Jenny L. Wiley, PhD - US grants

The International Cannabinoid Research Society (ICRS) is an international, not-for-profit professional scientific society incorporated in the United States in 1992. The annual ICRS Symposium on the annabinoids attracts a wide range of cannabinoid researchers with expertise ranging from molecular approaches in ligand design to human studies on the psychological effects of cannabis use. The format of ICRS Symposia has been designed to promote open communication and to foster collaborations among cannabinoid researchers. The 4-day conference provides an opportunity for all researchers (predoctoral, postdoctoral, young investigators, established investigators) to communicate recent important findings in an oral or poster presentation. The next ICRS meeting will take place in Tihany, Hungary in June 2006. This proposal is a competing continuation application for R13 support of the annual ICRS Symposia. The long-term goal of this project is to increase the number and diversity of cannabinoid researchers by recruiting and retaining young scientists trained in the cannabinoid field. To this end, we propose three specific aims: 1) The ICRS will sponsor a travel award grant program that will be used to increase participation of young cannabinoid researchers. Emphasis will be placed on support for students and young investigators from US racial and ethnic minorities. 2) The ICRS will sponsor a special session for young scientists at each annual Symposium. These sessions will focus on providing information on career options for students and young investigators that will allow continuation of their cannabinoid research, including information on programs that provide funding for pre- and postdoctoral students and young investigators, panel presentations on research in academic and government settings, and workshops on grant writing. 3) The ICRS will recognize outstanding predoctoral and postdoctoral student presentations. Given that marijuana abuse continues to be a significant problem in the U.S., it is crucial that research on the biological effects of cannabinoids continue. In addition, increasing evidence suggests that the endocannabinoid system (which may be disrupted by abuse of marijuana and other drugs of abuse) is involved in many diverse physiological functions. The goal of this R13 project is to encourage young scientists to continue research into this important area.

DESCRIPTION (provided by applicant): The endogenous cannabinoid system has been implicated in diverse biological processes as well as pathophysiologies including cachexia, obesity, dyslipidemia, nausea/vomiting, chronic pain, inflammation, drug dependence, neuropsychiatric disorders, multiple sclerosis, and cancer. The major guiding hypothesis of this project is that the complex physiological and pharmacological properties of the endocannabinoid system are due to both direct and indirect activation/inactivation of CB1, CB2 and as-yet-unidentified (CBx) receptors. To determine the contributions of each receptor/binding site to the physiological and pathological processes mediated by the endocannabinoid system, the proposed series of targeted experiments will evaluate the effects of receptor-selective ligands in animal models of specific disorders in which the endocannabinoid system has been implicated (e.g., substance abuse, psychiatric disorders, obesity and pain). Complementary in vitro studies will be used to verify selectivity of the ligands, as well as to investigate mechanisms involved in their in vivo effects. To date, most in vivo research has focused on delineation of cannabinoid effects mediated by orthosteric interaction with CB1 receptors. However, increasing evidence suggests that cannabinoid action at CB2 and CBx receptors, as well as allosteric modulation of CB1 receptors, may produce pharmacological profiles that are distinctly different from those produced by orthosteric CB1 agonists and antagonists. Hence, the proposed research will focus primarily on the effects of ligands that act via mechanisms other than the orthosteric site of the CB1 receptor. Aim 1 will focus on characterization of a novel class of 3-substituted pyrazole rimonabant analogs with in vivo effects mediated by non-CB1, non-CB2 mechanism(s). If these ligands activate a novel cannabinoid (CBx) receptor (as is hypothesized), delineation of their behavioral effects will not only determine potential therapeutic indications of the compounds, but also may suggest possible physiological roles of this receptor system. Aim 2 will examine the behavioral effects of CB2 receptor selective agonists in animal models of stimulant abuse and schizophrenia, disorders for which recent research has implicated CB2 receptor involvement. In Aim 3, allosteric modulators of the CB1 and CB2 receptors will be developed. Hypotheses to be examined in this aim are that allosteric modulators are likely to exhibit greater receptor selectivity and that they will have a side effect profile that lacks the marijuana-like intoxicatio effects of orthosteric CB1 receptor agonists. Whereas Aims 1-3 focus on the acute effects of cannabinoids, Aim 4 will examine the effects of sub-chronic administration, an important consideration since most therapeutic indications for which cannabinoids are being developed are chronic conditions. Increasing scientific knowledge of the contributory roles of different receptor mechanisms involved in endocannabinoid system functioning could enhance understanding and treatment of a wide variety of disorders in which dysregulation of the endocannabinoid system has been implicated, including substance abuse, psychiatric disorders, obesity, and various neurological disorders.

DESCRIPTION (provided by applicant): The endocannabinoid (eCB) system is one of several lipid signaling systems in the brain and in the body. Verified components of this system include two G-protein coupled receptors, their signaling pathways, two predominant endogenous ligands [anandamide (AEA) and 2-arachidonyl glycerol (2-AG)], and their synthetic and metabolic pathways. The system plays an important modulatory role in many crucial CNS processes (e.g., brain reward, appetite regulation, cognition). Consequently, it is not surprising that this system has been implicated in the pathophysiology of a variety of health problems related to these processes, including substance abuse, eating disorders, other types of addictive behavior, and psychiatric disorders. Although research in cells or tissues suggest that there are differences between AEA and 2-AG, examination of potential behavioral consequences of these differences is sparse. Yet, the health problems, for which dysregulation of eCBs in the CNS is most strongly implicated, are problems in which behavior is central. Hence, one of the first steps towards delineation of physiological role(s) that AEA and/or 2-AG may play in health problems such as substance abuse is to distinguish similarities and differences in effects of these two eCBs in pharmacologically selective and validated behavioral procedures relevant to cannabinoid abuse. To this end, two mouse models, drug discrimination and intracranial self-stimulation (ICSS), will be used (Aims 1 and 2). Drug discrimination is an animal model of the subjective effects of psychoactive drugs in humans whereas ICSS represents a method used to evaluate the effects of drugs and behavioral or genetic manipulations on brain reward processes. Each of these factors is known to play a strong role in substance abuse. In addition, brain reward processes undoubtedly are involved in other forms of addictive behavior such as binge eating. The primary guiding idea underlying the proposed studies is that finer distinctions among functions of individual eCBs will be facilitated by knowing the extent to which their behavioral endpoints differ. Further, selected pharmacodynamic mechanisms that may be responsible for differences in the behavioral profiles of these eCBs will be examined (Aim 3). Namely, the relative efficacies and potencies of AEA and 2-AG at a level signal transduction that is a proximal to the ligand-receptor interaction (G-protein activation) will be determined, as the nature of this interaction is associated with alterations in behavioral responses. Results of the proposed studies will enhance understanding of how the eCB system (and especially each of the two major eCBs, AEA and 2-AG) is involved in physiological and pathophysiological processes related to substance abuse. This knowledge, combined with the current rapid development of pharmacological tools to manipulate this system (e.g., inhibitors of eCB synthesis and metabolism), also has the potential to lead to more effective therapeutic agents for health problems related to dysregulation of the eCB system.

DESCRIPTION (provided by applicant): Smoking of cannabinoid-enhanced spices is an emerging substance abuse problem. Although JWH-018 and other indole-derived cannabinoids that have been identified in Spice products produce marijuana-like intoxication, they are structurally distinct from cannabinoids contained in the cannabis plant; hence, in many states, they are not currently illegal and are readily available over the internet and in headshops. Indole-derived cannabinoids were originally developed as research tools to investigate the nature of cannabinoid receptors; however, very little is known about their pharmacology, particularly their behavioral effects, as only a small sample of the over 200 compounds in the JWH series have been tested in preclinical in vivo models. The proposed research will synthesize and evaluate JWH-018 and related indole-derived cannabinoids in rodent models that have been used to investigate the abuse properties of cannabinoids. Emphasis will be placed on determination of the extent to which the in vivo pharmacological profile of this class of cannabinoids differs from that of ?9-tetrahydrocannabinol, the primary psychoactive substituent of the marijuana plant. Further, use of male and female rodents will allow examination of possible sex differences in the effects of these cannabinoids. Results of the studies proposed here will increase knowledge of the behavioral effects of these abused cannabinoids in both sexes, thereby providing a scientific basis for evaluation of potential health concerns associated with acute and repeated use of the compounds, including their physical dependence liability.

DESCRIPTION (provided by applicant): Epidemiological studies have shown that women and men differ in their patterns of substance abuse and dependence. Sex differences in the effects of D9-tetrahydrocannabinol (THC), the primary psychoactive substituent of marijuana, have been reported in humans and in rodent models; however, the mechanisms responsible for these differences are unclear and are the focus of the proposed research. Aims 1 and 2 will determine whether sex differences in the antinociceptive and dependence-related effects of THC, respectively, are modulated by gonadal hormones, estradiol, and progesterone in females and testosterone in males. Emphasis will be on the effects of repeated THC administration on behavior, as substance abuse, by definition, requires chronic use. Dependence-related behaviors include precipitated withdrawal (somatic and affective signs), THC discrimination (a rodent model of marijuana intoxication), and conditioned place preference/aversion (CPP/CPA), a rodent model to assess the degree to which drug effects are associated with pleasure or aversion. Sex differences in tolerance to the antinociceptive effects of THC, one of the potential therapeutic indications of cannabinoid-based medications, will also be evaluated. Aim 3 will characterize the pharmacokinetics of THC and will identify regional alterations in central CB1 and CB2 receptor densities and functioning and endocannabinoid levels that are associated with behavioral changes. Understanding these basic mechanisms underlying sex differences in antinociceptive and dependence-related effects of cannabinoids will facilitate development of sex-specific approaches to treat marijuana dependence and to use cannabinoid-based medications therapeutically, specifically in the treatment of pain and spasticity. In addition, increasing scientific knowledge of endocrinological mechanisms that affect endocannabinoid system functioning could enhance understanding and treatment of a wide variety of disorders, in which dysregulation of the endocannabinoid system has been implicated, including substance abuse, psychiatric disorders, obesity, and various neurological disorders.

DESCRIPTION (provided by applicant): Epidemiological studies have shown that women and men differ in their patterns of substance abuse and dependence. Sex differences in the effects of D9-tetrahydrocannabinol (THC), the primary psychoactive substituent of marijuana, have been reported in humans and in rodent models; however, the mechanisms responsible for these differences are unclear and are the focus of the proposed research. Aims 1 and 2 will determine whether sex differences in the antinociceptive and dependence-related effects of THC, respectively, are modulated by gonadal hormones, estradiol, and progesterone in females and testosterone in males. Emphasis will be on the effects of repeated THC administration on behavior, as substance abuse, by definition, requires chronic use. Dependence-related behaviors include precipitated withdrawal (somatic and affective signs), THC discrimination (a rodent model of marijuana intoxication), and conditioned place preference/aversion (CPP/CPA), a rodent model to assess the degree to which drug effects are associated with pleasure or aversion. Sex differences in tolerance to the antinociceptive effects of THC, one of the potential therapeutic indications of cannabinoid-based medications, will also be evaluated. Aim 3 will characterize the pharmacokinetics of THC and will identify regional alterations in central CB1 and CB2 receptor densities and functioning and endocannabinoid levels that are associated with behavioral changes. Understanding these basic mechanisms underlying sex differences in antinociceptive and dependence-related effects of cannabinoids will facilitate development of sex-specific approaches to treat marijuana dependence and to use cannabinoid-based medications therapeutically, specifically in the treatment of pain and spasticity. In addition, increasing scientific knowledge of endocrinological mechanisms that affect endocannabinoid system functioning could enhance understanding and treatment of a wide variety of disorders, in which dysregulation of the endocannabinoid system has been implicated, including substance abuse, psychiatric disorders, obesity, and various neurological disorders.

DESCRIPTION (provided by applicant): Epidemiological studies have shown that women and men differ in their patterns of substance abuse and dependence. Sex differences in the effects of D9-tetrahydrocannabinol (THC), the primary psychoactive substituent of marijuana, have been reported in humans and in rodent models; however, the mechanisms responsible for these differences are unclear and are the focus of the proposed research. Aims 1 and 2 will determine whether sex differences in the antinociceptive and dependence-related effects of THC, respectively, are modulated by gonadal hormones, estradiol, and progesterone in females and testosterone in males. Emphasis will be on the effects of repeated THC administration on behavior, as substance abuse, by definition, requires chronic use. Dependence-related behaviors include precipitated withdrawal (somatic and affective signs), THC discrimination (a rodent model of marijuana intoxication), and conditioned place preference/aversion (CPP/CPA), a rodent model to assess the degree to which drug effects are associated with pleasure or aversion. Sex differences in tolerance to the antinociceptive effects of THC, one of the potential therapeutic indications of cannabinoid-based medications, will also be evaluated. Aim 3 will characterize the pharmacokinetics of THC and will identify regional alterations in central CB1 and CB2 receptor densities and functioning and endocannabinoid levels that are associated with behavioral changes. Understanding these basic mechanisms underlying sex differences in antinociceptive and dependence-related effects of cannabinoids will facilitate development of sex-specific approaches to treat marijuana dependence and to use cannabinoid-based medications therapeutically, specifically in the treatment of pain and spasticity. In addition, increasing scientific knowledge of endocrinological mechanisms that affect endocannabinoid system functioning could enhance understanding and treatment of a wide variety of disorders, in which dysregulation of the endocannabinoid system has been implicated, including substance abuse, psychiatric disorders, obesity, and various neurological disorders.

PROJECT SUMMARY Tobacco and cannabis are two of the most commonly abused substances in the United States. Further, simultaneous cannabis and tobacco polysubstance use (CT-PSU) is not insubstantial and is trending upwards as cannabis use is legalized in an increasing number of states. This upward trend is of particular concern because, compared to use of cannabis without tobacco, CT-PSU is associated with greater cannabis dependence, psychosocial problems, and poorer cessation outcomes, as well as with additive health risks, relative to either substance alone. Determination of mechanisms underlying CT-PSU will require coordinated epidemiological, preclinical, and human laboratory studies. Yet, the models and measures in each discipline that are used most often to examine CT-PSU were developed to investigate single drug use and may not capture unique features of CT-PSU. In the proposed project, patterns of CT-PSU will be characterized through development and implementation of novel survey measures that focus specifically on CT-PSU (R21 phase). In addition, a procedure for inhaled nicotine self-administration in mice will be established and validated simultaneously with epidemiological data collection. Subsequently, results from both efforts will guide decisions regarding experimental parameters to be used to investigate the effects of ?9-tetrahydrocannabinol (THC; primary psychoactive substituent of cannabis) on inhaled nicotine?s reinforcing effects (i.e., ?chasing?) and the reinforcing effects of both drugs administered simultaneously (e.g., as in blunts or spliffs) in mice and in human tobacco smokers and non-smokers. This translationally relevant approach will result in the establishment of better models through which to investigate CT-PSU, with the ultimate goals of discovery of its underlying mechanisms and development of targeted treatment approaches.

PROJECT SUMMARY Cannabis sativa/indica (marijuana plant) is the most commonly abused substance in the United States, but is still classified as illegal to possess under federal law. Psychoactive phytocannabinoids such as ?9- tetrahydrocannabinol (THC) produce their effects on physiology and behavior through activation of CB1 and CB2 receptors. These receptors are part of the endocannabinoid system, one of several lipid signaling systems in the brain/body. Discovery of these mechanisms underlying cannabinoid action and characterization of their effects on physiology and behavior were due, in part, to research conducted in animals, and specifically, in rodent models. Future progress in this area also will involve findings derived from work with these models. Hence, fundamental knowledge of the advantages and limits of the models are crucial. Recently, we discovered an intriguing species X sex interaction in the effects of THC in rats and mice. Briefly, rats exhibited pronounced sex differences in physiological and behavioral responses following THC injection whereas female and male mice showed similar responses. The overall goals of the proposed project are to investigate mechanisms that may underlie these differences and to determine the extent to which a similar profile characterizes THC?s effects on physiology and behavior. Side-by-side comparisons of the physiological (temperature and cardiac and pulmonary function) and behavioral (discriminative stimulus) effects of THC and cannabis extracts will be made in female and male rats and mice. Differences in pharmacokinetics and receptor mechanisms will be characterized following parenteral injection routes of administration for rodent studies and after administration via more translationally relevant inhalational and oral routes of administration. The results of these studies will provide an empirical basis for choice of rodent model with which to investigate mechanisms underlying cannabis use.

PROJECT SUMMARY Throughout recorded history, humans have used natural products for medicinal purposes. With the onset of modern analytical and pharmaceutical methods, the active ingredients in some of these ?natural remedies? were identified and became foundational chemicals in drug development efforts. Among the numerous examples, aspirin (bark of the willow tree), capsaicin (chili peppers), and opioids (opium poppy plant) are notable for their analgesic properties. Similar to these other natural products, Cannabis sativa has long been used for medicinal purposes, including pain relief. The cannabis plant (Cannabis sativa/indica) contains over 100 phytocannabinoids as well as over 500 other identified chemicals, including the terpenoids that give cannabis its distinctive flavoring and aroma. Beyond ?9-THC and CBD, however, difficulties in identifying, isolating and purifying minor phytocannabinoids in quantities sufficient for in vivo evaluation have hampered adequate investigation of potential therapeutic uses of individual phytocannabinoids. Recently, we have secured access to purified minor cannabinoids synthesized via a patented biosynthetic process in quantities that allow for preclinical in vivo testing. The goal of the proposed project is to provide a comprehensive evaluation of these compounds alone, and in planned combinations, to determine their potential efficacy as analgesics. In addition, we will conduct a parallel investigation of selected terpenes that have been identified in the cannabis plant. Novel analgesic strategies are needed to combat over-reliance on opioids and the resulting secondary consequences of dependence and morbidity/mortality.

? DESCRIPTION (provided by applicant): Vaporization or smoking of synthetic cannabinoid-containing e-liquids or herbal formulations is a significant substance abuse problem. Although JWH-018 and other indole-derived cannabinoids that have been identified in these illicit products produce marijuana-like intoxication, they are structurally distinct from cannabinoids contained in the cannabis plant; hence, very little is known about the actual chemical exposures that occur during their use, or their pharmacology, particularly with regards to their actions at non-cannabinoid receptors and their behavioral and toxicological effects. Furthermore, the variety of structural scaffolds and analogs of synthetic cannabinoids that are being reported in these designer drug formulations continues to increase as individuals seek to become intoxicated and avoid detection, and the manufacturers and distributors of these chemicals and formulations appear to have little regard for the chemical reliability or safety of their products. Indeed, chemicals are frequently appearing in designer drug formulations that have not been previously described as cannabinoids in the scientific literature. Moreover, some of these compounds possess thermally unstable substituents of known carcinogenic activity that may be liberated during use. The proposed research will characterize synthetic cannabinoid formulations and the chemical exposures that occur during storage and use, and evaluate their pharmacological fate in in vitro and in vivo models. Emphasis will be placed on the identification of the compounds that are inhaled or produced during actual use scenarios, and the extent to which they can interact with cannabinoid CB1 and CB2 receptors, as well as with non-cannabinoid receptors, and the degree to which they produce in vivo pharmacological and toxicological effects in mouse models. The proposed studies will thereby increase knowledge of the structure-activity relationships at cannabinoid receptors and the behavioral and toxicological effects of these abused synthetic cannabinoid substances, and provide a scientific basis for evaluation of potential health concerns associated with use of these compounds.