cPLA2α-evoked formation of arachidonic acid and lysophospholipids is required for exocytosis in mouse pancreatic β-cells

Using capacitance measurements, we investigated the effects of intracellularly applied recombinant human cytosolic phospholipase A(2) (cPLA(2alpha)) and its lipolytic products arachidonic acid and lysophosphatidylcholine on Ca2+-dependent exocytosis in single mouse pancreatic beta-cells. cPLA(2alpha) dose dependently (EC50 = 86 nM) stimulated depolarization-evoked exocytosis by 450% without affecting the whole cell Ca2+ current or cytoplasmic Ca2+ levels. The stimulatory effect involved priming of secretory granules as reflected by an increase in the size of the readily releasable pool of granules from 70-80 to 280-300. cPLA(2alpha)-stimulated exocytosis was antagonized by the specific cPLA(2) inhibitor AACOCF(3). Ca2+-evoked exocytosis was reduced by 40% in cells treated with AACOCF(3) or an antisense oligonucleotide against cPLA(2alpha). The action of cPLA(2alpha) was mimicked by a combination of arachidonic acid and lysophosphatidylcholine (470% stimulation) in which each compound alone doubled the exocytotic response. Priming of insulin-containing secretory granules has been reported to involve Cl- uptake through ClC-3 Cl channels. Accordingly, the stimulatory action of cPLA(2alpha) was inhibited by the Cl- channel inhibitor DIDS and in cells pretreated with CIC-3 Cl- channel antisense oligonucleotides. We propose that cPLA(2alpha) has an important role in controlling the rate of exocytosis in beta-cells. This effect of cPLA(2alpha) reflects an enhanced transgranular Cl- flux, leading to an increase in the number of granules available for release, and requires the combined actions of arachidonic acid and lysophosphatidylcholine.