A new experimental model of ATP-sensitive K⁺ channel-independent insulinotropic action of glucose: a permissive role of cAMP for triggering of insulin release from rat pancreatic β-cells.

In pancreatic β-cells, glucose metabolism leads to closure of ATP sensitive K⁺ channels (K(ATP) channel) and Ca²⁺ influx, which is regarded as a required step for triggering of insulin release. Here, we demonstrate that glucose triggers rapid insulin release independent from its action on K(ATP) channels given the cellular cAMP is elevated. We measured insulin release from rat pancreatic islets by static and perifusion experiments. Changes in cytosolic free Ca²⁺ concentration ([Ca²⁺]i) were monitored using fura-2 loaded rat pancreatic β-cells. Glucose-induced insulin release was abolished when Ca²⁺ influx was inhibited by a combination of 250 μM diazoxide, an opener of K(ATP) channel, and 10 μM nifedipine, a blocker of L-type voltage-dependent Ca²⁺ channels. However, with both nifedipine and diazoxide, glucose induced a 5-fold increase in insulin release in the presence of 10 μM forskolin, an activator of adenylyl cyclase. In the presence of diazoxide, nifedipine, and forskolin, 22 mM glucose sharply increased the rate of insulin release within 2 min which peaked at 6 min: this was followed by a further gradual increase in insulin release. In contrast, it lowered [Ca(2+)]i with a nadir at 2-3 min followed by a gradual increase in [Ca²⁺]i. The glucose effect was mimicked by 20 mM α-ketoisocaproic acid, a mitochondrial fuel, and it was nullified by 2 mM sodium azide, an inhibitor of mitochondrial electron transport. Cerulenin, an inhibitor of protein acylation, decreased the glucose effect. In conclusion, a rise in [Ca²⁺]i through voltage-dependent Ca²⁺ channels is not mandatory for glucose-induced triggering of insulin release.