Gramicidin D is a potent insulin secretagogue: Dependence on extracellular calcium influx

Gramicidin D, a sodium ionophore, was identified as a potent insulin secretagogue in the mouse β-cell line, βTC3. Gramicidin D stimulated insulin secretion by 3.2-fold relative to control cells incubated with vehicle alone. Using ion-specific fluorescent probes, gramicidin D (1 μM) increased the intracellular concentrations of Na⁺ ([Na⁺]_i) and Ca²⁺ ([Ca²⁺]_i). By contrast, no changes in pH_i were detected in cells exposed to ionophore. The increase in [Ca²⁺]_i was biphasic and characterized by an initial peak at 1-2 minutes followed by a sustained second phase. The addition of EGTA (2 mM) to the extracellular medium abolished gramicidin D induced increase in [Ca²⁺]_i and insulin secretion. These parameters were also profoundly inhibited by the L-type Ca²⁺-channel inhibitor, verapamil (20 μM). These findings suggest that insulin secretion induced from βTC3 cells by gramicidin D is mediated via the promotion of Ca²⁺-influx.