Hydrogen sulfide (H(2)S) has been traditionally known for its toxic effects on living organisms. The role of H(2)S in the homeostatic regulation of pancreatic insulin metabolism has been unclear. The present study is aimed at elucidating the effect of endogenously produced H(2)S on pancreatic insulin release and its role in diabetes development. Diabetes development in Zucker diabetic fatty (ZDF) rats was evaluated in comparison with Zucker fatty (ZF) and Zucker lean (ZL) rats. Pancreatic H(2)S production and insulin release were also assayed. It was found that H(2)S was generated in rat pancreas islets, catalyzed predominantly by cystathionine gamma-lyase (CSE). Pancreatic CSE expression and H(2)S production were greater in ZDF rats than in ZF or ZL rats. ZDF rats exhibited reduced serum insulin level, hyperglycemia, and insulin resistance. Inhibition of pancreatic H(2)S production in ZDF rats by intraperitoneal injection of DL-propargylglycine (PPG) for 4 weeks increased serum insulin level, lowered hyperglycemia, and reduced hemoglobin A1c level (P<0.05). Although in ZF rats it also reduced pancreatic H(2)S production and serum H(2)S level, PPG treatment did not alter serum insulin and glucose level. Finally, H(2)S significantly increased K(ATP) channel activity in freshly isolated rat pancreatic beta-cells. It appears that insulin release is impaired in ZDF because of abnormally high pancreatic production of H(2)S. New therapeutic approach for diabetes management can be devised based on our observation by inhibiting endogenous H(2)S production from pancreas.