Adrenal chromaffin cells (CCs) are the main source of circulating catecholamines (CAs) that regulate the body response to stress. Release of CAs is controlled neurogenically by the activity of preganglionic sympathetic neurons through trains of action potentials (APs). APs in CCs are generated by robust depolarization following the activation of nicotinic and muscarinic receptors that are highly expressed in CCs. Bovine, rat, mouse, and human CCs also express a composite array of Na, K, and Ca channels that regulate the resting potential, shape the APs, and set the frequency of AP trains. AP trains of increasing frequency induce enhanced release of CAs. If the primary role of CCs is simply to relay preganglionic nerve commands to CA secretion, why should they express such a diverse set of ion channels? An answer to this comes from recent observations that, like in neurons, CCs undergo complex firing patterns of APs suggesting the existence of an intrinsic CC excitability (non-neurogenically controlled). Recent work has shown that CCs undergo occasional or persistent burst firing elicited by altered physiological conditions or deletion of pore-regulating auxiliary subunits. In this review, we aim to give a rationale to the role of the many ion channel types regulating CC excitability. We will first describe their functional properties and then analyze how they contribute to pacemaking, AP shape, and burst waveforms. We will also furnish clear indications on missing ion conductances that may be involved in pacemaking and highlight the contribution of the crucial channels involved in burst firing.

Mouse chromaffin cells (MCCs) generate action potential (AP) firing that regulates the Ca -dependent release of catecholamines (CAs). Recent findings indicate that MCCs possess a variety of spontaneous firing modes that span from the common 'tonic-irregular' to the less frequent 'burst' firing. This latter is evident in a small fraction of MCCs but occurs regularly when Nav1.3/1.7 channels are made less available or when the Slo1β2-subunit responsible for BK channel inactivation is deleted. Burst firing causes large increases of Ca -entry and potentiates CA release by ∼3.5-fold and thus may be a key mechanism for regulating MCC function. With the aim to uncover a physiological role for burst-firing we investigated the effects of acidosis on MCC activity. Lowering the extracellular pH (pH ) from 7.4 to 7.0 and 6.6 induces cell depolarizations of 10-15 mV that generate repeated bursts. Bursts at pH 6.6 lasted ∼330 ms, occurred at 1-2 Hz and caused an ∼7-fold increase of CA cumulative release. Burst firing originates from the inhibition of the pH-sensitive TASK-1/TASK-3 channels and from a 40% BK channel conductance reduction at pH 7.0. The same pH had little or no effect on Nav, Cav, Kv and SK channels that support AP firing in MCCs. Burst firing of pH 6.6 could be mimicked by mixtures of the TASK-1 blocker A1899 (300 nm) and BK blocker paxilline (300 nm) and could be prevented by blocking L-type channels by adding 3 μm nifedipine. Mixtures of the two blockers raised cumulative CA-secretion even more than low pH (∼12-fold), showing that the action of protons on vesicle release is mainly a result of the ionic conductance changes that increase Ca -entry during bursts. Our data provide direct evidence suggesting that MCCs respond to low pH with sustained depolarization, burst firing and enhanced CA-secretion, thus mimicking the physiological response of CCs to acute acidosis and hyperkalaemia generated during heavy exercise and muscle fatigue.

The accepted role of the protein Kv2.1 in arterial smooth muscle cells is to form K + channels in the sarcolemma. Opening of Kv2.1 channels causes membrane hyperpolarization, which decreases the activity of L-type Ca V 1.2 channels, lowering intracellular Ca 2+ ([Ca 2+ ] i ) and causing smooth muscle relaxation. A limitation of this model is that it is based exclusively on data from male arterial myocytes. Here, we used a combination of electrophysiology as well as imaging approaches to investigate the role of Kv2.1 channels in male and female arterial myocytes. We confirmed that Kv2.1 plays a canonical conductive role but found it also has a structural role in arterial myocytes to enhance clustering of Ca V 1.2 channels. Less than 1% of Kv2.1 channels are conductive and induce membrane hyperpolarization. Paradoxically, by enhancing the structural clustering and probability of Ca V 1.2-Ca V 1.2 interactions within these clusters, Kv2.1 increases Ca 2+ influx. These functional impacts of Kv2.1 depend on its level of expression, which varies with sex. In female myocytes, where expression of Kv2.1 protein is higher than in male myocytes, Kv2.1 has conductive and structural roles. Female myocytes have larger Ca V 1.2 clusters, larger [Ca 2+ ] i , and larger myogenic tone than male myocytes. In contrast, in male myocytes, Kv2.1 channels regulate membrane potential but not Ca V 1.2 channel clustering. We propose a model in which Kv2.1 function varies with sex: in males, Kv2.1 channels control membrane potential but, in female myocytes, Kv2.1 plays dual electrical and Ca V 1.2 clustering roles. This contributes to sex-specific regulation of excitability, [Ca 2+ ] i , and myogenic tone in arterial myocytes.

The cardiac cycle starts when an action potential is produced by pacemaking cells in the sino-atrial node. This cycle is repeated approximately 100,000 times in humans and 1 million times in mice per day, imposing a monumental metabolic demand on the heart, requiring efficient blood supply via the coronary vasculature to maintain cardiac function. Although the ventricular coronary circulation has been extensively studied, the relationship between vascularization and cellular pacemaking modalities in the sino-atrial node is poorly understood. Here, we tested the hypothesis that the organization of the sino-atrial node micro-vasculature varies regionally, reflecting local myocyte firing properties. We show that vessel densities are higher in the superior versus inferior sino-atrial node. Accordingly, sino-atrial node myocytes are closer to vessels in the superior versus inferior regions. Superior and inferior sino-atrial node myocytes produce stochastic subthreshold voltage fluctuations and action potentials. However, the intrinsic action potential firing rate of sino-atrial node myocytes is higher in the superior versus inferior node. Our data support a model in which the micro-vascular densities vary regionally within the sino-atrial node to match the electrical and Ca2+ dynamics of nearby myocytes, effectively determining the dominant pacemaking site within the node. In this model, the high vascular density in the superior sino-atrial node places myocytes with metabolically demanding, high frequency action potentials near vessels. The lower vascularization and electrical activity of inferior sino-atrial node myocytes could limit these cells to function to support sino-atrial node periodicity with sporadic voltage fluctuations via a stochastic resonance mechanism.

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 Ca2+ influx (‘window’ Ca2+ current). The increased ‘window current’ causes marked reduction of NaV 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 NaV 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’ Ca2+ 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’ Ca2+ current caused also decreased NaV 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 NaV 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.