Dichotomy of calcium 1.2 and calcium 1.3 L-type calcium channel function and modulation in GLP-1-mediated insulin exocytosis and ERK1/2 activation in pancreatic beta-cells

Activation of the Class B Gαs/Gαq-coupled Glucagon-Like Peptide-1 Receptor promotes multiple Ca2+-requiring processes that collectively enhance insulin secretion and proliferation of the pancreatic β-cell. In rats and humans, L-Type Voltage-Gated Ca2+ Channels (L-VGCCs) of the Cav1.2 (α1C) and Cav1.3 (α1D) subtypes are predominantly expressed, however their precise function and modulation by nutrients or second messengers remain unclear. Using a pharmacological knockdown system of Dihydropyridine-insensitive (DHPi) mutant channels, we examined the independent roles of Cav1.2 and Cav1.3 in two parallel processes mediated by the GLP-1: glucose-stimulated insulin secretion (GSIS) and ERK1/2 phosphorylation (GSEP). In the rat insulinoma INS-1 β-cells, GLP-1-potentiated GSIS and GSEP are preferentially coupled to Cav1.3. The GLP-1R – Cav1.3 cross-talk is mediated by spatiotemporal delimitation of cAMP by Phosphodiesterases, positive modulation of Cav1.3 by PKA and PKC, and efficient coupling of Cav1.3 with ER Ca2+ stores. The II-III interdomain loop of the Cav1.2 and Cav1.3 pore subunits are necessary but insufficient in mediating excitation-secretion coupling via both L-VGCCs. Overexpression of the peptides corresponding to the Cav1.2/II-III or the Cav1.3/II-III interdomain loops attenuated GLP-1-potentiated GSIS and GSEP, and reduced cAMP production by 50%, without affecting the response to forskolin. Isolation of detergent-insoluble membrane fractions from control INS-1 cells, and those that overexpress Cav1.2/II-III or the Ca v1.3/II-III showed a shift in distribution of Cav1.2 and Cav1.3 from lipid raft fractions to detergent-soluble non-lipid raft fractions, without altering the distribution of Caveolin-1. Thus, GLP-1 – potentiated GSIS and GSEP require the targeting of L-VGCCs to Caveolin-rich membrane microdomains in a manner that is facilitated by the II-III interdomain loop of the pore subunit. Lastly, we report the design and validation of an incretin-mimetic peptide derived from the autoactivation domain of the GLP-1R. The synthetic peptide showed cAMP-independent efficacy in potentiating GSIS and GSEP, and thus has potential as a novel class of Diabetes therapy