Paul Vezina - US grants

DESCRIPTION: (Applicant's Abstract) DOPAMINE NEUROTRANSMISSION AND AMPHETAMINE SENSITIZATION Changes in brain dopamine (DA) neurotransmission have been implicated in several disease processes including schizophrenia and substance abuse. One change posited to be common to both these disorders is the development of increasingly reactive or sensitized DA function. This has been suggested to result in unusually active levels of subcortical DA neurotransmission and in the ensuing expression of symptomatology. The experiments proposed in this application are part of a continuing research effort aimed at determining what neural events are involved in the development and in the behavioral expression of such enhancements in brain DA function. Psychomotor stimulant drugs, such as amphetamine and cocaine, increase extracellular levels of DA in the terminal and cell body regions of mesolimbic DA neurons and produce locomotor activation. Repeated exposure to these drugs sensitizes their locomotor effects so that subsequent reexposure to the drug produces greater behavioral activation than seen initially. Recent findings suggest that changes in midbrain DA neurotransmission may contribute importantly to both the induction and expression of this behavioral sensitization. The experiments proposed will investigate the effect of prior exposure to amphetamine and cocaine on pre- and postsynaptic aspects of DA neurotransmission in the rat nucleus accumbens (NAcc), the mesolimbic DA terminal field most implicated in mediation of the locomotor and motivational effects of psychomotor stimulants. As such, the experiments have three aims. AIM 1. To investigate how INDUCTION of sensitized locomotor and NAcc DA responding by amphetamine and cocaine is produced by assessing the contribution of their suggested recruitment of excitatory amino acids. AIM 2. To characterize the changes produced by amphetamine PRESYNAPTICALLY in mesolimbic DA neurons that may contribute to the EXPRESSION of sensitized locomotor responding. AIM 3. To determine whether amphetamine and cocaine produce changes POSTSYNAPTICALLY in DA and excitatory amino acid receptors in the NAcc that may also contribute to the EXPRESSION of sensitized locomotor responding.

The experiments proposed in this application are part of a continuing research effort by the applicant aimed at determining the neural basis of drug-induced sensitization in mesolimbic dopamine (DA) neurons and at understanding the nature and significance of its behavioral expression. Psychomotor stimulant and opiate drugs, both of which are self- administered by man and laboratory animals, increase the DA released by these neurons and produce locomotor activation. Repeated exposure to these drugs enhances or sensitizes both of these responses so that subsequent reexposure to the drug produces both greater behavioral activation and greater DA release than seen initially. Considerable evidence implicates mesolimbic DA neurons in the mediation of the rewarding properties of stimulant and opiate drugs. Changes in the functioning of these neurons may therefore be important in the initiation and maintenance of drug taking. Recently. a number of reports have appeared suggesting that exposure to drug regimens that produce behavioral sensitization also facilitates the acquisition of drug self-administration behavior. I propose, therefore, to investigate whether this facilitation is linked to sensitization in mesolimbic DA neurons. If it is, then facilitation should be accompanied by sensitized mesolimbic DA neuron responsivity to drug during acquisition and similarly influenced by the same manipulations that influence the induction of sensitization in these neurons. By studying the acquisition of self-administration of low doses of amphetamine in the rat, I will seek to answer three questions: A. Is facilitation of the acquisition of self-administration behavior accompanied by sensitized levels of DA overflow, as measured by in vivo microdialysis, in the terminal regions of mesolimbic DA neurons during acquisition? B. Does prior exposure to sensitizing injections of amphetamine into the cell body region of mesolimbic DA neurons facilitate the acquisition of self-administration behavior? C. Do manipulations that block the induction of sensitization to amphetamine, such as preceding injections with the D-1 DA receptor antagonist, SCH-23390, also block the facilitation of acquisition of self- administration behavior produced by prior exposure to the drug? The possibility of a link between sensitized DA function and liability to drug abuse could provide important insights into the neurobiology of drug dependence.

DESCRIPTION: (Adapted From The Applicant's Abstract) The experiments proposed in this application are part of a continuing research effort by the applicant aimed at determining the neural basis of drug-induced behavioral sensitization and at understanding the nature and significance of its expression. Psychomotor stimulants like the amphetamines as well as opiate drugs, both of which are self- administered by man and laboratory animals, produce locomotor activation. Repeated exposure to these drugs produces long-term enhancements in their ability to elicit locomotion so that subsequent re-exposure to the drug, weeks to months later, produces greater behavioral activation than seen initially. More importantly, prior exposure to such sensitizing regimens of amphetamine injections is also known to produce long lasting enhancements in animals' predisposition to subsequently self-administer the drug. Considerable evidence links mesoaccumbens dopamine neurons to the locomotion produced and the self-administration supported by psychomotor stimulants. Sensitization in this neuronal system also parallels the above long-term enhancements in locomotion and appears to be associated with enhancements in animals' predisposition to self-administer drug. Other findings indicate important contributions by the excitatory amino acid glutamate both to the INDUCTION and EXPRESSION of locomotor sensitization by amphetamines. Little is known, however, of the contribution of this neurotransmitter to the facilitation of drug taking produced by exposure to sensitizing drug regimens. By studying amphetamine self-administration in rats, the proposed experiments will attempt to elucidate this contribution by seeking to answer three questions: QUESTION 1: INDUCTION. Do manipulations known to block the induction of locomotor sensitization to amphetamine, such as preceding sensitizing injections with antagonists specific for ionotropic and metabotropic glutamate receptors, also block the facilitation of self-administration of the drug? QUESTION 2: EXPRESSION. Does prior exposure to amphetamine enhance extracellular levels of glutamate in the terminal field of mesoaccumbens dopamine neurons during the subsequent self-administration of the drug and reinstatement of this behavior after it has been extinguished? QUESTION 3: EXPRESSION. Does prior exposure to amphetamine enhance the ability of glutamate receptor subtype selective ligands in this site to subsequently influence the self-administration of the drug as well as its reinstatement after extinction?

DESCRIPTION (Adapted from applicant's abstract): Although approximately 20 million prescriptions for hypnotic medications are written each year, the molecular mechanisms by which they act, and the neuroanatomic site(s) at which they act. remain unclear. This problem is complicated by the wide range of pharmacologic classes of compounds which induce sleep, and it is uncertain whether these various agents differ in their mechanisms or site of action. The investigator has begun to build a case that a common molecular mechanism for several types of sleep-inducing compounds may involve interaction with the benzodiazepine (BZ)-GABAa receptor complex. The current proposal, based on the investigator's observations that microinjection of pentobarbital or the BZ hypnotic triazolam into the medial preoptic area (MPA) of the anterior hypothalamus induces sleep. addresses the anatomical site(s) of action of sleep-inducing drugs. The goal is to bring together these observations from the investigator's laboratory and those from other laboratories indicating similar effects from microinjections of ethanol and adenosine. The investigator shall explore these observations further, addressing the following questions: 1. Do these findings hold when all these compounds are examined in the same laboratory, under similar conditions, and do these various agents show clear dose-responsiveness? 2. Are these effects specific to the BZ-GABAa recognition site? 3. Are these effects specific to the MPA, or do they occur with injection into other structures? 4. Do they result as a consequence of altering hypothalamic temperature, or by a more direct mechanism? 5. Is an intact MPA necessary for sleep induction by these agents when given parenterally? In a broader sense, the goal of this proposal it to move beyond the conventional approach of simply identifying a hypnogenic compound, and instead to develop a series of critical questions which should be asked about any substance which alters sleep, including: 1) where does it act?; 2) on which receptors does it act?; 3) with which physiological processes does it interact?; and 4) which brain sites are critical for its hypnogenic effect? The investigator's goal, then, is both to characterize the properties of four classes of hypnogenic substances, and to begin to build a framework to make the currently diverse and complex literature on putative sleep substances more structured and comprehensible.

DESCRIPTION (provided by applicant): The overall objective of the NIDA Training Program in Drug Abuse Research at The University of Chicago is to provide both pre- and postdoctoral trainees with comprehensive educational and research experiences that will enable them to pursue distinguished research careers in areas impacting drug abuse. Drug abuse poses a serious threat to the health and well being of both individuals and society. Mental status, physical health, social function and economic productivity are compromised. Such factors can lead to an increase in crime and violence. To the credit of NIDA-sponsored research programs, considerable gains have been made over the last two decades. However, much more remains to be learned from the molecular to the social realm, especially if knowledge is to be translated into effective treatment. Existing programs in the area of drug abuse at The University of Chicago are designed to acquire knowledge about the social, behavioral, neuropharmacological, molecular and genetic factors relevant to the abuse of drugs. The aim is to increase our understanding of the etiology of and our ability to intervene in the treatment and prevention of drug abuse. Specific strengths of this program relate to the neuropharmacology, psychopharmacology, electrophysiology, genetics and molecular biology of drug abuse as well as the behavioral and subjective effects of drugs in humans. Several factors place the faculty submitting this application in an excellent position to continue to offer an outstanding training program. These factors include the strength of current interests and ongoing research into the problems of drug abuse, the diversity of approaches employed by individual faculty, the presence of a critical mass of creative scientists working on drug abuse related problems in a multidisciplinary setting as well as a curriculum that addresses the broad aspects of drug abuse ranging from the molecular bases of drug action to the consideration of ethical and social issues related to drug taking.

DESCRIPTION (provided by applicant): Psychomotor stimulant drugs such as amphetamine produce multiple effects. Notable among them is their ability to activate brain dopamine (DA) neurotransmission, increase locomotor activity and support self-administration in humans and laboratory animals. When repeatedly administered, their ability to produce these effects becomes enhanced so that re-exposure to the drug, weeks to months later, produces greater DA activation, locomotion, and work-output aimed at obtaining the drug. These findings support the proposal that sensitization of the appetitive effects of amphetamine and other psychostimulants promotes the pursuit and self-administration of these drugs and may underlie the transition from casual drug use to compulsive drug taking and abuse. A number of long-lasting neuroadaptations have been identified in forebrain regions like the nucleus accumbens (NAcc) that provide neuronal correlates for the expression of behavioral sensitization by amphetamine. Several lines of converging evidence now indicate a coordinated interaction between DA and glutamate in the expression of amphetamine sensitization, one in which the regulation of AMPA receptors is essential, and that is dependent on pre- and postsynaptic PKC signaling in the NAcc. This proposal builds on new data from the laboratory showing that DA can regulate AMPA receptor trafficking and function to enable the expression of behavioral sensitization. It also incorporates recently published findings showing new mechanisms by which PKC can directly and indirectly regulate AMPA receptor insertion and function. Together, these findings support the hypothesis that NAcc AMPA receptors activate medium spiny neurons and the ensuing motivated behavior the animal displays while Gq-coupled DA receptors enable the expression of sensitization by initiating PKC-mediated signaling to regulate AMPA receptor trafficking and function. To begin testing this hypothesis, the experiments outlined in this proposal will use a model of enhanced amphetamine self-administration and reinstatement to determine the role played by different PKC substrates in the NAcc in the expression of these sensitized behaviors. The proposed experiments exploit a multidisciplinary approach, using behavioral, biochemical, pharmacological, and viral-mediated gene transfer techniques. In Aims 1 and 2, viral-mediated gene transfer will be used to determine the contribution of PKC phosphorylation of GluR1 and neurogranin specifically in NAcc neurons to the expression of sensitization. In Aim 3, pharmacological inhibition of PKC will be used to assess its ability to enable the expression of amphetamine sensitization by regulating presynaptic DA overflow. By deciphering the neuroadaptations that underlie the expression of sensitization, we will increase our understanding of the mechanisms underlying the enhanced pursuit and self- administration of psychostimulant drugs and inform the development of therapeutic strategies aimed at reversing these maladaptive behaviors.

The function of the Administrative Core is to provide the Principal Investigator with the organizational, administrative, and secretarial support needed to accomplish the scientific goals of the Program Project. As outlined in the sections below, the Core will provide the infrastructure necessary to oversee the day-to-day management of the Program Project and to coordinate the preparation of rats and sharing of tissue between the different Projects. The Core will also function as a center for the sharing and integration of ideas and results generated by the different Projects. This latter function is critical because the timely and regular exchange and integration of ideas from all three Projects is vital to the decision making processes that impact the scientific direction of each Project as well as of the Program Project as a whole. The organizational structure of the Program Project and the relative position of the Administrative Core within the Program are outlined in Figure 1. The Administrative Core will be physically located within Dr. Vezina's office suite, adjacent to his laboratory in space allocated to the Department of Psychiatry, and in close proximity to the offices and laboratories of Drs. Green and McGehee.

Behavioral and dopaminergic sensitization by nicotine The aim of the following experiments is to determine how neuronal systems in brain are affected by chronic exposure to nicotine and to assess whether these effects can subsequently impact behaviors directed at obtaining the drug. Because nicotine, like other stimulants self-administered by rats and humans, is known to promote neurotransmission in the ascending midbrain dopamine (DA) systems, the proposed experiments will focus on this neurotransmitter known to be critical for the generation of appetitive behaviors such as locomotion and drug self-administration. Rats, like humans, will work to obtain nicotine. In addition, nicotine use is most often characterized by habitual, repeated intake over long periods of time. It is necessary therefore to gain a better understanding of the short- and long-term changes that are produced by prolonged exposure to the drug. This latter point is important because, while knowledge of the acute effects of nicotine on brain has increased somewhat, relatively little is known of the consequences for midbrain DA neurotransmission and the promotion of drug seeking behaviors of prolonged exposure to the drug either actively or passively. The following experiments will seek to elucidate these consequences by studying stimulant-induced locomotion, midbrain DA activation and self-administration in rats. Experiments in Aim 1 will characterize the effects of different nicotine exposure regimens on the induction of sensitization by nicotine. Those in Aim 2 will seek to identify the brain regions containing the nicotine acetylcholine receptor fields underlying the induction of sensitization. Finally, the impact of nicotine sensitization on the motivation to self-administer nicotine will be assessed in Aim 3. The results obtained will lead to a better understanding of the mechanisms underlying the pursuit of nicotine and the manner in which prolonged exposure to the drug may exacerbate drug-directed behaviors.

DESCRIPTION (provided by applicant): Tobacco use remains a major public health concern in the United States and abroad where it is a major factor in the initiation of illnesses leading to preventable death. Yet, despite such adverse consequences, the number of smokers continues to increase worldwide. Although considerable progress has been made in elucidating the neuronal mechanisms underlying several of the effects of nicotine, the active component of tobacco, the knowledge accumulated to date falls short of what is necessary to intervene successfully to reduce tobacco use. The experiments proposed in this program project will use a multi-disciplinary approach to assess the impact of repeated exposure to nicotine on nicotinic acetylcholine receptors (nAChRs), neuronal excitability and behavior. They have in common the goal of ultimately understanding how nicotine acts to initiate self-administration behaviors and how repeated exposure to nicotine produces changes that promote the continued expression of these behaviors. Nicotine produces its effects on motivated behavior by activating brain nAChRs. The ability of nicotine to up regulate these receptors can thus have a substantial impact on the ability of this drug to initiate such behaviors in animals subsequently re-exposed to it. Project 1 will aim to identify the molecular mechanisms underlying nicotine-induced up regulation of nAChRs and characterize this phenomenon in different brain regions of nicotine-exposed rats. As with other drugs of abuse, nicotine use relies in large part on its ability to increase midbrain dopamine transmission. Project 2 will characterize the effect of previous exposure to nicotine on the functional properties of nAChRs with access to these dopamine systems and assess their contribution to long-term potentiation of synapses onto dopamine cells. Repeated exposure to nicotine is known to lead to locomotor sensitization, yet little is known about how this effect is produced. Project 3 will identify the brain nAChR fields involved, characterize their role in the induction of sensitization of dopamine neuron reactivity and determine whether their repeated activation leads to increased motivation to self-administer nicotine.

DESCRIPTION (provided by applicant): Pathological gambling and drug addiction may share common neuronal substrates. Studies of cocaine- and alcohol-dependent individuals have revealed a higher prevalence of pathological gambling than seen in the general population often with gambling preceding drug addiction. Human imaging studies conducted during gambling and the performance of unpredictable reward tasks have also observed neuronal activation in the midbrain ventral tegmental area (VTA), a region intimately involved in the development and maintenance of addiction. Incentive sensitization provides a potential explanation for how this neuronal activation may underlie the transition from casual drug use to drug craving and abuse. In this model, repeated intermittent exposure to drugs, such as amphetamine, initiates neuroadaptations in the VTA that lead to sensitized locomotion and nucleus accumbens (NAcc) dopamine (DA) overflow in response to a drug challenge as well as enhanced amphetamine self-administration on a progressive ratio schedule. Sensitization has been observed following exposure to a number of drugs and cross-sensitization between drugs and non-drug reinforcers such as saccharin has been reported. Recently, we reported that exposure to conditions of gambling-like uncertain, rather than predictable, saccharin reinforcement may lead to the same neuroplastic changes in the brain that are produced by repeated intermittent psychostimulant exposure23. Non-deprived male rats were trained with escalating schedules of fixed-ratio (FR) or variable-ratio (VR) reinforcement for saccharin (FR: fixed relationship between presses and payout; VR: uncertain relationship between presses and payout). At the end of training, both groups worked reliably on the schedules and total saccharin intake did not differ between groups. Nonetheless, a threshold amphetamine challenge injection (0.5 mg/kg IP) administered two weeks after the last saccharin training session produced a significantly greater locomotor response in VR compared to FR rats. This finding suggests that the variable reinforcement schedule elicited neuroplastic changes similar to those produced by repeated psychostimulant exposure known to sensitize responding. In two aims, the proposed experiments will begin to characterize how repeated exposure to gambling-like unpredictable reinforcement promotes locomotor cross-sensitization to amphetamine and determine whether amphetamine self-administration and NAcc DA overflow are also subsequently enhanced. First, midbrain DA neuron reactivity will be assessed during responding for uncertain reward by measuring somatodendritic DA overflow in the VTA while rats respond for predictable or unpredictable saccharin reinforcement. Second, the long- lasting effects of exposure to uncertain reinforcement will be further characterized by testing whether this experience leads to both enhanced drug self-administration and NAcc DA overflow. These experiments have important implications for understanding the neuroadaptations that underlie different forms of addiction and are the first to investigate this potential transition from pathological gambling to drug addiction in an animal model.