Lawrence D. Middaugh - US grants

A substantial amount of clinical evidence indicates that alcohol during pregnancy can produce both morphological and behavioral abnormalities in offspring. Animal studies have extended these clinical findings by documenting both morphological and behavioral effects of prenatal ethanol exposure under conditions of nutritional and environmental control impossible in the humans. In addition to confirming clinical findings, animal studies suggest that the effects of prenatal ethanol exposure extend to the fully mature offspring. Although the animal studies have documented both the morphological and behavioral abnormalities, the former have utilized mice whereas the latter have utilized rats. The studies described in this proposal will extend previous work on the behavioral effects of prenatal ethanol exposure by utilizing another species and by examining the basis for previously reported abnormalities of these offspring on several behavioral tasks. The studies will use a mouse strain which has been documented to show the classic teratogenic effects of ethanol; hence if behavioral effects are also present, a more complete model of prenatal ethanol effects in humans will be provided. To help establish a behavioral mechanism for some of the task performance differences between prenatally ethanol exposed and control rats, studies are proposed to examine the role of stimulus and response parameters in perseverative behavior and abnormal performance on conditioned avoidance tasks commonly reported for these offspring. Since memory deficits might also contribute to the abnormal performance on many of the tasks and is of tremendous concern for humans but, largely ignored in the animal literature, studies are proposed to examine the effects of prenatal ethanol on memory. Finally, since animal studies suggest that some of the effects of prenatal ethanol exposure are age-dependent, studies are proposed to examine the effects of prenatal ethanol on the ontogeny of activity and passive-avoidance behavior. The ultimate goal of our research in this area is to provide a comprehensive model of the Fetal Alcohol Syndrome in mice which will enable examination of issues of clinical relevance such as assessment of intervention strategies and assessment of the interaction of ethanol with nutritional deficiency or other drugs.

biomedical equipment purchase;

A murine model is proposed for evaluating potential therapeutic agents for alcohol abuse as to their efficacy in reducing ethanol consumption and altering its interoceptive effects. The proposed experiments will utilize C57BL/6 mice, recognized for their consumption of ethanol, to assess ethanol consumption in a limited access paradigm and its interoceptive effect utilizing an operant ethanol discrimination paradigm after treatment with three antagonists; naltrexone, broad spectrum opioid receptor antagonist; naltrindole, a relatively specific delta-opioid receptor antagonist; and MDL 72222, a relatively specific 5-HT3 receptor antagonist. The following hypotheses and related specific aims will be examined. HYPOTHESIS 1 is that broad spectrum opioid, delta-opioid, and 5-HT3 antagonists will reduce ethanol consumption and its interoceptive effects. Thus, Specific Aim 1 is to determine the dose-responsive effects of naltrexone, naltrindole, and MDL 72222 on ethanol consumption and discrimination. These experiments will include time course evaluations to insure optimal intervals between antagonist injection and ethanol test, and sufficient doses to estimate ED50s for each antagonist. HYPOTHESIS 2 is that opioid and 5-HT receptor antagonists will act synergistically to reduce the consumption of ethanol and its interoceptive effect. Thus Specific Aim 2 is to compare the effects of naltexone + MDL 7222 and naltrindole + MDL 72222 combinations on ethanol consumption and discrimination with effects of each alone. Doses in the experiments will be selected to allow isobolographic evaluation for drug additivity and potentiation. HYPOTHESIS 3 is that withdrawal from chronic ethanol consumption will alter its interoceptive effects and that the alteration will be attenuated by opioid- and 5-HT3- antagonists. Specific Aim 3 is thus to determine if withdrawal from chronic ethanol exposure alters its discriminability and if such alterations can be counter-acted by naltrexone, naltrindole, or MDL 72222. The experiment involves training mice to discriminate ethanol and then a comparing groups of chronic- ethanol-exposed mice with controls on ethanol discrimination either in the presence or absence of antagonist. Finally, HYPOTHESIS 4 is that withdrawal from chronic exposure to opioid- and 5-HT3- antagonists will alter the interoceptive effects of ethanol. Specific Aim 4 is to determine if ethanol discrimination will be altered during withdrawal from chronic naltrexone, naltrindole, and MDL 72222. The experiment involves training mice to discriminate ethanol and then comparing groups chronically treated with vehicle or one of the antagonists during a re-test and reacquisition of the ethanol discrimination.

DESCRIPTION (provided by applicant): Most information about the neurobiology of substance abuse derives from experiments on rats. These models address several phenomena characteristic of substance abusing humans (e.g., self administration of drugs abused by humans, uncontrolled use of drugs, relapse upon presentation of interoceptive or exteroceptive cues related to the drug, etc) and have established the mesoaccumbens dopaminergic system as one of the important mediators of the reinforcing effects of drugs. Many of the findings have been confirmed in monkeys and more recently in mice. The desirability of using the mouse species in substance abuse research is prompted by the development of mutant mice which lack genes for specific proteins (i.e. "Gene Knockouts") important for neurotransmission in reward circuitry, as well as several well characterized inbred and transgenic strains. Technologies commonly used in substance abuse research on the rat such as jugular catheterization for i.v. self-administration and installation of indwelling cannulas for in vivo microdialysis have recently been down sized for use on mice. Using these techniques, the scant literature indicates that, like rats, mice self-administer most drugs which are abused by humans and the drugs appear to elicit similar increases in DA transmission in the anteroventral striatum as reported for rats. Notably absent from current literature using mice, however, are experiments to evaluate the effects of drug conditioned stimuli on behavior directed toward drug administration, models of relapse, and the effects of self administered drugs on DA systems. Thus, the proposed experiments will determine; 1) the extent to which cues present during lover responding for i.v. cocaine will maintain the behavior; 2) if the cocaine or cocaine conditioned cues will reinstate lever responding after extinction; and 3) if self-administered cocaine, and perhaps cocaine conditioned cues, will enhance extracellular DA consistent as seen in rats. The proposed experiments will initially be conducted on B6 mice, a strain which has been shown to self-administer i.v. cocaine. Additional experiments will be conducted on mutant mice with deletion of the DA D1 receptor gene to evaluate the role of the DA D1 receptor in the mediation of lever responding maintained by cocaine and cocaine conditioned stimuli, as well as reinstatement of 'cocaine-seeking behavior." These studies will significantly advance the use of the mouse species as a model for drug abuse and contribute toward establishing appropriate therapies for addiction.

SUBPROJECT ABSTRACT NOT AVAILABLE

The proposed experiments utilize a C57BL/6 murine model to determine the influence of potential therapeutic agents for alcohol abuse on the rewarding effect of ethanol and ethanol conditioned stimuli as well as accompanying changes in mesoaccumbens dopamine (DA). Experiments will test the general hypothesis that drugs which influence mesolimbic DA system function will 1) reduce the reinforcing effects of ethanol conditioned stimuli; and 2) attenuate the changes in mesoaccumbens DA function involved with mediating these effects. The drugs to be evaluated initially include SCH 39166, a dopamine D1 receptor antagonist, and drugs which indirectly influence mesolimbic system transmission via their action via their action on other neural systems. Indirect acting agents include: Nalmefene, which has specifically of the mu-opioid receptor; Ondansetron which has high specificity for the 5-HT3 receptor; Vigabatrin, a gamma-aminobutyric acid transaminase inhibitor, and Topiramate, an AMPA glutamate antagonist.. The compounds are used clinically, alter dopaminergic transmission, and preliminary data indicate they can reduce self administration of ethanol and other abused substances in rodents. Behavioral measures, all used during the initial grant period to evaluate naltrexone effects on ethanol reward, include: 1) lever responding for 12% ethanol and 2) voluntary ethanol consumption in a free access paradigm as measures of ethanol reward; and 3) ethanol place conditioning as a measure of ethanol conditioned stimuli effects. Possible adverse effects of the agents will be determined by examining their effects on food and water reward, on food and water ingestion, and whether or not they produce conditioned place aversion at doses which reduce ethanol reward. Proposed microdialysis experiments will assess changes in mesolimbic DA function associated with the rewarding effects of ethanol and ethanol conditioned stimuli, as well as their modulation by potential therapeutic agents. The proposed research contributes to the development of pharmacotherapy for alcohol abuse by examining the effect of compounds acting upon different transmitter systems on neurochemical and behavioral parameters associated with the regarding effects of ethanol and ethanol conditioned stimuli, and sets the stage for future studies.