Correlation between cytosolic free Ca2+ and insulin release in an insulin-secreting cell line

Changes in the concentration of free cytosolic Ca2+ [( Ca2+]i), membrane potential, and immunoreactive insulin release were measured in suspensions of RINm5F cells. [Ca2+]i was monitored with the intracellularly trapped fluorescent Ca2+ indicator quin 2. Changes in membrane potential were assessed with the fluorescent probe bisoxonol. The effects of depolarizing K+ concentrations and of the Ca2+ ionophore ionomycin were compared with those of the metabolizable stimuli glyceraldehyde and alanine. The mean resting [Ca2+]i was 105 nM +/- 6, n = 35. Of the substances tested, ionomycin caused the most marked increases of [Ca2+]i and insulin release. Membrane depolarization was evoked by K+, glyceraldehyde, or alanine and the agent stimulated insulin release and [Ca2+]i to a similar degree. However, the mechanism by which glyceraldehyde, on the one hand, and K+ and alanine, on the other, elevate [Ca2+]i appears different. Thus, verapamil, a blocker of voltage-sensitive Ca2+ channels, almost abolished the effects of K+ and alanine on [Ca2+]i and insulin release, whereas the responses to glyceraldehyde were only blocked by approximately 50%. Glyceraldehyde may thus, in addition to opening voltage-sensitive Ca2+ channels, alter cellular Ca2+ handling at another step. The present findings provide direct experimental evidence that secretagogue-induced insulin release is accompanied by a rise in [Ca2+]i.