The phytostilbene resveratrol has been shown to improve the metabolic state in animal models by increasing the insulin responsiveness of tissues and to directly affect insulin secretion from native beta-cells and insulinoma cells. We performed short (1 hour) and long term (24 to 72 hours) experiments to study the effect of the drug (10–100 mM) on stimulus-secretion-coupling, insulin release, proliferation and apoptosis of INS-1E cells. In whole cell patch clamp experiments resveratrol reversibly suppressed electrical activity elicited by high glucose, hypotonicity or tolbutamide by blocking L- and T-type calcium currents and swelling-dependent chloride currents and completely inhibited glucose-induced insulin release within 1 hour. Long term exposure to resveratrol at concentrations >50 M caused a time- and concentration-dependent inhibition of insulin release, cell cycle arrest in the S and G0/G1 phase, increased cell granularity and induced apoptotic volume decrease (AVD), phosphatidylserine exposure and caspase activation. The AMP-kinase (AMPK) inhibitor 6-[4-(2-piperidin-1-ylethoxy)phenyl]-3-pyridin-4-ylpyrazolo[1,5-a]pyrimidine (compound C; 10 mM) significantly inhibited cell proliferation and induced caspase activation within 48 hours but this effect was independent on the absence or presence of resveratrol suggesting that AMPK is not involved in mediating the proapoptotic effect of resveratrol in INS-1E cells. Immunoblotting revealed a concentration-dependent inhibition of Akt (PKB) phosphorylation by 50 and 100 mM resveratrol, which could be attenuated by addition of insulin (10 mM) to the culture medium. However, insulin did not alleviate resveratrol-induced inhibition of proliferation or caspase activation. We conclude that the antiproliferative/proapoptotic effect of resveratrol on INS-1E cells is due to negative interference with Akt signaling and disruption of auto/paracrine insulin signaling.