, research, and education. Dr. Kenneth I. Shine is president of the Institute of Medicine.

This study was supported under contract No. DC7C02 from the Executive Office of the President, Office of the National Drug Control Policy.

Additional copies of this report are available for sale from the National Academy Press, 2101 Constitution Avenue, N.W., Box 285, Washington, DC 20055. Call (800) 624-6242 or (202) 334-3313 (in the Washington metropolitan area) or visit the National Academy Press's online bookstore at http://www.nap.edu.

The full text of this report is available on line at http://www.nap.edu

For more information about the Institute of Medicine, visit the IOM home page at http://www2.nas.edu/iom.

Copyright 1999 by the National Academy of Sciences. All rights reserved.

Printed in the United States of America

The serpent has been a symbol of long life, healing, and knowledge among almost all cultures and religions since the beginning of recorded history. The image adopted as a logotype by the Institute of Medicine is based on a relief carving from ancient Greece, now held by the Staatliche Museum in Berlin.

---------------------------------------------------------------------------

Principal Investigators and Advisory Panel

JOHN A. BENSON, JR., co-Principal Investigator, Dean and Professor of Medicine, Emeritus, Oregon Health Sciences University School of Medicine, Portland, Oregon

STANLEY J. WATSON, JR., co-Principal Investigator, co-Director and Research Scientist, Mental Health Research Institute, University of Michigan, Ann Arbor, Michigan

STEVEN R. CHILDERS, Professor, Bowman Gray School of Medicine, Wake Forest University, Center for Neuroscience, Winston-Salem, North Carolina

J. RICHARD CROUT, Private Consultant, Bethesda, Maryland

THOMAS J. CROWLEY, Professor, University of Colorado, Health Sciences Center, Addiction Research and Treatments Services, Denver, Colorado

JUDITH FEINBERG, Professor, University of Cincinnati Medical Center, Division of Infectious Diseases, Department of Internal Medicine, Cincinnati, Ohio

HOWARD L. FIELDS, Professor, University of California in San Francisco, Neurology and Anesthesiology, San Francisco, California

DOROTHY HATSUKAMI, Professor, University of Minnesota, Department of Psychiatry, Minneapolis, Minnesota

ERIC B. LARSON, Medical Director, University of Washington Medical Center, Seattle, Washington

BILLY R. MARTIN, Professor, Virginia Commonwealth University, Department of Pharmacology, Richmond, Virginia

TIMOTHY VOLLMER, Professor, Yale School of Medicine, Yale MS Research Center, New Haven, Connecticut

iii ---------------------------------------------------------------------------

Study Staff

JANET E. JOY, Study Director DEBORAH O. YARNELL, Research Associate AMELIA B. MATHIS, Project Assistant CHERYL MITCHELL, Administrative Assistant (until September, 1998) THOMAS J. WETTERHAN, Research Assistant (until September, 1988) CONSTANCE M. PECHURA, Division Director (until April 1998) NORMAN GROSSBLATT, Manuscript Editor

Consultant

MIRIAM DAVIS

Section Staff

CHARLES H. EVANS, JR., Head, Health Sciences Section LINDA DEPUGH, Administrative Assistant CARLOS GABRIEL, Financial Associate

iv ---------------------------------------------------------------------------

REVIEWERS

This report has been reviewed in draft form by individuals chosen for their diverse perspectives and technical expertise, in accordance with procedures approved by the National Research Council's Report Review Committee. The purpose of this independent review is to provide candid and critical comments that will assist the Institute of Medicine in making the published report as sound as possible and to ensure that the report meets institutional standards for objectivity, evidence, and responsiveness to the study charge. The review comments and draft manuscript remain confidential to protect the integrity of the deliberative process. The committee wishes to thank the following individuals for their participation in the review of this report:

JAMES ANTHONY, Johns Hopkins University JACK BARCHAS, Cornell University Medical College SUMNER BURSTEIN, University of Massachusetts Medical School AVRAM GOLDSTEIN, Stanford University LESTER GRINSPOON, Harvard Medical School MILES HERKENHAM, National Institute of Mental Health, National Institutes of Health HERBERT KLEBER, Columbia University GEOFFREY LEVITT, Venable Attorneys at Law KENNETH MACKIE, University of Washington RAPHAEL MECHOULAM, Hebrew University CHARLES O'BRIEN, University of Pennsylvania JUDITH RABKIN, Columbia University ERIC VOTH, International Drug Strategy Institute

While the individuals listed above have provided constructive comments and suggestions, it must be emphasized that responsibility for the final content of this report rests entirely with the authoring committee and the Institute of Medicine.

v ---------------------------------------------------------------------------

Preface

Why study the medical value of marijuana now? What circumstances provoked this analysis and report? There have been a variety of influences since the IOM Report of 1982. First, advocates of personal choice with a growing distrust of scientific medicine sought alternatives congruent with their values about health and life. This view was expressed at the ballot box in recent state referenda. Proponents claimed their own "scientific evidence" of marijuana's safety and effectiveness. Others, distressed by the societal ravages of drug abuse, especially among young people, view legalized medical marijuana as a subterfuge enabling liberalization, the potential "gateway" to even more harmful substance abuse. Meanwhile, there have been remarkable and accelerating advances of relevant knowledge in molecular and behavioral neuroscience, in particular newly elaborated systems of transmitters, receptors, and antagonists all illuminating the physiological effects of cannabinoids, both those found in nature and those normally found in the brain. (Cannabinoids are the group of compounds related to THC, the primary psychoactive ingredient in marijuana.) The new science could inform policies responding to the public divide.

In January 1997, the White House Office of National Drug Control Policy (ONDCP) asked the Institute of Medicine to conduct a review of the scientific evidence to assess the potential health benefits and risks of marijuana and its constituent cannabinoids. That review began in August 1997 and culminates with this report.

Information for this study was gathered through analysis of the relevant scientific literature, scientific workshops, site visits to cannabis buyers' clubs and HIV/AIDS clinics, and extensive consultation with biomedical and social scientists. Three 2-day workshops -- in Irvine, California; New Orleans, Louisiana; and Washington, DC -- were open to the public and included scientific presentations and also reports, mostly from patients and their families, about their experiences with and perspectives on the medical use of marijuana. Scientific experts in various fields were selected to talk about the latest research on marijuana, cannabinoids, and related topics. In addition, advocates for and against the medical use of marijuana were invited to present scientific evidence in support of their positions. Finally, the Institute of Medicine appointed a panel of nine experts to advise the study team on technical issues.

Public outreach included setting up a Web site that provided information about the study and asked for input from the public. The Web site was open for

---------------------------------------------------------------------------

comments from November 1997 until November 1998. Some 130 organizations were invited to participate in the public workshops. Many people in the organizations --particularly those opposed to the medical use of marijuana -- felt that a public forum was not conducive to expressing their views; they were invited to communicate their opinions (and reasons for holding them) by mail or telephone. As a result, roughly equal numbers of persons and organizations opposed to and in favor of the medical use of marijuana were heard from.

Advances in cannabinoid science of the last 16 years have given rise to a wealth of new opportunities for the development of medically useful cannabinoid based drugs. The accumulated data suggest a variety of indications, particularly for pain relief, nausea, and appetite stimulation. For patients, such as those with AIDS or undergoing chemotherapy, who suffer simultaneously from severe pain, nausea, and appetite loss, cannabinoid drugs might offer broad spectrum relief not found in any other single medication.

Marijuana is not a completely benign substance. It is a powerful drug with a variety of effects. However, the harmful effects to individuals from the perspective of possible medical use of marijuana are not necessarily the same as the harmful physical effects of drug abuse.

Although marijuana smoke delivers THC and other cannabinoids to the body, it also delivers harmful substances, including most of those found in tobacco smoke. In addition, plants contain a variable mixture of biologically-active compounds and cannot be expected to provide a precisely defined drug effect. For those reasons, the report concludes that the future of cannabinoid drugs lies not in smoked marijuana, but in chemically-defined drugs that act on the cannabinoid systems that are a natural component of human physiology Until such drugs can be developed and made available for medical use, the report recommends interim solutions.

John A. Benson, Jr., M.D. Stanley J. Watson, Jr. M.D., Ph.D. Principal Investigators

---------------------------------------------------------------------------

Acknowledgments

This report covers such a broad range of disciplines -- neuroscience, pharmacology, immunology, drug abuse, drug laws, and a variety of medical specialties including neurology, oncology, infectious diseases, and ophthalmology -- that it would not have been complete without the generous support of many people. Our goal in preparing this report was to identify the solid ground of scientific consensus, and steer clear of the muddy distractions of opinions that are inconsistent with careful scientific analysis. To this end, we consulted extensively with experts in each of the disciplines covered in this report. We are deeply indebted to each of them.

Members of the Advisory Panel, selected because each is recognized as among the most accomplished in their respective disciplines (see list), provided guidance to the study team throughout the study -- from helping to lay the intellectual framework to reviewing early drafts of the report.

The following people wrote invaluable background papers for the report: Steven R. Childers, Paul Consroe, J. Richard Gralla, Howard Fields, Norbert Kaminski, Paul Kaufman, Thomas Klein, Donald Kotler, Richard Musty, Clara Sanudo-Pena, C. Robert Schuster, Stephen Sidney, Donald P. Tashkin, and J. Michael Walker.

Others provided expert technical commentary on draft sections of the report: Richard Bonnie, Keith Green, Frederick Fraunfelder, Andrea Hohmann, John McAnulty, Craig Nichols, John Nutt, and Robert Pandina.

Still others responded to many inquiries, provided expert counsel, or shared their unpublished data: Paul Consroe, Geoffrey Levitt, Richard Musty, David Pate, Roger Pertwee, Raphael Mechoulam. Clara Sanudo-Pena, Carl Soderstrom, J. Michael Walker, and Scott Yarnell.

Miriam Davis, consultant to the study team, provided excellent written material for the chapter on cannabinoid drug development.

The reviewers for the report (see list) provided extensive and constructive suggestions for improving the report. It was greatly enhanced by their thoughtful attentions.

Many of these people assisted us through many iterations of the report. All of them made contributions that were essential to the strength of the report. At the same time, it must be emphasized that responsibility for the final content of report rests entirely with the authors and the Institute of Medicine.

We would also like to thank the people who hosted our visits to their organizations. They were unfailingly helpful and generous with their time. Jeffrey Jones and members of the Oakland Cannabis Buyers' Cooperative, Denis Peron of

---------------------------------------------------------------------------

the San Francisco Cannabis Cultivators Club, Scott Imler staff at the Los Angeles Cannabis Resource Center, Victor Hernandez and members of Californians Helping Alleviate Medical Problems (CHAMPS), Michael Weinstein of the AIDS Health Care Foundation, and Marsha Bennett of the Louisiana State University Medical Center.

We also appreciate the many people who spoke at the public workshops or wrote to share their views on the medical use of marijuana (see appendix AA).

Jane Sanville, project officer for the study sponsor, was consistently helpful during the many negotiations and discussion held throughout study process.

Many IOM staff members provided much appreciated administrative, research, and intellectual support during the study. Robert Cook-Deegan, Marilyn Field, Constance Pechura, Daniel Quinn, Michael Stoto provided thoughtful and insightful comments on draft sections of the report. Others provided advice and consultation in many other aspects of the study process: Kathleen Stratton, Susan Fourt, Carolyn Fulco, Carlos Gabriel, Linda Kilroy, Catharyn Liverman, Clyde Behney, Dev Mani. As project assistant throughout the study, Amelia Mathis was tireless, gracious, and reliable.

Deborah Yarnell's contribution as Research Associate for this study was outstanding. She organized site visits, researched and drafted technical material for the report, and consulted extensively with relevant experts to ensure the technical accuracy of the text. The quality of her contributions throughout this study was exemplary.

Finally, the Principal Investigators on this study wish to personally thank Janet Joy for her deep commitment to the science and shape of this report. In addition, her help in integrating the entire data gathering and information organization of this report were nothing short of essential. Her knowledge of neurobiology, her sense of quality control, and her unflagging spirit over the 18 months illuminated the subjects and were indispensable to the study's successful completion.

---------------------------------------------------------------------------

EXECUTIVE SUMMARY

EXECUTIVE SUMMARY 2 EFFECTS OF ISOLATED CANNABINOIDS 3 RISKS ASSOCIATED WITH MEDICAL USE OF MARIJUANA6 USE OF SMOKED MARIJUANA 8

ES.1 ---------------------------------------------------------------------------

EXECUTIVE SUMMARY

Public opinion on the medical value of marijuana has been sharply divided. Some dismiss medical marijuana as a hoax that exploits our natural compassion for the sick; others claim it is a uniquely soothing medicine that has been withheld from patients through regulations based on false claims. Proponents of both views cite 'scientific evidence' to support their views and have expressed those views at the ballot box in recent state elections. In January 1997, the White House Office of National Drug Control Policy (ONDCP) asked the Institute of Medicine to conduct a review of the scientific evidence to assess the potential health benefits and risks of marijuana and its constituent cannabinoids (see box: Statement of Task). That review began in August 1997 and culminates with this report.

The ONDCP request came in the wake of state "medical marijuana" initiatives. In November 1996, voters in California and Arizona passed referenda designed to permit the use of marijuana as medicine. Although Arizona's referendum was invalidated five months later, the referenda galvanized a national response. In November 1998, voters in six states (Alaska, Arizona, Colorado, Nevada, Oregon, and Washington) passed ballot initiatives in support of medical marijuana. (The Colorado vote will not count, however, because after the vote was taken a court ruling determined there had not been enough valid signatures to place the initiative on the ballot.)

Can marijuana relieve health problems? Is it safe for medical use? Those straightforward questions are embedded in a web of social concerns, most of which lie outside the scope of this report. Controversies concerning the nonmedical use of marijuana spill over onto the medical marijuana debate and obscure the real state of scientific knowledge. In contrast with the many disagreements bearing on social issues, the study team found substantial consensus among experts in the relevant disciplines on the scientific evidence about potential medical uses of marijuana.

This report summarizes and analyzes what is known about the medical use of marijuana, it emphasizes evidence-based medicine (derived from knowledge and experience informed by rigorous scientific analysis), as opposed to belief-based medicine (derived from judgment, intuition, and beliefs untested by rigorous science).

Throughout this report, marijuana refers to unpurified plant substances, including leaves or flower tops whether consumed by ingestion or smoking. References to "the effects of marijuana" should be understood to include the composite effects of its various components; that is, the effects of THC, the primary psychoactive ingredient in marijuana, are included among its effects, but not all the effects of marijuana are necessarily due to THC. Cannabinoids are the group of compounds related to THC, whether found in the marijuana plant, in animals, or synthesized in chemistry laboratories.

ES.2 ---------------------------------------------------------------------------

Three focal concerns in evaluating the medical use of marijuana are:

*Evaluation of the effects of isolated cannabinoids. *Evaluation of the health risks associated with the medical use of marijuana. *Evaluation of the efficacy of marijuana.

EFFECTS OF ISOLATED CANNABINOIDS

Cannabinoid Biology

Much has been learned since a 1982 IOM Marijuana and Health report. Although it was clear then that most of the effects of marijuana were due to its actions on the brain, there was little information about how THC acted on brain cells (neurons), which cells were affected by THC, or even what general areas of the brain were most affected by THC. Additionally, too little was known about cannabinoid physiology to offer any scientific insights into the harmful or therapeutic effects of marijuana. That all changed with the identification and characterization of cannabinoid receptors in the 1980s and 1990s. During the last 16 years, science has advanced greatly and can tell us much more about the potential medical benefits of cannabinoids.

CONCLUSION: At this point, our knowledge about the biology of marijuana and cannabinoids allows us to make some general conclusions:

*Cannabinoids likely have a natural role in pain modulation, control of movement, and memory.

*The natural role of cannabinoids in immune systems is likely multifaceted and remains unclear.

*The brain develops tolerance to cannabinoids.

*Animal research demonstrates the potential for dependence, but this potential is observed under a narrower range of conditions than with benzodiazepines, opiates, cocaine, or nicotine.

*Withdrawal symptoms can be observed in animals, but appear to be mild compared to opiates or benzodiazepines, such as diazepam (Valium®).

ES.3 ---------------------------------------------------------------------------

CONCLUSION: The different cannabinoid receptor types found in the body appear to play different roles in normal human physiology. In addition, some effects of cannabinoids appear to be independent of those receptors. The variety of mechanisms through which cannabinoids can influence human physiology underlies the variety of potential therapeutic uses for drugs that might act selectively on different cannabinoid systems.

RECOMMENDATION 1: Research should continue into the physiological effects of synthetic and plant-derived cannabinoids and the natural function of cannabinoids found in the body. Because different cannabinoids appear to have different effects, cannabinoid research should include, but not be restricted to, effects attributable to THC alone.

Efficacy Of Cannabinoid Drugs

The accumulated data indicate a potential therapeutic value for cannabinoid drugs, particularly for symptoms such as pain relief, control of nausea and vomiting, and appetite stimulation. The therapeutic effects of cannabinoids are best established for THC, which is generally one of the two most abundant of the cannabinoids in marijuana. (Cannabidiol, the precursor of THC, is generally the other most abundant cannabinoid.)

The effects of cannabinoids on the symptoms studied are generally modest, and in most cases, there are more effective medications. However, people vary in their responses to medications and there will likely always be a subpopulation of patients who do not respond well to other medications. The combination of cannabinoid drug effects (anxiety reduction, appetite stimulation, nausea reduction, and pain relief) suggests that cannabinoids would be moderately well suited for certain conditions, such as chemotherapy-induced nausea and vomiting and AIDS wasting.

Defined substances, such as purified cannabinoid compounds, are preferable to plant products which are of variable and uncertain composition. Use of defined cannabinoids permits a more precise evaluation of their effects, whether in combination or alone. Medications that can maximize the desired effects of cannabinoids and minimize the undesired effects can very likely be identified.

Although most scientists who study cannabinoids agree that the pathways to cannabinoid drug development are clearly marked, there is no guarantee that the fruits of scientific research will be made available to the public for medical use. Cannabinoid-based drugs will only become available if public investment in cannabinoid drug research is sustained, and if there is enough incentive for private enterprise to develop and market such drugs.

ES.4 ---------------------------------------------------------------------------

CONCLUSION: Scientific data indicate the potential therapeutic value of cannabinoid drugs, primarily THC, for pain relief, control of nausea and vomiting, and appetite stimulation; smoked marijuana, however, is a crude THC delivery system that also delivers harmful substances.

RECOMMENDATION 2: Clinical trials of cannabinoid drugs for symptom management should be conducted with the goal of developing rapid-onset, reliable, and safe delivery systems.

Influence Of Psychological Effects On Therapeutic Effects

The psychological effects of THC and similar cannabinoids pose three issues for the therapeutic use of cannabinoid drugs. First, for some patients -- particularly older patients with no previous marijuana experience -- the psychological effects are disturbing. Those patients report experiencing unpleasant feelings and disorientation after being treated with THC, generally more severe for oral THC than for smoked marijuana. Second, for conditions such as movement disorders or nausea, in which anxiety exacerbates the symptoms, the anti-anxiety effects of cannabinoid drugs can influence symptoms indirectly. This can be beneficial or can create false impressions of the drug effect. Third, in cases where symptoms are multifaceted, the combination of THC effects might provide a form of adjunctive therapy; for example, AIDS wasting patients would likely benefit from a medication that simultaneously reduces anxiety, pain, and nausea while stimulating appetite.

CONCLUSION: The psychological effects of cannabinoids, such as anxiety reduction, sedation, and euphoria can influence their potential therapeutic value Those effects are potentially undesirable for certain patients and situations, and beneficial for others. In addition, psychological effects can complicate the interpretation of other aspects of the drug effect.

RECOMMENDATION 3: Psychological effects of cannabinoids such as anxiety reduction and sedation, which can influence medical benefits, should be evaluated in clinical trials.

ES.5 ---------------------------------------------------------------------------

RISKS ASSOCIATED WITH MEDICAL USE OF MARIJUANA

Physiological Risks

Marijuana is not a completely benign substance. It is a powerful drug with a variety of effects. However, except for the harms associated with smoking, the adverse effects of marijuana use are within the range of effects tolerated for other medications. The harmful effects to individuals from the perspective of possible medical use of marijuana are not necessarily the same as the harmful physical effects of drug abuse. When interpreting studies purporting to show the harmful effects of marijuana, it is important to keep in mind that the majority of those studies are based on smoked marijuana, and cannabinoid effects cannot be separated from the effects of inhaling smoke of burning plant material and contaminants.

For most people, the primary adverse effect of acute marijuana use is diminished psychomotor performance. It is, therefore, inadvisable to operate any vehicle or potentially dangerous equipment while under the influence of marijuana, THC, or any cannabinoid drug with comparable effects. In addition, a minority of marijuana users experience dysphoria, or unpleasant feelings. Finally, the short-term immunosuppressive effects are not well established but, if they exist, are not likely great enough to preclude a legitimate medical use.

The chronic effects of marijuana are of greater concern for medical use and fall into two categories: the effects of chronic smoking, and the effects of THC. Marijuana smoking is associated with abnormalities of cells lining the human respiratory tract. Marijuana smoke, like tobacco smoke, is associated with increased risk of cancer, lung damage, and poor pregnancy outcomes. Although cellular, genetic, and human studies all suggest that marijuana smoke is an important risk factor for the development of respiratory cancer, proof that habitual marijuana smoking does or does not cause cancer awaits the results of well-designed studies.

CONCLUSION: Numerous studies suggest that marijuana smoke is an important risk factor in the development of respiratory disease.

RECOMMENDATION 4: Studies to define the individual health risks of smoking marijuana should be conducted, particularly among populations in which marijuana use is prevalent.

ES.6 ---------------------------------------------------------------------------

Marijuana Dependence And Withdrawal

A second concern associated with chronic marijuana use is dependence on the psychoactive effects of THC Although few marijuana users develop dependence, some do. Risk factors for marijuana dependence are similar to those for other forms of substance abuse. In particular, antisocial personality and conduct disorders are closely associated with substance abuse.

CONCLUSION: A distinctive marijuana withdrawal syndrome has been identified, but it is mild and short-lived. The syndrome includes restlessness, irritability, mild agitation, insomnia, sleep EEG disturbance, nausea, and cramping.

Marijuana As A "Gateway" Drug

Patterns in progression of drug use from adolescence to adulthood are strikingly regular. Because it is the most widely used illicit drug, marijuana is predictably the first illicit drug most people encounter. Not surprisingly, most users of other illicit drugs have used marijuana first. In fact, most drug users begin with alcohol and nicotine before marijuana -- usually before they are of legal age.

In the sense that marijuana use typically precedes rather than follows initiation of other illicit drug use, it is indeed a "gateway" drug. But because underage smoking and alcohol use typically precede marijuana use, marijuana is not the most common, and is rarely the first, "gateway" to illicit drug use. There is no conclusive evidence that the drug effects of marijuana are causally linked to the subsequent abuse of other illicit drugs. An important caution is that data on drug use progression cannot be assumed to apply to the use of drugs for medical purposes. It does not follow from those data that if marijuana were available by prescription for medical use, the pattern of drug use would remain the same as seen in illicit use.

Finally, there is a broad social concern that sanctioning the medical use of marijuana might increase its use among the general population. At this point there are no convincing data to support this concern. The existing data are consistent with the idea that this would not be a problem if the medical use of marijuana were as closely regulated as other medications with abuse potential.

ES.7 ---------------------------------------------------------------------------

CONCLUSION: Present data on drug use progression neither support nor refute the suggestion that medical availability would increase drug abuse. However, this question is beyond the issues normally considered for medical uses of drugs, and should not be a factor in evaluating the therapeutic potential of marijuana or cannabinoids.

USE OF SMOKED MARIJUANA

Because of the health risks associated with smoking, smoked marijuana should generally not be recommended for long-term medical use. Nonetheless, for certain patients, such as the terminally ill or those with debilitating symptoms the long-term risks are not of great concern. Further, despite the legal, social, and health problems associated with smoking marijuana, it is widely used by certain patient groups.

RECOMMENDATION 5: Clinical trials of marijuana use for medical purposes should be conducted under the following limited circumstances: trials should involve only short-term marijuana use (less than six months); be conducted in patients with conditions for which there is reasonable expectation of efficacy; be approved by institutional review boards; and collect data about efficacy.

The goal of clinical trials of smoked marijuana would not be to develop marijuana as a licensed drug, but rather as a first step towards the possible development of nonsmoked, rapid-onset cannabinoid delivery systems. However, it will likely be many years before a safe and effective cannabinoid delivery system, such as an inhaler, will be available for patients. In the meantime there are patients with debilitating symptoms for whom smoked marijuana might provide relief The use of smoked marijuana for those patients should weigh both the expected efficacy of marijuana and ethical issues in patient care, including providing information about the known and suspected risks of smoked marijuana use.

ES.8 ---------------------------------------------------------------------------

RECOMMENDATION 6: Short-term use of smoked marijuana (less than six months) for patients with debilitating symptoms (such as intractable pain or vomiting) must meet the following conditions:

*failure of all approved medications to provide relief has been documented;

*the symptoms can reasonably be expected to be relieved by rapid onset cannabinoid drugs;

*such treatment is administered under medical supervision in a manner that allows for assessment of treatment effectiveness;

*and involves an oversight strategy comparable to an institutional review board process that could provide guidance within 24 hours of a submission by a physician to provide marijuana to a patient for a specified use.

Until a non-smoked, rapid-onset cannabinoid drug delivery system becomes available, we acknowledge that there is no clear alternative for people suffering from chronic conditions that might be relieved by smoking marijuana, such as pain or AIDS wasting. One possible approach is to treat patients as e-of-1 clinical trials, in which patients are fully informed of their status as experimental subjects using a harmful drug delivery system, and in which their condition is closely monitored and documented under medical supervision, thereby increasing the knowledge base of the risks and benefits of marijuana use under such conditions.

ES.9 ---------------------------------------------------------------------------

Statement of Task

The study will assess what is currently known, and not known about the medical use of marijuana. It will include a review of the science base regarding the mechanism of action of marijuana, an examination of the peer-reviewed scientific literature on the efficacy of therapeutic uses of marijuana, and the costs of using various forms of marijuana versus approved drugs for specific medical conditions (e.g., glaucoma, multiple sclerosis, wasting diseases, nausea, and palm).

The study will also include an evaluation of the acute and chronic effects of marijuana on health and behavior; a consideration of the adverse effects of marijuana use compared with approved drugs; an evaluation of the efficacy of different delivery systems for marijuana (e.g., inhalation vs. oral); and an analysis of the data concerning marijuana as a gateway drug; and an examination of the possible differences in the effects of marijuana due to age and type of medical condition.

Specific Issues

Specific issues to be addressed fall under three broad categories: the science base, therapeutic use, and economics.

Science Base *Review of neuroscience related to marijuana, particularly relevance of new studies on addiction and craving *Review of behavioral and social science base of marijuana use, particularly assessment of the relative risk of progression to other drugs following marijuana use *Review of the literature determining which chemical components of crude marijuana are responsible of possible therapeutic effects and for side effects

Therapeutic Use *Evaluation of any conclusions on the medical use of marijuana drawn by other groups *Efficacy and side-effects of various delivery systems for marijuana compared to existing medications for glaucoma, wasting syndrome, pain, nausea, or other symptoms *Differential effects of various forms of marijuana that relate to age or type of disease.

Economics *Costs of various forms of marijuana compared with costs of existing medications for glaucoma, wasting syndrome, pain, nausea, or other symptoms *Assessment of differences between marijuana and existing medications in terms of access and availability

These specific areas, along with the assessments described above will be integrated into a broad description and assessment of the available literature relevant to the medical use of marijuana.

ES.10 ---------------------------------------------------------------------------

Recommendations

Recommendation 1: Research should continue into the physiological effects of synthetic and plant-derived cannabinoids and the natural function of cannabinoids found in the body. Because different cannabinoids appear to have different effects, cannabinoid research should include, but not be restricted to effects attributable to THC alone

Scientific data indicate the potential therapeutic value of cannabinoid drugs for pain relief, control of nausea and vomiting, and appetite stimulation. This value would be enhanced by a rapid onset of drug effect.

Recommendation 2: Clinical trials of cannabinoid drugs for symptom management should be conducted with the goal of developing rapid-onset, reliable, and safe delivery systems.

The psychological effects of cannabinoids are probably important determinants of their potential therapeutic value. They can influence symptoms indirectly which could create false impressions of the drug effect or be beneficial as a form of adjunctive therapy.

Recommendation 3: Psychological effects of cannabinoids such as anxiety reduction and sedation, which can influence perceived medical benefits, should be evaluated in clinical trials.

Numerous studies suggest that marijuana smoke is an important risk factor in the development of respiratory diseases, but the data that could conclusively establish or refute this suspected link have not been collected.

ES.11 ---------------------------------------------------------------------------

Recommendation 4: Studies to define the individual health risks of smoking marijuana should be conducted, particularly among populations in which marijuana use is prevalent.

Because marijuana is a crude THC delivery system that also delivers harmful substances, smoked marijuana should generally not be recommended for medical use. Nonetheless, marijuana is widely used by certain patient groups, which raises both safety and efficacy issues.

Recommendation 5: Clinical trials of marijuana use for medical purposes should be conducted under the following limited circumstances: trials should involve only short-term marijuana use (less than six months); be conducted in patients with conditions for which there is reasonable expectation of efficacy; be approved y institutional review boards; and collect data about efficacy.

If there is any future for marijuana as a medicine, it lies in its isolated components, the cannabinoids and their synthetic derivatives. Isolated cannabinoids will provide more reliable effects than crude plant mixtures. Therefore, the purpose of clinical trials of smoked marijuana would not be to develop marijuana as a licensed drug, but such trials could be a first step towards the development of rapid-onset, nonsmoked cannabinoid delivery systems.

Recommendation 6: Short term use of smoked marijuana (less than six months) for patients with debilitating symptoms (such as intractable pain or vomiting) must meet the following conditions:

*failure of all approved medications to provide relief has been documented;

*the symptoms can reasonably be expected to be relieved by rapid-onset cannabinoid drugs;

*such treatment is administered under medical supervision in a manner that allows for assessment of treatment effectiveness;

*and involves an oversight strategy comparable to an institutional review board process that could provide guidance within 24 hours of a submission by a physician to provide marijuana to a patient for a specified use.

ES.12 ---------------------------------------------------------------------------

Chapter 1 --->>>

Preface and Executive Summary

Chapter 1 Introduction

Chapter 2 Cannabinoids and Animal Physiology

Chapter 3 First Do No Harm: Consequences of Marijuana

Chapter 4 The Medical Value of Marijuana and Related Substances

Chapter 5 Development of Cannabinoid Drugs

Appendix

INTRODUCTION

CHAPTER 1. INTRODUCTION 1 How This Study Was Conducted 3 Marijuana Today 4 Who Uses Medical Marijuana? 9 Cannabis and the cannabinoids18 Organization of the Report 23 References 24

1.1 ---------------------------------------------------------------------------

Chapter 1. Introduction

This report summarizes and analyzes what is known about the medical use of marijuana; it emphasizes evidence-based medicine (derived from knowledge and experience informed by rigorous scientific analysis), as opposed to belief-based medicine (derived from judgment, intuition, and beliefs untested by rigorous science).

Scientific data on controversial subjects are commonly misinterpreted, over interpreted, and misrepresented, and the medical marijuana debate is no exception We have tried to present the scientific studies in such a way as to reveal their strengths and limitations. One of the goals of this report is to help people to understand the scientific data, including the logic behind the scientific conclusions, so it goes into greater detail than previous reports on the subject. In many cases, we have explained why particular studies are inconclusive and what sort of evidence is needed to support particular claims about the harms or benefits attributed to marijuana. Ideally, this report will enable the thoughtful reader to interpret the new information about marijuana that will continue to emerge rapidly well after this report is published.

Can marijuana relieve health problems? Is it safe for medical use? Those straightforward questions are embedded in a web of social concerns, which lie outside the scope of this report. Controversies concerning non medical use of marijuana spill over onto the medical marijuana debate and tend to obscure the real state of scientific knowledge. In contrast with the many disagreements bearing on the social issues, the study team has found substantial consensus, among experts in the relevant disciplines, on the scientific evidence bearing on potential medical use.

This report analyzes science, not the law. As in any policy debate, the value of scientific analysis is that it can provide a foundation for further discussion. Distilling scientific evidence does not in itself solve a policy problem. What it can do is illuminate the common ground, bringing to light fundamental differences out of the shadows of misunderstanding and misinformation that currently prevail. Scientific analysis cannot be the end of the debate, but it should at least provide the basis for an honest and informed discussion.

Our analysis of the evidence and arguments concerning the medical use of marijuana focuses on the strength of the supporting evidence, and does not refer to the motivations of people who put forth the evidence and arguments. That is, it is not relevant to scientific validity whether an argument is put forth by someone who believes that all marijuana use should be legal or by someone who believes that any marijuana use is highly damaging to individual users and to society as a whole. Nor does this report comment on the degree to which scientific analysis is compatible with current regulatory policy. Although many have argued that current drug laws pertaining to marijuana are inconsistent with scientific data, it is important to understand that decisions about drug regulation are based on a variety of moral and social considerations, as well as on medical and scientific ones.

1.2 ---------------------------------------------------------------------------

Even when a drug is used only for medical purposes, value judgments affect policy decisions concerning its medical use. For example, the magnitude of a drug's expected medical benefit affects regulatory judgments about the acceptability of risks associated with its use. Also, although a drug is normally approved for medical use only on proof of its "safety and efficacy," patients with life-threatening conditions are sometimes (under protocols for "compassionate use") allowed access to unapproved drugs whose benefits and risks are uncertain. Value judgments play an even more substantial role in regulatory decisions concerning drugs, such as marijuana, that are sought and used for non medical purposes. Then policy-makers must take into account not only the risks and benefits associated with medical use, but also possible interactions between the regulatory arrangements governing medical use and the integrity of the legal controls set up to restrict non medical use.

It should be clear that many elements of drug control policy lie outside the realm of biology and medicine. Ultimately, the complex moral and social judgments that underlie drug control policy must be made by the American people and their elected officials A goal of this report is to evaluate the biological and medical factors that should be taken into account in making those judgments.

How This Study Was Conducted

Information was gathered through scientific workshops, site visits, analysis of the relevant scientific literature, and extensive consultation with biomedical and social scientists. The three 2-day workshops -- in Irvine, California, New Orleans, Louisiana; and Washington, DC -- were open to the public and included scientific presentations and reports, mostly from patients and their families, about their experiences with and perspectives on the medical use of marijuana. Scientific experts in various fields were selected to talk about the latest research on marijuana, cannabinoids, and related topics (listed in appendix A). The selection of the experts was based on recommendations by their peers, who ranked them among the most accomplished scientists and the most knowledgeable about marijuana and cannabinoids in their own fields. In addition, advocates for (John Morgan) and against (Eric A. Voth) the medical use of marijuana were invited to present scientific evidence in support of their positions.

Information presented at the scientific workshops was supplemented by analysis of the scientific literature, and evaluating the methods used in various studies and the validity of the authors' conclusions. Different kinds of clinical studies are useful in different ways: results of a controlled, double-blind study with adequate sample sizes can be expected to apply to the general population from which study subjects were drawn; an isolated case report can suggest further studies, but cannot be presumed to be broadly applicable; and survey data can be highly informative, but are generally limited by the need to rely on self-reports of drug use and on unconfirmed medical diagnoses. This report relies mainly on the most

1.3 ---------------------------------------------------------------------------

relevant and methodologically rigorous studies available, and treats the results of more limited studies cautiously. In addition, study results are presented in such a way as to allow thoughtful readers to judge the results themselves.

IOM appointed a panel of nine experts to advise the study team on technical issues. These included neurology and the treatment of pain (Howard Fields), regulation of prescription drugs (J. Richard Crout), AIDS wasting and clinical trials (Judith Feinberg), treatment and pathology of multiple sclerosis (Timothy Vollmer), drug dependence among adolescents (Thomas Crowley), varieties of drug dependence (Dorothy Hatsukami), internal medicine, health care delivery, and clinical epidemiology (Eric B. Larson), cannabinoids and marijuana pharmacology (Billy R. Martin), and cannabinoid neuroscience (Steven R. Childers).

Public outreach included setting up a Web site that provided information about the study and asked for input from the public. The Web site was open for comment from November 1997 until November 1998. Some 130 organizations were invited to participate in the public workshops. Many people in the organizations -- particularly those opposed to the medical use of marijuana -- felt that a public forum was not conducive to expressing their views; they were invited to communicate their opinions (and reasons for holding them) by mail or telephone. As a result, roughly equal numbers of persons and organizations opposed to and in favor of the medical use of marijuana were heard from.

The study team visited four cannabis buyers' clubs in California (the Oakland Cannabis Buyers' Cooperative, the San Francisco Cannabis Cultivators Club, the Los Angeles Cannabis Resource Center, and Californians Helping Alleviate Medical Problems, or CHAMPS) and two HIV-AIDS clinics (the AIDS Health Care Foundation in Los Angeles and the Louisiana State University Medical Center in New Orleans). We listened to many individual stories from the buyers' clubs about using marijuana to treat a variety of symptoms and heard clinical observations on the use of Marinol® to treat AIDS patients. Marinol® is the brand name for dronabinol, which is [Image]9-tetrahydrocannabinol (THC) in pill form and is available by prescription for the treatment of nausea associated with chemotherapy and AIDS wasting.

1.4 ---------------------------------------------------------------------------

Marijuana, Today

The Changing Legal Landscape

In the 20th century, marijuana has been used more for its euphoric effects than as a medicine. Its psychological and behavioral effects have concerned public officials since the drug first appeared in the southwestern and southern states during the first two decades of the century. By 1931, at least 29 states had prohibited use of the drug for non medical purposes.3 Marijuana was first regulated at the federal level by the Marijuana Tax Act of 1937, which required anyone producing, distributing, or using marijuana for medical purposes to register and pay a tax, and which effectively prohibited non medical use of the drug. Although the Act did not make medical use of marijuana illegal, it did make it expensive and inconvenient. In 1942, marijuana was removed from the U.S. Pharmacopoeia because it was believed to be a harmful and addictive drug that caused psychoses, mental deterioration, and violent behavior.

In the late 1960s and early 1970s, there was a sharp increase in marijuana use among adolescents and young adults. The current legal status of marijuana was established in 1970 with the passage of the Controlled Substances Act, which divided drugs into five Schedules and placed marijuana in Schedule I, the category for drugs with high potential for abuse and no accepted medical use (see appendix B. scheduling criteria). In 1972, the National Organization for the Reform of Marijuana Legislation (NORML), an organization which supports decriminalization of marijuana, unsuccessfully petitioned the Bureau of Narcotics and Dangerous Drugs to move marijuana from Schedule I to Schedule II. NORML argued that marijuana is therapeutic in numerous serious ailments, is less toxic, and in many cases more effective than conventional medicines.13 Thus, for 25 years, the medical marijuana movement has been closely linked with the marijuana-decriminalization movement, which has colored the debate. Many people criticized that association in their letters to IOM and during the public workshops of this study. The argument against the medical use of marijuana presented most often to the IOM study team is that "the medical marijuana movement is a Trojan horse"; that is, it is a deceptive tactic used by advocates of marijuana decriminalization who would exploit the public's sympathy for seriously ill patients.

Since NORML's petition in 1972, there have been a variety of legal decisions concerning marijuana. From 1973 to 1978, eleven states adopted statutes that decriminalized use of marijuana, although some of them recriminalized marijuana use in the 1980s and 1990s. During the 1970s, reports of the medical value of marijuana began to appear, particularly claims that marijuana relieved the nausea associated with chemotherapy. Health departments in six states conducted small studies to investigate the reports. When the AIDS epidemic spread in the 1980s, patients found that marijuana sometimes relieved their symptoms, most dramatically those associated with AIDS wasting. Over this period, a number of defendants

1.5 ---------------------------------------------------------------------------

charged with unlawful possession of marijuana claimed that they were using the drug to treat medical conditions and that violation of the law was therefore justified (the so-called "medical necessity" defense). Although most courts rejected these claims, some accepted them.8

Against that backdrop, voters in California and Arizona in 1996 passed two referendums that attempted to legalize the medical use of marijuana under particular conditions. Public support for patient access to marijuana for medical use appears substantial; public opinion polls taken during 1997 and 1998 generally report 60-70 percent of respondents in favor of allowing medical uses of marijuana.15 However, those referendums are at odds with federal laws regulating marijuana, and their implementation raises complex legal questions.

Despite the current level of interest, referendums and public discussions have not been well informed by carefully reasoned scientific debate. Although previous reports have all called for more research, the nature of the research that will be most helpful depends greatly on the specific health conditions to be addressed. And while there have been important recent advances in our understanding of the physiological effects of marijuana, few of the recent investigators have had the time or resources to permit detailed analysis. The results those advances, only now beginning to be explored, have significant implications for the medical marijuana debate.

Several months after the passage of the California and Arizona medical marijuana referendums, the Office of National Drug Control Policy (ONDCP) asked whether IOM would conduct a scientific review of the medical value of marijuana and its constituent compounds. In August 1997, IOM formally began the study and appointed John A. Benson Jr. and Stanley J. Watson Jr. to serve as principle investigators for the study. The charge to IOM was to review the medical use of marijuana and the harms and benefits attributed to it (details are given in appendix C).

1.6 ---------------------------------------------------------------------------

Medical Marijuana Legislation Among the States

The 1996 California referendum known as Proposition 215 allowed seriously ill Californians to obtain and use marijuana for medical purposes without criminal prosecution or sanction. A physician's recommendation is needed. Under the law, physicians cannot be punished or denied any right or privilege for recommending marijuana to patients who suffer from any illness for which marijuana will provide relief.

The 1996 Arizona referendum known as Proposition 200 was largely about prison reform but also gave physicians the option to prescribe controlled substances, including those in Schedule I (e.g., marijuana), to treat the disease or relieve the suffering of seriously or terminally ill patients. Five months after the referendum was passed, it was stalled when Arizona legislators voted that all prescription medications must be approved by the Food and Drug Administration, and marijuana is not so approved. In November 1998, Arizona voters passed a second referendum designed to allow physician's to prescribe marijuana as medicine, but this is still at odds with federal law.8 .

As of summer 1998, eight states -- California,, Connecticut, Louisiana, New Hampshire, Ohio, Vermont, Virginia, and Wisconsin -- had laws that permit physicians to prescribe marijuana for medical purposes or to allow a medical necessity defense.8 In November 1998, five states -- Arizona, Alaska, Oregon, Nevada, and Washington -- passed medical marijuana ballot initiatives. The District of Columbia also voted on a medical marijuana initiative, but was barred from counting the votes because an amendment designed to prohibit them from doing so was added to the federal appropriations bill, however, exit polls suggested that a majority of voters had approved the measure.

1.7 ---------------------------------------------------------------------------

Marijuana and Medicine

Marijuana plants have been used since antiquity for both herbal medication and intoxication. The current debate over the medical use of marijuana is essentially a debate over the value of its medicinal properties relative to the risk posed by its use.

Marijuana's use as an herbal remedy before the 20th century is well documented.1, 10, 11 However, modern medicine adheres to different standards from those used in the past. The question is not whether marijuana can be used as an herbal remedy, but rather how this remedy meets today's standards of efficacy and safety. We understand much more than previous generations about medical risks. Our society generally expects its licensed medications to be safe, reliable, and of proven efficacy, and contaminants and inconsistent ingredients in our health treatments are not tolerated. That refers not only to prescription and over-the-counter drugs, but also to the vitamin supplements and herbal remedies purchased at the grocery store. For example, the essential amino acid l-tryptophan was widely sold in health food stores as a natural remedy for insomnia until early 1990 when it became linked to an epidemic of a new and potentially fatal illness (eosinophilia-myalgia syndrome).9, 12 When it was removed from the market shortly thereafter, there was little protest, despite the fact that it was safe for the vast majority of the population. The 1536 cases and 27 deaths were later traced to contaminants in a batch produced by a single Japanese manufacturer.

Although few herbal medicines meet today's standards, they have provided the foundation for modern Western pharmaceuticals. Most current prescriptions have their roots either directly or indirectly in plant remedies.7 At the same time, most current prescriptions are synthetic compounds that are only distantly related to the natural compounds that led to their development. Digitalis was discovered in foxglove, morphine in poppies, and taxol in the yew tree. Even aspirin (acetylsalicylic acid) has its counterpart in herbal medicine: for many generations, American Indians relieved headaches by chewing the bark of the willow tree, which is rich in a related form of salicylic acid.

Although plants continue to be valuable resources for medical advances, drug development is likely to be less and less reliant on plants and more reliant on the tools of modern science. Molecular biology, bioinformatics software, and DNA array-based analyses of genes and chemistry are all beginning to yield great advances in drug discovery and development. Until recently, drugs could only be discovered; now they can be designed. Even the discovery process has been accelerated through the use of modern drug screening techniques. It is increasingly possible to identify or isolate the chemical compounds in a plant, determine which compounds are responsible for the plant's effects, and select the most effective and safe compounds either for use as purified substances or as tools to develop even more effective, safer, or less expensive compounds.

1.8 ---------------------------------------------------------------------------

Yet even as the modern pharmacological toolbox becomes more sophisticated and biotechnology yields an ever greater abundance of therapeutic drugs, people increasingly seek alternative, low-technology therapies.4, 5 In 1997, 46 percent of Americans sought nontraditional medicines and spent over 27 billion unreimbursed dollars; the total number of visits to alternative medicine practitioners appears to have exceeded the number of visits to primary care physicians. 5, 6 Recent interest in the medical use of marijuana coincides with this trend toward self-help and a search for "natural" therapies. Indeed, several people who spoke at the IOM public hearings in support of the medical use of marijuana said that they generally preferred herbal medicines to standard pharmaceuticals. However, few alternative therapies have been carefully and systematically tested for safety and efficacy, as is required for FDA-approved medications.2

Who Uses Medical Marijuana?

There have been no comprehensive surveys of the demographics and medical conditions of medical marijuana users, but a few reports provide some indication. In each case, survey results should be understood to reflect the situation in which they were conducted and are not necessarily characteristic of medical marijuana users as a whole. Respondents to surveys reported to the IOM study team were all members of "buyers' clubs," organizations that provide their members with marijuana, although not necessarily through direct cash transactions. The atmosphere of the marijuana buyers' clubs ranges from that of the comparatively formal and closely regulated Oakland Cannabis Buyers' Cooperative to that of a "country club for the indigent," as Dennis Peron described the San Francisco Cannabis Cultivators Club (SFCCC), which he directed.

John Mendelson, an internist and pharmacologist at the University of California, San Francisco (UCSF) Pain Management Center, surveyed 100 members of the SFCCC who were using marijuana at least weekly. Most of the respondents were unemployed men in their forties. Subjects were paid $50 to participate in the survey; this might have encouraged a greater representation of unemployed subjects. All subjects were tested for drug use. About half tested positive for marijuana only; the other half tested positive for drugs in addition to marijuana (23% for cocaine and 13% for amphetamines). The predominant disorder was AIDS, followed by roughly equal numbers of members who reported chronic pain, mood disorders, and musculoskeletal disorders (table 1.1).

1.9 ---------------------------------------------------------------------------

Table 1.1 Self-Reported Disorders Treated with Marijuana by Members of San Francisco Cannabis Cultivators Club

Disorder Number of Subjects HIV 60 Musculoskeletal disorders and arthritis 39 Psychiatric disorders (primarily depression) 27 Neurological disorders and nonmusculoskeletal pain syndromes 9 Gastrointestinal disorders (most often nausea) 7 Other disorders glaucoma, allergies, nephrolitiasis, and the skin manifestations of Reiter syndrome 7 Total disorders 149 Total number of respondents 100

1.10 ---------------------------------------------------------------------------

The membership profile of the San Francisco club was similar to that of the Los Angeles Cannabis Resource Center (LACRC), where 83% of the 739 patients were men, 45% were 36-45 years old, and 71% were HIV-positive. Table 1.2 shows a distribution of conditions somewhat different from that in SFCCC respondents, probably because of a different membership profile. For example, cancer is generally a disease that occurs late in life; 34 (4.7%) of LACRC members were over 55 years old; only 2% of survey respondents in the SFCCC study were over 55 years old.

Jeffrey Jones, executive director of the Oakland Cannabis Buyers' Cooperative, reported that its largest group of patients is HIV-positive men in their forties. The second largest group is patients with chronic pain.

Among the 42 people who spoke at the public workshops or wrote to the study team, only six identified themselves as members of marijuana buyers' clubs. Nonetheless, they presented a similar profile: HIV - AIDS was the predominant disorder, followed by chronic pain (table 1.3). All HIV-AIDS patients reported that marijuana relieved nausea and vomiting and improved their appetite. About half the patients who reported using marijuana for chronic pain also reported that it reduced nausea and vomiting.

1.11 ---------------------------------------------------------------------------

Table 1.2 Self-Reported Disorders Treated with Marijuana by Members of Los Angeles Cannabis Resource Center (LACRC) According to Center Staffa

Treated Disorder Number of Percent of Subjects Subjects HIVb 528 71

Cancer 40 5.4 Terminal cancer 10 1.4 Mood disorders (depression) 4 .05 Musculosketetal (multiple sclerosis, arthritis) 30 4.1 Chronic pain and back pain 33 4.5 Gastrointestinal 17 2.3 Neurological disorders (epilepsy, Tourette syndrome, brain trauma)7 0.9 Seizures or migrainec 13 1.8

Glaucoma 15 2.0 Miscellaneous 42 5.7 Total number 739 100

a Results are based on review of 739 individual records by LACRC staff members. In contrast with Mendelson's survey of SFCCC (table 1.1), only the primary disorder is indicated here. Membership in LACRC is contingent on a doctor's letter of acknowledgment, but diagnoses are not independently confirmed.

b HIV patients use marijuana to control nausea, increase appetite (to combat wasting), and relieve gastrointestinal distress caused by AIDS medications. These uses are not indicated separately.

c As described by LACRC staff; some of these cases might also be neurological disorders.

d Because of rounding error, percentages do not add up to 100.

1.12 ---------------------------------------------------------------------------

Table 1.3 Summary of Reports to IOM Study Team by 43 Individuals

Symptoms Dominant Disease AIDS(7) AIDS and cancer Cancer Testicular cancer Anorexia, nausea, vomitingCancer and multiple sclerosclerosis Thyroid conditione Migraine Wilsons disease Depression(2) Depression and anxiety(2) Mood disorders Manic depression(2) Posttraumatic stress Premenstrual syndrome Migraine Injury(2) Epilepsy and Postpolio syndrome Trauma and epilepsy Degenerative disk disease Pain Rheumatoid arthritis Nail-patella syndrome Reflex sympathetic dystrophy Gulf War chemical exposure Multiple congenital cartilaginous exostosis Histiocytosis X spasticitye Multiple sclerosis(3) Muscle Spasticity Paralysis Spinal cord injury Spasmodic torticollis Intraocular pressure Glaucoma Diarrhea Crohn's disease

Table 1.3. This table lists the people who reported to the IOM study team during the public workshops, or through letters, that they use marijuana as medicine, it should not be interpreted as a representative sample of the full spectrum of people who use marijuana as medicine. Each dominant disease represents an individual report.

e Not specified.

1.13 ---------------------------------------------------------------------------

Table 1.4 Primary Symptoms of 43 Individuals who Reported to IOM Study Team

. Symptom Frequency Multiple Symptoms Number who % of those who Primary Number of % of Total Reported reported Symptom Reportsf Symptoms (Primary) Primary Reported Additional Symptoms Symptoms Anorexia, nausea, 21 31 13 62 vomiting Diarrhea 4 6 3 75 Intraocular pressure 2 3 1 50 Mood disorders 12 18 7 58 Muscle spasticity 12 18 7 58 Pain 16 24 13 81 Totals 67 . 44 66

f Forty-three persons reporting; 20 reported relief from more than one symptom.

1.14 ---------------------------------------------------------------------------

Note that the medical conditions referred to are only those reported to the study team or to interviewers; they cannot be assumed to represent complete or accurate diagnoses. Michael Rowbotham, a neurologist at the UCSF Pain Management Center, noted that many pain patients referred to that center arrive with incorrect diagnoses or with pain of unknown origin. At that center, the patients who report medical benefit from marijuana report that marijuana does not reduce their pain but enables them to cope with it.

Most -- not all -- people who use marijuana to relieve medical conditions have previously used it recreationally. An estimated 95% of the LACRC members had used marijuana before joining the club. It is important to emphasize the absence of comprehensive information on marijuana use before its use for medical conditions. Frequency of prior use almost certainly depends on many factors, including membership in a buyers' club, membership in a population sector that uses marijuana more often than others (for example, men 20-30 years old), and the medical condition being treated with marijuana (for example, there are probably relatively fewer recreational marijuana users among cancer patients than among AIDS patients).

Patients who reported their experience with marijuana at the public workshops said that marijuana provided them with great relief from symptoms associated with disparate diseases and ailments, including AIDS wasting, spasticity from multiple sclerosis, depression, chronic pain, and nausea associated with chemotherapy. Their circumstances and symptoms were varied, and the IOM study team was not in a position to make medical evaluations or confirm diagnoses. Three representative cases presented to the IOM study team are presented here; the stories have been edited for brevity, but each case is presented in the patient's words and with the patient's permission.

1.15 ---------------------------------------------------------------------------

Boxes 1.1-1.3: Selected Cases From the Public Sessions

G.S. spoke at the IOM workshop in Louisiana about his use of marijuana first to combat AIDS wasting syndrome and later for relief from the side effects of AIDS medications.

Skin rashes, dry mouth, foul metallic aftertaste, numbness of the face, swelling of the limbs, fever spikes, headaches, dizziness, anemia, clinical depression, neuropathy so crippling that I could not type, so painful that the bed sheets felt like sandpaper, nausea so severe that I sometimes had to leave the dinner table to vomit, and diarrhea so unpredictable that I dared not leave the house without diapers.

These are some of the horrors that I endured in the last 10 years during my fight for life against the human immunodeficiency virus. But these ravages were not caused by HIV itself, or by any of the opportunistic infections that mark the steady progression of AIDS. Each of these nightmares was a side effect of one of the hundreds of medications I have taken to fight one infection after another on my way to a seemingly certain early grave.

Had you known me three years ago, you would not recognize me now. After years of final-stage AIDS, I had wasted to 130 lb. The purple Kaposi's sarcoma lesions were spreading. The dark circles under my eyes told of sleepless nights and half-waking days. I encountered passages of time marked by medication schedules, nausea, and diarrhea. I knew that I was dying. Every reflection shimmered with death, my ghost-like pallor in the mirror, the contained terror on the face of a bus passenger beside me, and most of all, the resigned sadness in my mother's eyes.

But still I was fortunate because along the way I rediscovered the ancient understanding of marijuana's medicinal benefit. So I smoked pot. Every day. The pot calmed my stomach against handfuls of pills. The pot made me hungry so that I could eat without a tube. The pot eased the pain of crippling neural side effects so that I could dial the phone by myself. The pot calmed my soul and allowed me to accept that I would probably die soon. Because I smoked pot I lived long enough to see the development of the first truly effective HIV therapies. I lived to gain 50 lb, regain my vigor, and celebrate my 35th birthday. I lived to sit on the bus without frightening the passenger beside me.

Even at this stage of my recovery, I take a handful of pills almost every day, and will probably continue to do so for the rest of my life. While I am grateful for the lifesaving pro/ease-inhibitor therapies, they bring with them a host of adverse reactions and undesirable side effects. Different patients experience different reactions, of course, but almost all patients experience some. Smoking marijuana relieves many of these side effects.

I am not one of the exceptional eight patients in the United States with legal permission to smoke marijuana. Every day I risk arrest, property forfeiture, fines, and imprisonment. But I have no choice, you see, just as I have no choice but to endure the side effects of these toxic medications. So many patients like me are breaking the law to enjoy relief that no other therapy provides.

I sit here, I believe, as living proof that marijuana can have a beneficial effect in staving off wasting. Every pound was a day. I figured that for every pound of body weight I could maintain, that was another day that I could live in hopes that some effective therapy would emerge.

1.16 ---------------------------------------------------------------------------

B.D. spoke at the IOM workshop in Louisiana She is one of eight patients who are legally allowed to smoke marijuana under a Compassionate Use Protocol She uses marijuana to relieve nausea, muscle spasticity, and pain associated with multiple sclerosis.

I was diagnosed with multiple sclerosis in 1988. Prior to that, I was an active person with ballet and swimming. I now have a swimming pool, so I swim each and every day, and I smoke marijuana. The government has given me the marijuana to smoke. Each month, I pick up a can filled with the marijuana cigarettes rolled by the government.

At one time, I weighed 85 lb and I now weigh 105. Twenty pounds is quite a bit to put on I could not walk. I did not have the appetite. I use a scooter now for distance. I can get around the house. I have a standard poodle who is kind of like an assistant dog. She is good at it. She helps me.

When I found out that there was a program to get marijuana from the government, I decided that was the answer. I was not a marijuana-smoker before that. In fact, I used to consider the people I knew who smoked the marijuana as undesirables. Now, I myself am an undesirable.

But it works. It takes away the backache. With multiple sclerosis, you can get spasms, and your leg will just go straight out and you cannot stop that leg. You may have danced all of your life and put the leg where you wanted it to be, but the MS takes that from you. So I use the swimming pool, and that helps a lot. The kicks are much less when I have smoked a marijuana cigarette. Since 1991, I've smoked 10 cigarettes a day. I do not take any other drugs. Marijuana seems to have been my helper. At one time, I did not think much of the people who smoke it. But when it comes to your health, it makes a big difference.

1.17 ---------------------------------------------------------------------------

J.H.spoke at the IOM workshop in Washington, DC. He was was seriously injured in an accident, suffers from a form of arthritis associated with abnormal activity of the sympathetic nervous system known as reflex sympathetic dystrophy, and has hepatitis C. He uses marijuana to relieve nausea from liver disease, pain, and muscle spasms.

I am 48 years old, married with two children. I am a veteran who served during the Vietnam War. I was exposed to hepatitis C in 1972 by a blood transfusion, which I needed because of a motor-vehicle accident that broke my back, ruined my right shoulder, my left thumb, and hand, and almost amputated my right leg at the knee. My hepatitis C was not diagnosed until 1997 -- after the disease had destroyed my pancreasg and I had four heart attacks, one angioplasty, and a minor stroke. In 1989, while at work, I was involved in an accident with a large soil-survey auger. My pelvis was crushed, and serious nerve damage was the result. I also have reflex sympathetic dystrophy, which is a neurological disease that has a tremendous amount of pain and muscle spasms.

I have reached what the doctors call end-stage liver disease from the hepatitis C. I have lost 85 lbs due to the severe bouts of nausea and vomiting. Every time I come home from a hospital stay, my 7-year-old asks if I got the liver transplant. I am on a transplant list, but I am not a candidate until I am seven days from death.

In October 1997, after trying four different antinausea medications, four of the doctors that I see told me to go to Europe and see a doctor and try medicinal cannabis. My primary-care doctor wrote me a letter to carry with my medical records asking that the doctor help me in any way that he can to alleviate the symptoms of the hepatitis C and the reflex sympathetic dystrophy. Those symptoms are severe nausea and pain from the hepatitis C and pain and muscle spasms from the neurological disease.

I went to Europe in November 1997, where I saw a doctor of internal medicine. He prescribed me cannabis, 1-2 g a day. I got the medicine and a pipe and tried it. Within two minutes of taking two puffs from the pipe, the nausea was gone. I don't think that I felt the high, although I was quite elated. In about 45 min., I was starving. Normally, I have a fear of eating because I vomit almost always after I eat or take a pill. I forgot about that, and I think I ate more that night than I had eaten in months. I did feel a little nauseated after about four hours, but I smoked two more puffs, and in about two hours I went to bed. The next morning, I felt hungry. During my nine-day stay in Europe, I was able to stay free of vomiting and the waves of nausea became less frequent.

I had experienced four years of pain control using Tegretol®, a drug used by epileptics to control seizures. Now, I can't use that medication because of the damage that it causes to my cirrhotic liver. When I smoked about two grams of marijuana a day, the nausea was gone and I was no longer losing weight. The pain was at an acceptable level. Sometimes, I still find it necessary to use an opiate pain-killer, but only when the pain is at its worst. Surprisingly, I lost an associated high within a few days. I also think the cannabis has an antidepressant effect on me, as I no longer have what I call the "poor me" feelings that I experienced after learning about the hepatitis C.

---------------------------------------------------------------------------

g This is an unlikely consequence of hepatitis C; it is more likely that the patient's liver was damaged.

1.18 ---------------------------------------------------------------------------

The variety of stories presented left the study team with a clear view of people's beliefs about how marijuana had helped them. But this collection of anecdotal data, although useful, is limited. We heard many positive stories, but no stories from people who had tried marijuana but found it ineffective. This is a fraction with an unknown denominator. For the numerator, we have a sample of positive responses for the denominator, we have no idea of the total number of people who have tried marijuana for medical purposes. Hence, it is impossible to estimate the clinical value of marijuana or cannabinoids in the general population based on anecdotal reports. Marijuana clearly seems to relieve some symptoms for some people even if only as a placebo effect. But what is the balance of harmful and beneficial effects? That is the essential medical question that can be answered only by careful analysis of data collected under controlled conditions.

Cannabis and the cannabinoids

Marijuana is the common name for Cannabis saliva, a hemp plant that grows throughout temperate and tropical climates. The most recent review of the constituents of marijuana lists 66 cannabinoids (table 1.5).16 But that does not mean that there are 66 different cannabinoid effects or interactions. Most of the cannabinoids are closely related; they fall into only 10 groups of closely related cannabinoids, many of which differ by only a single chemical moiety and might be midpoints along biochemical pathways -- that is, degradation products, precursors, or byproducts.16 18 [Image]9-tetrahydrocannabinol ([Image]9-THC) is the primary psychoactive ingredient; depending on the particular plant, either THC or cannabidiol is the most abundant cannabinoid in marijuana (figure 1.1) Throughout this report, THC is used to indicate [Image]9-THC. In the few cases where variants of THC are discussed, the full names are used. All the cannabinoids are lipophilic -- they are highly soluble in fatty fluids and tissues but not in water. Indeed, THC is so lipophilic that it is aptly described as "greasy".

Throughout this report, marijuana refers to unpurified plant extracts, including leaves and flower tops, regardless of how they are consumed -- whether by ingestion or by smoking. References to the effects of marijuana should be understood to include the composite effects of its various components; that is, the effects of THC are included among the effects of marijuana; but not all the effects of marijuana are necessarily due to THC. Discussions concerning cannabinoids refer only to those particular compounds and not to the plant extract. This distinction is important; it is often blurred or exaggerated.

cannabinoids are produced in epidermal glands on the leaves (especially the upper ones), stems, and the bracts that support the flowers of the marijuana plant. Although the flower itself has no epidermal glands, it has the highest cannabinoid content anywhere on the plant, probably because of the accumulation of resin

1.19 ---------------------------------------------------------------------------

secreted by the supporting bracteole (the small, leaf-like part below the flower). The amounts of cannabinoids and their relative abundance in a marijuana plant vary with growing conditions, including humidity, temperature, and soil nutrients (reviewed in Pate 199414). The chemical stability of cannabinoids in harvested plant material is also affected by moisture, temperature, sunlight, and storage. They degrade under any storage condition.

1.20 ---------------------------------------------------------------------------

Figure 1.1 Cannabinoid Biosynthesis

Cannabinoid Biosynthesis

Figure legend: Arrows indicate cannabinoid biosynthesis pathway; dark arrows indicate established pathways, the light gray arrow indicates a probable, but not well established, pathway (R. Mechoulam, personal communication, 1999).17 The great majority of studies reporting on effects of cannabinoids refer to THC; most of the rest are about CBD and CBN. Other cannabinoids found in marijuana do not appear to play an important role in effects of marijuana.

1.21 ---------------------------------------------------------------------------

Table 1.5 cannabinoids Identified in Marijuana

66 Cannabinoids Identified in Marijuana Number of cannabinoid Cannabinoid Group Common Variants known in each Abbreviation group [Image]9-tetrahydrocannabinol[Image]9-THC 9

[Image]8-Tetrahydrocannabinol[Image]8-THC 2

Cannabichromene CBC 5 Cannabicyclol CBL 3 Cannabidiol CBD 7 Cannabielsoin CBE 5 Cannabigerol CBG 6 Cannabinidiol CBND 2 Cannabinol CBN 7 Cannabitriol CBT 9 Miscellaneous types . 11

1.22 ---------------------------------------------------------------------------

Organization of the Report

Throughout the report, steps that might be taken to fill the gaps in understanding both the potential harms and benefits of marijuana and cannabinoid use will be identified. Those steps include identifying knowledge gaps, promising research directions, and potential therapies based on scientific advances in cannabinoid biology.

Chapter 2 reviews basic cannabinoid biology and provides a foundation to understand the medical value of marijuana or its constituent cannabinoids. In consideration of the physician's first rule, "first, do no harm," the potential harms attributed to the medical use of marijuana are reviewed before the potential medical benefits, chapter 3 reviews the risks posed by marijuana use, with emphasis on medical use.

Chapter 4 analyzes the most credible clinical data relevant to the medical use of marijuana. It reviews what is known about the physiological mechanisms underlying particular conditions (for example, chronic pain, vomiting, anorexia, and muscle spasticity), what is known about the cellular actions of cannabinoids, and the levels of proof needed to show that marijuana is an effective treatment for specific symptoms. It does not analyze the historical literature; history is informative in enumerating uses of marijuana, but it does not provide the sort of information needed for a scientifically sound evaluation of the efficacy and safety of marijuana for clinical use. Because marijuana is advocated primarily as affording relief from the symptoms of disease rather than as a cure, this chapter is organized largely by symptoms as opposed to disease categories. Finally, chapter 4 compares the conclusions of this report with those of other recent reports on the medical use of marijuana.

Chapter 5 describes the process of and analyzes the prospects for cannabinoid drug development.

1.23 ---------------------------------------------------------------------------

References

1. Abel EL. 1980 Marijuana: the first twelve thousand years. New York: Plenum Press.

2. Angell M, Kassirer JP. 1998. Alternative Medicine - The Risks of Untested and Unregulated Remedies. The New England Journal of Medicine 339:839-841.

3. Bonnie RJ, Whitebread II CH. 1974. The marihuana conviction: A history of marihuana prohibition in the United States. Charlottesville, VA: University Press of Virginia.

4. Eisenberg DM. 1997. Advising patients who seek alternative medical therapies. Annals of Internal Medicine 127:61-69.

5. Eisenberg DM, Davis RB, Ettner SL, Appel S. Wilkey S. Van Rompay M, Kessler RC. 1998. Trends in Alternative Medicine Use in the United States, 1990-1997: Results of a Follow-up National Survey. Journal of the American Medical Association 280:1569-1575

6. Eisenberg DM, Kessler RC, Foster C, Norlock FE, Calkins DR, Delbanco TL. 1993. Unconventional medicine in the United States: Prevalence, costs, and patterns of use. New England Journal of Medicine 328:246-252

7. Grifo F. Newman D, Fairfield A, Bhattacharya B. Grupenhoff JT 1997. The Origins of Prescription Drugs. In: Grifo F. Rosenthal J Editors Biodiversity and Human Health. Washington, D.C.: Island Press. Pp. 131-163.

8. Herstek J. 1998. Behavioral Health Issue Briefs. Medical Marijuana Washington, D.C.: Health Policy Tracking Service, National Conference of State Legislatures.

9. Kilbourne EM, Philen R M, Kamb M L, Falk H. 1996. Tryptophan produced by Showa Denko and epidemic eosinophilia-myalgia syndrome. Journal of Rheumatology Supplement 46:81-88. Comment on: Journal of Rheumatology Supplement 1996 46: 44-58 and 60-72; discussion 58-59.

10. Mathre ML, Editor. 1997. Cannabis in Medical Practice. Jefferson, North Carolina: MacFarland and Company, Inc.

11. Mechoulam R. 1986. The pharmacohistory of Cannabis Sativa. In: Mechoulam R Editor cannabinoids as therapeutic agents. Boca Raton, FL: CRC Press, Inc. Pp. 1-19.

12. Milburn DS,, Myers CW. 1991. Tryptophan toxicity: a pharmacoepidemiologic review of eosinophilia-myalgia syndrome. DICP 25: 1259- 1262.

13. NORML. The Medical Use of http://norml.org/medical/index.html (accessed July 9, 1998). Marijuana [WWW Document].URL

14. Pate DW.. 1994. Chemical ecology of cannabis. Journal of the International Hemp Association 1 :29,32-37.

15. Peterson K. 15 January 1997. Notes: Weighing in on a medical controversy; USA Today's Baby Boomer Panel. USA Today, sec. Life, p. 121).

1.24 ---------------------------------------------------------------------------

16. Ross SA, Elsohly MA. 1995. Constituents of Cannabis Sativa L. XXVIII a review of the natural constituents: 1980-1994. Zagazig Journal for Pharmaceutical Sciences 4:1-10.

17. Taura F. Morimoto S. Shoyama Y. 1995. First direct evidence for the mechanism of delta1-tetrahydrocannabinolic acid biosysnthesis. Journal of the American Chemical Society 117:9766-9767.

18. Turner CE, Elsohly MA, Boeren E.G. 1980. Constituents of Cannabis Sativa L. XVII. A review of the natural constituents. Journal of Natural Products 43: 169-234.

1.25 ---------------------------------------------------------------------------

Chapter 2 ---->>>>

CANNABINOIDS AND ANIMAL PHYSIOLOGY

CHAPTER 2. CANNABINOIDS AND ANIMAL PHYSIOLOGY 2 Introduction 2 Cannabinoid Receptors 9 The Endogenous Cannabinoid System 13 Sites of Action 21 Cannabinoid Receptors and Brain Functions25 Chronic Effects of THC 31 Cannabinoids and the Immune System 34 Conclusions and Recommendations 43 References 46

2.1 ---------------------------------------------------------------------------

Chapter 2. Cannabinoids and Animal Physiology

Introduction

Much has been learned since the publication of the 1982 IOM report on Marijuana and Health.a Although it was clear then that most of the effects of marijuana were due to its actions on the brain, there was little information about how THC acted on brain cells (neurons), which cells were affected by THC, or even what general areas of the brain were most affected by THC. Too little was known about cannabinoid physiology to offer any scientific insights into the harmful or therapeutic effects of marijuana. That is no longer true. During the last 16 years, there have been major advances in what basic science discloses about the potential medical benefits of cannabinoids, the group of compounds related to THC. Many variants are found in the marijuana plant, and other cannabinoids not found in the plant have been chemically synthesized. Sixteen years ago, it was still a matter of debate as to whether THC acted nonspecifically by affecting the fluidity of cell membranes or whether a specific pathway of action was mediated by a receptor that responded selectively to THC (table 2.1).

Basic science is the wellspring for developing new medications and is particularly important for understanding a drug that has as many effects as marijuana. Even committed advocates of the medical use of marijuana do not claim that all the effects of marijuana are desirable for every medical use. But they do claim that the combination of specific effects of marijuana enhances its medical value. An understanding of those specific effects is what basic science can provide. The multiple effects of marijuana can be singled out and studied with the goals of evaluating the medical value of marijuana and cannabinoids in specific medical conditions, as well as minimizing unwanted side effects. An understanding of the basic mechanisms through which cannabinoids affect physiology permits more strategic development of new drugs and designs for clinical trials that are most likely to yield conclusive results.

Research on cannabinoid biology offers new insights into clinical use, especially given the scarcity of clinical studies that adequately evaluate the medical value of marijuana. For example, despite the scarcity of substantive clinical data, basic science has made it clear that cannabinoids can affect pain transmission and specifically that, cannabinoids interact with the brain's endogenous opioid system, an important system for the medical treatment of pain (see chapter 4). --------------------------------------------------------------------------- a The field of neuroscience has grown substantially since the publication of the 1982 IOM report. The number of members in the Society for Neuroscience provide a rough measure of the growth in research and knowledge about the brain: as of the middle of 1998, there are over 27,000 members, more than triple the number in 1982.

2.2 ---------------------------------------------------------------------------

The cellular machinery that underlies the response of the body and brain to cannabinoids involves an intricate interplay of different systems. This chapter reviews the components of that machinery with enough detail to permit the reader to compare what is known about basic biology with specific indications proposed for marijuana. For some readers' that will be too much detail. Those readers who do not wish to read the entire chapter should, nonetheless, be mindful of the following key points in this chapter

· The most far-reaching of the recent advances in cannabinoid biology are the identification of two types of cannabinoid receptors (CB1 and CB2) and of anandamide, a substance naturally produced by the body that acts at the cannabinoid receptor, and has effects similar to those of THC. The CB1 receptor is found primarily in the brain, and mediates the psychological effects of THC. The CB2 receptor is associated with the immune system, its role remains unclear.

· The physiological roles of the brain cannabinoid system in humans are the subject of much active research, and not fully known; however, cannabinoids likely have a natural role in pain modulation, control of movement, and memory.

· Animal research has shown that the potential for cannabinoid dependence exists, and cannabinoid withdrawal symptoms can be observed. However, both appear to be mild compared to dependence and withdrawal seen with other drugs.

· Basic research in cannabinoid biology has revealed a variety of cellular pathways through which potentially therapeutic drugs could act on the cannabinoid system. In addition to the known cannabinoids, such drugs might include chemical derivatives of plant-derived cannabinoids or of endogenous cannabinoids such as anandamide, but would also include non-cannabinoid drugs that act on the cannabinoid system.

This chapter summarizes the basics of cannabinoid biology - as known today. It thus provides a scientific basis for interpreting claims founded on anecdotes and for evaluating the clinical studies of marijuana presented in chapter 4.

2.3 --------------------------------------------------------------------------- Table 2.1 Landmark Discoveries Since the 1982 IOM Report

Since the Previous IOM Report on Marijuana in 1982: A Decade of Landmark Discoveries Year Discovery Primary Investigators Potent cannabinoid agonists are M.R. Johnson and L.S. 1986developed the key to discovering the Melvin75 receptor

1988First conclusive evidence of specific A. Howlett and W. Devane36 cannabinoid receptors The cannabinoid brain receptor (CB1) L. Matsuda et al,107 and M. 1990is cloned, its DNA sequence is identified, and its location in the Herkenham et al60 brain is determined Anandamide is discovered - a naturally 1992occurring substance in the brain that R. Mechoulam and W. Devane37 acts on cannabinoid receptors A cannabinoid receptor is discovered outside the brain; this receptor (CB2) 1993 S. Munro112 is related to the brain receptor but is distinct

1994The first specific cannabinoid M.Rinaldi-Carmona132 antagonist, SR141716A, is developed The first cannabinoid antagonist, 1998SR144528, that can distinguish between M. Rinaldi-Carmona133 CB1 and CB2 receptors discovered.

2.4 ---------------------------------------------------------------------------

The Value of Animal Studies

Much of the research into the effects of cannabinoids on the brain is based on animal studies. Many speakers in the public workshops associated with this study argued that animal studies of marijuana are not relevant to humans. While animal studies are no substitute for clinical trials, they are a necessary complement. Ultimately, every biologically active substance exerts its effects at the cellular and molecular level, and at this level, the evidence has shown remarkable consistency among mammals, even those as different in body and mind as rats and humans. Animal studies typically provide information about how drugs work that would not be obtainable in clinical studies. At the same time, animal studies can never completely inform us about the full range of psychological and physiological effects of marijuana or cannabinoids on humans.

The Active Constituents of Marijuana

[Image]9-THC and [Image]8-THC are the only compounds in the marijuana plant that show all the psychoactive effects of marijuana. Because [Image]9-THC is much more abundant than [Image]8-THC, the psychoactivity of marijuana has been largely attributed to the effects of [Image]9-THC 11-OH-[Image]9-THC is the primary product of [Image]9-THC metabolism by the liver and is about three times as potent as [Image]9-THC.128

There have been considerably fewer experiments with cannabinoids other than [Image]9-THC although a few studies have been done to examine whether other cannabinoids modulate the effects of THC or mediate the non-psychological effects of marijuana. Cannabidiol (CBD) does not have the same psychoactivity as THC, but it was initially reported to attenuate the psychological response to THC in humans 81, 177 however, later studies reported that CBD did not attenuate the psychological effects of THC.11, 69 One double-blind study of eight volunteers reported that CBD can block the anxiety induced by high doses of THC (0.5 mg/kg).177 There are numerous anecdotal reports claiming that marijuana with relatively higher ratios of THC:CBD is less likely to induce anxiety in the user than marijuana with low THC:CBD ratios; but, taken together, the results published thus far are inconclusive.

The most significant effects of cannabidiol (CBD) seem to be its interference with drug metabolism in the liver, including [Image]9-THC metabolism.14, 114 CBD exerts this effect by inactivating cytochrome P450s, which are the most important class of enzymes that metabolize drugs. Like many P450 inactivators, CBD can also induce P450s after repeated doses.13 Experiments in which mice were treated with CBD followed by THC showed that CBD treatment was associated with a significant

2.5 ---------------------------------------------------------------------------

increase in brain levels of THC and its major metabolites, most likely because of its effects on decreasing the clearance rate of THC from the body 15

In mice, THC inhibits the release of luteinizing hormone (LH), the pituitary hormone that triggers the release of testosterone from the testis in males, this effect is increased when THC is given together with cannabinol or CBD.113

Cannabinol is considerably less active than THC in the brain, but studies of immune cells have shown that it can modulate immune function (see section on Cannabinoids and the Immune System). In mice, cannabinol lowers body temperature and increases sleep duration.175

The Pharmacological Toolbox

A researcher needs certain key tools in order to understand how a drug acts on the brain. To appreciate the importance of these tools, one must first understand some basic principles of drug action. All recent studies have indicated that the behavioral effects of THC are receptor-mediated.27 Neurons in the brain are activated when a compound binds to its receptor, which is a protein typically located on the cell surface. Thus, THC will exert its effects only after binding to its receptor. In general, a given receptor will accept only particular classes of compounds and will be unaffected by other compounds.

Compounds that activate receptors are called agonists. Binding to a receptor triggers an event or a series of events in the cell that results in a change in the cell's activity, its gene regulation, or the signals that it sends to neighboring cells (figure 2. 1). This agonist-induced process is called signal transduction.

Another tool for drug research, which only recently became available for cannabinoid research, are the receptor antagonists, so-called because they selectively bind to a receptor that would have otherwise been available for binding to some other compound or drug. Antagonists block the effects of agonists and are tools to identify receptor functions by showing what happens when a receptor's normal functions are blocked. Agonists and antagonists are both ligands; that is, they bind to receptors. Hormones, neurotransmitters, and drugs can all act as ligands. Morphine and naloxone provide a good example. A large dose of morphine acts as an agonist at opioid receptors in the brain and interferes with, or even arrests, breathing. Naloxone, a powerful opioid antagonist, blocks morphine's effects on opiate receptors, thereby allowing an overdose victim to resume breathing normally. Naloxone itself has no effect on breathing.

Another key tool involves identifying the receptor protein and determining how it works. That makes it possible to locate where a drug activates its receptor in the brain -both the general region of the brain and the cell type where the receptor is. The way to find a receptor for a drug in the brain is to make the receptor "visible" by attaching a radioactive or fluorescent marker to the drug so that it can be detected.

2.6 ---------------------------------------------------------------------------

Such markers show where in the brain it binds to the receptor, but this is not necessarily the part of the brain where the drug ultimately has its greatest effects.

Because drugs injected into animals must be dissolved in a water-based solution, it is easier to deliver water-soluble molecules than to deliver fat-soluble (lipophilic) molecules such as THC. THC is so lipophilic that it can stick to glass and plastic syringes used for injection. Because it is lipophilic, it readily enters cell membranes and thus can cross the blood brain barrier easily. (This barrier insulates the brain from many blood-borne substances.) Early cannabinoid research was hindered by the lack of potent cannabinoid ligands (THC binds to its cannabinoid receptors rather weakly) and because they were not readily water-soluble. The synthetic agonist, CP 55,940, which is more water-soluble than THC, became the first useful research tool for studying cannabinoid receptors because of its high potency, and the ability to label it with a radioactive molecule, which enabled researchers to trace its activity.

2.7 ---------------------------------------------------------------------------

Figure 2.1 Diagram of neuron with synapse

[Image]

How receptors work: Individual nerve cells, or neurons, both send and receive cellular signals to and from neighboring neurons, but for the purposes of this diagram only one activity is indicated for each cell. Neurotransmitter molecules (shown as black dots) are released from the neuron terminal and move across the gap between the "sending" and "receiving" neurons. A signal is transmitted to the receiving neuron when the neurotransmitters has bound to the receptor on its surface. The effects of a transmitted signal include:

·Changing the cell's permeability to ions such as calcium and potassium. ·Turning a particular gene on or off. ·Sending a signal to another neuron. ·Increasing or decreasing the responsiveness of the cell to other cellular signals.

Those effects can lead to cognitive, behavioral, or physiological changes, depending on which neuronal system is activated.

The expanded view of the synapse illustrates a variety of ligands, that is, molecules that bind to receptors. Anandamide is a substance produced by the body that binds to and activates cannabinoid receptors; it is an endogenous agonist. THC can also bind to and activate cannabinoid receptors, but is not naturally found in the body; it is an exogenous agonist. SR141617A binds to, but does not activate cannabinoid receptors. In this way, it prevents agonists, such as anandamide and THC, from activating cannabinoid receptors by binding to the receptors without activating them; SR141617A is an antagonist, but it is not normally produced in the body. Endogenous antagonists, that is, those normally produced in the body, might also exist, but none have been identified.

2.8 ---------------------------------------------------------------------------

Cannabinoid Receptors

The cannabinoid receptor is a typical member of the largest known family of receptors: the G-protein-coupled receptors with their distinctive pattern in which the receptor molecule spans the cell membrane seven times (figure 2.2). For excellent recent reviews of cannabinoid receptor biology, see Childers and Breivogel,27 Abood and Martin,1 Felder and Glass,43 and Pertwee.124 Cannabinoid receptor ligands bind reversibly (they bind to the receptor briefly and then dissociate) and stereoselectively (when there are molecules that are mirror images of each other, only one version activates the receptor). Thus far, two cannabinoid receptor subtypes (CB1 and CB2) have been identified, of which only CB1 is found in the brain.

The cell responds in a variety of ways when a ligand binds to the cannabinoid receptor (figure 2.3). The first step is activation of G-proteins, the first components of the signal-transduction pathway. That leads to changes in several intercellular components - such as cyclic AMP and calcium and potassium ions - which ultimately produce the changes in cell functions. The final result of cannabinoid receptor stimulation depends on the particular type of cell, the particular ligand, and the other molecules t hat might be competing for receptor binding sites. Different agonists vary in binding potency, which determines the effective dose of the drug, and efficacy, which determines the maximal strength of the signal that they transmit to the cell. The potency and efficacy of THC are both relatively lower than those of some synthetic cannabinoids; in fact, synthetic compounds are generally more potent and efficacious than endogenous agonists.

CB1 receptors are extraordinarily abundant in the brain. They are more abundant than most other G-protein-coupled receptors and ten times more abundant than mu opioid receptors, the receptors responsible for the effects of morphine.148

2.9 ---------------------------------------------------------------------------

Figure 2.2 Cannabinoid receptors

[Image]

Receptors are proteins, and proteins are made up of strings of amino acids. Each circle in the diagram represents one amino acid. The shaded bar represents the cell membrane, which like all cell membranes in animals, is largely composed of phospholipids. Like many receptors, the cannabinoid receptors span the cell membrane; some sections of the receptor protein are outside the cell membrane (extrucellular), some are inside (intracellular). THC, anandamide, and other known cannabinoid receptor agonists bind to the extracellular portion of the receptor, thereby activating the signal pathway inside the cell.

The CB1 molecule is larger than CB2. The receptor molecules are most similar in four of the seven regions where they are embedded in the cell membrane (known as the transmembrane regions). The intracellular loops of the two cannabinoid receptor sub-types are quite different, which might affect the cellular response to the ligand, because these loops are known to mediate G-protein signaling - that is, the next step in the cell signaling pathway after the receptor. Receptor homology between the two receptor sub-types is 44 percent for the full length protein, and 68 percent within the seven transmembrane regions. The ligand binding sites are typically defined by the extracellular loops and the transmembrane regions.

2.10 ---------------------------------------------------------------------------

Figure 2.3 How cannabinoids affected neuron signals

Figure legend. Intracellular events that happen when cannabinoid agonists bind to receptors. Cannabinoid receptors are embedded in the cell membrane where they are coupled to G-proteins (G) and the enzyme, adenylyl cyclase (AC). Receptors are activated when they bind with ligands such as anandamide or THC in this case. This triggers a variety of reactions including inhibition ((-)) of AC which decreases the production of cAMP and cellular activities dependent on cAMP, opening potassium (K+) channels which decreases cell firing, and closing calcium (Ca2+) channels which decreases the release of neurotransmitters. These changes can influence cellular communication.

[Image]

2.11 ---------------------------------------------------------------------------

The cannabinoid receptor in the brain is a protein referred to as CB1. The peripheral receptor (outside the nervous system), CB2, is most abundant on cells of the immune system and is not generally found in the brain.43, 124 Although no other receptor subtypes have been identified, there is a genetic variant known as CB1A (such variants are somewhat different proteins that have been produced by the same genes via alternative processing). In some cases, proteins produced via alternative splicing have different effects on cells. It is not yet known whether there are any functional differences between the two, but the structural differences raise the possibility.

CB1 and CB2 are similar, but not as similar as members of many other receptor families are to each other. On the basis of a comparison of the sequence of amino acids that make up the receptor protein, the similarity of the CB1 and CB2 receptors is 44 percent (figure 2.2). The differences between the two receptors indicate that it should be possible to design therapeutic drugs that would act only on one or the other receptor and thus would activate or attenuate (block) the appropriate cannabinoid receptors. This offers a powerful method for producing biologically selective effects. In spite of the difference between the receptor subtypes, most cannabinoid compounds bind with similar affinityb to both CB1 and CB2 receptors. One exception is the plant-derived compound, cannabinol, which shows greater binding affinity for CB2 than for CB1,112 although another research group has failed to substantiate that observation.129 Other exceptions include the synthetic compound, WIN 55,212-2, which shows greater affinity for CB2 than CB1, and the endogenous ligands, anandamide and 2-AG, which show greater affinity for CB1 than CB2.43 The search for compounds that bind to only one or the other of the cannabinoid receptor types has been under way for several years and has yielded a number of compounds that are useful research tools and have potential for medical use.

Cannabinoid receptors have been studied most in vertebrates, such as rats and mice. However, they are also found in invertebrates, such as leeches and mollusks.156 The evolutionary history of vertebrates and invertebrates diverged more than 500 million years ago, so cannabinoid receptors appear to have been conserved throughout evolution at least this long. This suggests that they serve an important and basic function in animal physiology. In general, cannabinoid receptor molecules are similar among different species.124 Thus, cannabinoid receptors likely fill many similar functions in a broad range of animals, including humans. --------------------------------------------------------------------------- b Affinity is a measure of how avidly a drug binds to a receptor. The higher the affinity of a drug, the higher its potency; that is, lower doses are needed to produce its effects.

2.12 ---------------------------------------------------------------------------

The Endogenous Cannabinoid System

For any drug for which there is a receptor, the logical question is, "Why does this receptor exist?" The short answer is that there is probably an endogenous agonist (that is, a compound that is naturally produced in the brain) that acts on that receptor. The long answer begins with a search for such compounds in the area of the body that produce the receptors and ends with a determination of the natural function of those compounds. So far, the search has yielded several endogenous compounds that bind selectively to cannabinoid receptors. The best studied of them are anandamide37 and arachidonyl glycerol (2-AG).108 However, their physiological roles are not yet known.

Initially, the search for an endogenous cannabinoid was based on the premise that its chemical structure would be similar to that of THC; that was reasonable, in that it was really a search for another "key" that would fit into the cannabinoid receptor "keyhole," thereby activating the cellular message system. One of the intriguing discoveries in cannabinoid biology was how chemically different THC and anandamide are. A similar search for endogenous opioids (endorphins) also revealed that their chemical structure is very different from the plant-derived opioids, opium and morphine.

Further research has uncovered a variety of compounds with quite different chemical structures that can activate cannabinoid receptors (table 2.2 and figure 2.4) It is not yet known exactly how anandamide and THC bind to cannabinoid receptors. Knowing this should permit more precise design of drugs that selectively activate the endogenous cannabinoid systems.

Anandamide

The first endogenous cannabinoid to be discovered was arachidonylethanolamine, named anandamide from the Sanskrit word ananda, meaning "bliss."37 Compared with THC, anandamide has only moderate affinity for CB1, and is rapidly metabolized by amidases (enzymes that remove amide groups). Despite its short duration of action, anandamide shares most of the pharmacological effects of THC37, 152 Rapid degradation of active molecules is a feature of neurotransmitter systems that allows them control of signal timing by regulating the abundance of signaling molecules. It creates problems for interpreting the results of many experiments and might explain why in vivo studies with anandamide injected into the brain have yielded conflicting results.

Anandamide appears to have both central (in the brain) and peripheral (in the rest of the body) effects. The precise neuroanatomical localization of anandamide and the enzymes that synthesize it are not yet known. This information will provide

2.13 ---------------------------------------------------------------------------

essential clues to the natural role of anandamide and an understanding of the brain circuits in which it is a neurotransmitter. The importance of knowing specific brain circuits that involve anandamide (and other endogenous cannabinoid ligands) is that such circuits are the pivotal elements for regulating specific brain functions, such as mood, memory, and cognition. Anandamide has been found in numerous regions of the human brain: hippocampus (and parahippocampic cortex), striatum, and cerebellum; but it has not been precisely identified with specific neuronal circuits. CB1 receptors are abundant in these regions, and this further implies a physiological role for endogenous cannabinoids in the brain functions controlled by these areas. But, substantial concentrations of anandamide are also found in the thalamus, an area of the brain that has relatively few CB1 receptors.124

Anandamide has also been found outside the brain. It has been found in spleen, which also has high concentrations of CB2 receptors; and small amounts have been detected in heart.44

In general, the affinity of anandamide for cannabinoid receptors is only one fourth to one-half that of THC (see table 2.3). The differences depend on the cells or tissue that are tested and on the experimental conditions, such as the binding assay used (reviewed by Pertwee124).

The molecular structure of anandamide is relatively simple, and it can be formed from arachidonic acid and ethanolamine. Arachidonic acid is a common precursor of a group of biologically active molecules known as eicosanoids, including prostaglandins.c Although anandamide can be synthesized in a variety of ways, the physiologically relevant pathway seems to be through enzymatic cleavage of N-arackidonyl-phosphatidyl-ethanolamine (NAPE), which yields anandamide and phosphatidic acid (reviewed by Childers and Breivogel27).

Anandamide can be inactivated in the brain via two mechanisms. In one, anandamide is enzymatically cleaved to yield arachidonic acid and ethanolamine- the reverse of what was initially proposed as its primary mode of synthesis. In the other, it is inactivated through neuronal uptake-i.e.., by being transported into the neuron, which prevents its continuing activation of neighboring neurons. --------------------------------------------------------------------------- c Eicosanoids all contain a chain of 20 carbon atoms, and are named after eikosi, the Greek word for twenty.

2.14 ---------------------------------------------------------------------------

Table 2.2 Compounds that bind to cannabinoid receptors

Compounds That Bind to Cannabinoid Receptorsd

Compound Properties -------------------------------------------------------------------------- Agonists (Receptor activators) Plant-derived compounds Main psychoactive cannabinoid in the marijuana plant; largely responsible for psychological and [Image]9-THC physiological effects. (Except in discussions of the different forms of THC, THC is used as a synonym for [Image]9-THC

Slightly less potent than [Image]9-THC and much [Image]8-THC less abundant in the marijuana plant, but otherwise similar. Bioactive compound formed when the body breaks down 11-OH-[Image]9-THC [Image]9-THC. Presumed to be responsible for some of the effects of marijuana. Cannabinoid agonists found in animals Found in animals ranging from mollusks to mammals. anandamide Appears to be primary endogenous cannabinoid (arachidonyl- agonist in mammals. Chemical structure very ethanolamide) different from plant cannabinoids, and related to prostaglandins. 2-AG Endogenous agonist. Structurally similar to (arachidonyl anandamide. More abundant but less potent than glycerol) anandamide. THC analogues Synthetic THC. Marketed in the US under the name Dronabinol Marinol® for nausea associated with chemotherapy and for AIDS-related wasting.

Nabilone THC analogue. Marketed in the UK under the name Cesamet® for the same indications as dronabionol.

CP 55,940 Synthetic cannabinoid; THC analogue; that is, it is structurally similar to THC Levonantradol THC analogue. THC analogue, 100-800 fold greater potency than HU-210 THC.97

Chemical structure unlike THC or anandamide Chemical structure different from known WIN-55,212 cannabinoids, but binds to both cannabinoid receptors. Chemically related to cyclo-oxygenase inhibitors, which include anti-inflammatory drugs. Antagonists (Receptor Blockers)

SR 141716A Synthetic CB1 antagonist, developed in 1994.132

SR 144528 Synthetic CB2 antagonist; developed in 1997.133

d Sources: Mechoulam et al. 1998 109, Felder and Glass 199843; BMA 199717

2.15 ---------------------------------------------------------------------------

Figure 2.4 Chemical structures of compounds that bind to cannabinoid receptors

[Image]

Figure Legend. Selected cannabinoid agonists, or molecules that bind to and activate cannabinoid receptors. THC is the primary psychoactive molecule found in marijuana. CP 55,940 is a THC-analogue; that is, its chemical structure is related to THC. Anandamide and 2-arachidonyl glycerol (2-AG) are endogenous molecules, meaning they are naturally produced in the body. Although the chemical structure of WIN 55,212 is very different from either THC or anandamide, it is also a cannabinoid agonists.

2.16 ---------------------------------------------------------------------------

Table 2.3 Comparison of cannabinoid receptor agonists

Potency can be measured in a variety of ways, from behavioral to physiological to cellular. This table shows potency in terms of receptor binding, which is the most broadly applicable to the many possible actions of cannabinoids. For example, anandamide binds to the cannabinoid receptor only about half as avidly as does THC. Measures of potency might include effects on activity (behavioral) or hypothermia (physiological).

The apparently low potency of 2-AG may, however, be misleading. A study published late in 1998, reports that 2-AG is found with two other, closely related compounds that, by themselves, are biologically inactive, but in the presence of those two compounds, 2-AG is only three times less active than THC.9 Further, 2-AG is much more abundant than anandamide, although the biological significance of this remains to be determined.

Receptor Binding in Brain Tissue124

Compound Potency Relative to THC CP 55,940 59 [Image]9-THC1

Anandamide 0.47 2-AG 0.08

2.17 ---------------------------------------------------------------------------

Other endogenous agonists

Several other endogenous compounds that are chemically related to anandamide and that bind to cannabinoid receptors have been discovered, one of which is 2-AG.108 2-AG is closely related to anandamide and is even more abundant in the brain. At time of this writing, all known endogenous cannabinoid receptor agonists (including anandamide) are eicosanoids, which are arachidonic acid metabolites. Arachidonic acid (a free fatty acid) is released via hydrolysis of membrane phospholipids.

Other, non-eicosanoid, compounds that bind cannabinoid receptors have recently been isolated from brain tissue, but they have not been identified, and their biological effects are under investigation. This is a fast-moving field of research, and no review over six months old can be fully up-to-date.

The endogenous compounds that bind to cannabinoid receptors probably perform a broad range of natural functions in the brain. This neural signaling system is rich and complex, and has many subtle variations' many of which await discovery. In the next few years, much more will likely by known about these naturally occurring cannabinoids.

Some effects of cannabinoid agonists are receptor-independent. For example, both THC and CBD can be neuroprotective through their antioxidative activity; that is, they can reduce the toxic forms of oxygen that are released when cells are under stress.54 Other likely examples of receptor-independent cannabinoid activity are modulation of activation of membrane-bound enzymes (e.g., ATPase), arachidonic acid release, and perturbation of membrane lipids. An important caution in interpreting those reports is that concentrations of THC or CBD used in cellular studies, such as these, are generally much higher than the concentrations of THC or CBD in the body that would likely be achieved by smoking marijuana.

Novel targets for therapeutic drugs

Drugs that alter the natural biology of anandamide, or other endogenous cannabinoids, might have therapeutic uses (table 2.4). For example, drugs that selectively inhibit neuronal uptake of anandamide would increase the brain's own natural cannabinoids and mimic some of the effects of THC. A number of important psychotherapeutic drugs act by inhibiting neurotransmitter uptake. For example, antidepressants like fluoxetine (Prozac®) inhibit serotonin uptake, and are known as selective serotonin re-uptake inhibitors, or SSRI's. Another way to alter levels of endogenous cannabinoids would be to develop drugs that act on the enzymes involved in anandamide synthesis. Some anti-hypertensive drugs work by inhibiting

2.18 ---------------------------------------------------------------------------

enzymes involved in the synthesis of endogenous hypertensive agents. Fore example, anti-converting enzyme (ACE) inhibitors are used in hypertensive patients to interfere with the conversion of angiotensin I, which is inactive, to the active hormone, angiotensin II

2.19 ---------------------------------------------------------------------------

Table 2.4. Cellular processes that can be targeted for drug development

TABLE LEGEND. Endogenous cannabinoids are part of a cellular signaling system. This table lists categories of natural processes that regulate such systems, and shows the results of altering those processes.

Cellular Processes That Can Be Targeted for Drug Development Drug action . Biological Result Synthesis of bioactive Block compounds is a continuous Weaker signal, due to synthesis process and is one means by decreased agonist which concentrations of that concentration compound are regulated. Chemical breakdown is one Inhibit method the body uses to Stronger signal, due to degradation inactivate endogenous increased agonist substances. concentration increased. Stronger signal, due to Facilitate Neuronal uptake is one of increased amount of time neuronal the natural ways in which a during which agonist is uptake receptor agonist is present in the synapse where inactivated. it can stimulate the receptor

2.20 ---------------------------------------------------------------------------

Sites of Action

Cannabinoid receptors are particularly abundant in some areas of the brain. The normal biology and behavior associated with these brain areas are consistent with the behavioral effects produced by cannabinoids (table 2.5 and figure 2.5). The highest receptor density is found in cells of the basal ganglia that project locally and to other brain regions. These cells include the substantia nigra pars reticulata, entopeduncular nucleus, and globus pallidus, regions that are generally involved in coordinating body movements. Patients with Parkinson and Huntington disease tend to have impaired functions in these regions.

CB1 receptors are also abundant in the putamen, part of the relay system within the basal ganglia that regulates body movements, the cerebellum, which coordinates body movements; the hippocampus, which is involved in learning, memory, and response to stress; and the cerebral cortex, which is concerned with the integration of higher cognitive functions.

CB1 receptors are found on various parts of neurons, including the axon, cell bodies, terminals, and dendrites.57, 165 Dendrites are generally the "receiving" part of a neuron, and receptors on axons or cell bodies generally modulate other signals. Axon terminals are the "sending" part of the neuron.

Cannabinoids like the inhibitory neurotransmitter [Image]-aminobutyric acid (GABA) -tend to inhibit neurotransmission, although the results are somewhat variable. In some cases, cannabinoids diminish the effects of the inhibitory neurotransmitter, [Image]-aminobutyric acid (GABA),144 in other cases, cannabinoids can augment the effects of GABA.120 The effect of activating a receptor depends on where it is found on the neuron: if cannabinoid receptors are presynaptic (on the "sending" side of the synapse) and inhibit the release Of GABA, cannabinoids would diminish GABA effects; the net effect would be stimulation. However, if cannabinoid receptors are postsynaptic (on the "receiving" side of the synapse) and on the same cell as GABA receptors, they will probably mimic the effects of GABA; in that case, the net effect would be inhibition.120, 144, 160

CB1 is the predominant brain cannabinoid receptor. CB2 receptors have not generally been found in the brain, but there is one isolated report suggesting some in mouse cerebellum.150 CB2 is found primarily on cells of the immune system. CB1 receptors are also found in immune cells, but CB2 is considerably more abundant there (table 2.6) (reviewed by Kaminski80 in 1998).

As can be appreciated in the next section, the presence of cannabinoid systems in key brain regions is strongly tied to the functions and pathology associated with those regions. The clinical value of cannabinoid systems is best understood in the context of the biology of these brain regions.

2.21 ---------------------------------------------------------------------------

Table 2.5 Brain regions in which cannabinoid receptors are abundante

Brain Region Functions Associated with Region Brain regions in which cannabinoid receptors are abundant

Basal ganglia Substantia nigra pars reticulate Entopeduncular nucleus Movement control Globus pallidus Putamen

Cerebellum Body-movement coordination Hippocampus Learning and memory, stress Cerebral cortex, especially cingulate, frontal, and parietal Higher cognitive functions regions Nucleus accumbens Reward center Brain regions in which cannabinoid brain receptors are moderately concentrated Body housekeeping functions Hypothalamus (body-temperature regulation, salt and water balance, reproductive function) Amygdala Emotional response, fear Spinal cord Peripheral sensation, including pain

Brain Stem Sleep and arousal, temperature regulation, motor control Central gray Analgesia Nucleus of the solitary tract Visceral sensation, nausea and vomiting

e Based on reviews by Pertwee 1997 124 and Herkenham 199557 This table will be accompanied by a figure.

2.22 ---------------------------------------------------------------------------

Figure 2.5. Location of brain regions in which cannabinoid receptors are abundant.

See table 2.5 for summary of functions associated with those regions.

[Image]

2.23 ---------------------------------------------------------------------------

Table 2.6 Summary table of cannabinoid receptors

. CB1 CB2 Effects of various cannabinoids [Image]9-THC Agonist Weak antagonist

Anandamide Agonist Agonist Agonist; greater affinity for Cannabinol (CBN) Weak agonist CB2 than for CB1

Cannabidiol (CBD) Does not bind to Does not bind to receptor receptor Receptor distribution Areas of greatest Immune system, especially B abundance Brain cells and natural killer cells

2.24 ---------------------------------------------------------------------------

Cannabinoid Receptors and Brain Functions

Motor effects

Marijuana affects psychomotor performance in humans. The effects depend both on the nature of the task and the experience with marijuana. In general, effects are clearest in steadiness (body sway and hand steadiness) and in motor tasks that require attention. The results of testing Cannabinoids in rodents are much clearer.

Cannabinoids clearly affect movement in rodents, but the effects depend on the dose: low doses stimulate and higher doses inhibit locomotion.111, 159 Cannabinoids mainly inhibit the transmission of neural signals, and they inhibit movement through their actions on the basal ganglia and cerebellum, where cannabinoid receptors are particularly abundant (figure 2.6a and 2.6b). Cannabinoid receptors are also found in the neurons that project from the striatum and subthalamic nucleus, which inhibit and stimulate movement, respectively.58, 101

Cannabinoids decrease both the inhibitory and stimulatory inputs to the substantia nigra, and therefore might provide dual regulation of movement at this nucleus. In the substantia nigra, Cannabinoids decrease transmission from both the striatum and the subthalamic nucleus.141 The globus pallidus has been implicated in mediating the cataleptic effects of large doses of Cannabinoids in rats.126 (Catalepsy is a condition of diminished responsiveness usually characterized by trancelike states and waxy rigidity of the muscles.) Several other brain regions - the cortex, the cerebellum, and the neural pathway from cortex to striatum - are also involved in the control of movement and contain abundant cannabinoid receptors.52, 59, 101 They are, therefore, possible additional sites that might underlie the effects of Cannabinoids on movement.

2.25 ---------------------------------------------------------------------------

Figure 2. 6a & b Diagrams showing motor regions of the brain

[Image]

Figure 2.6. Basal ganglia are a group of three brain regions, or nuclei - caudate, putamen, and globus pallidus. Figure 2.6a is a 3-dimensional view showing the location of those nuclei in the brain. Figure 2.6b shows those structures in a vertical cross-sectional view The major output pathways of the basal ganglia arise from the globus pallidus and pars reticula of the substantia nigra. Their main target is the thalamus.

2.26 ---------------------------------------------------------------------------

Memory effects

One of the primary effects of marijuana in humans is disruption of short-term memory.68 That is consistent with the abundance of CB1 receptors in the hippocampus, the brain region most closely associated with memory. The effects of THC resemble a temporary hippocampal lesion.63 Deadwyler and colleagues have demonstrated that cannabinoids decrease neuronal activity in the hippocampus and its inputs 23, 24, 83 In vitro, several cannabinoid ligands and endogenous cannabinoids can block the cellular processes associated with memory formation.29, 30, 116, 157, 163 Furthermore, cannabinoid agonists inhibit release of several neurotransmitters: acetylcholine from the hippocampus,49,50,51 norepinephrine from human and guinea pig (but not rat or mouse) hippocampal slices,143 and glutamate in cultured hippocampal cells.144 Cholinergic and noradrenergic neurons project into the hippocampus, but circuits within the hippocampus are glutamatergic.f Thus, cannabinoids could block transmission both into and within the hippocampus by blocking presynaptic neurotransmitter release.

Pain

After nausea and vomiting, chronic pain was the condition cited most often to the IOM study team as a medical use for marijuana. Recent research presented below has shown intriguing parallels with anecdotal reports of the modulating effects of cannabinoids on pain - both the effects of cannabinoids acting alone and the effects of their interaction with opioids.

Behavioral Studies

Cannabinoids reduce reactivity to acute painful stimuli in laboratory animals. In rodents, cannabinoids reduced the responsiveness to pain induced through various stimuli, including thermal, mechanical, and chemical stimuli.12, 19, 46, 72, 96, 154, 174 Cannabinoids were comparable with opiates in potency and efficacy in these expeniments. 12, 72

Cannabinoids are also effective in rodent models of chronic pain. Herzberg and coworkers found that cannabinoids can block allodynia and hyperalgesia --------------------------------------------------------------------------- fNeurons are often defined by the primary neurotransmitter released at their terminals. Thus, cholinergic neurons release acetylcholine, noradrenergic neurons release noradrenalin (also known as norepinephrine), and glutamergic neurons release glutamate.

2.27 ---------------------------------------------------------------------------

associated with neuropathic pain in rats.117 (Allodynia refers to pain elicited by stimuli that are normally innocuous; hyperalgesia refers to abnormally increased reactivity to pain.) This is an important advance, because chronic pain frequently results in a series of neural changes that increase suffering due to allodynia, hyperalgesia, and spontaneous pain; furthermore' some chronic pain syndromes are not amenable to therapy, even with the most powerful narcotic analgesics.10

Pain perception is controlled mainly by neurotransmitter systems within the central nervous system, and cannabinoids clearly play a role in the control of pain in those systems.45 However, pain-relieving and pain-preventing mechanisms also occur in peripheral tissues, and endogenous cannabinoids appear to play a role in peripheral tissues. Thus, the different cannabinoid receptor subtypes might act synergistically. Experiments in which pain is induced by injecting dilute formalin into a mouse's paw have shown that anandamide and palmitylethanolamide (PEA) can block peripheral pain.22, 73 22 Anandamide acts primarily at the CB1 receptor, whereas PEA has been proposed as a possible CB2 agonist, in short, there might be a biochemical basis for their independent effects. When injected together, the analgesic effect is stronger than that of either alone. That suggests an important strategy for the development of a new class of analgesic drug: a mixture of CB1 and CB2 agonists. Because there are few, if any, CB2 receptors in the brain, it might be possible to develop drugs that enhance the peripheral analgesic effect while minimizing the psychological effects.

2.28 ---------------------------------------------------------------------------

Neural sites of altered responsiveness to painful stimuli

The brain and spinal cord mediate cannabinoid analgesia. A number of brain areas participate in cannabinoid analgesia and support the role of descending pathways (neural pathways that project from the brain to the spinal cord).103, 105 Although more work is needed to produce a comprehensive map of the sites of cannabinoid analgesia, it is clear that the effects are limited to particular areas, most of which have an established role in pain.

Specific sites where cannabinoids act to affect pain processing include the periaqueductal gray,104 the rostral ventral medulla, 105, 110 and the thalamic nucleus submedius,102 the thalamic ventroposterolateral nucleus,102 dorsal horn of the spinal cord,64, 65 and peripheral sensory nerves.64, 65, 66, 131 Those nuclei also participate in opiate analgesia. Although similar to opiate analgesia, cannabinoid analgesia is not mediated by opioid receptors; morphine and cannabinoids sometimes act synergistically, and opioid antagonists generally have no effect on cannabinoid induced analgesia.171 However, a kappa-receptor antagonist has been shown to attenuate spinal, but not supraspinal, cannabinoid analgesia.153, 170, 171 (Kappa opioid receptors constitute one of the three major types of opioid receptors; the other two types are mu and delta receptors.)

2.29 ---------------------------------------------------------------------------

Neurophysiology and neurochemistry of cannabinoid analgesia

Because of the marked effects of cannabinoids on motor function, behavioral studies in animals alone cannot provide sufficient grounds for the conclusion that cannabinoids depress pain perception. Motor behavior is typically used to measure responses to pain, but this behavior is itself affected by cannabinoids. Thus, experimental results include an unmeasured combination of cannabinoid effects on motor and pain systems. The effects on specific neural systems, however, can be measured at the neurophysiological and neurochemical levels. Cannabinoids decrease the response of immediate-early genes (genes that are activated in the early or immediate stage of response to a broad range of cellular stimuli) to noxious stimuli in spinal cord, decrease response of pain neurons in the spinal cord, and decrease the responsiveness of pain neurons in the ventral posterolateral nucleus of the thalamus.67, 102 Those changes are mediated by cannabinoid receptors, selective for pain neurons, and unrelated to changes in skin temperature or depth of anesthesia, and they follow the time course of the changes in behavioral responses to painful stimuli, but not the time course of motor changes.67 Cannabinoids also modulate the responses of on-cells and off-cells in the rostral ventral medulla in a manner that is very similar to that of morphine.55, 110 These cells control pain transmission at the level of the spinal cord.

Endogenous cannabinoids modulate pain

Endogenous cannabinoids can modulate pain sensitivity, through both central and peripheral mechanisms. For example, animal studies have shown that pain sensitivity can be increased when endogenous cannabinoids are blocked from acting at CB1 receptors 22, 62, 110, 130, 158 Administration of cannabinoid antagonists in either the spinal cord 130 or paw 22 increase the sensitivity of animals to pain. In addition, there is evidence that cannabinoids also act at the site of injury to reduce peripheral inflammation.131

Current data suggest the endogenous cannabinoid analgesic system might offer protection against the long-lasting central hyperalgesia and allodynia that sometimes follow skin or nerve injuries 130, 158 These results raise the possibility that therapeutic interventions that alter the levels of endogenous cannabinoids might be useful for managing pain in humans.

2.30 ---------------------------------------------------------------------------

Chronic Effects of THC

Most substances of abuse produce tolerance, physical dependence, and withdrawal symptoms. Tolerance is the most common response to repetitive use of the same drug (not necessarily a drug of abuse) and is the condition in which, after repeated exposure to a drug, increasing doses are needed to achieve the same effect. Physical dependence develops as a result of tolerance (adaptation) produced by a resetting of homeostatic mechanisms in response to repeated drug use. It is important to reiterate that the phenomena of tolerance, dependence, and withdrawal are not associated uniquely with drugs of abuse. Many medications that are not addicting can produce these types of effects; examples of such medications include clonidine, propranolol, and tricyclic antidepressants. The following sections discuss what is known about the biological mechanisms that underlie on tolerance, reward, and dependence; clinical studies about those topics are discussed in chapter 3.

Tolerance

Chronic administration of cannabinoids to animals results in tolerance to many of the acute effects of THC, including memory disruption,34 decreased locomotion,2, 119 hypothermia,42, 125 neuroendocrine effects,134 and analgesia.4 Tolerance also develops to the cardiovascular and psychological effects of THC and marijuana in humans (see also discussion in chapter 3).55, 56, 76

Tolerance to cannabinoids appears to result from both pharmacokinetic (how the drug is absorbed, distributed, metabolized, and excreted) and pharmacodynamic (how the drug interacts with target cells) changes. Chronic treatment with the cannabinoid agonist, CP 55,940, increases the activity of the microsomal cytochrome P450 oxidative system.31 Because this is the system through which drugs are metabolized in the liver, this suggests pharrnacokinetic tolerance. Chronic cannabinoid treatments also produce changes in brain cannabinoid receptors and cannabinoid receptor mRNA levels, indicating that pharmacodynamic effects are important, as well.

Most studies have found that brain cannabinoid receptor levels usually decrease after prolonged exposure to agonists,42, 119, 136, 138 although some studies have reported increases 137 or no changes2 in receptor binding in brain. Differences among studies may be due to the particular agonist tested, the assay used, brain region examined, or treatment time. For example, the THC analogue, levonantradol, produces a greater desensitization of adenylyl cyclase inhibition than THC in cultured neuroblastoma cells,40 which may be explained by the efficacy differences between these two agonists 18, 147 Furthermore, a time course study revealed differences in the rates and magnitudes of receptor down-regulation across brain

2.31 ---------------------------------------------------------------------------

regions.16 These findings suggest that tolerance to different effects of cannabinoids develops at different rates

Chronic treatment with THC also produces variable effects on cannabinoid-mediated signal transduction systems. Chronic THC treatment produces significant desensitization of cannabinoid-activated G-proteins in a number of rat brain regions.147 Moreover, the time course of this desensitization varies across brain regions.16

It is difficult to extend the findings of these short-term animal studies to human marijuana use In order to simulate long-term use, the doses used in animal studies are higher than normally achieved by smoking marijuana. For example, the average human will feel "high" after a 0.06 mg/kg injection of THC,118 compared to 10-20 mg/kg/day used in many chronic studies in rats. At the same time, doses of marijuana needed to observe behavioral changes in rats (usually changes in locomotor behavior) are substantially higher than doses at which people feel "high." In addition, pharmacokinetics of THC distribution in the body are dramatically different between rats and humans, as well as being highly dependent on the THC delivery system - that is, whether it is inhaled, injected, or swallowed. Nevertheless, it is likely that some of the same biochemical adaptations to chronic cannabinoid administration occur in both laboratory animals and humans, but the magnitude of the effects in humans may be smaller in proportion to the respective doses used.

Reward and dependence

Experimental animals that are given the opportunity to self-administer cannabinoids generally do not choose to do so, which has led to the conclusion that they are not reinforcing and rewarding.38 However, behavioral95 and brain stimulation94 studies have shown that THC can be rewarding to animals. The behavioral study used a "place-preference" test, in which an animal is given repeated doses of a drug in one place, and is then given a choice between a place where it did not receive the drug and one where it did; the animals chose the place where they received the THC. These rewarding effects are highly dose-dependent. In all models studied, cannabinoids are only rewarding at mid-range; doses that are too low are not rewarding, doses that are too high can be aversive. Mice will self-administer the cannabinoid agonist, WIN 55,212, but only at low doses.106 This effect is specifically mediated by CB1 receptors, and indicates that stimulation of those receptors is rewarding to the mice. Antagonism of cannabinoid receptors is also rewarding in rats; in conditioned place-preference tests, animals show a preference for the place they receive the cannabinoid antagonist, SR141716A, at both low and high doses.140 Cannabinoids increase dopamine levels in the mesolimbic dopamine system of rats, a pathway associated with reinforcement.25, 39, 161 However, the

2.32 ---------------------------------------------------------------------------

mechanism by which THC increases dopamine levels appears to be different from that of other abused drugs 51 g (see chapter 3 for further discussion of reinforcement).

Physical dependence on cannabinoids has only been observed under experimental conditions of "precipitated withdrawal", in which animals are first treated chronically with cannabinoids and then given the CB1 antagonist, SR141716A.3, 166 The addition of the antagonist accentuates any withdrawal effect by competing with the agonist at receptor sites; that is, the antagonist helps to clear agonists off and keep them off receptor sites. This suggests that, under normal cannabis use, the long half-life and slow elimination from the body of THC, and the residual bioactivity of its metabolite, 11-OH -THC, may prevent significant abstinence symptoms. The precipitated withdrawal effects produced by SR141716A have some of the characteristics of opiate withdrawal, but are not affected by opioid antagonists and affect motor systems differently. An earlier study with monkeys also suggested that abrupt cessation of chronic THC is associated with withdrawal symptoms,8 Monkeys in that study were trained to work for food after which they were given THC on a daily basis; when the investigators stopped administering THC, the animals stopped working for food.

A study in rats indicated that the behavioral cannabinoid withdrawal syndrome correlates with stimulation of central amygdaloid corticotropin-releasing hormone release, consistent with the consequences of withdrawal from other abused drugs.135 However, the withdrawal syndrome for cannabinoids and the corresponding increase in corticotropin-releasing hormone are only observed following administration of the CB1 antagonist, SR 141716A, to cannabinoidtolerant animals;3, 166 The implications of data based on precipitated withdrawal in animals for human cannabinoid abuse have not been established.166 Furthermore, acute administration of THC also produces increases in corticotropin-releasing hormone and adrenocorticotropin release, both of which are stress-related hormones.71 This set of withdrawal studies may explain the generally aversive effects of cannabinoids in animals, and may indicate that the increase in corticotropin-releasing hormone is merely a rebound effect. Thus, while cannabinoids appear to be conforming to some of the neurobiological effects of other drugs abused by humans, the underlying mechanisms of these actions and their significance in determining the reinforcement and dependence liability of cannabinoids in humans remain undetermined.

---------------------------------------------------------------------------

g These increases in dopamine are due to increases in the firing rate of dopamine cells in the ventral tegmental area by [Image]9-THC47. However, these increases in firing rate in the ventral tegmental area could not be explained by increases in the firing of the A10 dopamine cell group, where other abused drugs have been shown to act51.

2.33 ---------------------------------------------------------------------------

Cannabinoids and the Immune System

The human body protects itself from invaders such as bacteria and viruses through the elaborate and dynamic network of organs and cells referred to as the immune system (see box on Cells of the Immune System).

Cannabinoids, especially THC, can modulate the function of immune cells in various ways - in some cases enhancing, and in others diminishing the immune response 85 (summarized in table 2.7). However, the natural function of the cannabinoids in the immune system is not known. Immune cells respond to cannabinoids in a variety of ways, depending upon experimental factors such as drug concentration, timing of drug delivery to leukocytes in relation to antigen stimulation, and the type of cell function analyzed. Although the chronic effects of cannabinoids on the immune system have not been studied, based on acute exposure studies in experimental animals it appears that the concentrations of THC which modulate immunological responses are higher than those required for psychoactivity.

2.34 ---------------------------------------------------------------------------

Table 2.7 Effects of Cannabinoids on the Immune System

Cell Types Drug Drug Tested or Concen- Tested Type Drug of tration a Result Reference Animal Experiment Luo, 1992; THC Lymphocytes Pross,1992* 2-AG and Higher doses Klein, 1985%; 11-OH-THC Splenocytes 0.1-30 µM suppress T cell Specter,1990& CBN in vitro proliferation Lee, 1995* Herring, 1998

Lymphocytes Lower doses Luo,1992; increase T cell THC and 0.1-25 µM proliferation in Lee,1995* 2-AG Splenocytes vitro Pross,1992*

Anandamide Splenocytes Little or no Lee,1995* in vitro 1-25 µM effect on T cell Devane,1992 proliferation THC, 11-OH-THC Splenocytes 3-30 µM Decrease B cell Klein,1985% AG-2 in vitro proliferation Lee,1995* THC CP 55,940 Lymphocytes 0.1-100nM Increase B cell WIN in vitro [0.0001-0.1 proliferation Derocq, 1995 55,212-2 µM]

THC Mice were >5mg/kg Antibody Baczynsky, 1983 injected with production was Schatz,1993 HU-210 drug >0.05 mg/kg suppressed Titishov,1989

THC Klein,1990 11-OH-THC Antibody Baczynasky,1983 CBD Splenocytes 1-30µM production was Kaminski,1994 CP55,940 in vitro suppressed Kaminski,1992 CBN Herring,1998 Repeated low doses or a high Rodents were 3mg/kg/day dose of THC THC injected with for 25days suppress the Patel,1985 drug 40mg/kg/day activity of Klein,1987 for 2 days natural killer cells Doses of >=10 µM suppress natural THC Natural killer cell Klein,1987 1l-OH-THC killer cells 0.1-32 µM cytolytic Luo,1989 in vitro activity, doses <10 µM produced no effect Variable doses Peritoneal of THC suppress Lopez-Cepero, 1986 THC macrophages 3-30 µM macophage Specter,1991 and monocytes functions in Tang,1992 vitro

2.35 THC suppresses >5mg/kg for normal immune Mice injected with 4 days or response, THCdrug; in one case, 50 mg/kg interferons failed Cabral,1986 CBDin vitro tests done every 5 to increase when Blanchard,1986 on spleens days for up exposed to cytokine to 8 weeks inducer, while CBD had no suppressive effect Increased <0.1 µM interferon Warzl, 1991 THCPeripheral blood production CBDmononuclear cells in Decreases vitro 30 µM interferon . production Both THC and CBD THCSplenocytes and T suppress IL-2 CBDcells in vitro 10 µM secretion and the Condie,1996 number of IL-2 transcripts

Phorbol myristate Increase in tumor necrosis factor THCacetate 10-20 µM production and IL-I Shivers, 1994 differentiated macrophage in vitro supernatant bioactiviy Increase processing and release of IL-I THCEndotoxin-activated 10-30 µM rather than Zhu, 1994 macrophages in vitro cellular production of the IL-I

THCPeritoneal 10-30 µM Increased IL-I Klein, 1990 macrophages in vitro bioactivity 8mg/kg Cytokine-mediated given septic shock and before and death occurs with after exposure to bacteria sublethal dose of Mice were injected infection the bacteria with drug and either Survival occurs, Klein, 1993 and THCsublethal or lethal < 5 mg/kg but with greater 1994 dose of Legionalla doses. or susceptiblity to Newton, 1994 pneumophilia one 8 mg/kg infection when or 4 mg/kg dose given challenged with before bacteria and death bacteria when challenged infection with a lethal dose of bacteria Two high doses of 100mg/kg THC potentiates the before and effects of herpes simplex and Immuno-deficient after virus enhances the infection THCmice injected with progression of Specter, 1991 drug and herpes death simplex virus 100 mg/kg before A single dose did virus not promote death infection

* cell density dependent; * mitogen dependent; % % serum dependent; & dependent on timing of drug exposure relative to mitogen exposure.

a Drug concentrations are given in the standard format of molarity (M). A one molar solution is the molecular weight of the compound (in grams) dissolved in 1 liter of water or other solvent. The molecular weight of THC is 314 so a 1 molar solution would be 314 grams of THC dissolved in 1 liter of solution, a 10 µM solution would be 3.14 mg THC/liter.

A 1-10 µM concentration will generally elicit a physiologically relevant response in immune cell cultures. Higher doses are often suspected of not being biologically meaningful, because they are a much larger dose than would ever be achieved in the body. The doses listed in this table are, for the most part, very high. See text for further discussion.

2.36 ---------------------------------------------------------------------------

Cells of the Immune System

The various organs of the immune system are positioned throughout the body and include bone marrow, thymus, lymph nodes and spleen. The cells of the immune system consist of white blood cells, or leukocytes, which are formed in the bone marrow from stem cells so-called because a great variety of cells descend from them (see below). There are two kinds of leukocytes: Lymphocytes and phagocytes. Lymphocytes consist of B cells, T cells,h and natural killer cells (NK), and the major phagocytes include monocytes, macrophages and neutrophils. Phagocytes have many important roles in the immune response, but most significantly they initiate these responses by engulfing and digesting foreign substances (e.g., bacteria, viruses, foreign proteins), or antigens, that enter the body. Once digested, the antigen is exposed to specialized Iymphocytes (i.e., B cells and T cells) so that antibodies and effector T cells can be produced to help destroy any remaining antigens in the body. Antibodies are proteins produced by B cells that bind to antigens and promote antigen destruction. effector T cells include killer T cells which attack and kill antigen laden cells, and helper T cells, which secrete special proteins called cytokines that promote antigen elimination. Natural killer cells are specialized Iymphocytes that are also activated by antigen to either kill infected targets or secrete immunoregulatory cytokines.

[Image] ---------------------------------------------------------------------------

h The B and T refer to where the cells mature, either in the bone marrow (B) or thymus (T).

2.37 ---------------------------------------------------------------------------

The CB2 receptor gene, which is not expressed in the brain, is particularly abundant in immune tissues, with an expression level 10-100 times higher than that of CB1 In spleen and tonsils, the CB2 mRNAi content is equivalent to that of CB1 mRNA in the brain.48 The rank order, from high to low, of CB2 mRNA levels in immune cells is B-cells > natural killer cells >> monocytes > polymorphonuclear neutrophil cells > T8 cells in T4 cells. In tonsils, the CB2 receptors appear to be restricted to B-lymphocyte-enriched areas. In contrast, CB1 receptors are mainly expressed in the central nervous system and, to a lower extent, in several peripheral tissues such as adrenal gland, heart, lung, prostate, uterus, ovary, testis, bone marrow, thymus and tonsils.

Cannabinoid Receptors and Intracellular Action in Immune Cells

CB2 appears to be the predominant gene expressed in resting leukocytes.78, 112 The level of CB1 gene activity is normally low in resting cells but increases with cell activation.32 Thus the CB1 receptor might be important only when immune responses are stimulated, but the physiological relevance of this observation remains to be determined. Some of the cannabinoid effects observed in immune systems, especially at high drug concentrations, are likely mediated through non-receptor mechanisms, but these have not yet been identified.4

Ligand binding to either the CB1 or CB2 receptors inhibits adenylate cyclase, an enzyme that is responsible for cAMP production, and is, thus, an integral aspect of intracellular signal transduction (see figure 2.3).53, 79, 91, 122, 139, 151, 167 Increases in intracellular cAMP concentrations lead to immune enhancement, while decreases lead to an inhibition of immune responses.77 Cannabinoids inhibit the rise in intracellular cAMP that normally results from leukocyte activation, and this might be the pathway through which cannabinoids suppress immune cell functions.28, 74, 167 In addition, cannabinoids activate other molecular pathways such as the nuclear factor-kB pathway and therefore these signals might be modified in drug treated immune cells.33, 74

i After a gene is transcribed it is often spliced and modified into mRNA, or message RNA. The CB-2 mRNA is the gene "message" that moves from the cell nucleus into the cytoplasm where it will be translated into the receptor protein.

2.38 ---------------------------------------------------------------------------

T and B Cells

When stimulated by antigen, lymphocytes (see box on Cells of the Immune System) first proliferate and then mature or differentiate to become potent effector cells, such as B cells that release antibodies or T cells that release cytokines. The normal T cell proliferation that is seen when human lymphocytes and mouse splenocytes (spleen cells) are exposed to antigens and mitogensj can be inhibited by THC, 11-OH-THC, cannabinol, and 2-AG, as well as synthetic cannabinoid agonists such as CP 55,940, WIN 55,212 and HU-210 61, 89, 93, 99, 127, 155 In contrast, one study testing anandamide revealed little or no effect on T-cell proliferation.93

However, these drug effects are variable, and depend on experimental conditions such as the experimental drug dose used, mitogen used, percent of serum in the culture, and timing of cannabinoid drug exposure. In general, lower doses of cannabinoids increase proliferation while higher doses suppress proliferation. Doses that are effective in suppressing immune function are typically greater than 10 µM in cell culture studies and greater than 5 mg/kg in whole animal studies.85 By comparison, at 0.05 mg/kg, people will experience the full psychoactive effects of THC; however, because of their high metabolic rates, small rodents frequently require drug doses that are 100-fold higher than doses needed for humans to achieve comparable drug effects. Thus, the immune effects of doses of cannabinoids higher than those ever experienced by humans, should be interpreted with caution.89, 93, 93, 127, 155

As with T cells, B cell proliferation can be suppressed by various cannabinoids, such as THC, 11-OH-THC and 2-AG, but B cell proliferation is more inhibited at lower drug concentrations than T cell proliferation.89, 93 Conversely, low doses of THC, CP 55,940 and WIN 55,212-2 increase B-cell proliferation in cultured human cells exposed to mitogen.35 This effect possibly involves the CB2 receptor, because the effect appears to be the same when the CB1 receptor was blocked by the antagonist, SR-141716A (which does not block the CB2 receptor). The reason for the differences in cell responsiveness to cannabinoids is probably due to differences in cell type and source; for example, B cells collected from mouse spleen might respond to cannabinoids somewhat differently than B cells from human tonsils.

jMitogens are substances that stimulate cell division (mitosis) and cell transformation.

2.39 ---------------------------------------------------------------------------

Natural Killer Cells

Repeated injections of relatively low doses of THC (3 mg/kg/day 121 k) or two injections of a high dose (40 mg/kg86) suppress the ability of natural killer (NK) cells to destroy foreign cells in rats and mice. THC can also suppress natural killer cell cytolytic activity in cell cultures; 11-OH-THC, is even more potent.86 In contrast, THC doses below 10µM had no effect on natural killer cell activity in mouse cell cultures.98

Macrophages

Macrophages perform various functions including phagocytosis (ingestion and destruction of foreign substances), cytolysis, antigen presentation to lymphocytes, and production of a variety of active proteins involved in destroying microorganisms, tissue repair and modulation of immune cells. Those functions can be suppressed by THC doses similar to those capable of modulating lymphocyte functions (see above).88, 109

Cytokines

Cytokines are proteins produced by immune cells. When released from the producing cell they can alter the function of other cells they come in contact with. In a sense, they are like hormones. Thus, cannabinoids can either increase or decrease cytokine production depending upon experimental conditions.

Certain cytokines, such as interferon-[Image] and interleukin-2 (IL-2) are produced by T helper-1 (Th1) cells. These cytokines help to activate cell-mediated immunity and the killer cells that eliminate microbes from the body (see Box on cells of the immune system). When injected into mice, THC suppresses the production of those cytokines that modulate the host response to infection (see below).115 Cannabinoids also modulate interferons induced by viral infection,21 as well as other interferon inducers.85 Furthermore, in human cell cultures, interferon production can be increased by low concentrations, but decreased by high concentrations of either THC or cannabidiol. 6 In addition to Th1 cytokines, cannabinoids also modulate the production of cytokines such as interleukin-1 (IL-1), tumor necrosis factor (TNF), and interleukin-6 (IL-6). 145, 176 At 8 mg/kg, THC can increase the in vivo mobilization of serum acute phase cytokines including IL-1, TNF, and IL-6.90 Finally, although these studies suggest that cannabinoids can induce an increase in cytokines, other studies suggest that they can also suppress cytokine production.85 The different results might be due to different cell culture conditions or because different cell lines were studied.

---------------------------------------------------------------------------

k While 3 mg/kg would be a high dose for humans (see table 3.1); in rodents, it is a low dose for immunological effects, and a moderate dose for behavioral effects.

2.40 ---------------------------------------------------------------------------

Antibody Production

Antibody production is an important measure of humoral immune function as contrasted with cellular (cell-mediated immunity). Antibody production can be suppressed in mice injected with relatively low doses of THC (>5 mg/kg) or HU210 (>0.05 mg/kg) and in mouse spleen cell cultures exposed to a variety of cannabinoids, including THC, 11-OH-THC, cannabinol, cannabidiol, CP 55,940, or HU-210.5, 6, 61, 78, 79, 84, 85, 142, 164 However, the inhibition of antibody response by cannabinoids was only observed when antibody-forming cells were exposed to T cell-dependent antigens (the responses require functional T cells and macrophages as accessory cells). Conversely, antibody responses to several T cell-independent antigens were not inhibited by THC, suggesting that the B cell is relatively insensitive to inhibition by cannabinoids.142

Resistance To Infection In Animals Exposed To Cannabinoids

Bacterial infections evaluated in mice demonstrated that THC can suppress resistance to infection, although the effect depends upon the dose and timing of drug administration. Mice pre-treated with THC (8 mg/kg) one day prior to infection with a sublethal dose of the pneumonia-causing bacteria, Legionella pneumophilia, and then treated again one day after the infection with THC, developed symptoms of cytokine-mediated septic shock and died; control mice that were not pre-treated with THC became immune to repeated infection and survived the bacterial challenge.90 If only one injection of THC was given or doses less than 5 mg/kg were used, all of the mice survived the initial infection, but failed to survive a subsequent challenge with a lethal dose of the bacteria; hence these mice failed to develop immune memory in response to the initial sublethal infection.87 Note that these are very high doses, and are considerably higher than doses experienced by marijuana users (see figure 3.1).115 In rats, doses of 4.0 mg/kg THC are aversive95

Few studies have been done to evaluate the effect of THC on viral infections, and this is an area that needs further study.20 Compared to healthy animals, THC might have greater immunosuppressive effects in animals whose immune systems are severely weakened. For example, a very high dose of THC (100 mg/kg) given twice, two days before and after herpes simplex virus infection, was shown to be a co-factor with herpes simplex virus in enhancing the progression to death in an immunodeficient mouse model infected with a leukemia virus.85 However, THC given as a single dose (100 mg/kg) two days before herpes simplex virus infection did not promote the progression to death in these animals. Hence, whether THC is immunosuppressive likely depends on the timing of THC exposure relative to an infection.

2.41 ---------------------------------------------------------------------------

Anti-inflammatory Effects

As discussed above, cannabinoid drugs can modulate the production of cytokines, which are central to inflammatory processes in the body. In addition, several studies have shown directly that cannabinoids can be anti-inflammatory. For example, in rats with autoimmune encephalomyelitis (an experimental model used to study multiple sclerosis), cannabinoids were shown to attenuate the signs and the symptoms of central nervous system damage.100, 172 (Some believe that nerve damage associated with multiple sclerosis is caused by an inflammatory reaction.) Likewise, the cannabinoid, HU-211, was shown to suppress brain inflammation that resulted from closed head injury 146 or infectious meningitis 7 in studies on rats. HU211 is a synthetic cannabinoid that does not bind to cannabinoid receptors, and is not psychoactive,7 thus, without direct evidence, the effects of marijuana cannot be assumed to include those of HU-211. CT-3, another atypical cannabinoid, suppresses acute and chronic joint inflammation in animals.178 It is a nonpsychoactive, synthetic derivative of 11-THC-oic acid (a breakdown product of THC), and does not appear to bind to cannabinoid receptors. 129 Cannabichromene, a cannabinoid found in marijuana, has also been reported to have anti-inflammatory properties.173 No mechanism of action for possible anti-inflammatory effects of cannabinoids has been identified and the effects of these atypical cannabinoids and effects of marijuana are not yet established.

It is interesting to note that two reports of cannabinoid-induced analgesia are based on the ability of the endogenous cannabinoids, anandamide and PEA, to reduce pain associated with local inflammation that was experimentally induced by subcutaneous injections of dilute formalin.22, 73 Both THC and anandamide can increase serum levels of ACTH and corticosterone in animals.169 Those hormones are involved in regulating many responses in the body, including those to inflammation. The possible link between experimental cannabinoid-induced analgesia and reported anti-inflammatory effects of cannabinoids is important for potential therapeutic uses of cannabinoid drugs, but has not yet been established.

Conclusions regarding effects on immune system

Based on cell culture and animal studies, cannabinoids have been established as immunomodulators - that is, they increase some immune responses and decrease others. The variable responses depend upon experimental factors such as drug dose, timing of delivery, and type of immune cell examined.

Cannabinoids affect multiple cellular targets within the immune system and a variety of effector functions. Many of the effects noted above appear to occur at

2.42 ---------------------------------------------------------------------------

concentrations of > 5 µM in vitro and > 5 mg/kg in vivo.l By comparison, a 5 mg injection of THC into a person (about 0.06 mg/kg) is enough to produce strong psychoactive effects. It should be emphasized, however, that little is known about the effects of chronic low dose exposure to cannabinoids on the immune system.

Another issue in need of further clarification involves the potential usefulness of cannabinoids as therapeutic agents in inflammatory diseases. Glucocorticoids have historically been used for these diseases, but non-psychotropic cannabinoids potentially have fewer side effects and might thus offer an improvement over glucocorticoids in treating inflammatory diseases.

Conclusions and Recommendations

Following the progress of the past fifteen years in understanding the effects of cannabinoids, research over the next decade is likely to reveal even more. It is interesting to compare how little we know about the cannabinoids with how much we know about the opiates (table 2.8). This table, in fact, suggests good reason for optimism about the future of cannabinoid drug development. Now that many of the basic tools of cannabinoid pharmacology and biology have been developed, one can expect to see rapid advances that can begin to match what is known for opiate systems in the brain.

Despite the tremendous progress in understanding the pharmacology and neurobiology of brain cannabinoid systems, this field is still in its early developmental stages. A key focus for future study is the neurobiology of endogenous cannabinoids. Establishing the precise brain localization - i.e.,., in which cells and where in those cannabinoids are found, cellular storage and release mechanisms, and uptake mechanisms will be crucial in determining the biological role of this system. Technology that will be crucial in establishing the biological significance of these systems will be broad based and include such research tools as the transgenic, or gene knockout mice, as have already been accomplished for various opioid receptor types.26 In 1997, both CB1 and CB2 receptor knockout mice were generated by a team of scientists at NIH, and a group in France has developed another strain of CB1 in receptor knockout mice.92 --------------------------------------------------------------------------- lIn vitro studies are those in which animal cells or tissue are removed and studied outside the animal; in vivo studies are those in which experiments are conducted in the whole animal.

2.43 ---------------------------------------------------------------------------

Table 2.8 Historical comparisons between cannabinoids and opiates

Comparisons between cannabinoids and opiates . Cannabinoids Opiates Pharmacological Discoveries 1973 (Pert and Snyder, Discovery of receptor 1988 (Howlett and Terenius, and Simon)123, existence Devane)36, 40 149, 162 Identification of 1994 SR141716A receptor antagonist (Rinaldi- Carmona)132 pre- 1973 Naloxone 1992 Anandamide 1975 Met- and Discovery of 1st (Devane And Leu-enkephalin (Hughes et endogenous ligand Mechoulam)37 al)70

1992 (Evans et al. and 1st Receptor cloned 1990 (Matsuda) 107 Kieffer et al.)41, 82

Natural functions of cannabinoid / opiate Unknown Pain, reproduction, mood, systems movement, and others

---------------------------------------------------------------------------

There are several research tools that will greatly aid such investigations - in particular, a greater selection of agonists and antagonists that permit discrimination between the activation of CB1 versus CB2 receptors; hydrophilic agonists (that can be delivered to animals or cells more effectively than hydrophobic compounds). In the area of drug development, future progress should continue to provide more specific agonists and antagonists for CB1 and CB2 receptors, with varying potential for therapeutic uses.

There are certain areas that will provide keys to a better understanding of the potential therapeutic value of cannabinoids. For example, basic biology indicates a role for cannabinoids in pain and control of movement, which is consistent with a possible therapeutic role in these areas. The evidence is relatively strong for the treatment of pain, and intriguingly, although less well-established, for movement disorders. The neuroprotective properties of cannabinoids might prove therapeutically useful, although it should be noted that this is a new area and other, better studied, neuroprotective drugs have not yet been shown to be therapeutically useful. Cannabinoid research is clearly relevant not only to drug abuse, but also to

2.44 ---------------------------------------------------------------------------

understanding basic human biology. Further, it offers the potential for the discovery and development of new, therapeutically useful drugs.

CONCLUSION: At this point, our knowledge about the biology of marijuana and cannabinoids allows us to make some general conclusions:

Cannabinoids likely have a natural role in pain modulation, control of movement, and memory.

The natural role of cannabinoids in immune systems is likely multifaceted and remains unclear.

The brain develops tolerance to cannabinoids.

Animal research demonstrates the potential for dependence, but this potential is observed under a narrower range of conditions than with benzodiazepines, opiates cocaine, or nicotine.

Withdrawal symptoms can be observed in animals, but appear to be mild compared to opiates or benzodiazepines, such as diazepam (Valium ®).

CONCLUSION: The different cannabinoid receptor types found in the body appear to play different roles in normal physiology. In addition, some effects of cannabinoids appear to be independent of those receptors. The variety of mechanisms through which cannabinoids can influence human physiology underlies the variety of potential therapeutic uses for drugs that might act selectively on different cannabinoid systems.

RECOMMENDATION: Research should continue into the physiological effects of synthetic and plant-derived cannabinoids and the natural function of cannabinoids found in the body. Because different cannabinoids appear to have different effects, cannabinoid research should include, but not be restricted to effects attributable to THC alone.

This chapter has summarized recent progress in understanding the basic biology of cannabinoids, and provides a foundation for the next two chapters which review studies on the potential health risks (chapter 3) and benefits of marijuana use (chapter 4).

2.45 ---------------------------------------------------------------------------

References

1. Abood ME, Martin BR. 1996. Molecular neurobiology of the cannabinoid receptor. International Review of Neurobiology 39:197-221.

2. Abood ME, Sauss C, Fan F. Tilton CL, Martin BR. 1993. Development of behavioral tolerance to delta 9- T HC without alteration of cannabinoid receptor binding or mRNA levels in whole brain. Pharmacology, Biochemistry and Behavior 46:575-9.

3. Aceto MD, Scates SM, Lowe JA, Martin BR. 1995. Cannabinoid precipitated withdrawal by the selective cannabinoid receptor antagonist, SR 141716A. European Journal of Pharmacology 282:R1-2.

4. Adams IB, Martin BR. 1996. Cannabis: pharmacology and toxicology in animals and humans. Addiction 91:1555-1614.

5. Baczynsky WO, Zimmerman AM. 1983a. Effects of delta-9-tetrahydrocannabinol, cannabinol, and cannabidiol on the immune system in mice: I. In vivo investigation of the primary ant secondary immune response Pharmocology 26: 1 - 11 .

6. Baczynsky WO, Zimmerman AM. 1983b. Effects of delta 9-tetrahydrocannabinol, cannabinol and cannabidiol on the immune system in mice. II. In vitro investigation using cultured mouse splenocytes. Pharmacology 26; 12-19.

7. Bass R. Engelhard D, Trembovler V, Shohami E. 1996. A novel nonpsychotropic cannabinoid, HU-211, in the treatment of experimental pneumococcal meningitis. Journal of Infectious Diseases 173:735-738.

8. Beardsley PM, Balster RL, Harris LS. 1986. Dependence on tetrahydrocannabinol in rhesus monkeys. Journal of Pharmacology and Experimental Therapeutics 239:311-319.

9. Ben-Shabat S. Fride E, Sheskin T. Tamiri T. Rhee MH, Vogel Z. Bisogno T. De Petrocellis L, Di Marzo V, Mechoulam R. 1998. An entourage effect: inactive endogenous fatty acid glycerol esters enhance 2-arachidonoyl-glycerol cannabinoid activity. European Journal of Pharmacology 353 :23-31.

10. Bennett GJ. 1994. Neuropathic Pain. In: Wall PD, Melzack R Editors Textbook of Pain. Edinburgh: Churchill Livingstone.

11. Bird KD, Boleyn T. Chesher GB, Jackson DM, Starmer GA, Teo RKC. 1980. Intercannabinoid and cannabinoid-ethanol interactions and their effects on human performance. Psychopharmacology 71:181-188.

12. Bloom AS, Dewey WL, Harris LS, Brosius KK. 1977. 9-nor-9b- hydroxyhexahydrocannabinol, a cannabinoid with potent antinociceptive activity: Comparisons with morphine. Journal of Pharmacology and Experimental Therapeutics 200:263-270.

13. Bornheim LM, Everhart ET, Li J. Correia MA. 1994. Induction and genetic regulation of mouse hepatic cytochrome P450 by cannabidiol. Biochemical Pharmacology (England) 48:161-171.

2.46 ---------------------------------------------------------------------------

14. Bornheim LM, Kim KY, Chen B., Correia MA. 1993. The effect of cannabidiol on mouse hepatic microsomal cytochrome P450-dependent anandamide metabolism. Biochemical and Biophysical Research Communications (United States) 197:740746.

15. Bornheim LM, Kim KY, Li J. Perotti BY, Benet LZ. 1995. Effect of cannabidiol pretreatment on the kinetics of tetrahydrocannabinol metabolites in mouse brain. Drug Metabolism and Disposition (United States) 23:825-831.

16. Breivogel CS, Sim LJ, Childers SR. 1997. Regional differences in cannabinoid receptor/G-protein coupling in rat brain. Journal of Pharmacology and Experimental Therapeutics 282:1632-1642.

17. British Medical Association. 1997. Therapeutic uses of cannabis. Amsterdam, The Netherlands: Harwood Academic Publishers.

18. Burkey THC Quock RM, Consroe P. Roeske WR, Yamamura H1. 1997. Delta-9 tetrahydrocannabinol is a partial agonist of cannabinoid receptors in mouse brain. European Journal of Pharmacology 323:R3-R4.

19. Buxbaum DM. 1972. Analgesic activity of D9-tetrahydrocannabinol in the rat and mouse. Psychopharmacology 25:275-280.

20. Cabral G A, Dove Pettit D A. 1998. Drugs and immunity: cannabinoids and their role in decreased resistance to infectious disease Journal of Neuroimmunology 83:116123.

21. Cabral GA, Lockmuller JC, Mishkin EM. 1986. Delta-9-tetrahydrocannabinol decreases alpha/beta interferon response to herpes simplex virus type 2 in the B6C3F1 mouse. Proceedings of the Society for Experimental Biology and Medicine 181 :305-311.

22. Calignano A, La Rana G. Giuffrida A, Piomelli D. 1998. Control of pain initiation by endogenous cannabinoids. Nature 394:277-281.

23. Campbell KA, Foster TC, Hampson RE, Deadwyler SA. 1986a. Delta-9- tetrahydrocannabinol differentially affects sensory-evoked potentials in the rat dentate gyrus. Journal of Pharmacology and Experimental Therapeutics 239:936--940.

24. Campbell KA, Foster TC, Hapson RE, Deadwyler SA. 1986b. Effects of delta-9- tetrahydrocannabinol on sensory-evoked discharges of granule cells in the dentate gyrus of behaving rats. Journal of Pharmacology and Experimental Therapeutics 239:941 -945.

25. Chen J. Marmur R. Pulles A, Paredes W. Gardner EL. 1993. Ventral segmental microinjection of delta-9-tetrahydrocannabinol enhances ventral segmental somatodendritic dopamine levels but not forebrain dopamine levels: evidence for local neural action by marijuana's psychoactive ingredient. Brain Research 621 :6570.

26. Childers SR. 1997. Opioid receptors: Pinning down the opiate targets. Current Biology 7:R695-R697.

2.47 ---------------------------------------------------------------------------

27. Childers SR, Breivogel CS. 1998. Cannabis and endogenous cannabinoid systems. Drug and Alcohol Dependence 51:173-187.

28. Coffey RG, Yamamoto Y. Shella E, Pross S. 1996. tetrahydrocannabinol inhibition of macrophage nitric oxide production. Biochemical Pharmacology 52:743-751.

29. Collins DR, Pertwee RG, Davies SN. 1994. The action of synthetic cannabinoids on the induction of long-term potentiation in the rat hippocampal slice. European Journal of Pharmacology 259:R7-R8.

30. Collins DR, Pertwee RG, Davies SN. 1995. Prevention by the cannabinoid antagonist, SR141716A, of cannabinoid-mediated blockade of long-term potentiation in the rat hippocampal slice. British Journal of Pharmacology 115:869-870.

31. Costa B. Parolaro D, Colleoni M. 1996. Chronic cannabinoid, CP-55,940, administration alters biotransformation in the rat. European Journal of Pharmacology 313: 17-24.

32. Daaka Y. Friedman H. Klein T. 1996. Cannabinoid receptor proteins are increased in Jurkat, human T-Cell line after mitogen activation. Journal of Pharmacology and Experimental Therapeutics 276:776-783.

33. Daaka Y. Zhu W. Friedman H. Klein T. 1997. Induction of interleukin-2 receptor a gene by [Image]9-tetrahydrocannabinol is mediated by nuclear factor kB and CB1 cannabinoid receptor. DNA and Cell Biology 16:301-309.

34. Deadwyler SA, Heyser CJ, Hampson RE. 1995. Complete adaptation to the memory disruptive effects of delta-9-THC following 35 days of exposure. Neuroscience Research Communications 17:9- 18.

35. Derocq JM, Segui M, Marchand J. Le Fur G. Casellas P. 1995. Cannabinoids enhance human B-cell growth at low nanomolar concentrations. FEBS Letters 369:177-182.

36. Devane WA, Dysarc FA, Johnson MR, Melvin LS, Howlett AC. 1988. Determination and characterization of a cannabinoid receptor in rat brain. Molecular Pharmacology 34:605-613.

37. Devane WA, Hanus L, Breuer A, Pertwee RG, Stevenson LA, Griffing F. Gibson D, Mandelbaum A, Etinger A, Mechoulam R. 1992. Isolation and structure of a brain constituent that binds to the cannabinoid receptor. Science 258: 1946- 1949.

38. Dewey WL. 1986. Cannabinoid pharamacology. Pharmacology Review 38:151-178.

39. Di Chiara G. Imperato A. 1988. Drugs abused by humans preferentially increase synaptic dopamine concentrations in the mesolimbic system of freely moving rats. Proceedings of the National Academy of Science, USA 85:5274-5278.

40. Dill JA, Howlett AC. 1988. Regulation of adenylate cyclase by chronic exposure to cannabimimetic drugs. Journal of Pharmacology and Experimental Therapeutics 244:1157-1163.

41. Evans DJ, Keith DEJ, Morrison H. Magendzo K, Edwards RH. 1992. Cloning of a delta opioid receptor by functional expression. Science 258:1952- 1955.

2.48 ---------------------------------------------------------------------------

42. Fan F. Tao Q. Abood ME, Martin BR. 1996. Cannabinoid receptor down-regulation without alteration of the inhibitory effect of CP 55,940 on adenylyl cyclase in the cerebellum of CP 55,940-tolerant mice. Brain Research 706: 13-20.

43. Felder CC, Glass M. 1998. Cannabinoid receptors and their endogenous agonists. Annual Reviews of Pharmacology and Toxicology 38: 179-200.

44. Felder CC, Nielsen A, Briley EM, Palkovits M, Priller J. Axelrod J. Nguyen DN, Richardson JM, Riggin RM, Koppel GA, Paul SM, Becker GW. 1996. Isolation and measurement of the endogenous cannabinoid receptor agonist, anandamide, in brain and peripheral tissues of human and rat. FEBS Letters 393:231 -5.

45. Fields HL. 1987. Pain. New York: McGraw-Hill.

46. Formukong EA, Evans AT, Evans FJ. 1988. Analgesic and anti-inflammatory activity of constituents of Cannabis sativa L. Inflammation 12:361-371.

47. French ED. 1997. Delta-9-tetrabydrocannabinol excites rat VTA dopamine neurons through activation of cannabinoid CB1 but not opioid receptors. Neuroscience Letters 226:159-162.

48. Galiegue S. Mary S. Marchand J. Dussossoy D, Carriere D, Carayon P. Bouaboula M, Shire D, Le Fur G. Casellas P. 1995. Expression of central and peripheral cannabinoid receptors in human immune tissues and leukocyte subpopulations. European Journal of Biochemistry 232:54-61

49. Gessa GL, Mascia MS, Casu MA, Carta G. 1997. Inhibition of hippocampal acetylcholine release by cannabinoids: reversal by SR 141716A. European Journal of Pharmacology 327:R1-2.

50. Gifford AN, Ashby Jr CR. 1996. Electrically evoked acetylcholine release from hippocampal slices is inhibited by the cannabinoid receptor agonist, WIN 55,212-2, and is potentiated by the cannabinoid antagonist, SR 141716A. Journal of Pharmacology and Experimental Therapeutics 277:1431-1436.

51. Gifford AN, Gardner EL, Ashby CRJ. 1997. The effect of intravenous administration of delta-9-tetrahydrocannabinol on the activity of A10 dopamine neurons recorded in vivo in anesthetized rats. Neuropsychobiology 36:96-99.

52. Glass M, Dragunow M, Faull RLM. 1997. Cannabinoid receptors in the human brain: a detailed anatomical and quantitative autoradiographic study in the fetal, neonatal and adult human brain. Neuroscience 77:299-318.

53. Hadden JW, Hadden EM, Haddox MK, Goldberg ND. 1972. Guanosine 3':5'-cyclic monophosphates: A possible intracellular mediator of mitogenic influences in Lymphocytes. Proceedings of the National Academy of Sciences 69:3024-3027.

54. Hampson AJ, Grimaldi M, Axelrod J. Wink D. 1998. Cannabidiol and (-)Delta-9- tetrahydrocannabinol are neuroprotective antioxidants. Proceedings of the National Academy of Science of the United States of America 95:8268-8273.

55. Haney M, Ward AS, Comer SD, Foltin RW, Fischman MW. 1999. Abstinence symptoms following oral THC administration to humans. Psychopharmacology 141:385-394.

2.49 ---------------------------------------------------------------------------

56. Haney M, Ward AS, Comer SD, Foltin RW, Fischman MW. 1999. Abstinence symptoms following smoked marijuana in humans. Psychopharmacology 141 :395-404.

57. Herkenham M. 1995. Localization of cannabinoid receptors in brain and periphery. In: Pertwee RG ed. Cannabinoid Receptors. Academic Press Limited. Pp. 145-166.

58. Herkenham M, Lynn AB, de Costa BR, Richfield EK. 1991a. Neuronal localization of cannabinoid receptors in the basal ganglia of the rat. Brain Research 547:267-274.

59. Herkenham M, Lynn AB, Johnson MR, Melvin LS, de Costa BR, Rice KC. 1991b. Characterization and localization of cannabinoid receptors in rat brain: a quantative in vitro autoradiographic study. Journal of Neuroscience 11 :563-583.

60. Herkenham M, Lynn AB, Little MD, Johnson MR, Melvin LS, de Costa BR, Rice KC. 1990. Cannabinoid receptor localization in the brain. Proceedings of the National Academy of Sciences of the United States of America 87: 1932- 1936.

61. Herring AC, Koh WS, Kaminski NE. 1998. Inhibition of the cyclic AMP signaling cascade and nuclear factor binding to CRE and kappa B elements by cannabinol, a minimally CNS-active cannabinoid. Biochemical Pharmacology 55:1013-1023.

62. Henberg U. Eliav E, Bennett GJ, Kopin IJ. 1997. The analgesic effects of R(+)-WIN 55,212-2 mesylate, a high affinity cannabinoid agonist, in a rat model of neuropathic pain. Neuroscience Letters 221: 157-60.

63. Heyser CJ, Hampson RE, Deadwyler SA. 1993. Effects of delta-9-tetrahydrocannabinol on delayed match to sample performance in rats: Alterations in short-term memory associated with changes in task specific firing of hippocampal cells. Journal of Pharmacology and Experimental Therapeutics 264:294-307.

64. Hohmann AG, Briley E M, Herkenham M. in press (1998). Pre- and postsynaptic distribution of cannabinoid and mu opioid receptors in rat spinal cord. Brain Research

65. Hohmann AG, Herkenham M. 1998. Regulation of cannabinoid and mu opioid receptor binding sites following neonatal capsaicin treatment. Neuroscience Letters 252: 1316.

66. Hohmann AG, Herkenham M. in press (1998). Localization of central cannabinoid CB1 receptor mRNA in neuronal subpopulations of rat dorsal root ganglia: A double- label in situ hybridization study. Neuroscience

67. Hohmann AG, Martin WJ, Tsou K, Walker JM 1995 Inhibition of noxious stimulus-evoked activity of spinal cord dorsal horn neurons by the cannabinoid WIN 55,212-2. Life Sciences 56:2111-2119.

68. Hollister LE. 1986. Health aspects of cannabis. Pharmacological Reviews 38:1-20.

69. Hollister LE, Gillespie BA. 1975. Interactions in man of delta-9-THC. II. Cannabinol and cannabidiol. Clinical Pharmacology and Therapeutics 18:80-83.

2.50 ---------------------------------------------------------------------------

70. Hughes J. Smith TW, Kosterlitz HW, Fothergill LA, Morgan BA, Morris HR. 1975. Identification of two related pentapeptides from the brain with potent opiate agonists activity. Nature 258:577-580.

71. Jackson AL, Murphy LL. 1997. Role of the hypothalamic-pituitary-adrenal axis in the suppression of luteinizing hormone release by delta-9-tetrahydrocannabinol. Neuroendocrinology 65:446-452.

72. Jacob J. Rarnabadran K, Campos-Medeiros M. 1981. A pharmacological analysis of levonantradol antinociception in mice. Journal of Clinical Pharmacology 21 :327S-333S.

73. Jaggar SI, Hasnie FS, Sellaturay S. Rice AS. 1998. The anti-hyperalgesic actions of the cannabinoid anandamide and the putative CB2 receptor agonist palmitoylethanolamide in visceral and somatic inflammatory pain. Pain 76:189199.

74. Jeon YJ, Yang K-H, Pulaski JT, Kaminski NE. 1996. Attenuation of inducible nitric oxide synthase gene expression by delta-9-tetrahydrocannabinol is mediated through the inhibition of nuclear factor-kB/Rel activation. Molecular Pharmacology 50:334341.

75. Johnson MR, Melvin LS. 1986. The discovery of non-classical cannabinoid analgesics. In: Mechoulam R editor Cannabinoids as therapeutic agents. Boca Raton, FL: CRC Press, Inc. Pp. 121-145.

76. Jones RT, Benowitz NL, Herning RI. 1981. Clinical relevance of cannabis tolerance and dependence. Journal of Clinical Pharmacology 21: 143S-152S.

77. Kaminski NE. 1996. Immune regulation by cannabinoid compounds through the inhibition of the cyclic AMP signaling cascade and altered gene expression. Biochemical Pharmacology 52:1133-1140.

78. Kaminski NE, Abood M E, Kessler F K, Martin B R. Schatz A R. 1992. ldentification of a functionally relevant cannabinoid receptor on mouse spleen cells that is involved in cannabinoid-mediated immune modulation. Molecular Pharmacology 42:736-742.

79. Kaminski NE, Koh WS, Yang KH, Lee M, Kessler FK. 1994 Suppression of the humoral immune response by cannabinoids is partially mediated through inhibition of adenylate cyclase by a pertussis toxin-sensitive G-protein coupled mechanism. Biochemical Pharmacology 48:1899-1908.

80. Kaminski NE. 1998. Regulation of cAMP cascade, gene expression and immune function by cannabinoid receptors. Journal of Neuroimmunology 83: 124- 132

81. Karniol IG, Shirakawa I, Kasinski N. Pfeferman A, Carlini EA. 1975. Cannabidiol interferes with the effects of delta-9-tetrahydrocannbinol in man. European Journal of Pharmacology 28:172-177.

82. Kieffer BL, Befort K, Gaveriaux-Ruff C, Hirth CG. 1992. The delta-opioid receptor: isolation of a cDNA by expression clining and pharmacological characterization. Proceedings of the National Academy of Sciences of the United States of America 89: 12048-12052.

2.51 ---------------------------------------------------------------------------

83. Kirby MT, Hampson RE, Deadwyler SA. 1995. Cannabinoids selectively decrease paired pulse perforant path synaptic potentials in the dentate gyrus in vitro. Brain Research 688:114-120.

84. Klein TW, Friedman H. 1990. Modulation of murine immune cell function by marijuana components. In: Watson R ed. Drugs of Abuse and immune Function. Boca Raton, FL: CRC Press.

85. Klein TW, Friedman H. Specter SC. 1998. Marijuana, immunity and infection. Journal of Neuroimmunology 83: 102- 115.

86. Klein TW, Newton C, Friedman H. 1987. Inhibition of natural killer cell function by marijuana components. Journal of Toxicology and Environmental Health 20:321-332.

87. Klein TW, Newton C, Friedman H. 1994. Resistance to Legionella pneumophila suppressed by the marijuana component, tetrahydrocannabinol Journal of Infectious Diseases 169:1177-1179.

88. Klein TW, Newton C, Friedman H. 1998. Cannabinoid receptors and immunity. Immunology Today 19:373-381.

89. Klein TW, Newton C, Widen R. Friedman H. l985 The effect of delta-9 tetrahydrocannabinol and 11-hydroxy-delta-9-tetrahydrocannabinol on T-lymphocyte and B-lymphocyte mitogen responses. Journal of Immunopharmacology 7:451-466.

90. Klein TW, Newton C, Widen R. Friedman H. 1993. Delta-9-tetrahydrocannabinol injection induces cytokine-mediated mortality of mice infected with Legionella pneumophila. Journal of Pharmacology and Experimental Therapeutics 267:635-640.

91. Koh WS, Yang KH, Kaminski NE. 1995. Cyclic AMP is an essential factor in immune responses. Biochemical and Biophysical Research Communications 206:703-709.

92. Ledent C, Valverde O. Cossu G. Petitet F. Aubert JF, Beslot F in Bohme GA, Imperato A, Pedrazzini T. Roques BP, Vassart G. Fratta W. Parmentier M. 1999. Unresponsiveness to cannabinoids and reduced addictive effects of opiates in CB1 receptor knockout mice. Science 283:401 -4 .

93. Lee M, Yang KH, Kaminski NE. 1995. Effects of putative cannabinoid receptor ligands, anandamide and 2-arachidonyl-glycerol, on immune function in B6C3F1 mouse splenocytes. Journal of Pharmacology and Experimental Therupeutics 275:529536.

94. Lepore M, Liu X, Savage V, Matalon D, Gardner EL. 1996. Genetic differences in delta 9- tetrahydrocannabinol-induced facilitation of brain stimulation reward as measured by a rate-frequency curve-shift electrical brain stimulation paradigm in three different rat strains. Life Sciences 58:365-372

95. Lepore M, Vorel SR, Lowinson J. Gardner EL. 1995. Conditioned place preference induced by delta 9-tetrahydrocannabinol: Comparison with cocaine, morphine, and food reward. Life Sciences 56:2073-2080.

2.52

96. Lichtman AH, Martin BR. 1991a. Spinal and supraspinal components of cannabinoid-induced antinociception. Journal of Pharmacology and Experimental Therapeutics 258:517-523.

97. Little PJ, Compton DR, Mechoulam R. Martin BR. 1989. Stereochemical effects of 11-OH- delta-8-THC-dimethylheptyl in mice and dogs. Pharmacology Biochemistry Behavior 32:661-666.

98. Lu F. Ou DW. 1989. Cocaine or delta-9-tetrahydrocannabinol does not affect cellular cytotoxicity in vitro. International Journal of Pharmacology 11: 849-852.

99. Luo YD, Patel MK, Wiederhold MD, Ou DW. 1992. Effects of cannabinoids and cocaine on the mitogen-induced transformations of lymphocytes of human and mouse origins. International Journal of lmmunopharmacology 14:49-56.

100. Lyman WD, Sonett JR, Brosnan CFER, Bornstein MB. 1989. Delta 9-tetrahydrocannabinol: a novel treatment fur experimental autoimmune encephalomyelitis. Journal of Neuroimmunology 23:73-81.

101. Mailleux P. Vanderhaeghen JJ. 1992. Distribution of neuronal cannabinoid receptor in the adult rat brain: a comparative receptor binding radioautography and in situ hybridization histochemistry. Neuroscience 48:655-668.

102. Martin WJ, Hohmann AG, Walker JM. 1996. Suppression of noxious stimulus-evoked activity in the ventral posterolateral nucleus of the thalamus by a cannabinoid agonist: Correlation between electrophysiological and antinociceptive effects. The Journal of Neuroscience 16:6601 -6611.

103. Martin WJ, Patrick SL, Coffin PO, Tsou K, Walker JM. 1995. An examination of the central sites of action of cannabinoid-induced antinociception in the rat. Life Sciences 56:2103-2109.

104. Martin W J. Patrick S L, Coffin PO, Tsou K, Walker J M. 1995. An examination of the central sites of action of cannabinoid-induced antinociception in the rat. Life Sciences 56:2103-2109.

105. Martin WJ, Tsou K, Walker JM. 1998. Cannabinoid receptor-mediated inhibition of the rat tail-flick reflex after microinjections into the rostral ventromedial medulla. Neuroscience Letters 242:33-36.

106. Martoletta MC, Cossu G. Fattore L, Gessa GL, Fratta W. 1998. Self-administration of the cannabinoid receptor agonist WIN 55,212-2 in drug-naive mice. Neuroscience 85 :327-330.

107. Matsuda L, Lolait SJ, Brownstein MJ, Young AC, Bonner TI. 1990. Structure of a cannabinoid receptor and functional expression of the cloned cDNA. Nature 346:561 -564.

108. Mechoulam R. Ben-Shabat S. Hanus L, Ligumsky M, Kaminski NSA, Gopher A, Almog S. Martin BR, Compton D, Pertwee RG, Griffin G. Bayewitch M, Barg in Vogel Z. 1995. Identification of an endogenous 2-monoglyceride, present in canine gut, that binds to cannabinoid receptors. Biochemical Pharmacology 50:83-90.

2.53

109. Mechoulam R. Hanus L, Fride E. 1998. Towards cannabinoid drugs - revisited. In: Ellis GP, Luscombe DK, Oxford AW eds. Progress in Medicinal Chemistry. v. 35. Amsterdam: Elsevier Science. Pp. 199-243.

110. Meng ID, Manning BH, Martin WJ, Fields HL. 1998. An analgesia circuit activated by cannabinoids. Nature 395:381-383.

111. Miller AS, Walker JM. 1996. Electrophysiological effects of a cannabinoid on neural activity in the globus pallidus. European Journal of Pharmacology 304:29-35.

112. Munro S. Thomas KL, Abu-Shaar M. 1993. Molecular characterization of a peripheral receptor for cannabinoids. Nature 365:61-65.

113. Murphy LL, Steger RW, Smith MS, Bartke A. 1990. Effects of delta-9- tetrahydrocannabinol cannabinol and cannabidiol, alone and in combinations, on luteinizing hormone and prolactin release and on hypothalamic neurotransmitters in the male rat. Neuroendocrinology 52:316-321.

1l4. Narimatsu S. Watanabe K, Matsunaga T., Yamamoto I, Imaoka S. Funae Y. Yoshimura H. 1993. Suppression of liver microsomal drug-metabolizing enzyme activities in adult female rats pretreated with cannabidiol Biological and Pharmaceutical Bulletin (Japan) 16:428-430.

115. Newton CA, Klein T. Friedman H. 1994. Secondary immunity to Legionella pneumophila and Th1 activity are suppressed by delta-9-tetrabydrocannabinol injection. Infection and immunity 62:4015-4020.

116. Norwicky AV, Teyler TJ, Vardaris RM. 1987. The modulation of long-term potentiation by delta-9-tetrahydrocannabinol in the rat hippocampus, in vitro. Brain Research Bulletin 19:663.

117. O'Leary D, Block RI, Flaum M, Boles Ponto LL, Watkins GL, Hichwa RD. 1998. Acute marijuana effects on rCBF and cognition: A PET study. Abstracts - Society for Neuroscience: 28th Annual Meeting. Los Angeles, CA, November 7-12, l998. Washington, DC: Society for Neuroscience.

118. Ohlsson A, Lindgren J-E, Wahlen A, Agurell S. Hollister L E, Gillespie HK. 1980. Plasma delta-9-tetrahydrocannabinol concentrations and clinical effects after oral and intravenous administration and smoking. Clinical Pharmacology and Therapeutics 28:409-416.

119. Oviedo A, Glowa J. Herkenham M. 1993. Chronic cannabinoid administration alters cannabinoid receptor binding in rat brain: a quantitative autoradiographic study. Brain Research 616:293-302

120. Pacheco MA, Ward SJ, Childers SR. 1993. Identification of cannabinoid receptors in cultures of rat cerebellar granule cells. Brain Research 603: 102- 110.

121. Patel V, Borysenko M, Kumar MSA, Millard WJ. 1985. Effects of acute and subchronic delta-9-tetrahydrocannabinol administration on the plasma catecholamine, beta- endorphin, and corticosterone levels and splenic natural killer cell activity in rats. Proceedings of the Society for Experimental Biology and Medicine 180:400-404.

2.54 ---------------------------------------------------------------------------

122. Pepe S. Ruggiero A, Tortora G. Ciaardiello F. Garbi C, Yokozaki H. Cho-Chung YS, Clair T. Skalhegg BS, Bianco AR. 1994. Flow cytometric detection of the RT alpha subunit of type-I cAMP-dependent protein kinase in human cells. Cytometry 15:7379.

123. Pert CB, Snyder SH. 1973. Opiate receptor: demonstration in nervous tissue. Science 179:1011-1014.

124. Pertwee RG. 1997b. Pharmacology of cannabinoid CB1 and CB2 receptors. Pharmacology and Therapeutics 74: 129-180.

125. Pertwee RG, Stevenson LA, Griffin G. 1993. Cross-tolerance between delta-9- tetrahydrocannabinol and the cannabimimetic agents, CP 55,940, WIN 55,212-2 and anandamide [published erratum appears in British Journal of Pharmacology 1994 Mar; 111(3):968]. British Journal of Pharmacology l10:1483-90.

126. Pertwee RG, Wickens AP. 1991. Enhancement by chlordiazepoxide of catalepsy induced in rats by intravenous or intrapallidal injections of enantiomeric cannabinoids. Neuropharmacology 30:237-244.

127. Pross SH, Nakano Y. Widen R. McHugh S. Newton C, Klein TW, Friedman H. 1992. Differing effects of delta-9-tetrahydrocannabinol (THC) on murine spleen cell populations dependent upon stimulators International Journal of Immunopharmacology 14:1019- 1027.

128. Razdan RK. 1986. Structure-activity relationships in cannabinoids. Pharmacology Review 38 75-149.

129. Rhee MH, Vogel Z. Barg J. Bayewitch M, Levy R. Hanus L, Breuer A, Mechoulam R. 1997. Cannabinol derivatives: binding to cannabinoid receptors and inhibition of adenylcyclase. Journal of Medicinal Chemistry 40:3228-3233.

130. Richardson JD, Aanonsen L, Hargreaves KM. 1998. Hypoactivity of the spinal cannabinoid system results in NMDA-dependent hyperalgesia. Journal of Neuroscience 18:451--457.

131. Richardson JD, Kilo S. Hargreaves KM. 1998. Cannabinoids reduce hyperalgesia and inflammation via interaction with peripheral CB1 receptors. Pain 75:111 - 119.

132. Rinaldi-Carmona M, Barth F. Heaulme M, Shire D, Calandra B. Congy C, Martinez S. Maruani J. Neliat G. Caput D, Ferrara P. Soubrie P. Breliere JC, Le Fur G. 1994. SR141716A, a potent and selective antagonist of the brain cannabinoid receptor. FEBS Letters 350:240-244.

133. Rinaldi-Carmona M, Barth F. Millan J. Defrocq J. Casellas P. Congy C, Oustric D, Sarran M, Bouaboula M, Calandra B. Portier M, Shire D, Breliere J. Le Fur G. 1998. SR144528, the first potent and selective antagonist of the CB2 cannabinoid receptor. Journal of Pharmacology and Experimental Therapeutics 284:644-650.

134. Rodriguez de Fonseca F. Fernandez-Ruiz JJ, Murphy LL, Eldridge JC, Steger RW, Bartke A. 1991. Effects of delta-9-tetrahydrocannabinol exposure on adrenal medullary function: evidence of an acute effect and development of tolerance in chronic treatments. Pharmacology, Biochemistry and Behavior 40:593-598

2.55 ---------------------------------------------------------------------------

135. Rodriguez de Fonseca F. Carrera MRA, Navarro M, Koob G. Weiss F. 1997. Activation of corticotropin-releasing factor in the limbic system during cannabinoid withdrawal [see comments Science 1997. 276:1967-1968]. Science 276:2050-2054.

136. Rodriguez de Fonseca F. Gorriti MA, Fernandez-Ruiz JJ, Palomo T. Ramos JA. 1994. Down-regulation of rat brain cannabinoid binding sites after chronic delta-9- tetrahydrocannabinol treatment. Pharmacology, Biochemistry and Behavior 47:33--40.

137. Romero J. Garcia L, Fernandez-Ruiz JJ, Cebeira M, Ramos JA 1995 Changes in rat brain cannabinoid binding sites after acute or chronic exposure to their endogenous agonist, anandamide, or to delta-9-tetrahydrocannabinol. Pharmacology, Biochemistry and Behavior 51:731 -737.

138. Romero J. Garcia-Palomero E, Castro JG, Garcia-Gil L, Ramos JA, Fernandez-Ruiz JJ. 1997. Effects of chronic exposure to delta-9-tetrahydrocannabinol on cannabinoid receptor binding and mRNA levels in several rat brain regions. Molecular Brain Research 46: 100-8.

139. Russell DH. 1978. Type I cyclic AMP-dependent protein kinase as a positive effector of growth. Advances in Cyclic Nucleotide Research 9:493-506.

140. Sanudo-Pena M C, Tsou K, Delay E R. Hohmann AG, Force M, Walker JM. 1997. Endogenous cannabinoids as an aversive or counter-rewarding system in the rat. Neuroscience Letters 223: 125- 128.

141. Sanudo-Pena MC, Walker JM. 1997. Role of the subthalamic nucleus in cannabinoid actions in the substantia nigra of the rat. Journal of Neurophysiology 77: 1635-1638.

142. Schatz AR, Koh WS, Kaminski, NE. 1993. Delta-9-tetrahydrocannabinol selectively inhibits T-cell dependent humoral immune responses through direct inhibition of accessory T-cell function. Immunopharmacology 26: 129- 137.

143. Schlicker E, Timm J. Zenter J. Goethert M. 1997. Cannabinoid CB1 receptor-mediated inhibition of noradrenaline release in the human and guinea-pig hippocampus. Naunyn-Schmiedeberg's Archives of Pharmacology 356:583-589.

144. Shen M, Piser TM, Seybold VS, Thayer SA. 1996. Cannabinoid receptor agonists inhibit glutamatergic synaptic transmission in rat hippocampal cultures. Journal of Neuroscience 16:4322-4334.

145. Shivers S C, Newton C, Friedman H. Klein TW. 1994. Delta 9-tetrahydrocannabinol (THC) modulates IL-1 bioactivity in human monocyte/macrophage cell lines. Life Sciences 54: 1281-1289.

146. Shohami E, Gallily R. Mechoulam R. Bass R. Ben-Hur T. 1997. Cytokine production in the brain following closed head injury: dexanabinol (HU-211) is a novel TNF-alpha inhibitor and an effective neuroprotectant. Journal of Neuroimmunology 72: 169--177.

147. Sim LJ, Hampson RE, Deadwyler SA, Childers SR. 1996. Effects of chronic treatment with delta-9-tetrahydrocannabinol on cannabinoid-stimulated [35S]GTPyS autoradiography in rat brain. Journal of Neuroscience 16:8057-8066.

2.56 ---------------------------------------------------------------------------

148. Sim LJ, Xiao R. Selley DE, Childers SR. 1996. Differences in G-protein activation by mu- and delta-opioid, and cannabinoid, receptors in rat striatum. European Journal of Pharmacology 307:97-105.

149. Simon EJ. 1973. In search of the opiate receptor. American Journal of Medical Sciences 266: 160- 168.

150. Skaper SD, Buriani A, Dal Toso R. Petrelli L, Romanello S. Facci L, Leon A. 1996. The ALIAmide palmitoylethanolamide and cannabinoids, but not anandamide, are protective in a delayed postglutamate paradigm of excitotoxic death in cerebellar granule neurons. Proceedings of the National Academy of Sciences of the United States of America 93:3984-9.

151. Smith JW, Steiner AL, Newberry WM, Parker CW. 1971. Cyclic adenosine 3',5'- monophosphate in human lymphocytes: Alteration after phytohemagglutinin. Journal of Clinical Investigation 50:432-441.

152. Smith PB, Compton DR, Welch SP, Razdan RK, Mechoulam R. Martin BR. 1994. The pharmacological activity of anandamide, a putative endogenous cannabinoid, in mice. Journal of Pharmacology and Experimental Therapeutics 270:219-227.

153. Smith PB, Welch SP, Martin BR. 1994. Interactions between delta 9-tetrahydrocannabinol and kappa opioids in mice. Journal of Pharmacology and Experimental Therapeutics 268:1381-1387.

154. Sofia RD, Nalepa SD, Harakal JJ, Vassar HB. 1973. Anti-edema and analgesic properties of delta-9-tetrahydrocannabinol (THC). Journal of Pharmacology and Experimental Therapeutics 186:646-655.

155. Specter S. Lancz C, Hazelden J. 1990. Marijuana and immunity: tetrahydrocannabinol mediated inhibition of lymphocyte blastogenesis. International Journal of Immunopharmacology 12;261-267.

156. Stefano G. Salzet B. Salzet M. 1997. Identification and characterization of the leech CNS cannabinoid receptor: Coupling to nitric oxide release. Brain Research 753:219--224.

157. Stella N. Schweitzer P. Piomelli D. 1997. A second endogenous cannabinoid that modulates long term potentiation. Nature 388:773-778.

158. Strangman NM, Patrick SL, Hohmann AG, Tsou K, Walker JM. 1998. Evidence for a role of endogenous cannabinoids in the modulation of acute and tonic pain sensitivity. Brain Research 813:. 323-328.

159. Sulcova C, Mechoulam R., Fride E. 1998. Biphasic effects of anandamide. Pharmacology, Biochemistry and Behavior 59:347-352.

160. Szabo B. Dorner L, Pfreundtner C, Norenberg W. Starke K. 1998. Inhibition of GABAergic inhibitory postsynaptic currents by cannabinoids in rat corpus straitum Neuroscience 85:395-403.

161. Tanda G. Pontieri FE, Di Chiara G. 1997. Cannabinoid and heroin activation of mesolimbic dopamine transmission by a common µ1 opioid receptor mechanism. Science 276:2048-2049.

2.57 ---------------------------------------------------------------------------

162. Terenius L. 1973. Characteristics of the "receptor" for narcotic analgesics in synaptic plasma membrane fraction from rat brain. Acta Pharmacologica Et Toxicologica 33:377-384.

163. Terranova JP, Michaud JC, Le Fur G. Soubrie P. 1995. Inhibition of long-term potentiation in rat hippocampal slice by anandamide and WIN 55.212-2: reversal by SR141716 A, a selective antagonist of CB1 cannabinoid receptors. Naunyn-Schmiedeberg's Archives of Pharmacology 352:576-579.

164. Titishov N. Mechoulam R. Zimmerman AM. 1989. Stereospecific effects of (-) and (+)-7- hydroxy-delta-6-tetrahydrocannabinol-dimethylheptyl on the immune system of mice. Pharmacology 39:337-349.

165. Tsou K, Brown S. Sanudo-Pena MC, Mackie K, Walker JM. 1998. Immunohistochemical distribution of cannabinoid CB1 receptors in the rat central nervous system. Neuroscience 83:393-411.

166. Tsou K, Patrick SL, Walker JM. 1995. Physical withdrawal in rats tolerant to delta-9- tetrahydrocannabinol precipitated by a cannabinoid receptor antagonist. European Journal of Pharmacology 280:R13-R15.

167. Watson PF, Krupinski J. Kempinski A, Frankenfield C. 1994. Molecular cloning and characterization of the type VII isoform of mammalian adenylyl cyclase expressed widely in mouse tissues and in S49 mouse Iymphoma cells. Journal of Biological Chemistry 269:28893-28898.

168. Watzl B. Scuder P. Watson RR. 1991. Marijuana components stimulate human peripheral blood mononuclear cell secretion of interferon-gamma and suppress interleukin-1 alpha in vitro. International Journal of Immunopharmology 13:1091-1097.

169. Weidenfeld J. Feldman S. Mechoulam R. 1994. Effect of the brain constituent anandamide, a cannabinoid receptor agonist, on the hypothalamo-pituitary-adrenal axis in the rat. Neuroendocrinology 59:110-112.

170. Welch SP. 1993. Blockade of cannabinoid-induced antinociception by norbinaltorphimine, but not N,N-diallyl-tyrosine-Aib-phenylalanine-leucine, ICI 174,864 or naloxone in mice. Journal of Pharmacology and Experimental Therapeutics 265:633-640.

171. Welch SP, Thomas C, Patrick GS. 1995. Modulation of cannabinoid-induced antinociception after intracerebroventricular versus intrathecal administration to mice: Possible mechanisms for interaction with morphine. Journal of Clinical and Experimental Therapeutics 272:310-321.

172. Wirguin I, Mechoulam R. Breuer A, Schezen E, Weidenfeld J. Brenner T. 1994. Suppression of experimental autoimmune encephalomyelitis by cannabinoids. Immunopharmacology 28 :209-214.

173. Wirth PW, Watson ES, ElSohly M, Turner CE, Murphy JC. 1980. Anti-inflammatory properties of cannabichromene. Life Sciences 26:1991-1995.

174. Yaksh TL. 1981. The antinociceptive effects of intrathecally administered levonantradol and desacetyl-levonantradol in the rat. Journal of Clinical Pharmacology 21 :334S--340S.

2.58 ---------------------------------------------------------------------------

175. Yoshida H. Usami N. Ohishi Y. Watanabe K, Yamamoto I, Yoshimura H. 1995. Synthesis and pharmacological effects in mice of halogenated cannabinol derivatives. Chemical and Pharmaceutical Bulletin 42:335-337.

176. Zhu W. Newton C, Daaka Y. Friedman H. Klein TW. 1994. Delta 9-tetrahydrocannabinol enhances the secretion of interleukin I from endotoxin-stimulated macrophages. Journal of Pharmacology and Experimental Therapeutics 270: 1334-1339.

177. Zuardi AW, Shirakawa I, Finkelfarb E, Karniol IG. 1982. Action of cannabidiol on the anxiety and other effects produced by delta 9-THC in normal subjects. Psychopharmacology (Berl) 76:245-250.

178. Zurier R.B., Rossetti RG, Lane JH, Goldberg JM, Hunter SA, Burstein SH 1998. Dimethylheptyl-THC- 11 oic acid: A non-psychoactive antiinflmnmatory agent with a cannabinoid template structure. Arthritis and Rheumatism 41:163- 170.

2.59 ---------------------------------------------------------------------------

Chapter 3 ==>>

Please visit the web site for the remaining chapters.