monensin and Insulinoma

Plasma membrane vesicles from a glucose-responsive insulinoma exhibited properties consistent with the presence of a membrane Na+/Ca2+ exchange. The exchange was rapid, reversible, and was dependent on the external Ca2+ concentration (Km = 4.1 +/- 1.1 microM). External Na+ inhibited the uptake in a dose-dependent manner (IC50 = 15 mM). Dissipation of the Na+ gradient by 10 microM monensin decreased Na+/Ca2+ exchange from 0.74 +/- 0.17 nmoles/mg protein/s to 0.11 +/- 0.05 nmoles/mg protein/s. Exchange was not influenced by veratridine, tetrodotoxin and ouabain, or by modifiers of cAMP. No effect was seen using the calcium channel blockers, nitrendipine or nifedipine. Glucose had no direct effect on Na+/Ca2+ exchange, while glyceraldehyde, glyceraldehyde-3-phosphate and dihydroxyacetone inhibited the exchange. Na+ induced efflux of calcium was seen in Ca2+ loaded vesicles and was half maximal at [Na+] of 11.1 +/- 0.75 mM. Ca2+ efflux was dependent on [Na+], with a Hill coefficient of 2.7 +/- 0.07 indicating that activation of Ca2+ release involves a minimum of three sites. The electrogenicity of this exchange was demonstrated using the lipophilic cation tetraphenylphosphonium [( 3H]-TPP), a membrane potential sensitive probe. [3H]-TPP uptake increased transiently during Na+/Ca2+ exchange indicating that the exchange generated a membrane potential. These results show that Na+/Ca2+ exchange operates in the beta cell and may be an important regulator of intracellular free Ca2+ concentrations.

Topics: Animals; Calcium; Cell Membrane; Glyceraldehyde; Insulinoma; Ion Pumps; Male; Monensin; Onium Compounds; Organophosphorus Compounds; Rats; Rats, Inbred Strains; Sodium; Sodium-Potassium-Exchanging ATPase

The In-R1-G9 cell line is one of the clones derived from the In-111-R1 hamster insulinoma cell line and produces glucagon. The secretory responses of In-R1-G9 cells were further examined to characterize the nature of the cells. Vincristine had no effect on glucagon secretion and colchicine enhanced glucagon secretion slightly after a short incubation. Two calmodulin inhibitors, trifluoperazine and chlorpromazine, did not affect glucagon secretion. Monensin at 10(-8) M suppressed glucagon secretion by 50%. Secretion of glucagon was calcium-dependent. The addition of A23187 to the incubation medium resulted in a 180% increase over control for 1 h and calcium deprivation from the medium suppressed glucagon secretion markedly. Theophylline, a phosphodiesterase inhibitor, caused a 230% increase in glucagon secretion. An experiment using cycloheximide suggested that newly synthesized glucagon appears in the medium at 30 min. This cell line should be useful for various experiments in many fields of research.

Topics: Adenoma, Islet Cell; Animals; Calcium; Clone Cells; Colchicine; Cricetinae; Cycloheximide; Cytoplasmic Granules; Glucagon; Insulinoma; Monensin; Pancreatic Neoplasms; Tumor Cells, Cultured