Effects of methyl beta-cyclodextrin on EDHF responses in pig and rat arteries; association between SK(Ca) channels and caveolin-rich domains

M Absi; M P Burnham; A H Weston; E Harno; M Rogers; G Edwards

doi:10.1038/sj.bjp.0707222

Effects of methyl beta-cyclodextrin on EDHF responses in pig and rat arteries; association between SK(Ca) channels and caveolin-rich domains

M Absi, M P Burnham, A H Weston, E Harno, M Rogers, G Edwards

Research output: Contribution to journal › Article › peer-review

Abstract

BACKGROUND AND PURPOSE: The small and intermediate conductance, Ca2+-sensitive K+ channels (SK(Ca) and IK(Ca), respectively) which are pivotal in the EDHF pathway may be differentially activated. The importance of caveolae in the functioning of IK(Ca) and SK(Ca) channels was investigated.

EXPERIMENTAL APPROACH: The effect of the caveolae-disrupting agent methyl-beta-cyclodextrin (MbetaCD) on IK(Ca) and SK(Ca) localization and function was determined.

KEY RESULTS: EDHF-mediated, SK(Ca)-dependent myocyte hyperpolarizations evoked by acetylcholine in rat mesenteric arteries (following blockade of IK(Ca) with TRAM-34) were inhibited by MbetaCD. Hyperpolarizations evoked by direct SK(Ca) channel activation (using NS309 in the presence of TRAM-34) were also inhibited by MbetaCD, an effect reversed by cholesterol. In contrast, IK(Ca)-dependent hyperpolarizations (in the presence of apamin) were unaffected by MbetaCD. Similarly, in porcine coronary arteries, EDHF-mediated, SK(Ca)-dependent (but not IK(Ca)-dependent) endothelial cell hyperpolarizations evoked by substance P were inhibited by MbetaCD. In mesenteric artery homogenates subjected to sucrose-density centrifugation, caveolin-1 and SK3 (SK(Ca)) proteins but not IK1 (IK(Ca)) protein migrated to the buoyant, caveolin-rich fraction. MbetaCD pretreatment redistributed caveolin-1 and SK3 proteins into more dense fractions. In immunofluorescence images of porcine coronary artery endothelium, SK3 (but not IK1) and caveolin-1 were co-localized. Furthermore, caveolin-1 immunoprecipitates prepared from native porcine coronary artery endothelium contained SK3 but not IK1 protein.

CONCLUSIONS AND IMPLICATIONS: These data provide strong evidence that endothelial cell SK(Ca) channels are located in caveolae while the IK(Ca) channels reside in a different membrane compartment. These studies reveal cellular organisation as a further complexity in the EDHF pathway signalling cascade.

Original language	English
Pages (from-to)	332-340
Number of pages	9
Journal	British Journal of Pharmacology
Volume	151
Issue number	3
DOIs	https://doi.org/10.1038/sj.bjp.0707222
Publication status	Published - Jun 2007

Keywords

Acetylcholine
Animals
Arteries
Biological Factors
Blotting, Western
Caveolae
Caveolins
Coronary Vessels
Dose-Response Relationship, Drug
Endothelium, Vascular
In Vitro Techniques
Indoles
Male
Membrane Potentials
Mesenteric Arteries
Oximes
Potassium Channels, Calcium-Activated
Pyrazoles
Rats
Rats, Wistar
Swine
Vasodilator Agents
beta-Cyclodextrins

Access to Document

10.1038/sj.bjp.0707222

Cite this

@article{dbb967fb0f13434a9f27a3e7e736bf6f,

title = "Effects of methyl beta-cyclodextrin on EDHF responses in pig and rat arteries; association between SK(Ca) channels and caveolin-rich domains",

abstract = "BACKGROUND AND PURPOSE: The small and intermediate conductance, Ca2+-sensitive K+ channels (SK(Ca) and IK(Ca), respectively) which are pivotal in the EDHF pathway may be differentially activated. The importance of caveolae in the functioning of IK(Ca) and SK(Ca) channels was investigated.EXPERIMENTAL APPROACH: The effect of the caveolae-disrupting agent methyl-beta-cyclodextrin (MbetaCD) on IK(Ca) and SK(Ca) localization and function was determined.KEY RESULTS: EDHF-mediated, SK(Ca)-dependent myocyte hyperpolarizations evoked by acetylcholine in rat mesenteric arteries (following blockade of IK(Ca) with TRAM-34) were inhibited by MbetaCD. Hyperpolarizations evoked by direct SK(Ca) channel activation (using NS309 in the presence of TRAM-34) were also inhibited by MbetaCD, an effect reversed by cholesterol. In contrast, IK(Ca)-dependent hyperpolarizations (in the presence of apamin) were unaffected by MbetaCD. Similarly, in porcine coronary arteries, EDHF-mediated, SK(Ca)-dependent (but not IK(Ca)-dependent) endothelial cell hyperpolarizations evoked by substance P were inhibited by MbetaCD. In mesenteric artery homogenates subjected to sucrose-density centrifugation, caveolin-1 and SK3 (SK(Ca)) proteins but not IK1 (IK(Ca)) protein migrated to the buoyant, caveolin-rich fraction. MbetaCD pretreatment redistributed caveolin-1 and SK3 proteins into more dense fractions. In immunofluorescence images of porcine coronary artery endothelium, SK3 (but not IK1) and caveolin-1 were co-localized. Furthermore, caveolin-1 immunoprecipitates prepared from native porcine coronary artery endothelium contained SK3 but not IK1 protein.CONCLUSIONS AND IMPLICATIONS: These data provide strong evidence that endothelial cell SK(Ca) channels are located in caveolae while the IK(Ca) channels reside in a different membrane compartment. These studies reveal cellular organisation as a further complexity in the EDHF pathway signalling cascade.",

keywords = "Acetylcholine, Animals, Arteries, Biological Factors, Blotting, Western, Caveolae, Caveolins, Coronary Vessels, Dose-Response Relationship, Drug, Endothelium, Vascular, In Vitro Techniques, Indoles, Male, Membrane Potentials, Mesenteric Arteries, Oximes, Potassium Channels, Calcium-Activated, Pyrazoles, Rats, Rats, Wistar, Swine, Vasodilator Agents, beta-Cyclodextrins",

author = "M Absi and Burnham, {M P} and Weston, {A H} and E Harno and M Rogers and G Edwards",

year = "2007",

month = jun,

doi = "10.1038/sj.bjp.0707222",

language = "English",

volume = "151",

pages = "332--340",

journal = "British Journal of Pharmacology",

issn = "1476-5381",

publisher = "John Wiley & Sons Ltd",

number = "3",

}

TY - JOUR

T1 - Effects of methyl beta-cyclodextrin on EDHF responses in pig and rat arteries; association between SK(Ca) channels and caveolin-rich domains

AU - Absi, M

AU - Burnham, M P

AU - Weston, A H

AU - Harno, E

AU - Rogers, M

AU - Edwards, G

PY - 2007/6

Y1 - 2007/6

N2 - BACKGROUND AND PURPOSE: The small and intermediate conductance, Ca2+-sensitive K+ channels (SK(Ca) and IK(Ca), respectively) which are pivotal in the EDHF pathway may be differentially activated. The importance of caveolae in the functioning of IK(Ca) and SK(Ca) channels was investigated.EXPERIMENTAL APPROACH: The effect of the caveolae-disrupting agent methyl-beta-cyclodextrin (MbetaCD) on IK(Ca) and SK(Ca) localization and function was determined.KEY RESULTS: EDHF-mediated, SK(Ca)-dependent myocyte hyperpolarizations evoked by acetylcholine in rat mesenteric arteries (following blockade of IK(Ca) with TRAM-34) were inhibited by MbetaCD. Hyperpolarizations evoked by direct SK(Ca) channel activation (using NS309 in the presence of TRAM-34) were also inhibited by MbetaCD, an effect reversed by cholesterol. In contrast, IK(Ca)-dependent hyperpolarizations (in the presence of apamin) were unaffected by MbetaCD. Similarly, in porcine coronary arteries, EDHF-mediated, SK(Ca)-dependent (but not IK(Ca)-dependent) endothelial cell hyperpolarizations evoked by substance P were inhibited by MbetaCD. In mesenteric artery homogenates subjected to sucrose-density centrifugation, caveolin-1 and SK3 (SK(Ca)) proteins but not IK1 (IK(Ca)) protein migrated to the buoyant, caveolin-rich fraction. MbetaCD pretreatment redistributed caveolin-1 and SK3 proteins into more dense fractions. In immunofluorescence images of porcine coronary artery endothelium, SK3 (but not IK1) and caveolin-1 were co-localized. Furthermore, caveolin-1 immunoprecipitates prepared from native porcine coronary artery endothelium contained SK3 but not IK1 protein.CONCLUSIONS AND IMPLICATIONS: These data provide strong evidence that endothelial cell SK(Ca) channels are located in caveolae while the IK(Ca) channels reside in a different membrane compartment. These studies reveal cellular organisation as a further complexity in the EDHF pathway signalling cascade.

AB - BACKGROUND AND PURPOSE: The small and intermediate conductance, Ca2+-sensitive K+ channels (SK(Ca) and IK(Ca), respectively) which are pivotal in the EDHF pathway may be differentially activated. The importance of caveolae in the functioning of IK(Ca) and SK(Ca) channels was investigated.EXPERIMENTAL APPROACH: The effect of the caveolae-disrupting agent methyl-beta-cyclodextrin (MbetaCD) on IK(Ca) and SK(Ca) localization and function was determined.KEY RESULTS: EDHF-mediated, SK(Ca)-dependent myocyte hyperpolarizations evoked by acetylcholine in rat mesenteric arteries (following blockade of IK(Ca) with TRAM-34) were inhibited by MbetaCD. Hyperpolarizations evoked by direct SK(Ca) channel activation (using NS309 in the presence of TRAM-34) were also inhibited by MbetaCD, an effect reversed by cholesterol. In contrast, IK(Ca)-dependent hyperpolarizations (in the presence of apamin) were unaffected by MbetaCD. Similarly, in porcine coronary arteries, EDHF-mediated, SK(Ca)-dependent (but not IK(Ca)-dependent) endothelial cell hyperpolarizations evoked by substance P were inhibited by MbetaCD. In mesenteric artery homogenates subjected to sucrose-density centrifugation, caveolin-1 and SK3 (SK(Ca)) proteins but not IK1 (IK(Ca)) protein migrated to the buoyant, caveolin-rich fraction. MbetaCD pretreatment redistributed caveolin-1 and SK3 proteins into more dense fractions. In immunofluorescence images of porcine coronary artery endothelium, SK3 (but not IK1) and caveolin-1 were co-localized. Furthermore, caveolin-1 immunoprecipitates prepared from native porcine coronary artery endothelium contained SK3 but not IK1 protein.CONCLUSIONS AND IMPLICATIONS: These data provide strong evidence that endothelial cell SK(Ca) channels are located in caveolae while the IK(Ca) channels reside in a different membrane compartment. These studies reveal cellular organisation as a further complexity in the EDHF pathway signalling cascade.

KW - Acetylcholine

KW - Animals

KW - Arteries

KW - Biological Factors

KW - Blotting, Western

KW - Caveolae

KW - Caveolins

KW - Coronary Vessels

KW - Dose-Response Relationship, Drug

KW - Endothelium, Vascular

KW - In Vitro Techniques

KW - Indoles

KW - Male

KW - Membrane Potentials

KW - Mesenteric Arteries

KW - Oximes

KW - Potassium Channels, Calcium-Activated

KW - Pyrazoles

KW - Rats

KW - Rats, Wistar

KW - Swine

KW - Vasodilator Agents

KW - beta-Cyclodextrins

U2 - 10.1038/sj.bjp.0707222

DO - 10.1038/sj.bjp.0707222

M3 - Article

C2 - 17450174

SN - 1476-5381

VL - 151

SP - 332

EP - 340

JO - British Journal of Pharmacology

JF - British Journal of Pharmacology

IS - 3

ER -

Effects of methyl beta-cyclodextrin on EDHF responses in pig and rat arteries; association between SK(Ca) channels and caveolin-rich domains

Abstract

Keywords

Access to Document

Fingerprint

Cite this