Characterization of a Ca2+ release-activated nonselective cation current regulating membrane potential and [Ca2+]i oscillations in transgenically derived beta-cells.

Although stimulation of insulin secretion by glucose is regulated by coupled oscillations of membrane potential and intracellular Ca2+ ([Ca2+]i), the membrane events regulating these oscillations are incompletely understood. In the presence of glucose and tetraethylammonium, transgenically derived beta-cells (betaTC3-neo) exhibit coupled voltage and [Ca2+]i oscillations strikingly similar to those observed in normal islets in response to glucose. Using these cells as a model system, we investigated the membrane conductance underlying these oscillations. Alterations in delayed rectifier or Ca2+-activated K+ channels were excluded as a source of the conductance oscillations, as they are completely blocked by tetraethylammonium. ATP-sensitive K+ channels were also excluded, since the ATP-sensitive K+ channel blocker tolbutamide substituted for glucose in inducing [Ca2+]i oscillations. Thapsigargin, which depletes intracellular Ca2+ stores, and maitotoxin, an activator of nonselective cation channels, both converted the glucose-dependent [Ca2+]i oscillations into a sustained elevation. On the other hand, both SKF 96365, a blocker of Ca2+ store-operated channels, and external Na+ removal suppressed the glucose-stimulated [Ca2+]i oscillations. Maitotoxin activated a nonselective cation current in betaTC3 cells that was attenuated by removal of extracellular Na+ and by SKF 96365, in the same manner to a current activated in mouse beta-cells following depletion of intracellular Ca2+ stores. Currents similar to these are produced by the mammalian trp-related channels, a gene family that includes Ca2+ store-operated channels and inositol 1,4,5-trisphosphate-activated channels. We found several of the trp family genes were expressed in betaTC3 cells by reverse transcriptase polymerase chain reaction using specific primers, but by Northern blot analysis, mtrp-4 was the predominant message expressed. We conclude that a conductance underlying glucose-stimulated oscillations in beta-cells is provided by a Ca2+ store depletion-activated nonselective cation current, which is plausibly encoded by homologs of trp genes.