Depolarization by transient receptor potential melastatin 4 in pancreatic alpha-cells regulates glucagon secretion

The Transient Receptor Potential Melastatin 4 protein (TRPM4) is a member of the TRP family of ion channels that is expressed in both electrically excitable and non-excitable cells. Functional studies revealed that TRPM4 significantly impacts Ca2+ signals in both immune and pancreatic β-cells, which is important for cellular processes such as hormone secretion. However, its role in glucagon secreting α-cells has not been reported. Type 2 Diabetes Mellitus is often associated with increased glucagon levels; yet, the exact mechanism controlling its secretion is not known. In pancreatic α-cells, an increase in intracellular Ca2+ concentration causes glucagon secretion. We hypothesize that TRPM4 is important for glucagon secretion in α-cells by controlling intracellular Ca2+ signals. In this study, we investigated TRPM4 expression in the α-cell lines INR1G9 (hamster) and αTC1-6 (mouse) and characterized the channel using the patch-clamp technique. By RT-PCR we identified TRPM4-transcripts in both cell lines examined. Furthermore, patch-clamp recordings with increasing intracellular Ca2+ concentrations resulted in a dose-dependent activation of TRPM4-like currents. The greatest depolarizing currents were obtained with 3μM Ca2+ concentration. The current-voltage relationship (I/V) resembled those previously described for TRPM4. In addition, we demonstrated the voltage dependency of the channel, where negative potentials inhibited and positive potentials increased channel activity. Finally, replacement of Na+ ions in the extracellular solution with N-methyl-D-glucamine significantly reduced the inward currents and caused a hyperpolarizing shift in the I/V, which affirms that the channel is Na+ permeable. These data demonstrate that TRPM4 is present and functional in pancreatic α-cells and suggest a potential role for the channel in glucagon secretion and glucose homeostasis. The role of TRPM4 in glucagon secretion was assessed using a stable TRPM4 knockdown αTC1-6 cell line. Calcium-imaging and glucagon secretion experiments revealed a relationship between the decreased intracellular Ca2+ concentration and glucagon secretion in TRPM4 knockdown cells compared to controls. These results indicate that depolarization by TRPM4 plays an important role in glucagon secretion and perhaps glucose homeostasis. Elucidation of the glucagon secretion pathway could lead to a treatment for hyperglucagonemia associated with Type 2 Diabetes