The gamma-aminobutyric acid type A (GABA(A)) receptor is an important pharmacological target of ethanol. The effect of ethanol withdrawal on the expression of the alpha(2) subunit of this receptor was examined with rat cerebellar granule cells in primary culture. Long-term exposure of these cells to ethanol (100 mM, 5 days) did not affect the abundance of the mRNA for the alpha(2) subunit, as revealed by an RNase protection assay. In contrast, subsequent ethanol withdrawal for 3 h induced a marked increase in the amount of this mRNA (2.6-fold) as well as in that of the encoded polypeptide (2.2-fold), the latter revealed by immunoblot analysis. Exposure of the cells to gamma-hydroxybutyric acid (100 mM) during ethanol withdrawal prevented the increase in the amounts of both the alpha(2) mRNA and polypeptide, whereas similar treatment with diazepam (10 microM) blocked the increase in the abundance of the alpha(2) polypeptide but not that in the amount of the alpha(2) mRNA. The effect of gamma-hydroxybutyric acid was not blocked by the competitive GABA(B) receptor antagonist SCH 50911(10 microM). Given that the alpha(2) subunit of the GABA(A) receptor mediates the anxiolytic action of benzodiazepines, its up-regulation during discontinuation of long-term ethanol exposure might be relevant to the therapeutic efficacy of these drugs in the treatment of anxiety associated with ethanol withdrawal.