The effect of glucagon on ACTH secretion was studied in anaesthetized rats injected with either saline (0·1 ml i.m.) or glucagon (0·02 mg/kg i.m.). For the first 90 min after glucagon injection, plasma ACTH fell by 50% from the basal value of 23 ±4 pmol/l (mean ± s.e.m.) to 11 ±2 (P=0·011), after which an abrupt return to baseline occurred (120 min value: 26 ± 2 pmol/l). In saline injected rats, the baseline ACTH value was not significantly different from either the 90 min value or the 120 min value (27 ±3 vs 21 ± 4 and 24 ± 3 pmol/l respectively; P>0·10). Plasma glucose after glucagon peaked at 11·6 ± 1·1 mmol/l by 15 min but subsequently fell rapidly, attaining the baseline by 60 min. Insulin levels increased sharply after glucagon, from 381 ±78 pmol/l to 3172 ±668 pmol/l at 15 min, and plateaued at approximately 1000 pmol/l thereafter. No changes in glucose or insulin were seen in saline injected rats. The magnitude of suppression of ACTH after glucagon was not affected either by sustained hyperinsulinaemia (≃ 1400 pmol/l), induced with continuous glucose infusion to maintain plasma glucose>12 mmol/l, or by pretreatment with the long-acting somatostatin analogue octreotide (50 μg/kg s.c.). However, the return to baseline between 100 and 120 min was prevented both by hyperinsulinaemia induced with sustained hyperglycaemia, and by octreotide. It is postulated that glucagon may inhibit ACTH secretion either by a direct effect on the hypothalamus or indirectly through insulin, which is known to stimulate endogenous somatostatin release.