Subplasmalemmal Ca(2+) measurements in mouse pancreatic beta cells support the existence of an amplifying effect of glucose on insulin secretion.

aims/hypothesisGlucose-induced insulin secretion is attributed to a rise of beta cell cytosolic free [Ca(2+)] ([Ca(2+)](c)) (triggering pathway) and amplification of the action of Ca(2+). This concept of amplification rests on observations that glucose can increase Ca(2+)-induced insulin secretion without further elevating an imposed already high [Ca(2+)](c). However, it remains possible that this amplification results from an increase in [Ca(2+)] just under the plasma membrane ([Ca(2+)](SM)), which escaped detection by previous measurements of global [Ca(2+)](c). This was the hypothesis that we tested here by measuring [Ca(2+)](SM).

methodsThe genetically encoded Ca(2+) indicators D3-cpv (untargeted) and LynD3-cpv (targeted to plasma membrane) were expressed in clusters of mouse beta cells. LynD3-cpv was also expressed in beta cells within intact islets. [Ca(2+)](SM) changes were monitored using total internal reflection fluorescence microscopy. Insulin secretion was measured in parallel.

resultsBeta cells expressing D3cpv or LynD3cpv displayed normal [Ca(2+)] changes and insulin secretion in response to glucose. Distinct [Ca(2+)](SM) fluctuations were detected during repetitive variations of KCl between 30 and 32-35 mmol/l, attesting to the adequate sensitivity of our system. When the amplifying pathway was evaluated (high KCl + diazoxide), increasing glucose from 3 to 15 mmol/l consistently lowered [Ca(2+)](SM) while stimulating insulin secretion approximately two fold. Blocking Ca(2+) uptake by the endoplasmic reticulum largely attenuated the [Ca(2+)](SM) decrease produced by high glucose but did not unmask localised [Ca(2+)](SM) increases. CONCLUSIONS/

interpretationGlucose can increase Ca(2+)-induced insulin secretion without causing further elevation of beta cell [Ca(2+)](SM). The phenomenon is therefore a true amplification of the triggering action of Ca(2+).