Calbindin-D-28k controls [Ca2+](i) and insulin release - Evidence obtainedfrom calbindin-D-28k knockout mice and beta cell lines

The role of the calcium-binding protein, calbindin-D-28k in potassium/depol arization-stimulated increases in the cytosolic free Ca2+ concentration ([C a2+](i)) and insulin release was investigated in pancreatic islets from cal bindin-D-28k nullmutant mice (knockouts; KO) or wild type mice and beta cel l lines stably transfected and overexpressing calbindin. Using single islet s from KO mice and stimulation with 45 mM KCl, the peak of [Ca2+](i) was 3. 5-fold greater in islets from KO mice compared with wild type islets (p < 0 .01) and [Ca2+](i) remained higher during the plateau phase. In addition to the increase in [Ca2+](i) in response to KCl there was also a significant increase in insulin release in islets isolated from KO mice. Evidence for m odulation by calbindin of [Ca2+](i) and insulin release was also noted usin g beta cell lines. Rat calbindin was stably expressed in beta TC-3 and beta HC-13 cells. In response to depolarizing concentrations of K+, insulin rel ease was decreased by 45-47% in calbindin expressing beta TC cells and was decreased by 70-80% in calbindin expressing beta HC cells compared with ins ulin release from vector transfected beta TC or beta HC cells (p < 0.01). I n addition, the K+-stimulated intracellular calcium peak was markedly inhib ited in calbindin expressing beta HC cells compared with vector transfected cells (225 nM versus 1,100 nm, respectively). Buffering of the depolarizat ion-induced rise in [Ca2+](i) was also observed in calbindin expressing bet a TC cells. In summary, our findings, using both isolated islets from calbi ndin-D-28k KO mice and beta cell lines, establish a role for calbindin in t he modulation of depolarization-stimulated insulin release and suggest that calbindin can control the rate of insulin release via regulation of [Ca2+] (i).