Two types of peptides derived from the neurotoxin GsMTx4 inhibit a mechanosensitive potassium channel by modifying the mechanogate.

Atrial fibrillation is the most common sustained cardiac arrhythmia in humans. Current atrial fibrillation antiarrhythmic drugs have limited efficacy and carry the risk of ventricular proarrhythmia. GsMTx4, a mechanosensitive channel-selective inhibitor, has been shown to suppress arrhythmias through the inhibition of stretch-activated channels (SACs) in the heart. The cost of synthesizing this peptide is a major obstacle to clinical use. Here, we studied two types of short peptides derived from GsMTx4 for their effects on a stretch-activated big potassium channel (SAKcaC) from the heart. Type I, a 17-residue peptide (referred to as Pept 01), showed comparable efficacy, whereas type II (i.e., Pept 02), a 10-residue peptide, exerted even more potent inhibitory efficacy on SAKcaC compared with GsMTx4. We identified through mutagenesis important sequences required for peptide functions. In addition, molecular dynamics simulations revealed common structural features with a hydrophobic head followed by a positively charged protrusion that may be involved in peptide channel-lipid interactions. Furthermore, we suggest that these short peptides may inhibit SAKcaC through a specific modification to the mechanogate, as the inhibitory effects for both types of peptides were mostly abolished when tested with a mechano-insensitive channel variant (STREX-del) and a nonmechanosensitive big potassium (mouse Slo1) channel. These findings may offer an opportunity for the development of a new class of drugs in the treatment of cardiac arrhythmia generated by excitatory SACs in the heart.

Blocking the BKCa channel induces NF-κB nuclear translocation by increasing nuclear calcium concentration†.

Nuclear factor kappa B (NF-κB) transcriptionally regulates several genes involved in initiating uterine contractions. A key factor controlling NF-κB activity is its translocation to the nucleus. In myometrial smooth muscle cells (MSMCs), this translocation can be stimulated by the inflammatory molecule lipopolysaccharide (LPS) or by blocking the potassium calcium-activated channel subfamily M alpha 1 (KCNMA1 or BKCa) with paxilline (PAX). Here, we sought to determine the mechanism by which blocking BKCa causes NF-κB-p65 translocation to the nucleus in MSMCs. We show that LPS- and PAX-induced NF-κB-p65 translocation are similar in that neither depends on several mitogen-activated protein kinase pathways, but both require increased intracellular calcium (Ca2+). However, the nuclear transport inhibitor wheat germ agglutinin prevented NF-κB-p65 nuclear translocation in response to LPS but not in response to PAX. Blocking BKCa located on the plasma membrane resulted in a transient NF-κB-p65 nuclear translocation that was not sufficient to induce expression of its transcriptional target, suggesting a role for intracellular BKCa. We report that BKCa also localizes to the nucleus and that blocking nuclear BKCa results in an increase in nuclear Ca2+ in MSMCs. Together, these data suggest that BKCa localized on the nuclear membrane plays a key role in regulating nuclear Ca2+ and NF-κB-p65 nuclear translocation in MSMCs.

KCa1.1 K+ Channel Inhibition Overcomes Resistance to Antiandrogens and Doxorubicin in a Human Prostate Cancer LNCaP Spheroid Model.

Several types of K+ channels play crucial roles in tumorigenicity, stemness, invasiveness, and drug resistance in cancer. Spheroid formation of human prostate cancer (PC) LNCaP cells with ultra-low attachment surface cultureware induced the up-regulation of cancer stem cell markers, such as NANOG, and decreased the protein degradation of the Ca2+-activated K+ channel KCa1.1 by down-regulating the E3 ubiquitin ligase, FBXW7, compared with LNCaP monolayers. Accordingly, KCa1.1 activator-induced hyperpolarizing responses were larger in isolated cells from LNCaP spheroids. The pharmacological inhibition of KCa1.1 overcame the resistance of LNCaP spheroids to antiandrogens and doxorubicin (DOX). The protein expression of androgen receptors (AR) was significantly decreased by LNCaP spheroid formation and reversed by KCa1.1 inhibition. The pharmacological and genetic inhibition of MDM2, which may be related to AR protein degradation in PC stem cells, revealed that MDM2 was responsible for the acquisition of antiandrogen resistance in LNCaP spheroids, which was overcome by KCa1.1 inhibition. Furthermore, a member of the multidrug resistance-associated protein subfamily of ABC transporters, MRP5 was responsible for the acquisition of DOX resistance in LNCaP spheroids, which was also overcome by KCa1.1 inhibition. Collectively, the present results suggest the potential of KCa1.1 in LNCaP spheroids, which mimic PC stem cells, as a therapeutic target for overcoming antiandrogen- and DOX-resistance in PC cells.

Ca2+ -activated K+ channel KCa 1.1 as a therapeutic target to overcome chemoresistance in three-dimensional sarcoma spheroid models.

The large-conductance Ca2+ -activated K+ channel KCa 1.1 plays a pivotal role in tumor development and progression in several solid cancers. The three-dimensional (3D) in vitro cell culture system is a powerful tool for cancer spheroid formation, and mimics in vivo solid tumor resistance to chemotherapy in the tumor microenvironment (TME). KCa 1.1 is functionally expressed in osteosarcoma and chondrosarcoma cell lines. KCa 1.1 activator-induced hyperpolarizing responses were significantly larger in human osteosarcoma MG-63 cells isolated from 3D spheroid models compared with in those from adherent 2D monolayer cells. The present study investigated the mechanisms underlying the upregulation of KCa 1.1 and its role in chemoresistance using a 3D spheroid model. KCa 1.1 protein expression levels were significantly elevated in the lipid-raft-enriched compartments of MG-63 spheroids without changes in its transcriptional level. 3D spheroid formation downregulated the expression of the ubiquitin E3 ligase FBXW7, which is an essential contributor to KCa 1.1 protein degradation in breast cancer. The siRNA-mediated inhibition of FBXW7 in MG-63 cells from 2D monolayers upregulated KCa 1.1 protein expression. Furthermore, a treatment with a potent and selective KCa 1.1 inhibitor overcame the chemoresistance of the MG-63 and human chondrosarcoma SW-1353 spheroid models to paclitaxel, doxorubicin, and cisplatin. Among several multidrug resistance ATP-binding cassette transporters, the expression of the multidrug resistance-associated protein MRP1 was upregulated in both spheroids and restored by the inhibition of KCa 1.1. Therefore, the pharmacological inhibition of KCa 1.1 may be an attractive new strategy for acquiring resistance to chemotherapeutic drugs in the TME of KCa 1.1-positive sarcomas.

Piezo1 and BKCa channels in human atrial fibroblasts: Interplay and remodelling in atrial fibrillation.

AIMS: Atrial Fibrillation (AF) is an arrhythmia of increasing prevalence in the aging populations of developed countries. One of the important indicators of AF is sustained atrial dilatation, highlighting the importance of mechanical overload in the pathophysiology of AF. The mechanisms by which atrial cells, including fibroblasts, sense and react to changing mechanical forces, are not fully elucidated. Here, we characterise stretch-activated ion channels (SAC) in human atrial fibroblasts and changes in SAC- presence and activity associated with AF. METHODS AND RESULTS: Using primary cultures of human atrial fibroblasts, isolated from patients in sinus rhythm or sustained AF, we combine electrophysiological, molecular and pharmacological tools to identify SAC. Two electrophysiological SAC- signatures were detected, indicative of cation-nonselective and potassium-selective channels. Using siRNA-mediated knockdown, we identified the cation-nonselective SAC as Piezo1. Biophysical properties of the potassium-selective channel, its sensitivity to calcium, paxilline or iberiotoxin (blockers), and NS11021 (activator), indicated presence of calcium-dependent 'big potassium channels' (BKCa). In cells from AF patients, Piezo1 activity and mRNA expression levels were higher than in cells from sinus rhythm patients, while BKCa activity (but not expression) was downregulated. Both Piezo1-knockdown and removal of extracellular calcium from the patch pipette resulted in a significant reduction of BKCa current during stretch. No co-immunoprecipitation of Piezo1 and BKCa was detected. CONCLUSIONS: Human atrial fibroblasts contain at least two types of ion channels that are activated during stretch: Piezo1 and BKCa. While Piezo1 is directly stretch-activated, the increase in BKCa activity during mechanical stimulation appears to be mainly secondary to calcium influx via SAC such as Piezo1. During sustained AF, Piezo1 is increased, while BKCa activity is reduced, highlighting differential regulation of both channels. Our data support the presence and interplay of Piezo1 and BKCa in human atrial fibroblasts in the absence of physical links between the two channel proteins.

BKCa Channel Inhibition by Peripheral Nerve Injury Is Restored by the Xanthine Derivative KMUP-1 in Dorsal Root Ganglia.

This study explored whether KMUP-1 improved chronic constriction injury (CCI)-induced BKCa current inhibition in dorsal root ganglion (DRG) neurons. Rats were randomly assigned to four groups: sham, sham + KMUP-1, CCI, and CCI + KMUP-1 (5 mg/kg/day, i.p.). DRG neuronal cells (L4-L6) were isolated on day 7 after CCI surgery. Perforated patch-clamp and inside-out recordings were used to monitor BKCa currents and channel activities, respectively, in the DRG neurons. Additionally, DRG neurons were immunostained with anti-NeuN, anti-NF200 and anti-BKCa. Real-time PCR was used to measure BKCa mRNA levels. In perforated patch-clamp recordings, CCI-mediated nerve injury inhibited BKCa currents in DRG neurons compared with the sham group, whereas KMUP-1 prevented this effect. CCI also decreased BKCa channel activity, which was recovered by KMUP-1 administration. Immunofluorescent staining further demonstrated that CCI reduced BKCa-channel proteins, and KMUP-1 reversed this. KMUP-1 also changed CCI-reduced BKCa mRNA levels. KMUP-1 prevented CCI-induced neuropathic pain and BKCa current inhibition in a peripheral nerve injury model, suggesting that KMUP-1 could be a potential agent for controlling neuropathic pain.

Loss-of-function BK channel mutation causes impaired mitochondria and progressive cerebellar ataxia.

Despite a growing number of ion channel genes implicated in hereditary ataxia, it remains unclear how ion channel mutations lead to loss-of-function or death of cerebellar neurons. Mutations in the gene KCNMA1, encoding the α-subunit of the BK channel have emerged as responsible for a variety of neurological phenotypes. We describe a mutation (BKG354S) in KCNMA1, in a child with congenital and progressive cerebellar ataxia with cognitive impairment. The mutation in the BK channel selectivity filter dramatically reduced single-channel conductance and ion selectivity. The BKG354S channel trafficked normally to plasma, nuclear, and mitochondrial membranes, but caused reduced neurite outgrowth, cell viability, and mitochondrial content. Small interfering RNA (siRNA) knockdown of endogenous BK channels had similar effects. The BK activator, NS1619, rescued BKG354S cells but not siRNA-treated cells, by selectively blocking the mutant channels. When expressed in cerebellum via adenoassociated virus (AAV) viral transfection in mice, the mutant BKG354S channel, but not the BKWT channel, caused progressive impairment of several gait parameters consistent with cerebellar dysfunction from 40- to 80-d-old mice. Finally, treatment of the patient with chlorzoxazone, a BK/SK channel activator, partially improved motor function, but ataxia continued to progress. These studies indicate that a loss-of-function BK channel mutation causes ataxia and acts by reducing mitochondrial and subsequently cellular viability.

Targeting BKCa Channels in Migraine: Rationale and Perspectives.

Large (big)-conductance calcium-activated potassium (BKCa) channels are expressed in migraine-related structures such as the cranial arteries, trigeminal ganglion and trigeminal spinal nucleus, and they play a substantial role in vascular tonus and neuronal excitability. Using synthetic BKCa channels openers was associated with headache as a frequent adverse effect in healthy volunteers. Additionally, BKCa channels are downstream molecules in migraine signalling pathways that are activated by several compounds known to provoke migraine, including calcitonin gene-related peptide (CGRP), pituitary adenylate cyclase-activating polypeptide (PACAP) and glyceryl trinitrate (GTN). Also, there is a high affinity and a close coupling between BKCa channels and ATP-sensitive potassium (KATP) channels, the role of which has recently been established in migraine pathophysiology. These observations raise the question as to whether direct BKCa channel activation can provoke migraine in migraine patients, and whether the BKCa channel could be a potential novel anti-migraine target. Hence, randomized and placebo-controlled clinical studies on BKCa channel openers or blockers in migraine patients are needed.

Globotriaosylceramide-induced reduction of KCa1.1 channel activity and activation of the Notch1 signaling pathway in skin fibroblasts of male Fabry patients with pain.

BACKGROUND: Fabry disease (FD) is an X-linked lysosomal storage disorder that leads to cellular globotriaosylceramide (Gb3) accumulation due to mutations in the gene encoding α-galactosidase A. Trigger-induced acral burning pain is an early FD symptom of unknown pathophysiology. We aimed at investigating the potential role of skin fibroblasts in nociceptor sensitization. PATIENTS AND METHODS: We enrolled 40 adult FD patients and ten healthy controls, who underwent a 6-mm skin punch biopsy at the lower leg. Dermal fibroblasts were cultivated and analyzed for Gb3 load. Fibroblast electrical activity was assessed using patch-clamp analysis at baseline and upon incubation with agalsidase-α for 24 h. We investigated gene expression of CC motif chemokine ligand 2 (CCL2), Ca2+activated K+-channel 1.1 (KCa1.1), interferone-γ (IFN-γ), transforming growth factor-ß1 (TGF-ß1), and transmembrane receptor notch homolog 1 (Notch1) using quantitative real-time-PCR, and protein levels of KCa1.1 by ELISA. Gene expression was determined at baseline and after fibroblast stimulation with tumor necrosis factor-α (TNF), modeling inflammation as a common pain trigger in FD. RESULTS: Total Gb3 load was higher in FD fibroblasts than in control fibroblasts (p < .01). Upon increase of intracellular Ca2+ concentrations, we detected differential electrical activity of KCa1.1 in fibroblasts obtained from patients with FD. Gene expression (p < .05) and protein levels of KCa1.1 (p < .05) were higher in fibroblasts from FD patients compared to control fibroblasts, whereas electric channel activity was lower in FD fibroblasts. After incubation with agalsidase-α, we observed an over-proportionate increase of KCa1.1 activity in FD fibroblasts reaching 7-fold the currents of control cells (p < .01). Gene expression studies revealed higher mRNA levels of CCL2, INF-γ, and Notch1 in FD fibroblasts compared to controls at baseline and after TNF incubation (p < .05 each), while TGF-ß1 was higher in FD fibroblasts only after incubation with TNF (p < .05). CONCLUSIONS: Gb3 deposition in skin fibroblasts may impair KCa1.1 activity and activate the Notch1 signaling pathway. The resulting increase in pro-inflammatory mediator expression may contribute to cutaneous nociceptor sensitization as a potential mechanism of FD-associated pain.

Inhibition of BKCa negatively alters cardiovascular function.

Large conductance calcium and voltage-activated potassium channels (BKCa ) are transmembrane proteins, ubiquitously expressed in the majority of organs, and play an active role in regulating cellular physiology. In the heart, BKCa channels are known to play a role in regulating the heart rate and protect it from ischemia-reperfusion injury. In vascular smooth muscle cells, the opening of BKCa channels results in membrane hyperpolarization which eventually results in vasodilation mediated by a reduction in Ca2+ influx due to the closure of voltage-dependent Ca2+ channels. Ex vivo studies have shown that BKCa channels play an active role in the regulation of the function of the majority of blood vessels. However, in vivo role of BKCa channels in cardiovascular function is not completely deciphered. Here, we have evaluated the rapid in vivo role of BKCa channels in regulating the cardiovascular function by using two well-established, rapid-acting, potent blockers, paxilline and iberiotoxin. Our results show that BKCa channels are actively involved in regulating the heart rate, the function of the left and right heart as well as major vessels. We also found that the effect on BKCa channels by blockers is completely reversible, and hence, BKCa channels can be exploited as potential targets for clinical applications for modulating heart rate and cardiac contractility.

Targeting KCa1.1 Channels with a Scorpion Venom Peptide for the Therapy of Rat Models of Rheumatoid Arthritis.

Fibroblast-like synoviocytes (FLSs) are a key cell type involved in rheumatoid arthritis (RA) progression. We previously identified the KCa1.1 potassium channel (Maxi-K, BK, Slo 1, KCNMA1) as a regulator of FLSs and found that KCa1.1 inhibition reduces disease severity in RA animal models. However, systemic KCa1.1 block causes multiple side effects. In this study, we aimed to determine whether the KCa1.1 ß1-3-specific venom peptide blocker iberiotoxin (IbTX) reduces disease severity in animal models of RA without inducing major side effects. We used immunohistochemistry to identify IbTX-sensitive KCa1.1 subunits in joints of rats with a model of RA. Patch-clamp and functional assays were used to determine whether IbTX can regulate FLSs through targeting KCa1.1. We then tested the efficacy of IbTX in ameliorating disease in two rat models of RA. Finally, we determined whether IbTX causes side effects including incontinence or tremors in rats, compared with those treated with the small-molecule KCa1.1 blocker paxilline. IbTX-sensitive subunits of KCa1.1 were expressed by FLSs in joints of rats with experimental arthritis. IbTX inhibited KCa1.1 channels expressed by FLSs from patients with RA and by FLSs from rat models of RA and reduced FLS invasiveness. IbTX significantly reduced disease severity in two rat models of RA. Unlike paxilline, IbTX did not induce tremors or incontinence in rats. Overall, IbTX inhibited KCa1.1 channels on FLSs and treated rat models of RA without inducing side effects associated with nonspecific KCa1.1 blockade and could become the basis for the development of a new treatment of RA.

The role of BKCa in endometrial cancer HEC-1-B cell proliferation and migration.

BKCa is a large conductance calcium activated potassium channel ubiquitously expressed in various cell types. Accumulating evidence demonstrates that BKCa is aberrantly expressed in many malignancies, involving in cancerous behaviors such as cell proliferation and migration. In this study, we investigated the functional role of BKCa in endometrial cancer HEC-1-B cells. Overexpression of BKCa by plasmid transfection enhanced endometrial cancer cell proliferation and migration. Conversely, silence of BKCa by lentivirus mediated RNAi system not only inhibited proliferation and migration but also impaired tumor growth in vivo. Patch clamp assay identified the BKCa currents in HEC-1-B cells, which was supported by the observation of channel activation or inhibition in response to the specific opener (NS1619) or blocker (IBTX) of BKCa. Moreover, NS1619 significantly increased cell proliferation and migration while IBTX exhibited the opposite effects. In summary, these data suggested an important role of BKCa in proliferation and migration of endometrial cancer HEC-1-B cells. Thus, BKCa may be established as a potential therapeutic target in endometrial cancer.

Methylglyoxal Impairs ß2-Adrenoceptor-Mediated Vasodilatory Mechanisms in Rat Retinal Arterioles.

Methylglyoxal, a highly reactive dicarbonyl compound, is formed as a by-product of glycolysis and plays an important role in the pathogenesis of diabetic complications, including diabetic retinopathy. However, it remains to be determined how methylglyoxal affects the regulatory mechanisms of retinal blood flow. In this study, we examined the effects of methylglyoxal on ß2-adrenoceptor-mediated vasodilatory mechanisms in rat retinal arterioles. The retinal vasodilator responses were assessed by measuring the diameter of retinal arterioles in the fundus images. Intravitreal injection of methylglyoxal significantly diminished the vasodilation of retinal arterioles induced by the ß2-adrenoceptor agonist salbutamol. The vasodilator effect of BMS-191011, a large-conductance Ca2+-activated K+ (BKCa) channel opener, on retinal arterioles was also attenuated by methylglyoxal. In contrast, methylglyoxal had no significant effect on retinal vasodilator response to forskolin. Methylglyoxal attenuated retinal vasodilator response to salbutamol under blockade of BKCa channels with iberiotoxin, an inhibitor of the channels. These results suggest that methylglyoxal attenuates ß2-adrenoceptor-mediated retinal vasodilation by impairing the coupling of the ß2-adrenoceptor to the guanine nucleotide-binding protein (Gs protein) and the function of the BKCa channel. Increased methylglyoxal in the eyes may contribute to the impairment of regulatory mechanisms of retinal blood flow in patients with diabetic retinopathy.

Functional BKCa channel in human resident cardiac stem cells expressing W8B2.

Recently, a new population of resident cardiac stem cells (CSCs) positive for the W8B2 marker has been identified. These CSCs are considered to be an ideal cellular source to repair myocardial damage after infarction. However, the electrophysiological profile of these cells has not been characterized yet. We first establish the conditions of isolation and expansion of W8B2+ CSCs from human heart biopsies using a magnetic sorting system followed by flow cytometry cell sorting. These cells display a spindle-shaped morphology, are highly proliferative, and possess self-renewal capacity demonstrated by their ability to form colonies. Besides, W8B2+ CSCs are positive for mesenchymal markers but negative for hematopoietic and endothelial ones. RT-qPCR and immunostaining experiments show that W8B2+ CSCs express some early cardiac-specific transcription factors but lack the expression of cardiac-specific structural genes. Using patch clamp in the whole-cell configuration, we show for the first time the electrophysiological signature of BKCa current in these cells. Accordingly, RT-PCR and western blotting analysis confirmed the presence of BKCa at both mRNA and protein levels in W8B2+ CSCs. Interestingly, BKCa channel inhibition by paxilline decreased cell proliferation in a concentration-dependent manner and halted cell cycle progression at the G0/G1 phase. The inhibition of BKCa also decreased the self-renewal capacity but did not affect migration of W8B2+ CSCs. Taken together, our results are consistent with an important role of BKCa channels in cell cycle progression and self-renewal in human cardiac stem cells.

A voltage-dependent K+ channel in the lysosome is required for refilling lysosomal Ca2+ stores.

The resting membrane potential (Δψ) of the cell is negative on the cytosolic side and determined primarily by the plasma membrane's selective permeability to K+ We show that lysosomal Δψ is set by lysosomal membrane permeabilities to Na+ and H+, but not K+, and is positive on the cytosolic side. An increase in juxta-lysosomal Ca2+ rapidly reversed lysosomal Δψ by activating a large voltage-dependent and K+-selective conductance (LysoKVCa). LysoKVCa is encoded molecularly by SLO1 proteins known for forming plasma membrane BK channels. Opening of single LysoKVCa channels is sufficient to cause the rapid, striking changes in lysosomal Δψ. Lysosomal Ca2+ stores may be refilled from endoplasmic reticulum (ER) Ca2+ via ER-lysosome membrane contact sites. We propose that LysoKVCa serves as the perilysosomal Ca2+ effector to prime lysosomes for the refilling process. Consistently, genetic ablation or pharmacological inhibition of LysoKVCa, or abolition of its Ca2+ sensitivity, blocks refilling and maintenance of lysosomal Ca2+ stores, resulting in lysosomal cholesterol accumulation and a lysosome storage phenotype.

Estradiol-modified prolactin secretion independently of action potentials and Ca2+ and blockade of outward potassium currents in GH3 cells.

Estrogens facilitate prolactin (PRL) secretion acting on pituitary cells. In GH3 cells, estradiol induces acute action potentials and oscillations of intracellular Ca2+ associated with the secretagogue function. Estradiol modulates several ion channels which may affect the action potential rate and the release of PRL in lactotroph cells, which might depend on its concentration. The aims were to characterize the acute effect of supraphysiological concentrations of estradiol on Ca2+ and noninactivating K+ currents and measure the effect on the spontaneous action potentials and PRL release in the somatolactotroph cell line, GH3. Electrophysiological studies were carried out by voltage- and current-clamp techniques and ELISA determination of PRL secretion. Pharmacological concentrations of estradiol (above 1 µM), without a latency period, blocked Ca2+ channels and noninactivating K+ currents, including the large-conductance voltage- and Ca2+-activated K+ channels (BK), studied in whole-cell nystatin perforated and in excided inside-out patches of GH3 and CHO cells, transiently transfected with the human α-pore forming subunit of BK. The effect on BK was contrary to the agonist effect associated with the regulatory ß1-subunits of the BK, which GH3 cells lack, but its transient transfection did not modify the noninactivating current blockade, suggesting a different mechanism of regulation. Estradiol, at the same concentration range, acutely decreased the frequency of action potentials, an expected effect as consequence of the Ca2+ channel blockade. Despite this, PRL secretion initially increased, followed by a decrease in long-term incubations. This suggests that, in GH3 cells, supraphysiological concentrations of estradiol modulating PRL secretion are partially independent of extracellular Ca2+ influx.

Down-Regulation of Ca2+-Activated K⁺ Channel KCa1.1 in Human Breast Cancer MDA-MB-453 Cells Treated with Vitamin D Receptor Agonists.

Vitamin D (VD) reduces the risk of breast cancer and improves disease prognoses. Potential VD analogs are being developed as therapeutic agents for breast cancer treatments. The large-conductance Ca2+-activated K⁺ channel KCa1.1 regulates intracellular Ca2+ signaling pathways and is associated with high grade tumors and poor prognoses. In the present study, we examined the effects of treatments with VD receptor (VDR) agonists on the expression and activity of KCa1.1 in human breast cancer MDA-MB-453 cells using real-time PCR, Western blotting, flow cytometry, and voltage-sensitive dye imaging. Treatments with VDR agonists for 72 h markedly decreased the expression levels of KCa1.1 transcripts and proteins in MDA-MB-453 cells, resulting in the significant inhibition of depolarization responses induced by paxilline, a specific KCa1.1 blocker. The specific proteasome inhibitor MG132 suppressed VDR agonist-induced decreases in KCa1.1 protein expression. These results suggest that KCa1.1 is a new downstream target of VDR signaling and the down-regulation of KCa1.1 through the transcriptional repression of KCa1.1 and enhancement of KCa1.1 protein degradation contribute, at least partly, to the antiproliferative effects of VDR agonists in breast cancer cells.

KCNMA1, a pore-forming α-subunit of BK channels, regulates insulin signalling in mature adipocytes.

KCNMA1 is a pore-forming α-subunit of the large conductance Ca2+ - and voltage-activated K+ channels, referred to as BK channels, which play key roles in various physiological functions. However, the role of KCNMA1 in mature adipocytes remains unclear. In this study, we reveal that kcnma1 expression is downregulated in white adipose tissue of mice fed a high-fat diet and in hypertrophied adipocytes. Furthermore, inhibition of kcnma1 expression or treatment with a BK channel blocker attenuated insulin-induced Akt phosphorylation in mature adipocytes. These results strongly indicate that KCNMA1 contributes to the regulation of insulin signalling in mature adipocytes.

Contraception: Search for an Ideal Unisex Mechanism by Targeting Ion Channels.

Targeting sperm ion channels and other sperm-specific proteins is an effective way to develop unisex contraceptives, as they should have decreased side effects. This Science & Society summarizes the current advances in human sperm physiology in attempts to evaluate what would be appropriate targets for unisex contraceptives.

BK channels in microglia are required for morphine-induced hyperalgesia.

Although morphine is a gold standard medication, long-term opioid use is associated with serious side effects, such as morphine-induced hyperalgesia (MIH) and anti-nociceptive tolerance. Microglia-to-neuron signalling is critically involved in pain hypersensitivity. However, molecules that control microglial cellular state under chronic morphine treatment remain unknown. Here we show that the microglia-specific subtype of Ca(2+)-activated K(+) (BK) channel is responsible for generation of MIH and anti-nociceptive tolerance. We find that, after chronic morphine administration, an increase in arachidonic acid levels through the µ-opioid receptors leads to the sole activation of microglial BK channels in the spinal cord. Silencing BK channel auxiliary ß3 subunit significantly attenuates the generation of MIH and anti-nociceptive tolerance, and increases neurotransmission after chronic morphine administration. Therefore, microglia-specific BK channels contribute to the generation of MIH and anti-nociceptive tolerance.