Deletion of the α2δ-1 calcium channel subunit increases excitability of mouse chromaffin cells

Abstract

High voltage-gated Ca²⁺ channels (HVCCs) shape the electrical activity and control hormone release in most endocrine cells. HVCCs are multi-subunit protein complexes formed by the pore-forming α₁ and the auxiliary β, α₂δ and γ subunits. Four genes code for the α₂δ isoforms. At the mRNA level, mouse chromaffin cells (MCCs) express predominantly the CACNA2D1 gene coding for the α₂δ-1 isoform. Here we show that α₂δ-1 deletion led to ∼60% reduced HVCC Ca²⁺ influx with slower inactivation kinetics. Pharmacological dissection showed that HVCC composition remained similar in α₂δ-1^-/- MCCs compared to wild-type (WT), demonstrating that α₂δ-1 exerts similar functional effects on all HVCC isoforms. Consistent with reduced HVCC Ca²⁺ influx, α₂δ-1^-/- MCCs showed reduced spontaneous electrical activity with action potentials (APs) having a shorter half-maximal duration caused by faster rising and decay slopes. However, the induced electrical activity showed opposite effects with α₂δ-1^-/- MCCs displaying significantly higher AP frequency in the tonic firing mode as well as an increase in the number of cells firing AP bursts compared to WT. This gain-of-function phenotype was caused by reduced functional activation of Ca²⁺-dependent K⁺ currents. Additionally, despite the reduced HVCC Ca²⁺ influx, the intracellular Ca²⁺ transients and vesicle exocytosis or endocytosis were unaltered in α₂δ-1^-/- MCCs compared to WT during sustained stimulation. In conclusion, our study shows that α₂δ-1 genetic deletion reduces Ca²⁺ influx in cultured MCCs but leads to a paradoxical increase in catecholamine secretion due to increased excitability. KEY POINTS: Deletion of the α₂δ-1 high voltage-gated Ca²⁺ channel (HVCC) subunit reduces mouse chromaffin cell (MCC) Ca²⁺ influx by ∼60% but causes a paradoxical increase in induced excitability. MCC intracellular Ca²⁺ transients are unaffected by the reduced HVCC Ca²⁺ influx. Deletion of α₂δ-1 reduces the immediately releasable pool vesicle exocytosis but has no effect on catecholamine (CA) release in response to sustained stimuli. The increased electrical activity and CA release from MCCs might contribute to the previously reported cardiovascular phenotype of patients carrying α₂δ-1 loss-of-function mutations.

Keywords: CACNA2D1; CaV; HVCC; MCCs; adrenaline; calcium channel; catecholamine; chromaffin cell; α2δ.