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1
Zhu, Shu, Darren D. Browning, Richard E. White, David Fulton, and Scott A. Barman. "Mutation of protein kinase C phosphorylation site S1076 on α-subunits affects BKCa channel activity in HEK-293 cells." American Journal of Physiology-Lung Cellular and Molecular Physiology 297, no. 4 (October 2009): L758—L766. http://dx.doi.org/10.1152/ajplung.90518.2008.
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Large conductance, calcium- and voltage-activated potassium (BKCa) channels are important modulators of pulmonary vascular smooth muscle membrane potential, and phosphorylation of BKCa channels by protein kinases regulates pulmonary arterial smooth muscle function. However, little is known about the effect of phosphorylating specific channel subunits on BKCa channel activity. The present study was done to determine the effect of mutating protein kinase C (PKC) phosphorylation site serine 1076 (S1076) on transfected human BKCa channel α-subunits in human embryonic kidney (HEK-293) cells, a heterologous expression system devoid of endogenous BKCa channels. Results showed that mutating S1076 altered the effect of PKC activation on BKCa channels in HEK-293 cells. Specifically, the phospho-deficient mutation BKCa-α(S1076A)/β1 attenuated the excitatory effect of the PKC activator phorbol myristate acetate (PMA) on BKCa channels, whereas the phospho-mimetic mutation BKCa-α(S1076E)/β1 increased the excitatory effect of PMA on BKCa channels. In addition, the phospho-null mutation S1076A blocked the activating effect of cGMP-dependent protein kinase G (PKG) on BKCa channels. Collectively, these results suggest that specific putative PKC phosphorylation site(s) on human BKCa channel α-subunits influences BKCa channel activity, which may subsequently alter pulmonary vascular smooth muscle function and tone.
2
Zhao, Guiling, Zachary P. Neeb, M. Dennis Leo, Judith Pachuau, Adebowale Adebiyi, Kunfu Ouyang, Ju Chen, and Jonathan H. Jaggar. "Type 1 IP3 receptors activate BKCa channels via local molecular coupling in arterial smooth muscle cells." Journal of General Physiology 136, no. 3 (August 16, 2010): 283–91. http://dx.doi.org/10.1085/jgp.201010453.
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Plasma membrane large-conductance Ca2+-activated K+ (BKCa) channels and sarcoplasmic reticulum inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) are expressed in a wide variety of cell types, including arterial smooth muscle cells. Here, we studied BKCa channel regulation by IP3 and IP3Rs in rat and mouse cerebral artery smooth muscle cells. IP3 activated BKCa channels both in intact cells and in excised inside-out membrane patches. IP3 caused concentration-dependent BKCa channel activation with an apparent dissociation constant (Kd) of ∼4 µM at physiological voltage (−40 mV) and intracellular Ca2+ concentration ([Ca2+]i; 10 µM). IP3 also caused a leftward-shift in BKCa channel apparent Ca2+ sensitivity and reduced the Kd for free [Ca2+]i from ∼20 to 12 µM, but did not alter the slope or maximal Po. BAPTA, a fast Ca2+ buffer, or an elevation in extracellular Ca2+ concentration did not alter IP3-induced BKCa channel activation. Heparin, an IP3R inhibitor, and a monoclonal type 1 IP3R (IP3R1) antibody blocked IP3-induced BKCa channel activation. Adenophostin A, an IP3R agonist, also activated BKCa channels. IP3 activated BKCa channels in inside-out patches from wild-type (IP3R1+/+) mouse arterial smooth muscle cells, but had no effect on BKCa channels of IP3R1-deficient (IP3R1−/−) mice. Immunofluorescence resonance energy transfer microscopy indicated that IP3R1 is located in close spatial proximity to BKCa α subunits. The IP3R1 monoclonal antibody coimmunoprecipitated IP3R1 and BKCa channel α and β1 subunits from cerebral arteries. In summary, data indicate that IP3R1 activation elevates BKCa channel apparent Ca2+ sensitivity through local molecular coupling in arterial smooth muscle cells.
3
Xie, Man-Jiang, Yu-Guang Ma, Fang Gao, Yun-Gang Bai, Jiu-Hua Cheng, Yao-Ming Chang, Zhi-Bin Yu, and Jin Ma. "Activation of BKCa channel is associated with increased apoptosis of cerebrovascular smooth muscle cells in simulated microgravity rats." American Journal of Physiology-Cell Physiology 298, no. 6 (June 2010): C1489—C1500. http://dx.doi.org/10.1152/ajpcell.00474.2009.
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Cerebral arterial remodeling is one of the critical factors in the occurrence of postspaceflight orthostatic intolerance. We hypothesize that large-conductance calcium-activated K+ (BKCa) channels in vascular smooth muscle cells (VSMCs) may play an important role in regulating cerebrovascular adaptation during microgravity exposure. The aim of this work was to investigate whether activation of BKCa channels is involved in regulation of apoptotic remodeling of cerebral arteries in simulated microgravity rats. In animal studies, Sprague-Dawley rats were subjected to 1-wk hindlimb unweighting to simulate microgravity. Alterations of BKCa channels in cerebral VSMCs were investigated by patch clamp and Western blotting; apoptosis was assessed by electron microscopy and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick-end labeling (TUNEL). To evaluate the correlation of BKCa channel and apoptosis, channel protein and cell nucleus were double-stained. In cell studies, hSloα+β1 channel was coexpressed into human embryonic kidney 293 (HEK293) cells to observe the effects of BKCa channels on apoptosis. In rats, enhanced activities and expression of BKCa channels were found to be correlated with increased apoptosis in cerebral VSMCs after simulated microgravity. In transfected HEK293 cells, activation of cloned BKCa channel induced apoptosis, whereas inhibition of cloned BKCa channel decreased apoptosis. In conclusion, activation of BKCa channels is associated with increased apoptosis in cerebral VSMCs of simulated microgravity rats.
4
Ling, Shizhang, Jian-Zhong Sheng, and Andrew P. Braun. "The calcium-dependent activity of large-conductance, calcium-activated K+ channels is enhanced by Pyk2- and Hck-induced tyrosine phosphorylation." American Journal of Physiology-Cell Physiology 287, no. 3 (September 2004): C698—C706. http://dx.doi.org/10.1152/ajpcell.00030.2004.
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Recent results showing that large-conductance, calcium-activated K+ (BKCa) channels undergo direct tyrosine phosphorylation in the presence of c-Src tyrosine kinase have suggested the involvement of these channels in Src-mediated signaling pathways. Given the important role for c-Src in integrin-mediated signal transduction, we have examined the potential regulation of BKCa channels by proline-rich tyrosine kinase 2 (Pyk2), a calcium-sensitive tyrosine kinase activated upon integrin stimulation. Transient coexpression of murine BKCa channels with either wild-type Pyk2 or hematopoietic cell kinase (Hck), a Src-family kinase, led to an enhancement of BKCa channel activity over the range of 1–10 μM free calcium, whereas coexpression with catalytically inactive forms of either kinase did not significantly alter BKCa gating compared with channels expressed alone. In the presence of either wild-type Pyk2 or Hck, BKCa α-subunits were found to undergo tyrosine phosphorylation, as determined by immunoprecipitation and Western blotting strategies. However, tyrosine phosphorylation of the BKCa α-subunit was not detected for channels expressed alone or together with inactive forms of either Pyk2 or Hck. Interestingly, wild-type, but not inactive, Pyk2 was also present in BKCa channel immunoprecipitates, suggesting that Pyk2 may coassociate with the BKCa channel complex after phosphorylation. Collectively, the observed modulation and phosphorylation of BKCa channels by Pyk2 and a Src-family kinase may reflect a general cellular mechanism by which G protein-coupled receptor and/or integrin activation leads to the regulation of membrane ion channels.
5
Kim, Eun Young, Jae Mi Suh, Yu-Hsin Chiu, and Stuart E. Dryer. "Regulation of podocyte BKCa channels by synaptopodin, Rho, and actin microfilaments." American Journal of Physiology-Renal Physiology 299, no. 3 (September 2010): F594—F604. http://dx.doi.org/10.1152/ajprenal.00206.2010.
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Mechanosensitive large-conductance Ca2+-activated K+ channels encoded by the Slo1 gene (BKCa channels) are expressed in podocytes. Here we show that BKCa channels reciprocally coimmunoprecipitate with synaptopodin (Synpo) in mouse glomeruli, in mouse podocytes, and in a heterologous expression system (HEK293T cells) in which these proteins are transiently expressed. Synpo and Slo1 colocalize along the surface of the glomerular basement membrane in mouse glomeruli. Synpo interacts with BKCa channels at COOH-terminal domains that overlap with an actin-binding domain on the channel molecule that is necessary for trafficking of BKCa channels to the cell surface. Moreover, addition of exogenous β-actin to mouse podocyte lysates reduces BKCa-Synpo interactions. Coexpression of Synpo increases steady-state surface expression of BKCa channels in HEK293T cells. However, Synpo does not affect the stability of cell surface BKCa channels, suggesting a primary effect on the rate of forward trafficking, and Synpo coexpression does not affect BKCa gating. Conversely, stable knockdown of Synpo expression in mouse podocyte cell lines reduces steady-state surface expression of BKCa channels but does not affect total expression of BKCa channels or their gating. The effects of Synpo on surface expression of BKCa are blocked by inhibition of Rho signaling in HEK293T cells and in podocytes. Functional cell surface BKCa channels in podocytes are also reduced by sustained (2 h) but not acute (15 min) depolymerization of actin with cytochalasin D. Synpo may regulate BKCa channels through its effects on actin dynamics and by modulating interactions between BKCa channels and regulatory proteins of the podocyte slit diaphragm.
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Werner, Matthias E., Andrea L. Meredith, Richard W. Aldrich, and Mark T. Nelson. "Hypercontractility and impaired sildenafil relaxations in the BKCa channel deletion model of erectile dysfunction." American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 295, no. 1 (July 2008): R181—R188. http://dx.doi.org/10.1152/ajpregu.00173.2008.
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Erectile dysfunction (ED) can be elicited by a variety of pathogenic factors, particularly impaired formation of and responsiveness to nitric oxide (NO) and the downstream effectors soluble guanylate cyclase (sGC) and cGMP-dependent protein kinase I (PKGI). One important target of PKGI in smooth muscle is the large-conductance, Ca2+-activated potassium (BKCa) channel. In our previous report ( 42 ), we demonstrated that deletion of the BKCa channel in mice induced force oscillations and led to reduced nerve-evoked relaxations and ED. In the current study, we used this ED model to explore the role of the BKCa channel in the NO/sGC/PKGI pathway. Electrical field stimulation (EFS)-induced contractions of corpus cavernosum smooth muscle strips were significantly enhanced in the absence of BKCa channel function. In strips precontracted with phenylephrine, EFS-induced relaxations were converted to contractions by inhibition of sGC, and this was further enhanced by loss of BK channel function. Sildenafil-induced relaxations were decreased to a similar extent by inhibition of sGC or BKCa channels. At concentrations >1 μM, sildenafil caused relaxations independent of inhibition of sGC or BKCa channels. Sildenafil did not affect the enhanced force oscillations that were induced by the loss of BKCa channel function. Yet, these oscillations could be completely eliminated by blocking L-type voltage-dependent Ca2+ channels (VDCCs). These results suggest that therapeutically relevant concentrations of sildenafil act through cGMP and BKCa channels, and loss of BKCa channel function leads to hypercontractility, which depends on VDCCs and cannot be modified by the cGMP pathway.
7
Choi, Chang-Rok, Eun-Jin Kim, Tae Hyun Choi, Jaehee Han, and Dawon Kang. "Enhancing Human Cutaneous Wound Healing through Targeted Suppression of Large Conductance Ca2+-Activated K+ Channels." International Journal of Molecular Sciences 25, no. 2 (January 9, 2024): 803. http://dx.doi.org/10.3390/ijms25020803.
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The modulation of K+ channels plays a crucial role in cell migration and proliferation, but the effect of K+ channels on human cutaneous wound healing (CWH) remains underexplored. This study aimed to determine the necessity of modulating K+ channel activity and expression for human CWH. The use of 25 mM KCl as a K+ channel blocker markedly improved wound healing in vitro (in keratinocytes and fibroblasts) and in vivo (in rat and porcine models). K+ channel blockers, such as quinine and tetraethylammonium, aided in vitro wound healing, while Ba2+ was the exception and did not show similar effects. Single-channel recordings revealed that the Ba2+-insensitive large conductance Ca2+-activated K+ (BKCa) channel was predominantly present in human keratinocytes. NS1619, an opener of the BKCa channel, hindered wound healing processes like proliferation, migration, and filopodia formation. Conversely, charybdotoxin and iberiotoxin, which are BKCa channel blockers, dramatically enhanced these processes. The downregulation of BKCa also improved CWH, whereas its overexpression impeded these healing processes. These findings underscore the facilitative effect of BKCa channel suppression on CWH, proposing BKCa channels as potential molecular targets for enhancing human cutaneous wound healing.
8
Barman, Scott A., Shu Zhu, and Richard E. White. "Protein kinase C inhibits BKCa channel activity in pulmonary arterial smooth muscle." American Journal of Physiology-Lung Cellular and Molecular Physiology 286, no. 1 (January 2004): L149—L155. http://dx.doi.org/10.1152/ajplung.00207.2003.
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Signaling mechanisms that elevate cyclic AMP (cAMP) activate large-conductance, calcium- and voltage-activated potassium (BKCa) channels in pulmonary vascular smooth muscle and cause pulmonary vasodilatation. BKCa channel modulation is important in the regulation of pulmonary arterial pressure, and inhibition (closing) of the BKCa channel has been implicated in the development of pulmonary vasoconstriction. Protein kinase C (PKC) causes pulmonary vasoconstriction, but little is known about the effect of PKC on BKCa channel activity. Accordingly, studies were done to determine the effect of PKC activation on cAMP-induced BKCa channel activity using patch-clamp studies in pulmonary arterial smooth muscle cells (PASMC) of the fawn-hooded rat (FHR), a recognized animal model of pulmonary hypertension. Forskolin (10 μM), a stimulator of adenylate cyclase and an activator of cAMP, opened BKCa channels in single FHR PASMC, which were blocked by the PKC activators phorbol 12-myristate 13-acetate (100 nM) and thymeleatoxin (100 nM). The inhibitory response by thymeleatoxin on forskolin-induced BKCa channel activity was blocked by Gö-6983, which selectively blocks the α, β, δ, γ, and ζ PKC isozymes, and Gö-6976, which selectively inhibits PKC-α, PKC-β, and PKC-μ, but not by rottlerin, which selectively inhibits PKC-δ. Collectively, these results indicate that activation of specific PKC isozymes inhibits cAMP-induced activation of the BKCa channel in pulmonary arterial smooth muscle, which suggests a unique signaling pathway to modulate BKCa channels and subsequently cAMP-induced pulmonary vasodilatation.
9
Khan, Raheela N., Stephen K. Smith, J. J. Morrison, and Michael L. J. Ashford. "Ca2+ dependence and pharmacology of large-conductance K+ channels in nonlabor and labor human uterine myocytes." American Journal of Physiology-Cell Physiology 273, no. 5 (November 1, 1997): C1721—C1731. http://dx.doi.org/10.1152/ajpcell.1997.273.5.c1721.
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Two populations, Ca2+-dependent (BKCa) and Ca2+-independent K+ (BK) channels of large conductance were identified in inside-out patches of nonlabor and labor freshly dispersed human pregnant myometrial cells, respectively. Cell-attached recordings from nonlabor myometrial cells frequently displayed BKCa channel openings characterized by a relatively low open-state probability, whereas similar recordings from labor tissue displayed either no channel openings or consistently high levels of channel activity that often exhibited clear, oscillatory activity. In inside-out patch recordings, Ba2+ (2–10 mM), 4-aminopyridine (0.1–1 mM), and Shaker B inactivating peptide (“ball peptide”) blocked the BKCa channel but were much less effective on BK channels. Application of tetraethylammonium to inside-out membrane patches reduced unitary current amplitude of BKCa and BK channels, with dissociation constants of 46 mM and 53 μM, respectively. Tetraethylammonium applied to outside-out patches decreased the unitary conductance of BKCa and BK channels, with dissociation constants of 423 and 395 μM, respectively. These results demonstrate that the properties of human myometrial large-conductance K+channels in myocytes isolated from laboring patients are significantly different from those isolated from nonlaboring patients.
10
Hou, Shangwei, Stefan H. Heinemann, and Toshinori Hoshi. "Modulation of BKCa Channel Gating by Endogenous Signaling Molecules." Physiology 24, no. 1 (February 2009): 26–35. http://dx.doi.org/10.1152/physiol.00032.2008.
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Large-conductance Ca2+- and voltage-activated K+ (BKCa, MaxiK, or Slo1) channels are expressed in almost every tissue in our body and participate in many critical functions such as neuronal excitability, vascular tone regulation, and neurotransmitter release. The functional versatility of BKCa channels owes in part to the availability of a spectacularly wide array of biological modulators of the channel function. In this review, we focus on modulation of BKCa channels by small endogenous molecules, emphasizing their molecular mechanisms. The mechanistic information available from studies on the small naturally occurring modulators is expected to contribute to our understanding of the physiological and pathophysiological roles of BKCa channels.
11
Hannigan, K. I., R. J. Large, E. Bradley, M. A. Hollywood, G. P. Sergeant, N. G. McHale, and K. D. Thornbury. "Effect of a novel BKCa opener on BKCa currents and contractility of the rabbit corpus cavernosum." American Journal of Physiology-Cell Physiology 310, no. 4 (February 15, 2016): C284—C292. http://dx.doi.org/10.1152/ajpcell.00273.2015.
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Large-conductance Ca2+-activated K+ (BKCa) channels are thought to play a key role in the regulation of corpus cavernosum smooth muscle (CCSM) excitability. Few BKCa channel openers have been accepted for clinical development. The effect of the novel BKCa channel opener GoSlo-SR5-130 on electrical activity in isolated rabbit CCSM cells and mechanical activity in strips of rabbit CCSM was examined. Single-channel currents were observed in inside-out patches. These channels were sensitive to Ca2+, blocked by penitrem A, and had a conductance of 291 ± 20 pS ( n = 7). In the presence of GoSlo-SR5-130, the number of open BKCa channels increased. Using voltage-ramp protocols, GoSlo-SR5-130 caused currents to activate at more negative potentials in a concentration-dependent manner, shifting the half-maximal activation voltage potential to the left on the voltage axis. Therefore, BKCa channels were open within the physiological range of membrane potentials in the presence of GoSlo-SR5-130. GoSlo-SR5-130 also resulted in an increase in the activity of spontaneous transient outward currents in myocytes isolated from CCSM, and this effect was reversed by iberiotoxin. In current-clamp mode, GoSlo-SR5-130 hyperpolarized the cell membrane. Isometric tension recording of strips of rabbit corpus cavernosum showed that GoSlo-SR5-130 inhibited spontaneous contractions in a concentration-dependent manner. This effect was reversed in the presence of iberiotoxin, suggesting that GoSlo-SR5-130 exerts its effect through BKCa channels. These findings suggest that GoSlo-SR5-130 is an effective tool for the study of BKCa channels and that these channels can modulate CCSM activity and are possible targets for the treatment of erectile dysfunction.
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Chang, Wei-Ting, and Sheng-Nan Wu. "Effective Activation of BKCa Channels by QO-40 (5-(Chloromethyl)-3-(Naphthalen-1-yl)-2-(Trifluoromethyl)Pyrazolo [1,5-a]pyrimidin-7(4H)-one), Known to Be an Opener of KCNQ2/Q3 Channels." Pharmaceuticals 14, no. 5 (April 21, 2021): 388. http://dx.doi.org/10.3390/ph14050388.
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QO-40 (5-(chloromethyl)-3-(naphthalene-1-yl)-2-(trifluoromethyl) pyrazolo[1,5-a]pyrimidin-7(4H)-one) is a novel and selective activator of KCNQ2/KCNQ3 K+ channels. However, it remains largely unknown whether this compound can modify any other type of plasmalemmal ionic channel. The effects of QO-40 on ion channels in pituitary GH3 lactotrophs were investigated in this study. QO-40 stimulated Ca2+-activated K+ current (IK(Ca)) with an EC50 value of 2.3 μM in these cells. QO-40-stimulated IK(Ca) was attenuated by the further addition of GAL-021 or paxilline but not by linopirdine or TRAM-34. In inside-out mode, this compound added to the intracellular leaflet of the detached patches stimulated large-conductance Ca2+-activated K+ (BKCa) channels with no change in single-channel conductance; however, there was a decrease in the slow component of the mean closed time of BKCa channels. The KD value required for the QO-40-mediated decrease in the slow component at the mean closure time was 1.96 μM. This compound shifted the steady-state activation curve of BKCa channels to a less positive voltage and decreased the gating charge of the channel. The application of QO-40 also increased the hysteretic strength of BKCa channels elicited by a long-lasting isosceles-triangular ramp voltage. In HEK293T cells expressing α-hSlo, QO-40 stimulated BKCa channel activity. Overall, these findings demonstrate that QO-40 can interact directly with the BKCa channel to increase the amplitude of IK(Ca) in GH3 cells.
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Niloy, Sayeman Islam, Yue Shen, Lirong Guo, Stephen T. O’Rourke, and Chengwen Sun. "Loss of IP3R-BKCa Coupling Is Involved in Vascular Remodeling in Spontaneously Hypertensive Rats." International Journal of Molecular Sciences 24, no. 13 (June 30, 2023): 10903. http://dx.doi.org/10.3390/ijms241310903.
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Mechanisms by which BKCa (large-conductance calcium-sensitive potassium) channels are involved in vascular remodeling in hypertension are not fully understood. Vascular smooth muscle cell (VSMC) proliferation and vascular morphology were compared between hypertensive and normotensive rats. BKCa channel activity, protein expression, and interaction with IP3R (inositol 1,4,5-trisphosphate receptor) were examined using patch clamp, Western blot analysis, and coimmunoprecipitation. On inside-out patches of VSMCs, the Ca2+-sensitivity and voltage-dependence of BKCa channels were similar between hypertensive and normotensive rats. In whole-cell patch clamp configuration, treatment of cells with the IP3R agonist, Adenophostin A (AdA), significantly increased BKCa channel currents in VSMCs of both strains of rats, suggesting IP3R-BKCa coupling; however, the AdA-induced increases in BKCa currents were attenuated in VSMCs of hypertensive rats, indicating possible IP3R-BKCa decoupling, causing BKCa dysfunction. Co-immunoprecipitation and Western blot analysis demonstrated that BKCa and IP3R proteins were associated together in VSMCs; however, the association of BKCa and IP3R proteins was dramatically reduced in VSMCs of hypertensive rats. Genetic disruption of IP3R-BKCa coupling using junctophilin-2 shRNA dramatically augmented Ang II-induced proliferation in VSMCs of normotensive rats. Subcutaneous infusion of NS1619, a BKCa opener, to reverse BKCa dysfunction caused by IP3R-BKCa decoupling significantly attenuated vascular hypertrophy in hypertensive rats. In summary, the data from this study demonstrate that loss of IP3R-BKCa coupling in VSMCs induces BKCa channel dysfunction, enhances VSMC proliferation, and thus, may contribute to vascular hypertrophy in hypertension.
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Sansom, Steven C., Rong Ma, Pamela K. Carmines, and David A. Hall. "Regulation of Ca2+-activated K+ channels by multifunctional Ca2+/calmodulin-dependent protein kinase." American Journal of Physiology-Renal Physiology 279, no. 2 (August 1, 2000): F283—F288. http://dx.doi.org/10.1152/ajprenal.2000.279.2.f283.
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Activation of mesangial cells by ANG II provokes release of intracellular Ca2+ stores and subsequent Ca2+influx through voltage-gated channels, events that are reflected by a large transient increase in intracellular concentration [Ca2+]i followed by a modest sustained elevation in [Ca2+]i. These ANG II-induced alterations in [Ca2+]i elicit activation of large Ca2+-activated K+ channels (BKCa) in a negative-feedback manner. The mechanism of this BKCa feedback response may involve the direct effect of intracellular Ca2+ on the channel and/or channel activation by regulatory enzymes. The present study utilized patch-clamp and fura 2 fluorescence techniques to assess the involvement of multifunctional calcium calmodulin kinase II (CAMKII) in the BKCa feedback response. In cell-attached patches, KN62 (specific inhibitor of CAMKII) either abolished or reduced to near zero the ANG II-induced BKCa feedback response. This phenomenon did not reflect direct effects of KN62 on the BKCa channel, because this agent alone did not significantly alter BKCa channel activity in inside-out patches. KN62 also failed to alter either the transient peak or sustained plateau phases of the [Ca2+]i response to ANG II. In inside-out patches (1 μM Ca2+ in bath), calmodulin plus ATP activated BKCa channels in the presence but not the absence of CAMKII. These observations are consistent with the postulate that CAMKII is involved in the BKCa feedback response of mesangial cells, acting to potentiate the influence of increased [Ca2+]i on the BKCa channel or a closely associated regulator of the channel. An additional effect of CAMKII to activate a voltage-gated Ca2+ channel cannot be ruled out by these experiments.
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Yokoshiki, Hisashi, Takashi Seki, Masanori Sunagawa, and Nicholas Sperelakis. "Inhibition of Ca2+-activated K+ channels by tyrosine phosphatase inhibitors in rat mesenteric artery." Canadian Journal of Physiology and Pharmacology 78, no. 9 (September 1, 2000): 745–50. http://dx.doi.org/10.1139/y00-042.
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To investigate the possible regulation of large-conductance Ca2+-activated K+ channels (BKCa) by tyrosine phosphatases (Tyr-PPs), single-channel currents of myocytes from rat mesenteric artery were recorded in open cell-attached patches. Two structurally different Tyr-PP inhibitors, sodium orthovanadate (Na3VO4) and dephostatin, were used. The channels (236 pS) evoked at +40 mV and pCa 6, were significantly inhibited by 1 mM Na3VO4 (-81 ± 3%, n = 10; P < 0.005). Similarly, 100 µM dephostatin strongly inhibited the BKCa channels (-80 ± 7%, n = 7 ; P < 0.05). Therefore, BKCa channels in vascular smooth muscle cells may be regulated by tyrosine phosphatase-dependent signal transduction pathways, whose inhibition could attenuate the channel activity.Key words: Ca2+-activated K+ channel, vascular smooth muscle, tyrosine phosphatase, vanadate, dephostatin.
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Paisansathan, Chanannait, Haoliang Xu, Francesco Vetri, Moises Hernandez, and Dale A. Pelligrino. "Interactions between adenosine and K+ channel-related pathways in the coupling of somatosensory activation and pial arteriolar dilation." American Journal of Physiology-Heart and Circulatory Physiology 299, no. 6 (December 2010): H2009—H2017. http://dx.doi.org/10.1152/ajpheart.00702.2010.
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Multiple, perhaps interactive, mechanisms participate in the linkage between increased neural activity and cerebral vasodilation. In the present study, we assessed whether neural activation-related pial arteriolar dilation (PAD) involved interactions among adenosine (Ado) A2 receptors (A2Rs), large-conductance Ca2+-operated K+ (BKCa) channels, and inward rectifier K+ (Kir) channels. In rats with closed cranial windows, we monitored sciatic nerve stimulation (SNS)-induced PAD in the absence or presence of pharmacological blockade of A2Rs (ZM-241385), ecto-5′-nucleotidase (α,β-methylene-adenosine diphosphate), BKCa channels (paxilline), and Kir channels (BaCl2). Individually, these interventions led to 53–66% reductions in SNS-induced PADs. Combined applications of these blockers led to little or no further repression of SNS-induced PADs, suggesting interactions among A2Rs and K+ channels. In the absence of SNS, BaCl2 blockade of Kir channels produced 52–80% reductions in Ado and NS-1619 (BKCa channel activator)-induced PADs. In contrast, paxilline blockade of BKCa channels was without effect on dilations elicited by KCl (Kir channel activator) and Ado suffusions, indicating that Ado- and NS-1619-associated PADs involved Kir channels. In addition, targeted ablation of the superficial glia limitans was associated with a selective 60–80% loss of NS-1619 responses, suggesting that the BKCa channel participation (and paxilline sensitivity) derived largely from channels within the glia limitans. Additionally, blockade of either PKA or adenylyl cyclase caused markedly attenuated pial arteriolar responses to SNS and, in the absence of SNS, responses to Ado, KCl, and NS-1619. These findings suggested a key, possibly permissive, role for A2R-linked cAMP generation and PKA-induced K+ channel phosphorylation in somatosensory activation-evoked PAD.
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Lu, Te-Ling, Zi-Han Gao, Shih-Wei Li, and Sheng-Nan Wu. "High Efficacy by GAL-021: A Known Intravenous Peripheral Chemoreceptor Modulator that Suppresses BKCa-Channel Activity and Inhibits IK(M) or Ih." Biomolecules 10, no. 2 (January 25, 2020): 188. http://dx.doi.org/10.3390/biom10020188.
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GAL-021 has recently been developed as a novel breathing control modulator. However, modifications of ionic currents produced by this agent remain uncertain, although its efficacy in suppressing the activity of big-conductance Ca2+-activated K+ (BKCa) channels has been reported. In pituitary tumor (GH3) cells, we found that the presence of GAL-021 decreased the amplitude of macroscopic Ca2+-activated K+ current (IK(Ca)) in a concentration-dependent manner with an effective IC50 of 2.33 μM. GAL-021-mediated reduction of IK(Ca) was reversed by subsequent application of verteporfin or ionomycin; however, it was not by that of diazoxide. In inside-out current recordings, the addition of GAL-021 to the bath markedly decreased the open-state probability of BKCa channels. This agent also resulted in a rightward shift in voltage dependence of the activation curve of BKCa channels; however, neither the gating charge of the curve nor single-channel conductance of the channel was changed. There was an evident lengthening of the mean closed time of BKCa channels in the presence of GAL-021, with no change in mean open time. The GAL-021 addition also suppressed M-type K+ current with an effective IC50 of 3.75 μM; however, its presence did not alter the amplitude of erg-mediated K+ current, or mildly suppressed delayed-rectifier K+ current. GAL-021 at a concentration of 30 μM could also suppress hyperpolarization-activated cationic current. In HEK293T cells expressing α-hSlo, the addition of GAL-021 was also able to suppress the BKCa-channel open probabilities, and GAL-021-mediated suppression of BKCa-channel activity was attenuated by further addition of BMS-191011. Collectively, the GAL-021 effects presented herein do not exclusively act on BKCa channels and these modifications on ionic currents exert significant influence on the functional activities of electrically excitable cells occurring in vivo.
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Riddle, Melissa A., Jennifer M. Hughes, and Benjimen R. Walker. "Role of caveolin-1 in endothelial BKCa channel regulation of vasoreactivity." American Journal of Physiology-Cell Physiology 301, no. 6 (December 2011): C1404—C1414. http://dx.doi.org/10.1152/ajpcell.00013.2011.
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A novel vasodilatory influence of endothelial cell (EC) large-conductance Ca2+-activated K+ (BKCa) channels is present following in vivo exposure to chronic hypoxia (CH) and may exist in other pathological states. However, the mechanism of channel activation that results in altered vasoreactivity is unknown. We tested the hypothesis that CH removes an inhibitory effect of the scaffolding domain of caveolin-1 (Cav-1) on EC BKCa channels to permit activation, thereby affecting vasoreactivity. Experiments were performed on gracilis resistance arteries and ECs from control and CH-exposed (380 mmHg barometric pressure for 48 h) rats. EC membrane potential was hyperpolarized in arteries from CH-exposed rats and arteries treated with the cholesterol-depleting agent methyl-β-cyclodextrin (MBCD) compared with controls. Hyperpolarization was reversed by the BKCa channel antagonist iberiotoxin (IBTX) or by a scaffolding domain peptide of Cav-1 (AP-CAV). Patch-clamp experiments documented an IBTX-sensitive current in ECs from CH-exposed rats and in MBCD-treated cells that was not present in controls. This current was enhanced by the BKCa channel activator NS-1619 and blocked by AP-CAV or cholesterol supplementation. EC BKCa channels displayed similar unitary conductance but greater Ca2+ sensitivity than BKCa channels from vascular smooth muscle. Immunofluorescence imaging demonstrated greater association of BKCa α-subunits with Cav-1 in control arteries than in arteries from CH-exposed rats, although fluorescence intensity for each protein did not differ between groups. Finally, AP-CAV restored myogenic and phenylephrine-induced constriction in arteries from CH-exposed rats without affecting controls. AP-CAV similarly restored diminished reactivity to phenylephrine in control arteries pretreated with MBCD. We conclude that CH unmasks EC BKCa channel activity by removing an inhibitory action of the Cav-1 scaffolding domain that may depend on cellular cholesterol levels.
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Barman, Scott A., Shu Zhu, and Richard E. White. "PKC activates BKCa channels in rat pulmonary arterial smooth muscle via cGMP-dependent protein kinase." American Journal of Physiology-Lung Cellular and Molecular Physiology 286, no. 6 (June 2004): L1275—L1281. http://dx.doi.org/10.1152/ajplung.00259.2003.
Full textAbstract:
Normally, signaling mechanisms that activate large-conductance, calcium- and voltage-activated potassium (BKCa) channels in pulmonary vascular smooth muscle cause pulmonary vasodilatation. BKCa-channel modulation is important in the regulation of pulmonary arterial pressure, and inhibition (decrease in the opening probability) of the BKCa channel has been implicated in the development of pulmonary vasoconstriction. Protein kinase C (PKC) causes pulmonary vasoconstriction, but little is known about the effect of PKC on BKCa-channel activity in pulmonary vascular smooth muscle. Accordingly, studies were done to determine the effect of PKC on BKCa-channel activity using patch-clamp studies in pulmonary arterial smooth muscle cells (PASMCs) of the Sprague-Dawley rat. The PKC activators phorbol myristate acetate (PMA) and thymeleatoxin opened BKCa channels in single Sprague-Dawley rat PASMC. The activator response to both PMA and thymeleatoxin on BKCa-channel activity was blocked by Gö-6983, which selectively blocks PKC-α, -δ, -γ, and -ζ, and by rottlerin, which selectively inhibits PKC-δ. In addition, the specific cyclic GMP-dependent protein kinase antagonist KT-5823 blocked the responses to PMA and thymelatoxin, whereas the specific cyclic AMP-dependent protein kinase blocker KT-5720 had no effect. In isolated pulmonary arterial vessels, both PMA and forskolin caused vasodilatation, which was inhibited by KT-5823, Gö-6983, or the BKCa-channel blocker tetraethylammonium. The results of this study indicate that activation of specific PKC isozymes increases BKCa-channel activity in Sprague-Dawley rat PASMC via cyclic GMP-dependent protein kinase, which suggests a unique signaling mechanism for vasodilatation.
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Szteyn, Kalina, and Harpreet Singh. "BKCa Channels as Targets for Cardioprotection." Antioxidants 9, no. 8 (August 17, 2020): 760. http://dx.doi.org/10.3390/antiox9080760.
Full textAbstract:
The large-conductance calcium- and voltage-activated K+ channel (BKCa) are encoded by the Kcnma1 gene. They are ubiquitously expressed in neuronal, smooth muscle, astrocytes, and neuroendocrine cells where they are known to play an important role in physiological and pathological processes. They are usually localized to the plasma membrane of the majority of the cells with an exception of adult cardiomyocytes, where BKCa is known to localize to mitochondria. BKCa channels couple calcium and voltage responses in the cell, which places them as unique targets for a rapid physiological response. The expression and activity of BKCa have been linked to several cardiovascular, muscular, and neurological defects, making them a key therapeutic target. Specifically in the heart muscle, pharmacological and genetic activation of BKCa channels protect the heart from ischemia-reperfusion injury and also facilitate cardioprotection rendered by ischemic preconditioning. The mechanism involved in cardioprotection is assigned to the modulation of mitochondrial functions, such as regulation of mitochondrial calcium, reactive oxygen species, and membrane potential. Here, we review the progress made on BKCa channels and cardioprotection and explore their potential roles as therapeutic targets for preventing acute myocardial infarction.
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Mizutani, Hiroya, Hisao Yamamura, Makoto Muramatsu, Yumiko Hagihara, Yoshiaki Suzuki, and Yuji Imaizumi. "Modulation of Ca2+ oscillation and melatonin secretion by BKCa channel activity in rat pinealocytes." American Journal of Physiology-Cell Physiology 310, no. 9 (May 1, 2016): C740—C747. http://dx.doi.org/10.1152/ajpcell.00342.2015.
Full textAbstract:
The pineal glands regulate circadian rhythm through the synthesis and secretion of melatonin. The stimulation of nicotinic acetylcholine receptor due to parasympathetic nerve activity causes an increase in intracellular Ca2+ concentration and eventually downregulates melatonin production. Our previous report shows that rat pinealocytes have spontaneous and nicotine-induced Ca2+ oscillations that are evoked by membrane depolarization followed by Ca2+ influx through voltage-dependent Ca2+ channels (VDCCs). These Ca2+ oscillations are supposed to contribute to the inhibitory mechanism of melatonin secretion. Here we examined the involvement of large-conductance Ca2+-activated K+ (BKCa) channel conductance on the regulation of Ca2+ oscillation and melatonin production in rat pinealocytes. Spontaneous Ca2+ oscillations were markedly enhanced by BKCa channel blockers (1 μM paxilline or 100 nM iberiotoxin). Nicotine (100 μM)-induced Ca2+ oscillations were also augmented by paxilline. In contrast, spontaneous Ca2+ oscillations were abolished by BKCa channel opener [3 μM 12,14-dichlorodehydroabietic acid (diCl-DHAA)]. Under whole cell voltage-clamp configurations, depolarization-elicited outward currents were significantly activated by diCl-DHAA and blocked by paxilline. Expression analyses revealed that the α and β3 subunits of BKCa channel were highly expressed in rat pinealocytes. Importantly, the activity of BKCa channels modulated melatonin secretion from whole pineal gland of the rat. Taken together, BKCa channel activation attenuates these Ca2+ oscillations due to depolarization-synchronized Ca2+ influx through VDCCs and results in a recovery of reduced melatonin secretion during parasympathetic nerve activity. BKCa channels may play a physiological role for melatonin production via a negative-feedback mechanism.
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Gururaja Rao, Shubha, Piotr Bednarczyk, Atif Towheed, Kajol Shah, Priyanka Karekar, Devasena Ponnalagu, Haley N. Jensen, et al. "BKCa (Slo) Channel Regulates Mitochondrial Function and Lifespan in Drosophila melanogaster." Cells 8, no. 9 (August 21, 2019): 945. http://dx.doi.org/10.3390/cells8090945.
Full textAbstract:
BKCa channels, originally discovered in Drosophila melanogaster as slowpoke (slo), are recognized for their roles in cellular and organ physiology. Pharmacological approaches implicated BKCa channels in cellular and organ protection possibly for their ability to modulate mitochondrial function. However, the direct role of BKCa channels in regulating mitochondrial structure and function is not deciphered. Here, we demonstrate that BKCa channels are present in fly mitochondria, and slo mutants show structural and functional defects in mitochondria. slo mutants display an increase in reactive oxygen species and the modulation of ROS affected their survival. We also found that the absence of BKCa channels reduced the lifespan of Drosophila, and overexpression of human BKCa channels in flies extends life span in males. Our study establishes the presence of BKCa channels in mitochondria of Drosophila and ascertains its novel physiological role in regulating mitochondrial structural and functional integrity, and lifespan.
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Li, N., R. Shi, Y. Ye, Y. Zhang, Y. Zhang, Z. Wang, Y. Gu, Y. Yin, D. Chen, and J. Tang. "Aging-induced down-regulation of Pka/Bkca pathway in rat cerebral arteries." Physiological Research 71, no. 6 (November 25, 2022): 811–23. http://dx.doi.org/10.33549/physiolres.934944.
Full textAbstract:
The incidence of cerebrovascular diseases increases significantly with aging. This study aimed to test the hypothesis that aging may influence the protein kinase A (PKA)-dependent vasodilation via RyR/BKCa pathway in the middle cerebral arteries (MCA). Male Sprague-Dawley rats were randomly divided into control (4-6 month-old) and aged (24-month-old) groups. The functions of MCA and ion channel activities in smooth muscle cells were examined using myograph system and patch-clamp. Aging decreased the isoproterenol/forskolin-induced relaxation in the MCA. Large-conductance Ca2+-activated-K+ (BKCa) channel inhibitor, iberiotoxin, significantly attenuated the forskolin-induced vasodilatation and hyperpolarization in the young group, but not in the aged group. The amplitude and frequency of spontaneous transient outward currents (STOCs) were significantly decreased in the aged group. Single channel recording revealed that the mean open time of BKCa channels were decreased, while an increased mean closed time of BKCa channels were found in the aged group. The Ca2+/voltage sensitivity of the channels was decreased accompanied by reduced BKCa α and β1-subunit, the expression of RyR2, PKA-Cα and PKA-Cβ subunits were also declined in the aged group. Aging induced down-regulation of PKA/BKCa pathway in cerebral artery in rats. The results provides new information on further understanding in cerebrovascular diseases resulted from age-related cerebral vascular dysfunction.
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Idres, Sarah, Germain Perrin, Valérie Domergue, Florence Lefebvre, Susana Gomez, Audrey Varin, Rodolphe Fischmeister, Véronique Leblais, and Boris Manoury. "Contribution of BKCa channels to vascular tone regulation by PDE3 and PDE4 is lost in heart failure." Cardiovascular Research 115, no. 1 (June 23, 2018): 130–44. http://dx.doi.org/10.1093/cvr/cvy161.
Full textAbstract:
Abstract Aims Regulation of vascular tone by 3′,5′-cyclic adenosine monophosphate (cAMP) involves many effectors including the large conductance, Ca2+-activated, K+ (BKCa) channels. In arteries, cAMP is mainly hydrolyzed by type 3 and 4 phosphodiesterases (PDE3, PDE4). Here, we examined the specific contribution of BKCa channels to tone regulation by these PDEs in rat coronary arteries, and how this is altered in heart failure (HF). Methods and results Concomitant application of PDE3 (cilostamide) and PDE4 (Ro-20-1724) inhibitors increased BKCa unitary channel activity in isolated myocytes from rat coronary arteries. Myography was conducted in isolated, U46619-contracted coronary arteries. Cilostamide (Cil) or Ro-20-1724 induced a vasorelaxation that was greatly reduced by iberiotoxin (IBTX), a BKCa channel blocker. Ro-20-1724 and Cil potentiated the relaxation induced by the β-adrenergic agonist isoprenaline (ISO) or the adenylyl cyclase activator L-858051 (L85). IBTX abolished the effect of PDE inhibitors on ISO but did not on L85. In coronary arteries from rats with HF induced by aortic stenosis, contractility and response to acetylcholine were dramatically reduced compared with arteries from sham rats, but relaxation to PDE inhibitors was retained. Interestingly, however, IBTX had no effect on Ro-20-1724- and Cil-induced vasorelaxations in HF. Expression of the BKCa channel α-subunit, of a 98 kDa PDE3A and of a 80 kDa PDE4D were lower in HF compared with sham coronary arteries, while that of a 70 kDa PDE4B was increased. Proximity ligation assays demonstrated that PDE3 and PDE4 were localized in the vicinity of the channel. Conclusion BKCa channels mediate the relaxation of coronary artery induced by PDE3 and PDE4 inhibition. This is achieved by co-localization of both PDEs with BKCa channels, enabling tight control of cAMP available for channel opening. Contribution of the channel is prominent at rest and on β-adrenergic stimulation. This coupling is lost in HF.
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Herrera, Gerald M., Thomas J. Heppner, and Mark T. Nelson. "Voltage dependence of the coupling of Ca2+ sparks to BKCa channels in urinary bladder smooth muscle." American Journal of Physiology-Cell Physiology 280, no. 3 (March 1, 2001): C481—C490. http://dx.doi.org/10.1152/ajpcell.2001.280.3.c481.
Full textAbstract:
Large-conductance Ca2+-dependent K+(BKCa) channels play a critical role in regulating urinary bladder smooth muscle (UBSM) excitability and contractility. Measurements of BKCa currents and intracellular Ca2+ revealed that BKCa currents are activated by Ca2+ release events (Ca2+ sparks) from ryanodine receptors (RyRs) in the sarcoplasmic reticulum. The goals of this project were to characterize Ca2+ sparks and BKCa currents and to determine the voltage dependence of the coupling of RyRs (Ca2+ sparks) to BKCachannels in UBSM. Ca2+ sparks in UBSM had properties similar to those described in arterial smooth muscle. Most Ca2+ sparks caused BKCa currents at all voltages tested, consistent with the BKCa channels sensing ∼10 μM Ca2+. Membrane potential depolarization from −50 to −20 mV increased Ca2+ spark and BKCacurrent frequency threefold. However, membrane depolarization over this range had a differential effect on spark and current amplitude, with Ca2+ spark amplitude increasing by only 30% and BKCa current amplitude increasing 16-fold. A major component of the amplitude modulation of spark-activated BKCa current was quantitatively explained by the known voltage dependence of the Ca2+ sensitivity of BKCa channels. We, therefore, propose that membrane potential, or any other agent that modulates the Ca2+sensitivity of BKCa channels, profoundly alters the coupling strength of Ca2+ sparks to BKCa channels.
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Dimitropoulou, Christiana, Guichun Han, Allison W. Miller, Mariela Molero, Leslie C. Fuchs, Richard E. White, and Gerald O. Carrier. "Potassium (BKCa) currents are reduced in microvascular smooth muscle cells from insulin-resistant rats." American Journal of Physiology-Heart and Circulatory Physiology 282, no. 3 (March 1, 2002): H908—H917. http://dx.doi.org/10.1152/ajpheart.00382.2001.
Full textAbstract:
Insulin resistance (IR) syndrome is associated with impaired vascular relaxation; however, the underlying pathophysiology is unknown. Potassium channel activation causes vascular smooth muscle hyperpolarization and relaxation. The present study determined whether a reduction in large conductance calcium- and voltage-activated potassium (BKCa) channel activity contributes to impaired vascular relaxation in IR rats. BKCa channels were characterized in mesenteric microvessels from IR and control rats. Macroscopic current density was reduced in myocytes from IR animals compared with controls. In addition, inhibition of BKCachannels with tetraethylammonium (1 mM) or iberiotoxin (100 nM) was greater in myocytes from control (70%) compared with IR animals (∼20%). Furthermore, activation of BKCa channels with NS-1619 was three times more effective at increasing outward current in cells from control versus IR animals. Single channel and Western blot analysis of BKCa channels revealed similar conductance, amplitude, voltage sensitivity, Ca2+ sensitivity, and expression density between the two groups. These data provide the first direct evidence that microvascular potassium currents are reduced in IR and suggest a molecular mechanism that could account for impaired vascular relaxation in IR.
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Darkow, D. J., L. Lu, and R. E. White. "Estrogen relaxation of coronary artery smooth muscle is mediated by nitric oxide and cGMP." American Journal of Physiology-Heart and Circulatory Physiology 272, no. 6 (June 1, 1997): H2765—H2773. http://dx.doi.org/10.1152/ajpheart.1997.272.6.h2765.
Full textAbstract:
Estrogens are proposed to exert protection against cardiovascular disease, and evidence now suggests that this protection involves a direct vasodilatory effect. We have shown previously that estrogen relaxes endothelium-denuded porcine coronary arteries by opening the large-conductance calcium- and voltage-activated potassium (BKCa) channel of myocytes through guanosine 3',5'-cyclic monophosphate (cGMP)-dependent phosphorylation (35). The present study confirms these results and now demonstrates that this mechanism involves production of nitric oxide (NO). S-nitroso-N-acetylpenicillamine (SNAP), an NO donor, or 8-bromo-cGMP mimicked the effect of estrogen on BKCa channels. Furthermore, inhibition of NO synthase (NOS) attenuated estrogen- or tamoxifen-induced BKCa-channel activity, and this effect was disinhibited by L-arginine. Inhibition of guanylyl cyclase activity blocked the stimulatory effect of estrogen, SNAP, or L-arginine on BKCa channels. Furthermore, 17 beta-estradiol stimulated accumulation of nitrite and cGMP in coronary myocytes. Therefore, we propose that the vasodilatory effect of estrogen on the coronary circulation is mediated by NO. A portion of the beneficial cardiovascular effects of estrogen may be attributed to relaxation of vascular smooth muscle by a process that involves NO- and cGMP-dependent stimulation of BKCa channels.
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Shieh, D. B., S. R. Yang, X. Y. Shi, Y. N. Wu, and S. N. Wu. "Properties of BKCa Channels in Oral Keratinocytes." Journal of Dental Research 84, no. 5 (May 2005): 468–73. http://dx.doi.org/10.1177/154405910508400513.
Full textAbstract:
Keratinocytes are important for epithelial antimicrobial barrier function. The activity of ion channels can affect the proliferation of keratinocytes. Little is known about Ca2+-activated K+ currents in these cells. Ion currents in normal human oral keratinocytes were characterized with a patch-clamp technique. In whole-cell configuration, depolarizing pulses evoked K+ outward currents ( IK) in oral keratinocytes. Iberiotoxin (200 nM) and paxilline (1 μM) suppressed IK; however, neither apamin (200 nM) nor 5-hydroxydecanoate (30 μM) had any effects on it. Caffeic acid phenethyl ester, a compound of honeybee propolis, increased IK with an EC50 value of 12.8 ± 1.2 μM. In inside-out patches, a BKCa channel was observed in keratinocytes, but not in oral squamous carcinoma (OCE-M1) cells. Caffeic acid phenethyl ester or cinnamyl-3,4-dihydroxy-α-cyanocinnamate applied to the intracellular surface of a detached patch increased BKCa-channel activity. The results demonstrate that the properties of BKCa channels in normal human oral keratinocytes are similar to those described in other types of cells. Caffeic acid derivatives can also stimulate BKCa-channel activity directly.
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Chen, Yin-Chia, Chia-Lung Shih, Chao-Liang Wu, Yi-Hsien Fang, Edmund Cheung So, and Sheng-Nan Wu. "Exploring the Impact of BKCa Channel Function in Cellular Membranes on Cardiac Electrical Activity." International Journal of Molecular Sciences 25, no. 3 (January 26, 2024): 1537. http://dx.doi.org/10.3390/ijms25031537.
Full textAbstract:
This review paper delves into the current body of evidence, offering a thorough analysis of the impact of large-conductance Ca2+-activated K+ (BKCa or BK) channels on the electrical dynamics of the heart. Alterations in the activity of BKCa channels, responsible for the generation of the overall magnitude of Ca2+-activated K+ current at the whole-cell level, occur through allosteric mechanisms. The collaborative interplay between membrane depolarization and heightened intracellular Ca2+ ion concentrations collectively contribute to the activation of BKCa channels. Although fully developed mammalian cardiac cells do not exhibit functional expression of these ion channels, evidence suggests their presence in cardiac fibroblasts that surround and potentially establish close connections with neighboring cardiac cells. When cardiac cells form close associations with fibroblasts, the high single-ion conductance of these channels, approximately ranging from 150 to 250 pS, can result in the random depolarization of the adjacent cardiac cell membranes. While cardiac fibroblasts are typically electrically non-excitable, their prevalence within heart tissue increases, particularly in the context of aging myocardial infarction or atrial fibrillation. This augmented presence of BKCa channels’ conductance holds the potential to amplify the excitability of cardiac cell membranes through effective electrical coupling between fibroblasts and cardiomyocytes. In this scenario, this heightened excitability may contribute to the onset of cardiac arrhythmias. Moreover, it is worth noting that the substances influencing the activity of these BKCa channels might influence cardiac electrical activity as well. Taken together, the BKCa channel activity residing in cardiac fibroblasts may contribute to cardiac electrical function occurring in vivo.
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Guntur, Divya, Horst Olschewski, Péter Enyedi, Réka Csáki, Andrea Olschewski, and Chandran Nagaraj. "Revisiting the Large-Conductance Calcium-Activated Potassium (BKCa) Channels in the Pulmonary Circulation." Biomolecules 11, no. 11 (November 3, 2021): 1629. http://dx.doi.org/10.3390/biom11111629.
Full textAbstract:
Potassium ion concentrations, controlled by ion pumps and potassium channels, predominantly govern a cell′s membrane potential and the tone in the vessels. Calcium-activated potassium channels respond to two different stimuli-changes in voltage and/or changes in intracellular free calcium. Large conductance calcium-activated potassium (BKCa) channels assemble from pore forming and various modulatory and auxiliary subunits. They are of vital significance due to their very high unitary conductance and hence their ability to rapidly cause extreme changes in the membrane potential. The pathophysiology of lung diseases in general and pulmonary hypertension, in particular, show the implication of either decreased expression and partial inactivation of BKCa channel and its subunits or mutations in the genes encoding different subunits of the channel. Signaling molecules, circulating humoral molecules, vasorelaxant agents, etc., have an influence on the open probability of the channel in pulmonary arterial vascular cells. BKCa channel is a possible therapeutic target, aimed to cause vasodilation in constricted or chronically stiffened vessels, as shown in various animal models. This review is a comprehensive collation of studies on BKCa channels in the pulmonary circulation under hypoxia (hypoxic pulmonary vasoconstriction; HPV), lung pathology, and fetal to neonatal transition, emphasising pharmacological interventions as viable therapeutic options.
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Li, Yan, Jin Bai, Yi-hua Yang, Naoto Hoshi, and Dong-bao Chen. "Hydrogen Sulfide Relaxes Human Uterine Artery via Activating Smooth Muscle BKCa Channels." Antioxidants 9, no. 11 (November 13, 2020): 1127. http://dx.doi.org/10.3390/antiox9111127.
Full textAbstract:
Opening of large conductance calcium-activated and voltage-dependent potassium (BKCa) channels hyperpolarizes plasma membranes of smooth muscle (SM) to cause vasodilation, underling a key mechanism for mediating uterine artery (UA) dilation in pregnancy. Hydrogen sulfide (H2S) has been recently identified as a new UA vasodilator, yet the mechanism underlying H2S-induced UA dilation is unknown. Here, we tested whether H2S activated BKCa channels in human UA smooth muscle cells (hUASMC) to mediate UA relaxation. Multiple BKCa subunits were found in human UA in vitro and hUASMC in vitro, and high β1 and γ1 proteins were localized in SM cells in human UA. Baseline outward currents, recorded by whole-cell and single-channel patch clamps, were significantly inhibited by specific BKCa blockers iberiotoxin (IBTX) or tetraethylammonium, showing specific BKCa activity in hUASMC. H2S dose (NaHS, 1–1000 µM)-dependently potentiated BKCa currents and open probability. Co-incubation with a Ca2+ blocker nifedipine (5 µM) or a chelator (ethylene glycol-bis (β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA), 5 mM) did not alter H2S-potentiated BKCa currents and open probability. NaHS also dose-dependently relaxed phenylephrine pre-constricted freshly prepared human UA rings, which was inhibited by IBTX. Thus, H2S stimulated human UA relaxation at least partially via activating SM BKCa channels independent of extracellular Ca2+.
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Wang, Wei, Haixia Huang, Dongyan Hou, Ping Liu, Hua Wei, Xiaosuo Fu, and Weizhen Niu. "Mechanosensitivity of STREX-lacking BKCa channels in the colonic smooth muscle of the mouse." American Journal of Physiology-Gastrointestinal and Liver Physiology 299, no. 6 (December 2010): G1231—G1240. http://dx.doi.org/10.1152/ajpgi.00268.2010.
Full textAbstract:
Stretch sensitivity of Ca2+-activated large-conductance K+ channels (BKCa) has been observed in a variety of cell types and considered to be a potential mechanism in mechanoelectric transduction (MET). Mechanical stress is a major stimulator for the smooth muscle in the gastrointestinal (GI) tract. However, much about the role and mechanism of MET in GI smooth muscles remains unknown. The BKCa shows a functional diversity due to intensive Slo I alternative splicing and different α/β-subunit assembly in various cells. The stress-regulated exon (STREX) insert is suggested to be an indispensable domain for the mechanosensitivity of BKCa. The purpose of this study was to determine whether the BKCa in colonic myocytes of the adult mouse is sensitive to mechanical stimulation and whether the STREX insert is a crucial segment for the BKCa mechanosensitivity. The α- and β1-subunit mRNAs and the α-subunit protein of the BKCa channels were detected in the colonic muscularis. We found that the BKCa STREX-lacking variant was abundantly expressed in the smooth muscle, whereas the STREX variant was not detectable. We demonstrated that the STREX-lacking BKCa channels were also sensitive to membrane stretch. We suggest that in addition to the STREX domain, there are other additional structures in the channel responsible for mechanically coupling with the cell membrane.
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Bao, Lin, and Daniel H. Cox. "Gating and Ionic Currents Reveal How the BKCa Channel's Ca2+ Sensitivity Is Enhanced by its β1 Subunit." Journal of General Physiology 126, no. 4 (September 26, 2005): 393–412. http://dx.doi.org/10.1085/jgp.200509346.
Full textAbstract:
Large-conductance Ca2+-activated K+ channels (BKCa channels) are regulated by the tissue-specific expression of auxiliary β subunits. β1 is predominately expressed in smooth muscle, where it greatly enhances the BKCa channel's Ca2+ sensitivity, an effect that is required for proper regulation of smooth muscle tone. Here, using gating current recordings, macroscopic ionic current recordings, and unitary ionic current recordings at very low open probabilities, we have investigated the mechanism that underlies this effect. Our results may be summarized as follows. The β1 subunit has little or no effect on the equilibrium constant of the conformational change by which the BKCa channel opens, and it does not affect the gating charge on the channel's voltage sensors, but it does stabilize voltage sensor activation, both when the channel is open and when it is closed, such that voltage sensor activation occurs at more negative voltages with β1 present. Furthermore, β1 stabilizes the active voltage sensor more when the channel is closed than when it is open, and this reduces the factor D by which voltage sensor activation promotes opening by ∼24% (16.8→12.8). The effects of β1 on voltage sensing enhance the BKCa channel's Ca2+ sensitivity by decreasing at most voltages the work that Ca2+ binding must do to open the channel. In addition, however, in order to fully account for the increase in efficacy and apparent Ca2+ affinity brought about by β1 at negative voltages, our studies suggest that β1 also decreases the true Ca2+ affinity of the closed channel, increasing its Ca2+ dissociation constant from ∼3.7 μM to between 4.7 and 7.1 μM, depending on how many binding sites are affected.
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Braun, Andrew P. "Ammonium ion enhances the calcium-dependent gating of a mammalian large conductance, calcium-sensitive K+ channel." Canadian Journal of Physiology and Pharmacology 79, no. 11 (November 1, 2001): 919–23. http://dx.doi.org/10.1139/y01-076.
Full textAbstract:
We observed that the current amplitude and activation of expressed, mouse brain large conductance, calcium-sensitive K+ channels (BKCa channels) may be reversibly enhanced following addition of low concentrations of the weakly permeant cation NH4+ to the cytoplasmic face of the channel in excised, inside-out membrane patches from HEK 293 cells. Conductance-voltage relations were left-shifted along the voltage axis by addition of NH4Cl in a concentration-dependent manner, with an EC50 of 18.5 mM. Furthermore, this effect was observed in the presence of cytosolic free calcium (~1 µM), but was absent in a cytosolic bath solution containing nominally zero free calcium (e.g., 5 mM EGTA only), a condition under which these channels undergo largely voltage-dependent gating. Recordings of single BKCa channel events indicated that NH4+ increased the channel open probability of single channel activity ~3-fold, but did not alter the amplitude of single channel currents. These findings suggest that the calcium-sensitive gating of mammalian BKCa channels may be modified by other ions present in cytosolic solution.Key words: potassium channel, calcium, modulation, electrophysiology.
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Lynch, Fiona M., Sarah B. Withers, Zhihong Yao, Matthias E. Werner, Gill Edwards, Arthur H. Weston, and Anthony M. Heagerty. "Perivascular adipose tissue-derived adiponectin activates BKCa channels to induce anticontractile responses." American Journal of Physiology-Heart and Circulatory Physiology 304, no. 6 (March 15, 2013): H786—H795. http://dx.doi.org/10.1152/ajpheart.00697.2012.
Full textAbstract:
This study aims to identify the potential mechanisms by which perivascular adipose tissue (PVAT) reduces tone in small arteries. Small mesenteric arteries from wild-type and large-conductance Ca2+-activated K+ (BKCa) channel knockout mice were mounted on a wire myograph in the presence and absence of PVAT, and contractile responses to norepinephrine were assessed. Electrophysiology studies were performed in isolated vessels to measure changes in membrane potential produced by adiponectin. Contractile responses from wild-type mouse small arteries were significantly reduced in the presence of PVAT. This was not observed in the presence of a BKCa channel inhibitor or with nitric oxide synthase (NOS) inhibition or in BKCa or adiponectin knockout mice. Solution transfer experiments demonstrated the presence of an anticontractile factor released from PVAT. Adiponectin-induced vasorelaxation and hyperpolarization in wild-type arteries were not evident in the absence of or after inhibition of BKCa channels. PVAT from BKCa or adiponectin knockout mice failed to elicit an anticontractile response in wild-type arteries. PVAT releases adiponectin, which is an anticontractile factor. Its effect on vascular tone is mediated by activation of BKCa channels on vascular smooth muscle cells and adipocytes and by endothelial mechanisms.
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Hannah, Rachael M., Kathryn M. Dunn, Adrian D. Bonev, and Mark T. Nelson. "Endothelial SKCa and IKCa Channels Regulate Brain Parenchymal Arteriolar Diameter and Cortical Cerebral Blood Flow." Journal of Cerebral Blood Flow & Metabolism 31, no. 5 (December 22, 2010): 1175–86. http://dx.doi.org/10.1038/jcbfm.2010.214.
Full textAbstract:
Calcium-sensitive potassium (KCa) channels have been shown to modulate the diameter of cerebral pial arteries; however, little is known regarding their roles in controlling cerebral parenchymal arterioles (PAs). We explored the function and cellular distribution of small-conductance (SKCa) and intermediate-conductance (IKCa) KCa channels and large-conductance KCa (BKCa) channels in endothelial cells (ECs) and smooth muscle cells (SMCs) of PAs. Both SKCa and IKCa channels conducted the outward current in isolated PA ECs (current densities, ~20 pA/pF and ~28 pA/pF at + 40 mV, respectively), but these currents were not detected in PA SMCs. In contrast, BKCa currents were prominent in PA SMCs (~ 154pA/pF), but were undetectable in PA ECs. Pressurized PAs constricted to inhibition of SKCa (~ 16%) and IKCa (~ 16%) channels, but were only modestly affected by inhibition of BKCa channels (~ 5%). Blockade of SKCa and IKCa channels decreased resting cortical cerebral blood flow (CBF) by ~ 15%. NS309 (6,7-dichloro-1H-indole-2,3-dione3-oxime), a SKCa/IKCa channel opener, hyperpolarized PA SMCs by ~ 27 mV, maximally dilated pressurized PAs, and increased CBF by ~ 40%. In conclusion, these data show that SKCa and IKCa channels in ECs profoundly modulate PA tone and CBF, whereas BKCa channels in SMCs only modestly influence PA diameter.
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Fallet, Rachel W., Joseph P. Bast, Keiji Fujiwara, Naohito Ishii, Steven C. Sansom, and Pamela K. Carmines. "Influence of Ca2+-activated K+ channels on rat renal arteriolar responses to depolarizing agonists." American Journal of Physiology-Renal Physiology 280, no. 4 (April 1, 2001): F583—F591. http://dx.doi.org/10.1152/ajprenal.2001.280.4.f583.
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Experiments were performed to evaluate the hypothesis that opening of Ca2+-activated K+ channels (BKCachannels) promotes juxtamedullary arteriolar dilation and curtails constrictor responses to depolarizing agonists. Under baseline conditions, afferent and efferent arteriolar lumen diameters averaged 23.4 ± 0.9 ( n = 36) and 22.8 ± 1.1 ( n= 13) μm, respectively. The synthetic BKCa channel opener NS-1619 evoked concentration-dependent afferent arteriolar dilation. BKCa channel blockade (1 mM tetraethylammonium; TEA) decreased afferent diameter by 15 ± 3% and prevented the dilator response to 30 μM NS-1619. ANG II (10 nM) decreased afferent arteriolar diameter by 44 ± 4%, a response that was reduced by 30% during NS-1619 treatment; however, TEA failed to alter afferent constrictor responses to either ANG II or arginine vasopressin. Neither NS-1619 nor TEA altered agonist-induced constriction of the efferent arteriole. Thus, although the BKCa channel agonist was able to curtail afferent (but not efferent) arteriolar constrictor responses to ANG II, BKCa channel blockade did not allow exaggerated agonist-induced arteriolar constriction. These observations suggest that the BKCa channels evident in afferent arteriolar smooth muscle do not provide a prominent physiological brake on agonist-induced constriction under our experimental conditions.
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Qian, Lingling, Xiaoyu Liu, and Ruxing Wang. "Role of BKCa channels in diabetic vascular complications." Chinese Medical Journal 127, no. 9 (May 5, 2014): 1775–81. http://dx.doi.org/10.3760/cma.j.issn.0366-6999.20132503.
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Objective This review focuses on the role of the large conductance calcium-activated potassium (BKCa) channels in diabetic vascular complications. Data sources Relevant articles published in English or Chinese from 1981 to present were selected from PubMed. The search terms were “BKCa channels” and “diabetes”. Important references from selected articles were also retrieved. Study selection Articles regarding the role of BKCa channels in diabetic vascular complications and relevant mechanisms were selected. Results The BKCa channels are abundantly expressed in vascular smooth cells and play an important role in regulation of vascular tone. Multiple studies indicated that the expression and function of BKCa channels are altered by different mechanisms in diabetic vascular diseases such as coronary arterial disease, cerebral arterial disease, and diabetic retinopathy. Conclusion BKCa channels may play an important role in diabetic vascular complications and may be an effective therapeutic target for relieving and reducing the burden of diabetic vascular complications.
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Campbell, William B., Blythe B. Holmes, John R. Falck, Jorge H. Capdevila, and Kathryn M. Gauthier. "Regulation of potassium channels in coronary smooth muscle by adenoviral expression of cytochrome P-450 epoxygenase." American Journal of Physiology-Heart and Circulatory Physiology 290, no. 1 (January 2006): H64—H71. http://dx.doi.org/10.1152/ajpheart.00516.2005.
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Epoxyeicosatrienoic acids (EETs) are endothelium-derived cytochrome P-450 (CYP) metabolites of arachidonic acid that relax vascular smooth muscle by large-conductance calcium-activated potassium (BKCa) channel activation and membrane hyperpolarization. We hypothesized that if smooth muscle cells (SMCs) had the capacity to synthesize EETs, endogenous EET production would increase BKCa channel activity. Bovine coronary SMCs were transduced with adenovirus coding the CYP Bacillus megaterium -3 (F87V) (CYP BM-3) epoxygenase that metabolizes arachidonic acid exclusively to 14( S),15( R)-EET. Adenovirus containing the cytomegalovirus promoter- Escherichia coli β-galactosidase was used as a control. With the use of an anti-CYP BM-3 (F87V) antibody, a 124-kDa immunoreactive protein was detected only in CYP BM-3-transduced cells. Protein expression increased with increasing amounts of virus. When CYP BM-3-transduced cells were incubated with [14C]arachidonic acid, HPLC analysis detected 14,15-dihydroxyeicosatrienoic acid (14,15-DHET) and 14,15-EET. The identity of 14,15-EET and 14,15-DHET was confirmed by mass spectrometry. In CYP BM-3-transduced cells, methacholine (10−5 M) increased 14,15-EET release twofold and BKCa channel activity fourfold in cell-attached patches. Methacholine-induced increases in BKCa channel activity were blocked by the CYP inhibitor 17-octadecynoic acid (10−5 M). 14( S),15( R)-EET was more potent than 14( R),15( S)-EET in relaxing bovine coronary arteries and activating BKCa channels. Thus CYP BM-3 adenoviral transduction confers SMCs with epoxygenase activity. These cells acquire the capacity to respond to the vasodilator agonist by synthesizing 14( S),15( R)-EET from endogenous arachidonic acid to activate BKCa channels. These studies indicate that 14( S),15( R)-EET is a sufficient endogenous activator of BKCa channels in coronary SMCs.
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Sweet, Tara-Beth, and Daniel H. Cox. "Measuring the Influence of the BKCa β1 Subunit on Ca2+ Binding to the BKCa Channel." Journal of General Physiology 133, no. 2 (January 12, 2009): 139–50. http://dx.doi.org/10.1085/jgp.200810129.
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The large-conductance Ca2+-activated potassium (BKCa) channel of smooth muscle is unusually sensitive to Ca2+ as compared with the BKCa channels of brain and skeletal muscle. This is due to the tissue-specific expression of the BKCa auxiliary subunit β1, whose presence dramatically increases both the potency and efficacy of Ca2+ in promoting channel opening. β1 contains no Ca2+ binding sites of its own, and thus the mechanism by which it increases the BKCa channel's Ca2+ sensitivity has been of some interest. Previously, we demonstrated that β1 stabilizes voltage sensor activation, such that activation occurs at more negative voltages with β1 present. This decreases the work that Ca2+ must do to open the channel and thereby increases the channel's apparent Ca2+ affinity without altering the real affinities of the channel's Ca2+ binding sites. To explain the full effect of β1 on the channel's Ca2+ sensitivity, however, we also proposed that there must be effects of β1 on Ca2+ binding. Here, to test this hypothesis, we have used high-resolution Ca2+ dose–response curves together with binding site–specific mutations to measure the effects of β1 on Ca2+ binding. We find that coexpression of β1 alters Ca2+ binding at both of the BKCa channel's two types of high-affinity Ca2+ binding sites, primarily increasing the affinity of the RCK1 sites when the channel is open and decreasing the affinity of the Ca2+ bowl sites when the channel is closed. Both of these modifications increase the difference in affinity between open and closed, such that Ca2+ binding at either site has a larger effect on channel opening when β1 is present.
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Kang, Lori S., SeJeong Kim, James M. Dominguez, Amy L. Sindler, Gregory M. Dick, and Judy M. Muller-Delp. "Aging and muscle fiber type alter K+ channel contributions to the myogenic response in skeletal muscle arterioles." Journal of Applied Physiology 107, no. 2 (August 2009): 389–98. http://dx.doi.org/10.1152/japplphysiol.91245.2008.
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Aging diminishes myogenic tone in arterioles from skeletal muscle. Recent evidence indicates that both large-conductance Ca2+-activated (BKCa) and voltage-dependent (KV) K+ channels mediate negative feedback control of the myogenic response. Thus we tested the hypothesis that aging increases the contributions of KV and BKCa channels to myogenic regulation of vascular tone. Because myogenic responsiveness differs between oxidative and glycolytic muscles, we predicted that KV and BKCa channel contributions to myogenic responsiveness vary with fiber type. Myogenic responses of first-order arterioles from the gastrocnemius and soleus muscles of 4- and 24-mo-old Fischer 344 rats were evaluated in the presence and absence of 4-aminopyridine (5 mM) or iberiotoxin (30 nM), inhibitors of KV and BKCa, respectively. 4-Aminopyridine enhanced myogenic tone with aging and normalized age-related differences in both muscle types. By contrast, iberiotoxin eliminated age-related differences in soleus arterioles and had no effect in gastrocnemius vessels. KV1.5 is an integral component of KV channels in vascular smooth muscle; therefore, we determined the relative protein expression of KV1.5, as well as BKCa, in soleus and gastrocnemius arterioles. Immunoblot analysis revealed no differences in KV1.5 protein with aging or between variant fiber types, whereas BKCa protein levels declined with age in arterioles from both muscle groups. Collectively, these results suggest that the contribution of BKCa to myogenic regulation of vascular tone changes with age in soleus muscle arterioles, whereas increased KV channel expression and negative feedback regulation of myogenic tone increases with advancing age in arterioles from both oxidative and glycolytic muscles.
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Cordeiro, Brenda, Dmitry Terentyev, and Richard T. Clements. "BKCa channel activation increases cardiac contractile recovery following hypothermic ischemia/reperfusion." American Journal of Physiology-Heart and Circulatory Physiology 309, no. 4 (August 15, 2015): H625—H633. http://dx.doi.org/10.1152/ajpheart.00818.2014.
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Mitochondrial Ca2+-activated large-conductance K+ (BKCa) channels are thought to provide protection during ischemic insults in the heart. Rottlerin (mallotoxin) has been implicated as a potent BKCa activator. The purpose of this study was twofold: 1) to investigate the efficacy of BKCa channel activation as a cardioprotective strategy during ischemic cardioplegic arrest and reperfusion (CP/R) and 2) to assess the specificity of rottlerin for BKCa channels. Wild-type (WT) and BKCa knockout (KO) mice were subjected to an isolated heart model of ischemic CP/R. A mechanism of rottlerin-induced cardioprotection was also investigated using H9c2 cells subjected to in vitro CP/reoxygenation and assessed for mitochondrial membrane potential and reactive oxygen species (ROS) production. CP/R decreased left ventricular developed pressure, positive and negative first derivatives of left ventricular pressure, and coronary flow (CF) in WT mice. Rottlerin dose dependently increased the recovery of left ventricular function and CF to near baseline levels. BKCa KO hearts treated with or without 500 nM rottlerin were similar to WT CP hearts. H9c2 cells subjected to in vitro CP/R displayed reduced mitochondrial membrane potential and increased ROS generation, both of which were significantly normalized by rottlerin. We conclude that activation of BKCa channels rescues ischemic damage associated with CP/R, likely via effects on improved mitochondrial membrane potential and reduced ROS generation.
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Borbouse, Léna, Gregory M. Dick, Shinichi Asano, Shawn B. Bender, U. Deniz Dincer, Gregory A. Payne, Zachary P. Neeb, Ian N. Bratz, Michael Sturek, and Johnathan D. Tune. "Impaired function of coronary BKCa channels in metabolic syndrome." American Journal of Physiology-Heart and Circulatory Physiology 297, no. 5 (November 2009): H1629—H1637. http://dx.doi.org/10.1152/ajpheart.00466.2009.
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The role of large-conductance Ca2+-activated K+ (BKCa) channels in regulation of coronary microvascular function is widely appreciated, but molecular and functional changes underlying the deleterious influence of metabolic syndrome (MetS) have not been determined. Male Ossabaw miniature swine consumed for 3–6 mo a normal diet (11% kcal from fat) or an excess-calorie atherogenic diet that induces MetS (45% kcal from fat, 2% cholesterol, 20% kcal from fructose). MetS significantly impaired coronary vasodilation to the BKCa opener NS-1619 in vivo (30–100 μg) and reduced the contribution of these channels to adenosine-induced microvascular vasodilation in vitro (1–100 μM). MetS reduced whole cell penitrem A (1 μM)-sensitive K+ current and NS-1619-activated (10 μM) current in isolated coronary vascular smooth muscle cells. MetS increased the concentration of free intracellular Ca2+ and augmented coronary vasoconstriction to the L-type Ca2+ channel agonist BAY K 8644 (10 pM–10 nM). BKCa channel α and β1 protein expression was increased in coronary arteries from MetS swine. Coronary vascular dysfunction in MetS is related to impaired BKCa channel function and is accompanied by significant increases in L-type Ca2+ channel-mediated coronary vasoconstriction.
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Rosenfeld, Charles R., David N. Cornfield, and Timothy Roy. "Ca2+-activated K+ channels modulate basal and E2β-induced rises in uterine blood flow in ovine pregnancy." American Journal of Physiology-Heart and Circulatory Physiology 281, no. 1 (July 1, 2001): H422—H431. http://dx.doi.org/10.1152/ajpheart.2001.281.1.h422.
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Uterine blood flow (UBF) increases >30-fold during ovine pregnancy. During the last trimester, this reflects vasodilation, which may be due to placentally derived estrogens. In nonpregnant ewes, estradiol-17β (E2β) increases UBF >10-fold by activating nitric oxide synthase and large conductance calcium-dependent potassium channels (BKCa). To determine whether BKCa channels modulate basal and E2β-induced increases in UBF, studies were performed in near-term pregnant ewes with uterine artery flow probes and catheters for intra-arterial infusions of tetraethylammonium (TEA), a selective BKCa channel antagonist at <1 mM, in the absence or presence of E2β (1 μg/kg iv). Uterine arteries were collected to measure BKCa channel mRNA. TEA (0.15 mM) decreased basal UBF ( P < 0.0001) 40 ± 8% and 55 ± 7% ( n = 11) at 60 and 90 min, respectively, and increased resistance 175 ± 48% without affecting ( P > 0.1) mean arterial pressure (MAP), heart rate, or contralateral UBF. Systemic E2β increased UBF 30 ± 6% and heart rate 13 ± 1% ( P ≤ 0.0001, n = 13) without altering MAP. Local TEA (0.15 mM) inhibited E2β-induced increases in UBF without affecting increases in heart rate (10 ± 4%; P = 0.006). BKCa channel mRNA was present in uterine artery myocytes from pregnant and nonpregnant ewes. Exponential increases in ovine UBF in late pregnancy may reflect BKCa channel activation, which may be mediated by placentally derived estrogens.
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Savalli, Nicoletta, Andrei Kondratiev, Sarah Buxton de Quintana, Ligia Toro, and Riccardo Olcese. "Modes of Operation of the BKCa Channel β2 Subunit." Journal of General Physiology 130, no. 1 (June 25, 2007): 117–31. http://dx.doi.org/10.1085/jgp.200709803.
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The β2 subunit of the large conductance Ca2+- and voltage-activated K+ channel (BKCa) modulates a number of channel functions, such as the apparent Ca2+/voltage sensitivity, pharmacological and kinetic properties of the channel. In addition, the N terminus of the β2 subunit acts as an inactivating particle that produces a relatively fast inactivation of the ionic conductance. Applying voltage clamp fluorometry to fluorescently labeled human BKCa channels (hSlo), we have investigated the mechanisms of operation of the β2 subunit. We found that the leftward shift on the voltage axis of channel activation curves (G(V)) produced by coexpression with β2 subunits is associated with a shift in the same direction of the fluorescence vs. voltage curves (F(V)), which are reporting the voltage dependence of the main voltage-sensing region of hSlo (S4-transmembrane domain). In addition, we investigated the inactivating mechanism of the β2 subunits by comparing its properties with the ones of the typical N-type inactivation process of Shaker channel. While fluorescence recordings from the inactivated Shaker channels revealed the immobilization of the S4 segments in the active conformation, we did not observe a similar feature in BKCa channels coexpressed with the β2 subunit. The experimental observations are consistent with the view that the β2 subunit of BKCa channels facilitates channel activation by changing the voltage sensor equilibrium and that the β2-induced inactivation process does not follow a typical N-type mechanism.
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Zhao, T., H. Zhang, C. Jin, F. Qiu, Y. Wu, and L. Shi. "Melatonin mediates vasodilation through both direct and indirect activation of BKCa channels." Journal of Molecular Endocrinology 59, no. 3 (October 2017): 219–33. http://dx.doi.org/10.1530/jme-17-0028.
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Melatonin, synthesized primarily by the pineal gland, is a neuroendocrine hormone with high membrane permeability. The vascular effects of melatonin, including vasoconstriction and vasodilation, have been demonstrated in numerous studies. However, the mechanisms underlying these effects are not fully understood. Large-conductance Ca2+-activated K+ (BKCa) channels are expressed broadly on smooth muscle cells and play an important role in vascular tone regulation. This study explored the mechanisms of myocyte BKCa channels and endothelial factors underlying the action of melatonin on the mesenteric arteries (MAs). Vascular contractility and patch-clamp studies were performed on myocytes of MAs from Wistar rats. Melatonin induced significant vasodilation on MAs. In the presence of Nω-nitro-l-arginine methyl ester (l-NAME), a potent endothelial oxide synthase (eNOS) inhibitor, melatonin elicited concentration-dependent relaxation, with lowered pIC50. The effect of melatonin was significantly attenuated in the presence of BKCa channel blocker iberiotoxin or MT1/MT2 receptor antagonist luzindole in both (+) l-NAME and (−) l-NAME groups. In the (+) l-NAME group, iberiotoxin caused a parallel rightward shift of the melatonin concentration–relaxation curve, with pIC50 lower than that of luzindole. Both inside-out and cell-attached patch-clamp recordings showed that melatonin significantly increased the open probability, mean open time and voltage sensitivity of BKCa channels. In a cell-attached patch-clamp configuration, the melatonin-induced enhancement of BKCa channel activity was significantly suppressed by luzindole. These findings indicate that in addition to the activation of eNOS, melatonin-induced vasorelaxation of MAs is partially attributable to its direct (passing through the cell membrane) and indirect (via MT1/MT2 receptors) activation of the BKCa channels on mesenteric arterial myocytes.
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Bai, Bing, Nanjuan Lu, Wei Zhang, Jinghan Lin, Tingting Zhao, Shanshan Zhou, Elona Khasanova, and Liming Zhang. "Inhibitory Effects of Genistein on Vascular Smooth Muscle Cell Proliferation Induced by Ox-LDL: Role of BKCa Channels." Analytical Cellular Pathology 2020 (December 13, 2020): 1–12. http://dx.doi.org/10.1155/2020/8895449.
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Background. Oxidized low-density lipoprotein (Ox-LDL) is a crucial pathogenic factor for vascular diseases, which can induce the proliferation of vascular smooth muscle cells (VSMCs). Genistein is the main component of soybean isoflavone. Genistein has a variety of pharmacological properties in the treatment of vascular diseases and a promising clinical application. Large-conductance calcium-activated potassium (BKCa) channels are the primary type of potassium channels in VSMCs, which regulate various biological functions of VSMCs. However, whether genistein exerts an antiproliferation effect on Ox-LDL-stimulated VSMCs remains unclear. The current study is aimed at elucidating the effect of genistein on the Ox-LDL-stimulated proliferation of VSMCs and its possible molecular mechanism, especially the electrophysiological mechanism related to BKCa channels. Methods. Monoculture VSMC was obtained by an acute enzyme-dispersing method. The proliferation of cells was measured by CCK-8, cell cycle, and proliferating cell nuclear antigen (PCNA) expression. The BKCa whole-cell currents were measured by patch-clamp. Results. Ox-LDL treatment induced the proliferation of VSMCs, upregulated the BKCa protein expression, and increased the density of BKCa currents, while genistein significantly inhibited these effects caused by Ox-LDL. BKCa channels exerted a regulatory role in the proliferation of VSMCs in response to Ox-LDL. The inhibition of BKCa channels suppressed Ox-LDL-stimulated VSMC proliferation, while the activation of BKCa channels showed the opposite effect. Moreover, genistein suppressed the activity of BKCa, including protein expression and current density in a protein tyrosine kinase- (PTK-) dependent manner. Conclusion. This study demonstrated that genistein inhibited the Ox-LDL-mediated proliferation of VSMCs by blocking the cell cycle progression; the possible molecular mechanism may be related to PTK-dependent suppression of BKCa channels. Our results provided novel ideas for the application of genistein in the treatment of vascular diseases and proposed a unique insight into the antiproliferative molecular mechanism of genistein.
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Storer, RJ, DC Immke, R. Yin, and PJ Goadsby. "Large Conductance Calcium-Activated Potassium Channels (BKCa) Modulate Trigeminovascular Nociceptive Transmission." Cephalalgia 29, no. 12 (December 2009): 1242–58. http://dx.doi.org/10.1111/j.1468-2982.2009.01849.x.
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Migraine is a common, disabling, neurological problem whose acute management would benefit from the development of purely neurally acting therapies. The trigeminocervical complex is pivotal in nociceptive signaling in migraine, and is an accepted target for putative antimigraine agents. Whole-cell patch-clamp or extracellular recordings were made of trigeminal neurons identified in rat brainstem slices. Bath application of the large conductance calcium-activated potassium (BKCa) channel opener NS1619 caused a dramatic decrease of cell firing that could be reversed by the co-application of iberiotoxin. NS1619 hyperpolarized the resting membrane potential and reduced the frequency of spontaneous action potentials in these neurons. These data suggest the presence of BKCa channels in the trigeminocervical complex. In vivo in cat L-glutamate-evoked firing was facilitated in nociceptive neurons, also responding to stimulation of the superior sagittal sinus, in the trigeminal nucleus caudalis by the BKCa peptide antagonists, iberiotoxin and slotoxin. Of units tested, 70% responded to microiontophoretic application of the blockers, identifying a subpopulation of trigeminal neurons expressing toxin-sensitive BKCa channels. NS1619 inhibited 74% of cells tested, and this was reversed by slotoxin, suggesting that the action of NS1619 in these cells was mediated through BKCa channels. These data are consistent with the presence of BKCa channels in the trigeminal nucleus caudalis that are potential targets for the development of antimigraine treatments, and may also offer insights into receptor mechanisms involved in sensitization and thus allodynia, in migraine.
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Kim, Eun Young, Shengwei Zou, Lon D. Ridgway, and Stuart E. Dryer. "β1-Subunits Increase Surface Expression of a Large-Conductance Ca2+-Activated K+ Channel Isoform." Journal of Neurophysiology 97, no. 5 (May 2007): 3508–16. http://dx.doi.org/10.1152/jn.00009.2007.
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Auxiliary (beta) subunits of large-conductance Ca2+-activated K+ (BKCa) channels regulate the gating properties of the functional channel complex. Here we show that an avian β1-subunit also stimulates the trafficking of BKCa channels to the plasma membrane in HEK293T cells and in a native population of developing vertebrate neurons. One C-terminal variant of BKCa α-subunits, called the VEDEC isoform after its five last residues, is largely retained in intracellular compartments when it is heterologously expressed in HEK293T cells. A closely related splice variant, called QEERL, shows high levels of constitutive trafficking to the plasma membrane. Co-expression of β1-subunits with the VEDEC isoform resulted in a large increase in surface BKCa channels as assessed by cell-surface biotinylation assays, whole cell recordings of membrane current, and confocal microscopy in HEK293T cells. Co-expression of β1-subunits slowed the gating kinetics of BKCa channels, as reported previously. Consistent with this, overexpression of β1-subunits in a native cell type that expresses intracellular VEDEC channels, embryonic day 9 chick ciliary ganglion neurons, resulted in a significant increase in macroscopic Ca2+-activated K+ current. Both the cytoplasmic N- and C-terminal domains of avian β1 are able to bind directly to VEDEC and QEERL channels. However, overexpression of the N-terminal domain by itself is sufficient to stimulate trafficking of VEDEC channels to the plasma membrane, whereas overexpression of either the cytoplasmic C-terminal domain or the extracellular loop domain did not affect surface expression of VEDEC.
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Zhang, Q., Y. Bai, Z. Yang, J. Tian, and Z. Meng. "The molecular mechanism of the effect of sulfur dioxide inhalation on the potassium and calcium ion channels in rat aortas." Human & Experimental Toxicology 35, no. 4 (June 24, 2015): 418–27. http://dx.doi.org/10.1177/0960327115591375.
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This study investigated the molecular mechanism of the effect of sulfur dioxide (SO2) on the expression of adenosine triphosphate (ATP)-sensitive potassium ion (K+; KATP) channel, big-conductance calcium ion (Ca2+)-activated K+ (BKCa) channel, and L-type (L-Ca2+) channel subunits in rat aortas with quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blot. The results showed that the messenger RNA and protein levels of the KATP channel subunits Kir6.1, Kir6.2, and sulfonylurea receptor 2B (SUR2B) of rat aortas were significantly increased by SO2 at 14 mg/m3, whereas the levels of SUR2A were not changed. SO2 at all the treated concentrations markedly raised the expression of the BKCa channel subunits α and β1. SO2 at 14 mg/m3 significantly decreased the expression of the L-Ca2+ channel Cav1.2 and Cav1.3. The histological examination of rat aorta tissues showed moderate injury of tunica media in the presence of SO2 at 14 mg/m3. These suggest that SO2 can activate the KATP and BKCa channels by upregulating the expression of Kir6.1, Kir6.2, SUR2B, BKCa α, and BKCa β1, while inhibit the L-Ca2+ channels by downregulating the expression of Cav1.2 and Cav1.3 in rat aortas. The molecular mechanism of SO2-induced vasorelaxant effect might be linked to the changes in expression of these channel subunits, which plays an important role in the pathogenesis of SO2-associated cardiovascular diseases.