The Strex-Insert in a Stretch-Activated BK Channel from Chick Heart is not Enough for Conferring GsMTx-4 Inhibitory Effect on mSlo1

High-conductance apical K+(BK) channels are present in surface colonocytes of mammalian (including human) colon. Their location makes them well fitted to contribute to the excessive intestinal K+losses often associated with infective diarrhea. Since many channel proteins are regulated by phosphorylation, we evaluated the roles of protein kinase A (PKA) and phosphatases in the modulation of apical BK channel activity in surface colonocytes from rat distal colon using patch-clamp techniques, having first increased channel abundance by chronic dietary K+enrichment. We found that PKA activation using 50 μmol/l forskolin and 5 mmol/l 3-isobutyl-1-methylxanthine stimulated BK channels in cell-attached patches and the catalytic subunit of PKA (200 U/ml) had a similar effect in excised inside-out patches. The antidiarrheal peptide somatostatin (SOM; 2 μmol/l) had a G protein-dependent inhibitory effect on BK channels in cell-attached patches, which was unaffected by pretreatment with 10 μmol/l okadaic acid (an inhibitor of protein phosphatase type 1 and type 2A) but completely prevented by pretreatment with 100 μmol/l Na+orthovanadate and 10 μmol/l BpV (inhibitors of phosphoprotein tyrosine phosphatase). SOM also inhibited apical BK channels in surface colonocytes in human distal colon. We conclude that cAMP-dependent PKA activates apical BK channels and may enhance colonic K+losses in some cases of secretory diarrhea. SOM inhibits apical BK channels through a phosphoprotein tyrosine phosphatase-dependent mechanism, which could form the basis of new antidiarrheal strategies.

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Peroxynitrite reversibly inhibits Ca2+-activated K+ channels in rat cerebral artery smooth muscle cells

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.2000.278.6.h1883 ◽

2000 ◽

Vol 278 (6) ◽

pp. H1883-H1890 ◽

Cited By ~ 63

Author(s):

Anna K. Brzezinska ◽

Debebe Gebremedhin ◽

William M. Chilian ◽

Balaraman Kalyanaraman ◽

Stephen J. Elliott

Keyword(s):

Single Channel ◽

Cerebral Arteries ◽

Ionic Current ◽

Inhibitory Effect ◽

Bk Channels ◽

Bk Channel ◽

Channel Activity ◽

Open Probability ◽

Whole Cell ◽

Cell Current

Peroxynitrite (ONOO−) is a contractile agonist of rat middle cerebral arteries. To determine the mechanism responsible for this component of ONOO−bioactivity, the present study examined the effect of ONOO− on ionic current and channel activity in rat cerebral arteries. Whole cell recordings of voltage-clamped cells were made under conditions designed to optimize K+ current. The effects of iberiotoxin, a selective inhibitor of large-conductance Ca2+-activated K+ (BK) channels, and ONOO− (10–100 μM) were determined. At a pipette potential of +50 mV, ONOO− inhibited 39% of iberiotoxin-sensitive current. ONOO− was selective for iberiotoxin-sensitive current, whereas decomposed ONOO− had no effect. In excised, inside-out membrane patches, channel activity was recorded using symmetrical K+solutions. Unitary currents were sensitive to increases in internal Ca2+ concentration, consistent with activity due to BK channels. Internal ONOO− dose dependently inhibited channel activity by decreasing open probability and mean open times. The inhibitory effect of ONOO− could be overcome by reduced glutathione. Glutathione, added after ONOO−, restored whole cell current amplitude to control levels and reverted single-channel gating to control behavior. The inhibitory effect of ONOO− on membrane K+ current is consistent with its contractile effects in isolated cerebral arteries and single myocytes. Taken together, our data suggest that ONOO− has the potential to alter cerebral vascular tone by inhibiting BK channel activity.

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Regulation of endothelial BK channels by heme oxygenase-derived carbon monoxide and caveolin-1

AJP Cell Physiology ◽

10.1152/ajpcell.00356.2011 ◽

2012 ◽

Vol 303 (1) ◽

pp. C92-C101 ◽

Cited By ~ 12

Author(s):

Melissa A. Riddle ◽

Benjimen R. Walker

Keyword(s):

Carbon Monoxide ◽

Heme Oxygenase ◽

Barometric Pressure ◽

Inhibitory Effect ◽

Bk Channels ◽

Bk Channel ◽

Channel Activity ◽

Outward Currents ◽

In Cells

A novel vasodilatory influence of endothelial cell (EC) large-conductance Ca2+-activated K+ (BK) channels is present after 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. Previously, we demonstrated that inhibition of either BK channels or heme oxygenase (HO) restores vasoconstrictor reactivity after CH. Additionally, administration of the scaffolding domain of caveolin (Cav)-1 inhibits EC BK activity and restores vasoconstrictor reactivity in this setting. These results led us to hypothesize that CH exposure results in a loss in Cav-1 inhibition of EC BK channels, resulting in their activation by HO-derived carbon monoxide (CO). Experiments were conducted on freshly dispersed aortic ECs from control and CH-exposed (barometric pressure: 380 mmHg for 48 h) rats. In electrophysiology experiments, outward currents were greater in cells from CH rats as well as from cells from control rats treated with the cholesterol-depleting agent methyl-β-cyclodextrin. These enhanced currents were returned to control by HO inhibition. Channel activity could be restored by the CO donor CO-releasing molecule (CORM)-2 during HO inhibition. Administration of the Cav-1 scaffolding domain eliminated BK currents in cells from CH rats, and current was not restored by the addition of CORM-2. Colocalization experiments in ECs from control and CH rats demonstrated an association between HO-2, Cav-1, and BK. We conclude that EC BK channel activity is HO dependent in the absence of the inhibitory effect of the Cav-1 scaffolding domain.

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Bethanidine increased Na+ and Ca2+ currents and caused a positive inotropic effect in heart cells

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y88-033 ◽

1988 ◽

Vol 66 (2) ◽

pp. 190-196 ◽

Cited By ~ 5

Author(s):

G. Bkaily ◽

M. D. Payet ◽

M. Benabderrazik ◽

J.-F. Renaud ◽

R. Sauvé ◽

...

Keyword(s):

Positive Inotropic Effect ◽

Inhibitory Effect ◽

Inotropic Effect ◽

Heart Cells ◽

Dependent Manner ◽

Isolated Atria ◽

Cell Current ◽

Chick Heart ◽

Embryonic Chick Heart ◽

Dose Dependent

The effects of bethanidine sulphate, a pharmacological analog of the cardiac antibrillatory drug, bretylium tosylate, were studied on action potentials (APs) and K+, Na+, and Ca2+ currents of single cultured embryonic chick heart cells using the whole-cell current clamp and voltage clamp technique. Extracellular application of bethanidine (3 × 10−4 M) increased the overshoot and the duration of the APs and greatly decreased the outward K+ current (IK) and potentiated the inward fast Na+ currents (INa) and the inward slow calcium current (ICa). However, intracellular introduction of bethanidine (10−4 M) blocked INa. In isolated atria of rat, bethanidine increased the force of contraction in a dose-dependent manner. These findings suggest that when applied extracellularly, bethanidine exerts a potentiating effect on the myocardial fast Na+ current and slow Ca2+ current and an inhibitory effect of IK. The positive inotropic effect of bethanidine could be due, at least in part, to an increase of Ca2+ influx via the slow Ca2+ channel and the Na–Ca exchange. It is suggested that the decrease of IK by bethanidine may account for its antifibrillatory action.

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3R1030 Effects of amphipaths on a stretch activated BK channel(SAKCa) cloned from chick heart

Seibutsu Butsuri ◽

10.2142/biophys.42.s206_3 ◽

2002 ◽

Vol 42 (supplement2) ◽

pp. S206

Author(s):

Z. Qi ◽

K. Naruse ◽

M. Sokabe

Keyword(s):

Bk Channel ◽

Chick Heart

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Techniques for cryoultramicrotomy of propane jet frozen biological samples

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010014292x ◽

1986 ◽

Vol 44 ◽

pp. 260-261

Author(s):

B. Craig ◽

L. Hawkey ◽

A. LeFurgey

Keyword(s):

Freeze Fracture ◽

Freeze Substitution ◽

Heart Cells ◽

Crystal Formation ◽

X Ray ◽

Transmission Electron ◽

Scanning Transmission ◽

Chick Heart ◽

Embryonic Chick Heart ◽

A New Technique

Ultra-rapid freezing followed by cryoultramicrotomy is essential for the preservation of diffusible elements in situ within cells prior to scanning transmission electron microscopy and quantitative energy dispersive x-ray microanalysis. For cells or tissue fragments in suspension and for monolayer cell cultures, propane jet freezing provides cooling rates greater than 30,000°C/sec with regions up to 40μm in thickness free of significant ice crystal formation. While this method of freezing has frequently been applied prior to freeze fracture or freeze substitution, it has not been widely utilized prior to cryoultramicrotomy and subsequent x-ray microanalytical studies. This report describes methods devised in our laboratory for cryosectioning of propane jet frozen kidney proximal tubule suspensions and cultured embryonic chick heart cells, in particular a new technique for mounting frozen suspension specimens for sectioning. The techniques utilize the same specimen supports and sample holders as those used for freeze fracture and freeze substitution and should be generally applicable to any cell suspension or culture preparation.

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