Increased gluconeogenesis has been suggested to account for all of the increase in basal glucose production in patients with non-insulin-dependent diabetes mellitus (NIDDM). We studied the effect of inhibition of gluconeogenesis with ethanol on total hepatic glucose output (HGO) in patients with NIDDM. Fourteen patients with NIDDM (mean +/- SE age 61 +/- 2 yr, fasting plasma glucose 11.4 +/- 0.8 mM; body mass index 27 +/- 1 kg/m2) were studied twice after an overnight fast, once during ethanol administration (blood ethanol approximately 12 mM) and once during saline administration. Total HGO rate was measured with [3H]glucose. Inhibition of gluconeogenesis by ethanol was followed qualitatively with [U-14C]lactate (n = 8) and [U-14C]glycerol (n = 6) as tracers. Ethanol inhibited gluconeogenesis from lactate by 71 +/- 5% (0.5 +/- 0.2 vs. 1.8 +/- 0.1 mumol glucose.kg-1.min-1, 240-300 min, P less than 0.001; ethanol vs. saline, P less than 0.001) and from glycerol by 65 +/- 6% (0.8 +/- 0.2 vs. 2.3 +/- 0.6 mumol glucose.kg.min-1, P less than 0.001). Total HGO rate remained unchanged and averaged 12.8 +/- 1.8 and 11.8 +/- 2.1 mumol.kg-1.min-1 in the saline and ethanol studies, respectively (NS). We concluded that inhibition of gluconeogenesis by ethanol does not decrease total HGO in patients with NIDDM. Our results suggest the existence of a regulatory mechanism in the liver that maintains constant total HGO despite inhibition of gluconeogenesis.