| 10212237 |
Maechler P, Kennedy ED, Sebo E, Valeva A, Pozzan T, Wollheim CB: Secretagogues modulate the calcium concentration in the endoplasmic reticulum of insulin-secreting cells. J Biol Chem. 1999 Apr 30;274(18):12583-92.
Studies in aequorin-expressing intact and permeabilized ins-1 cells.. The precise regulation of the Ca2+ concentration in the endoplasmic reticulum ([Ca2+] er) is important for protein processing and signal transduction. In the pancreatic beta-cell, dysregulation of [Ca2+] er may cause impaired insulin secretion. The Ca2+-sensitive photoprotein aequorin mutated to lower its Ca2+ affinity was stably expressed in the endoplasmic reticulum (ER) of rat insulinoma INS-1 cells. The steady state [Ca2+] er was 267 +/- 9 microM. Both the Ca2+-ATPase inhibitor cyclopiazonic acid and 4-chloro-m-cresol, an activator of ryanodine receptors, caused an almost complete emptying of ER Ca2+. The inositol 1,4,5-trisphosphate generating agonists, carbachol, and ATP, reduced [Ca2+] er by 20-25%. Insulin secretagogues that raise cytosolic [Ca2+] by membrane depolarization increased [Ca2+] er in the potency order K+ >> glucose > leucine, paralleling their actions in the cytosolic compartment. Glucose, which augmented [Ca2+] er by about 25%, potentiated the Ca2+-mobilizing effect of carbachol, explaining the corresponding observation in cytosolic [Ca2+]. The filling of ER Ca2+ by glucose is not directly mediated by ATP production as shown by the continuous monitoring of cytosolic ATP in luciferase expressing cells. Both glucose and K+ increase [Ca2+] er, but only the former generated whereas the latter consumed ATP. Nonetheless, drastic lowering of cellular ATP with a mitochondrial uncoupler resulted in a marked decrease in [Ca2+] er, emphasizing the requirement for mitochondrially derived ATP above a critical threshold concentration. Using alpha-toxin permeabilized cells in the presence of ATP, glucose 6-phosphate did not change [Ca2+] er, invalidating the hypothesis that glucose acts through this metabolite. Therefore, insulin secretagogues that primarily stimulate Ca2+ influx, elevate [Ca2+] er to ensure beta-cell homeostasis. |
1(0,0,0,1) |