Previous studies have shown that chronic benzodiazepine treatment reduces the in vivo sensitivity of dorsal raphe neurons (DRN) to microiontophoretically applied gamma-aminobutyric acid (GABA). We have examined sensitivity of DRN in vitro using a modified midbrain slice technique which allows side-by-side analysis of slices from control and chronic diazepam-treated rats. GABA sensitivity of raphe neurons was reduced in slices from rats treated for 3 weeks with diazepam, compared to control sensitivity. Thus, GABA subsensitivity following chronic diazepam treatment appears to be dependent on changes intrinsic to the midbrain area.