Activation of KATP channels by H2S in rat insulin‐secreting cells and the underlying mechanisms

Abstract

H 2 S is an important gasotransmitter, generated in mammalian cells from l‐cysteine metabolism. As it stimulates K ATP channels in vascular smooth muscle cells, H 2 S may also function as an endogenous opener of K ATP channels in INS‐1E cells, an insulin‐secreting cell line. In the present study, K ATP channel currents in INS‐1E cells were recorded using the whole‐cell and single‐channel recording configurations of the patch‐clamp technique. K ATP channels in INS‐1E cells have a single‐channel conductance of 78 pS. These channels were activated by diazoxide and inhibited by gliclazide. ATP (3 m m) in the pipette solution inhibited K ATP channels in INS‐1E cells. Significant amount of H 2 S was produced from INS‐1E cells in which the expression of cystathinonie gamma‐lyase (CSE) was confirmed. After INS‐1E cells were transfected with CSE‐targeted short interfering RNA (CSE‐siRNA) or treated with dl‐propargylglycine (PPG; 1–5 m m) to inhibit CSE, endogenous production of H 2 S was abolished. Increase in extracellular glucose concentration significantly decreased endogenous production of H 2 S in INS‐1E cells, and increased insulin secretion. After transfection of INS‐1E cells with adenovirus containing the CSE gene (Ad‐CSE) to overexpress CSE, high glucose‐stimulated insulin secretion was virtually abolished. Basal K ATP channel currents were significantly reduced after incubating INS‐1E cells with a high glucose concentration (16 m m) or lowering endogenous H 2 S level by CSE‐siRNA transfection. Under these conditions, exogenously applied H 2 S significantly increased whole‐cell K ATP channel currents at concentrations equal to or lower than 100 μ m. H 2 S (100 μ m) markedly increased open probability by more than 2‐fold of single K ATP channels (inside‐out recording) in native INS‐1E cells (n= 4, P < 0.05). Single‐channel conductance and ATP sensitivity of K ATP channels were not changed by H 2 S. In conclusion, endogenous H 2 S production from INS‐1E cells varies with in vivo conditions, which significantly affects insulin secretion from INS‐1E cells. H 2 S stimulates K ATP channels in INS‐1E cells, independent of activation of cytosolic second messengers, which may underlie H 2 S‐inhibited insulin secretion from these cells. Interaction among H 2 S, glucose and the K ATP channel may constitute an important and novel mechanism for the fine control of insulin secretion from pancreatic β‐cells.