Search

Calcium influx via the L‐type voltage‐gated Cav1.2 calcium channel in smooth muscle cells regulates vascular contraction. Calcium channel blockers (CCBs) are widely used to treat hypertension by inhibiting Cav1.2 channels. Using the vascular smooth muscle cell line, A7r5 and primary culture of cerebral vascular smooth muscle cells, we found that the expression and function of Cav1.2 channels are downregulated during hypoxia. Furthermore, hypoxia induces structural changes in Cav1.2 channels via alternative splicing. The expression of exon 9* is upregulated, whereas exon 33 is downregulated. Such structural alterations of Cav1.2 channels are caused by the decreased expression of RNA‐binding proteins RNA‐binding protein fox‐1 homolog 1 and 2 (RbFox1 and RbFox2). Overexpression of RbFox1 and RbFox2 prevents hypoxia‐induced exon 9* inclusion and exon 33 exclusion. Importantly, such structural alterations of the Cav1.2 channel partly contribute to the enhanced sensitivity of Cav1.2 to isradipine (a CCB) under hypoxia. Overexpression of RbFox1 and RbFox2 successfully reduces isradipine sensitivity in hypoxic smooth muscle cells. Our results suggest a new strategy to manage ischemic diseases such as stroke and myocardial infarction. [ABSTRACT FROM AUTHOR]

Pre-exercise passive heating attenuates muscle damage caused by eccentric exercise in rats where the induction of heat shock proteins (HSPs) confers a myoprotective effect. We investigated whether pre-exercise hot water immersion (HWI) confers similar benefits in humans. Eleven recreational male athletes were immersed in 41°C water up to 60 min or until rectal temperatures reached 39.5°C. After a 6 h rest, the participants performed an eccentric downhill run for 1 h at −4% gradient to induce muscle damage. An endurance capacity test at 75% VO2max was conducted 18 h later. The control trial was similar except that participants were immersed at 34°C. Blood samples were collected to assess HSPs levels, creatine kinase, and lactate dehydrogenase activities. Plasma eHSP70 was higher post-immersion in HWI trials (1.3 ± 0.4 vs 1.1 ± 0.4; p = 0.005). Plasma eHSP27 was higher before (p = 0.049) and after (p = 0.015) endurance test in HWI. Leukocytic p-HSP27 was increased 18 h after HWI (0.97 ± 0.14 vs 0.67 ± 0.11; p = 0.04). Creatine kinase and lactate dehydrogenase activities were increased by 3-fold and 1.5-fold, respectively, after endurance test in HWI but did not differ across trials (p > 0.05). Mean heart rates were higher during eccentric run and endurance test in HWI as compared to control (p < 0.05). Endurance capacity was similar between trials (57.3 ± 11.5 min vs 55.0 ± 13.5 min; p = 0.564). Pre-exercise heating increased the expression of plasma eHSPs and leukocytic p-HSP27 but did not reduce muscle damage nor enhance endurance capacity. [ABSTRACT FROM AUTHOR]

Background: L-type CaV1.2 calcium channel, the primary gateway for Ca2+ influx in smooth muscles, is widely regulated by multiple posttranslational modifications, such as protein kinase-mediated phosphorylation and nitric oxide-induced S-nitrosylation. However, the effect of S-nitrosylation on CaV1.2 channel function and its role in arterial contractility are not well understood.Methods: Electrophysiological recordings, Ca2+ and confocal imaging, and biochemical assays were used to functionally characterize S-nitrosylated CaV1.2 channels in vitro, while pressure myography and tail-cuff blood pressure measurement were conducted to evaluate the physiological effects of CaV1.2 S-nitrosylation ex vivo and in vivo.Results: S-nitrosylation significantly reduced the CaV1.2 current density by promoting lysosomal degradation that leads to decreased levels of total and surface CaV1.2 channel proteins in a CaVβ-independent manner and reducing the open probability of CaV1.2 channel. Mechanistically, the Cys1180 and Cys1280 residues within CaV1.2 channel have been determined as the molecular targets for S-nitrosylation as substitution of either Cys1180 or Cys1280 for serine resulted in substantial reduction of S-nitrosylation levels. Of note, CaV1.2 S-nitrosylation levels were significantly reduced in arteries isolated from both spontaneously hypertensive rats and patients with pulmonary hypertension. Moreover, mouse resistance arteries incubated with S-nitrosocysteine displayed much lower contractility and spontaneously hypertensive rats injected with S-nitrosocysteine also showed significantly reduced blood pressure, suggesting that reduced S-nitrosylation contributes to the upregulation of CaV1.2 channel activity in hypertensive arteries.Conclusions: This study provides strong evidence that S-nitrosylation-mediated downregulation of CaV1.2 channels is via 2 distinctive mechanisms and the findings offer potential pathways for therapeutic inventions in hypertension. [ABSTRACT FROM AUTHOR]

Calcium influx via the L-type voltage-gated Ca v 1.2 calcium channel in smooth muscle cells regulates vascular contraction. Calcium channel blockers (CCBs) are widely used to treat hypertension by inhibiting Ca v 1.2 channels. Using the vascular smooth muscle cell line, A7r5 and primary culture of cerebral vascular smooth muscle cells, we found that the expression and function of Ca v 1.2 channels are downregulated during hypoxia. Furthermore, hypoxia induces structural changes in Ca v 1.2 channels via alternative splicing. The expression of exon 9* is upregulated, whereas exon 33 is downregulated. Such structural alterations of Ca v 1.2 channels are caused by the decreased expression of RNA-binding proteins RNA-binding protein fox-1 homolog 1 and 2 (RbFox1 and RbFox2). Overexpression of RbFox1 and RbFox2 prevents hypoxia-induced exon 9* inclusion and exon 33 exclusion. Importantly, such structural alterations of the Ca v 1.2 channel partly contribute to the enhanced sensitivity of Ca v 1.2 to isradipine (a CCB) under hypoxia. Overexpression of RbFox1 and RbFox2 successfully reduces isradipine sensitivity in hypoxic smooth muscle cells. Our results suggest a new strategy to manage ischemic diseases such as stroke and myocardial infarction., (© 2024 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

A missense mutation c.1220C>G of KCN2A gene was recently identified in an infant with epilepsy. KCNA2 encodes K V 1.2 subunits that form voltage-gated potassium channels (VGKC) via tetrameric assembly. The mutation results in amino acid change P407R at the highly conserved PVP motif. Functional characterization revealed that mutant K V 1.2&#95;P407R subunits formed loss-of-function channels and suppressed both K V 1.2 and K V 1.1 channel activities. Hetero-tetrameric assembly of the K V 1.2&#95;P407R subunits with other neuronal voltage-gated potassium channels of Shaker subfamily could lead to general deficit of repolarizing potassium current and potentially underlie the enhanced seizure susceptibility. Indeed, expression of human K V 1.2&#95;P407R in early postnatal rat cortical neurons or genetically engineered hESC-derived neurons disclosed broadening of action potential duration and early afterdepolarization (EAD), associating with reduced potassium current. We hypothesize that Gapmer antisense oligonucleotides (ASOs) targeted to c.1220C>G mutation will selectively degrade the mutant mRNA while allowing the remaining wild-type (WT) subunits to form functional channels. As a proof of principle, delivery of Gapmer packaged in lipid nanoparticle into cortical neurons selectively suppressed K V 1.2&#95;P407R over the WT protein expression, reversing the broadening of action potential duration, abrogating the EAD and leading to overall increase in potassium current., Competing Interests: The authors declare no competing interests., (© 2024 The Author(s).)

L-type voltage-gated calcium channels are involved in multiple physiological functions. Currently available antagonists do not discriminate between L-type channel isoforms. Importantly, no selective blocker is available to dissect the role of L-type isoforms Ca v 1.2 and Ca v 1.3 that are concomitantly co-expressed in the heart, neuroendocrine and neuronal cells. Here we show that calciseptine, a snake toxin purified from mamba venom, selectively blocks Ca v 1.2 -mediated L-type calcium currents (I CaL ) at concentrations leaving Ca v 1.3-mediated I CaL unaffected in both native cardiac myocytes and HEK-293T cells expressing recombinant Ca v 1.2 and Ca v 1.3 channels. Functionally, calciseptine potently inhibits cardiac contraction without altering the pacemaker activity in sino-atrial node cells, underscoring differential roles of Ca v 1.2- and Ca v 1.3 in cardiac contractility and automaticity. In summary, calciseptine is a selective L-type Ca v 1.2 Ca 2+ channel blocker and should be a valuable tool to dissect the role of these L-channel isoforms., (© 2024. The Author(s).)