Pancreatic islets were cultured for 24 h in medium containing either low (1.4), normal (5.5), or high (16.7 mM) glucose, and then insulin secretion was measured at the end of 1 h incubation at 37 degrees C. Insulin release in the absence of glucose was 64 +/- 20, 152 +/- 11, and 284 +/- 30 pg.islet-1.h-1 (mean +/- SE, n = 6, G1.4 and G16.7 vs. G.5.5, P < 0.05) and the response to 22 mM glucose stimulation was 640 +/- 136, 2460 +/- 276, and 1890 +/- 172 pg.islet-1.h-1, respectively (n = 6, G1.4 vs. G5.5, P < 0.01, G16.7 vs. G5.5, P = 0.065). The 50% maximal response of insulin secretion (increment over baseline) was reached at an average glucose concentration of 9.9 +/- 0.7 mM in islets preexposed to G5.5, and at glucose 13.3 +/- 0.9 and 4.8 +/- 0.4 mM (P < 0.05 in respect to G5.5) in islets preexposed to G1.4 and G16.7, respectively. To investigate the molecular mechanism responsible for this altered glucose sensitivity, we measured, in parallel experiments, the kinetic characteristics of glucose transport, glucokinase, and glucose utilization. Glucose transport was measured by evaluating 3-O-methylglucose uptake. The apparent Km of the low-affinity transporter (GLUT2) was 16.6 +/- 2.4 mM in isolated pancreatic cells cultured at 5.5 mM glucose.(ABSTRACT TRUNCATED AT 250 WORDS)