Control of Insulin Release by Transient Receptor Potential Melastatin 3 (TRPM3) Ion Channels

Abstract

Background/aims: The release of insulin in response to increased levels of glucose in the blood strongly depends on Ca²⁺ influx into pancreatic beta cells by the opening of voltage-gated Ca²⁺ channels. Transient Receptor Potential Melastatin 3 proteins build Ca²⁺ permeable, non-selective cation channels serving as pain sensors of noxious heat in the peripheral nervous system. TRPM3 channels are also strongly expressed in pancreatic beta cells that respond to the TRPM3 agonist pregnenolone sulfate with Ca²⁺ influx and increased insulin release. Therefore, we hypothesized that in beta cells TRPM3 channels may contribute to pregnenolone sulfate- as well as to glucose-induced insulin release.

Methods: We used INS-1 cells as a beta cell model in which we analysed the occurrence of TRPM3 isoformes by immunoprecipitation and western blotting and by cloning of RT-PCR amplified cDNA fragments. We applied pharmacological as well as CRISPR/Cas9-based strategies to analyse the interplay of TRPM3 and voltage-gated Ca²⁺ channels in imaging experiments (FMP, Fura-2) and electrophysiological recordings. In immunoassays, we examined the contribution of TRPM3 channels to pregnenolone sulfate- and glucose-induced insulin release. To confirm our findings, we generated beta cell-specific Trpm3-deficient mice and compared their glucose clearance with the wild type in glucose tolerance tests.

Results: TRPM3 channels triggered the activity of voltage-gated Ca²⁺ channels and both channels together contributed to insulin release after TRPM3 activation. Trpm3-deficient INS-1 cells lacked pregnenolone sulfate-induced Ca²⁺ signals just like the pregnenolone sulfate-induced insulin release. Both, glucose-induced Ca²⁺ signals and the glucose-induced insulin release were strongly reduced. Accordingly, Trpm3-deficient mice displayed an impaired decrease of the blood sugar concentration after intraperitoneal or oral administration of glucose.

Conclusion: The present study suggests an important role for TRPM3 channels in the control of glucose-dependent insulin release.

Keywords: Transient receptor potential M3 channels (TRPM3); Calcium; Glucose-stimulated insulin secretion; CRISPR/Cas; INS-1; Trpm3 knockout.