Comparative studies of sedative action and tolerance on GABA(A) receptor-mediated chloride influx in brai

The work described in this thesis is an examination of the effects of ethanol, diazepam and pentobarbital on GABA-mediated $sp{36}$Cl$sp-$ uptake into rat and mouse brain microsacs, in naive animals, and in those which had undergone chronic treatment with ethanol.;In manual (3 sec incubation) experiments, diazepam and pentobarbital both consistently enhanced GABA-mediated chloride uptake into microsacs, while bicuculline and picrotoxin antagonized it. However, ethanol never had any effect except at a very high concentration (600 mM).;Diazepam and pentobarbital were shown, by use of the quench-flow machine (using 21-500 msec incubations), to interact differently at the GABA$sb{rm A}$ receptor. Pentobarbital preincubation with microsacs had no additional effect unless GABA was also added in the preincubate. The effect of diazepam, in contrast, was greatly increased by preincubation, either with or without GABA. Ethanol, even when preincubated with the microsacs $pm$ GABA, had no effect.;A study involving the chronic treatment of rats with ethanol was undertaken in an attempt to correlate biochemical measures of tolerance at the GABA$sb{rm A}$-receptor/chloride channel complex with behavioural measures of tolerance to the motor-incoordinating effects of ethanol, as well as behavioural and biochemical cross tolerance to diazepam and pentobarbital. In addition, the development of behavioural and biochemical tolerance and cross tolerance to diazepam and pentobarbital was compared in two different paradigms of tolerance development, one involving a major contribution of intoxicated practice (on the moving belt) and one which did not involve a learning component. The chronic tolerance/cross tolerance experiments demonstrated that the behaviourally-augmented cross tolerance to diazepam, seen after chronic ethanol treatment, may have a correlate on the biochemical level. Diazepam enhancement of GABA-mediated chloride uptake was decreased in animals made tolerant to ethanol by means of the intoxicated-practice paradigm, but only if animals received a dose of ethanol one hour before sacrifice. No evidence was found of tolerance to ethanol or cross tolerance to pentobarbital occurring at the GABA$sb{rm a}$-receptor. Our data does not support the idea of the GABA$sb{rm A}$-receptor/chloride channel complex being a primary site of ethanol action, or of tolerance development.