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High voltage-gated Ca 2+ 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 α 1 and the auxiliary β, α 2 δ and γ subunits. Four genes code for the α 2 δ isoforms. At the mRNA level, mouse chromaffin cells (MCCs) express predominantly the CACNA2D1 gene coding for the α 2 δ-1 isoform. Here we show that α 2 δ-1 deletion led to ∼60% reduced HVCC Ca 2+ influx with slower inactivation kinetics. Pharmacological dissection showed that HVCC composition remained similar in α 2 δ-1 -/- MCCs compared to wild-type (WT), demonstrating that α 2 δ-1 exerts similar functional effects on all HVCC isoforms. Consistent with reduced HVCC Ca 2+ influx, α 2 δ-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 α 2 δ-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 Ca 2+ -dependent K + currents. Additionally, despite the reduced HVCC Ca 2+ influx, the intracellular Ca 2+ transients and vesicle exocytosis or endocytosis were unaltered in α 2 δ-1 -/- MCCs compared to WT during sustained stimulation. In conclusion, our study shows that α 2 δ-1 genetic deletion reduces Ca 2+ influx in cultured MCCs but leads to a paradoxical increase in catecholamine secretion due to increased excitability. KEY POINTS: Deletion of the α 2 δ-1 high voltage-gated Ca 2+ channel (HVCC) subunit reduces mouse chromaffin cell (MCC) Ca 2+ influx by ∼60% but causes a paradoxical increase in induced excitability. MCC intracellular Ca 2+ transients are unaffected by the reduced HVCC Ca 2+ influx. Deletion of α 2 δ-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 α 2 δ-1 loss-of-function mutations., (© 2024 The Author(s). The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Key Points: Tymothy syndrome (TS) is a multisystem disorder featuring cardiac arrhythmias, autism and adrenal gland dysfunction that originates from a de novo point mutation in the gene encoding the Cav1.2 (CACNA1C) L-type channel. To study the role of Cav1.2 channel signals in autism, the autistic TS2-neo mouse has been generated bearing the G406R point-mutation associated with TS type-2. Using heterozygous TS2-neo mice, we report that the G406R mutation reduces the rate of inactivation and shifts leftward the activation and inactivation of L-type channels, causing marked increase of resting Ca 2+ influx ('window' Ca 2+ current). The increased 'window current' causes marked reduction of Na V channel density, switches normal tonic firing to abnormal burst firing, reduces mitochondrial metabolism, induces cell swelling and decreases catecholamine release. Overnight incubations with nifedipine rescue Na V channel density, normal firing and the quantity of catecholamine released. We provide evidence that chromaffin cell malfunction derives from altered Cav1.2 channel gating., Abstract: L-type voltage-gated calcium (Cav1) channels have a key role in long-term synaptic plasticity, sensory transduction, muscle contraction and hormone release. A point mutation in the gene encoding Cav1.2 (CACNA1C) causes Tymothy syndrome (TS), a multisystem disorder featuring cardiac arrhythmias, autism spectrum disorder (ASD) and adrenal gland dysfunction. In the more severe type-2 form (TS2), the missense mutation G406R is on exon 8 coding for the IS6-helix of the Cav1.2 channel. The mutation causes reduced inactivation and induces autism. How this occurs and how Cav1.2 gating-changes alter cell excitability, neuronal firing and hormone release on a molecular basis is still largely unknown. Here, using the TS2-neo mouse model of TS we show that the G406R mutation altered excitability and reduced secretory activity in adrenal chromaffin cells (CCs). Specifically, the TS2 mutation reduced the rate of voltage-dependent inactivation and shifted leftward the activation and steady-state inactivation of L-type channels. This markedly increased the resting 'window' Ca 2+ current that caused an increased percentage of CCs undergoing abnormal action potential (AP) burst firing, cell swelling, reduced mitochondrial metabolism and decreased catecholamine release. The increased 'window' Ca 2+ current caused also decreased Na V channel density and increased steady-state inactivation, which contributed to the increased abnormal burst firing. Overnight incubation with the L-type channel blocker nifedipine rescued the normal AP firing of CCs, the density of functioning Na V channels and their steady-state inactivation. We provide evidence that CC malfunction derives from the altered Cav1.2 channel gating and that dihydropyridines are potential therapeutics for ASD., (© 2019 University of Torino. The Journal of Physiology © 2019 The Physiological Society.)

Voltage-gated calcium channels represent the sole mechanism converting electrical signals of excitable cells into cellular functions such as contraction, secretion and gene regulation. Specific voltage-sensing domains detect changes in membrane potential and control channel gating. Calcium ions entering through the channel function as second messengers regulating cell functions, with the exception of skeletal muscle, where Ca V 1.1 essentially does not function as a channel but activates calcium release from intracellular stores. It has long been known that calcium currents are dispensable for skeletal muscle contraction. However, the questions as to how and why the channel function of Ca V 1.1 is curtailed remained obscure until the recent discovery of a developmental Ca V 1.1 splice variant with normal channel functions. This discovery provided new means to study the molecular mechanisms regulating the channel gating and led to the understanding that in skeletal muscle, calcium currents need to be restricted to allow proper regulation of fibre type specification and to prevent mitochondrial damage., (© 2016 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)