The effect of diazepam withdrawal (2 mg/kg/day) on release of [3H]-5-hydroxytryptamine(5-HT) and [14C]-GABA from rat cortical and hippocampal slices was studied. No changes in [14C]-GABA release (basal, K(+)-evoked, uptake) from slices of either region were observed. Similarly, all parameters of [3H]-5-HT release were unchanged in cortical slices. However, during diazepam withdrawal, depolarised [3H]-5-HT release from hippocampal slices was raised with no changes in basal release or uptake into the slices being found. This increase could be prevented by in vivo administration of 1 mg/kg baclofen--this dose having no significant effect on [3H]-5-HT release from hippocampal slices of control rats. To further investigate this effect, 45Ca2+ uptake into hippocampal synaptosomes was examined and found to be increased during withdrawal. This was blocked by in vitro addition of 10 microM (-)baclofen, which had no effect on 45Ca2+ uptake in controls. Inhibition of 45Ca2+ uptake by (-)baclofen was also enhanced in nonwithdrawn diazepam-treated rats, but not in rats treated acutely with diazepam. The results from both studies indicate that chronic diazepam treatment increases neuronal sensitivity to baclofen. These results are discussed with reference to the anxiogenic state during diazepam withdrawal and a recent report of reversal of this behaviour by baclofen.