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

AIMS/HYPOTHESIS: Glucose-induced insulin secretion is attributed to a rise of beta cell cytosolic free [Ca(2+)] (Ca(2+)) (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+). However, it remains possible that this amplification results from an increase in [Ca(2+)] just under the plasma membrane (Ca(2+)), which escaped detection by previous measurements of global Ca(2+). This was the hypothesis that we tested here by measuring Ca(2+).

METHODS

The 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+) changes were monitored using total internal reflection fluorescence microscopy. Insulin secretion was measured in parallel.

RESULTS

Beta cells expressing D3cpv or LynD3cpv displayed normal [Ca(2+)] changes and insulin secretion in response to glucose. Distinct Ca(2+) 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+) while stimulating insulin secretion approximately two fold. Blocking Ca(2+) uptake by the endoplasmic reticulum largely attenuated the Ca(2+) decrease produced by high glucose but did not unmask localised Ca(2+) increases.

CONCLUSIONS/INTERPRETATION: Glucose can increase Ca(2+)-induced insulin secretion without causing further elevation of beta cell Ca(2+). The phenomenon is therefore a true amplification of the triggering action of Ca(2+).