Biobehavioural Pharmacology: CAMH Research Annual Report 2002

The research goals of the Biobehavioural Pharmacology Section are: to understand the underlying behavioural and neurobiological mechanisms that initiate and maintain alcohol dependence; and to use this understanding to explore therapeutic agents for treating alcohol dependence. The majority of our research focuses on issues related to alcohol's reinforcing ability and relapse to alcohol drinking behaviour with an emphasis on the role of stress in relapse. We continue to explore the role of specific central neurochemical systems in regulating these behavioural processes, in addition to examining the possible role of genetic factors involved in problem drinking and concurrent problems with other substances, such as nicotine.

During the past year, we have increased our knowledge of the role of specific neurochemical systems within the brain that regulate alcohol consumption in general, as well as identified potential systems that may predispose an individual to problem drinking. We have begun to explore sex differences in susceptibility to alcohol's effects, with a particular emphasis on cognitive function, which is an increasing public health issue. We hope our research will, in time, help develop therapeutic agents, identify potential risk factors, and provide research-based information on alcohol's effects on the brain and its ability to function.

Stress and Relapse to Alcohol

Relapse is a major challenge in treating alcohol dependence. Exposure to stressful situations has been identified as a key factor in relapse to alcohol use. A major research effort in the section attempts to elucidate neurochemical mechanisms underlying stress-induced relapse to alcohol.

Over the past year, we have advanced our understanding of the mechanisms of stress-induced relapse. Using our animal model, we found that the brain neuropeptide involved in the co-ordination of stress responses, corticotrophin releasing factor (CRF), plays a critical role in stress-induced relapse to alcohol. Administration of a crf receptor antagonist can block stress-induced relapse to alcohol; these crf receptor antagonists may potentially be used to treat alcohol dependence.

CRF might control relapse to alcohol by interacting with a specific serotonergic pathway in the brain. Such a serotonergic pathway has been shown to play a significant role in inhibitory control of behaviour. A number of CRF receptor antagonists are currently being developed for treating anxiety and depression.

Co-Abuse of Alcohol and Nicotine

Another finding is the mechanism underlying the co-abuse of alcohol and tobacco. We have shown previously that nicotine can enhance alcohol self-administration in experimental animals, and treatment with a nicotinic receptor antagonist can reduce alcohol consumption. Genetics have been shown to play a critical role in problem alcohol use in humans. We have found that there might be a common genetic determination for alcohol and nicotine abuse. Animals selectively bred for high alcohol consumption also self-administer more nicotine than those bred for low alcohol consumption.

Of most interest to our researchers is our finding on the effect of exposure to nicotine on alcohol consumption. Animals exposed to nicotine for a short period during their adolescence have a much higher preference for alcohol when tested during adulthood. This effect of nicotine is age-dependent, as similar treatment with nicotine in adult animals did not affect alcohol consumption when tested three months later.

This finding is consistent with the notion that nicotine might act as a "gateway" drug.

5-HT Receptor Subtypes and Alcohol Reinforcement Processes

Multiple neurotransmitter systems help to modulate the impact that alcohol has on the behaviours linked to problem alcohol use and alcohol's dependence liability. We have been selectively manipulating central neurotransmitter function in animal models of alcohol drinking behaviour. We hope to further our understanding of the neurobiological mechanisms underlying excessive alcohol consumption.

Studies in humans and animals suggest an association between the central neurotransmitter, 5-HT, and problem alcohol use and dependence. We continue our work to assess how modulating activity at various 5-HT receptor subtypes affects alcohol self-administration behaviour.

One receptor of particular interest is the 5-HT1B receptor. Human studies have suggested that a locus that predisposes people to antisocial alcoholism is linked to the 5-HT1B receptor gene. Over the past year, we have clearly demonstrated that 5-HT1B receptors play an important role in regulating alcohol intake in our animal models. So far, this effect of the 5-HT1B receptors appears limited to substances with pharmacological activity, such as alcohol and cocaine, because consumption of other fluids and other general behaviours are less sensitive to effects of 5-HT1B receptor manipulations.

We continue to study two brain areas, the amygdala and the ventral tegmental area, that may be important in mediating 5-HT1B receptor effects on alcohol intake. Results to date suggest that activation of 5-HT1B receptors within the ventral tegmental area lead to a decrease in alcohol intake, while in the amygdala, the same manipulation leads to an enhancement of alcohol intake.

These differential findings in discrete brain regions demonstrate that the regulatory effect of 5-HT1B receptors within the brain is site-specific. Furthermore, the data on the amygdala is particularly intriguing, as very few reported pharmacological manipulations have increased alcohol intake in animal models. Our findings may suggest that the amygdala exerts an important modifying influence on alcohol consumption, under normal circumstances, that can be reversed by activation of 5-HT1B receptors within this area.

We are continuing this line of research in the hopes of better understanding the neural circuitry important in regulating drinking behaviour.

GABAa Receptor Subunits, Drinking Behaviour and Voluntary Intake

Compelling evidence suggests that central gabaergic systems play an important role in regulating alcohol's effects, particularly those effects mediated via the GABAa receptor.

We continue to investigate regional differences in the expression of the GABAa receptor subunits. These differences have been demonstrated in the brains of high-alcohol preferring rats, and humans with drinking problems. These differences might represent one of the neurobiological factors underlying problem alcohol use.

Previously, we found significant differences in the GABAa receptor between animals with a propensity to self-administer alcohol and those without. We found support in reports of altered brain, cerebral spinal fluid and plasma gaba levels associated with alcohol dependence and withdrawal and in reports of altered GABAa receptor binding, and region- and subunit-specific changes in gabaa receptors, in the brains of people who are alcohol-dependent. A tentative link between various GABAa receptor subunit genes and a risk for alcoholism may be related to differences in the expression of alcohol's behavioural effects.

This ongoing project combines behavioural and biological approaches to investigate if higher levels of GABAa receptor subunits within discrete brain loci are a predictor and/or a consequence of high-alcohol drinking behaviour. Much of the research on neurochemical effects of alcohol has used force administration. We consider potentially crucial differences between "self-administered" versus "experimenter-administered" alcohol, as different neurochemical systems may be involved in voluntary drug-seeking behaviour and in forced intake, the former being more analogous to drug-taking by humans.

The data generated thus far demonstrate that regional differences in GABAa receptor expression and subunit conformation also affect the binding profile of some pharmacological agents that interact with this receptor complex, including muscimol, flunitrazepam and diazepam, but not others, such as zolpidem. There appears to be a complex interaction between inherent alcohol preference, alcohol drinking history and the binding ligand employed. Currently, we are analysing and interpreting the extensive database generated over the last year. These data will provide important insights not only into the genetic and non-genetic GABAa receptor influences on alcohol preference and consumption, but also on potential interactions with and/or influences over other clinically used pharmacological agents that interact with this receptor complex, such as the benzodiazepines.

Sex Differences in Susceptibility to Alcohol-Induced Cognitive Deficits

We are studying the differences in alcohol's effects on brain function in men and women. In our work, we hope to unravel some of the gender differences in susceptibility to alcohol-induced cognitive impairments and provide new knowledge about the role of GABAa receptors in these impairments. Ultimately, this research could help identify risk and/or protective factors for alcoholism specific to women and help develop effective treatment and prevention strategies. Over the past year, we began to explore the long-term effects of alcohol exposure on cognitive function and behaviour, with specific emphasis on examining potential differences in susceptibility between males and females.

Clinical evidence suggests that women are more vulnerable to the negative effects of alcohol than men are, even when doses are adjusted for body composition. In addition to being more susceptible to alcohol-related diseases (e.g., liver damage), women appear to be more sensitive to the cognitive deficits induced by long-term alcohol exposure. For example, women are more impaired during tasks demanding divided attention or delayed recall, even when blood alcohol levels are comparable to those of men. Interestingly, in tests of psychomotor performance, males and females have been consistently reported to be equally impaired following alcohol ingestion.

Despite the urgency of attending to such critical sex differences, alcohol research has customarily employed males (humans and animals) to avoid the hormonal confounds introduced by the female estrous cycle. However, research to date has found no consistent evidence that alcohol-induced impairments differ over the female menstrual cycle.

Furthermore, in animal studies where sex differences have been studied, investigators have not considered sex differences in the pharmacokinetic profile of alcohol. Females have generally been exposed to equivalent, or even higher, doses compared to their male counterparts during these studies. Functionally equivalent doses of alcohol were not administered in these studies, as females (both animal and human) generally achieve higher blood alcohol levels due to sex differences in body composition. This limits the interpretability of these data.

Our recent studies were designed to examine sex differences in susceptibility to ethanol-induced impairments of cognition. In these studies, the doses are titrated to ensure that equivalent blood ethanol levels are achieved in both sexes over an extended period of time. We are currently testing the following hypotheses:

Female rats will require lower doses of ethanol to be administered to achieve similar blood ethanol levels to that observed in their male counterparts.
On measures of psychomotor performance, female and male rats will be equally impaired following acute and chronic exposure to equivalent functional levels of ethanol.
Females will show greater impairments on measures of spatial learning and delayed recall compared to their male counterparts when exposed to equivalent levels of ethanol over an extended period of time. These sex differences will not necessarily be observable during acute ethanol exposure, but will be observable both in the ethanol-free state following chronic ethanol exposure, and following a subsequent ethanol challenge test.
Females and males will not show impairments under experimental conditions that assess sensorimotor ability and motivation during the spatial learning and delayed recall tasks, demonstrating that differences in performances between the sexes are due to cognitive deficits, and not other factors such as their inability to complete the task due to motor deficits, visual deficits and overall motivation to perform the task.
Chronic exposure to functionally equivalent levels of ethanol in male and female rats will result in differential alterations in gabaa receptor subunit expression in a sex-, brain region- and subunit-specific manner. Furthermore, these differences in brain regions implicated in cognition are correlated with the deficits observed on measures of spatial learning and delayed recall.

Our data thus far support our hypothesis that sex differences in some, but not all, behaviours occur when equivalent blood alcohol levels are maintained over an extended period of time. This first step in the project confirms the validity of our test regimen. Over the coming year, we will expand on this pilot data and specifically explore sex-related differences in alcohol's long-term effects on cognitive function.