Ignition of Calcium Sparks in Arterial and Cardiac Muscle Through Caveolae

Löhn, Matthias; Fürstenau, Michael; Sagach, Victoriya; Elger, Marlies; Schulze, Wolfgang; Luft, Friedrich C.; Haller, Hermann; Gollasch, Maik

doi:10.1161/01.res.87.11.1034

Cited by 156 publications

(138 citation statements)

References 31 publications

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“…Because adrenergic contractions are unaffected, it is unlikely that the contractile mechanism is involved. Furthermore, as also reported by Löhn et al, 31 L-type channel activation by membrane depolarization seems to function normally in cholesterol-depleted preparations, because responses to depolarization by high K ϩ were unaffected. Thus, the impaired 5-HT-induced elevation of [Ca 2ϩ ] i after extraction of cholesterol seems to be due to a defect upstream from Ca 2ϩ channel activation, presumably leading to reduced membrane depolarization after agonist stimulation.…”

Section: Discussionsupporting

confidence: 80%

“…The basal tone, although sensitive to extracellular Ca 2ϩ , was resistant to a variety of Ca 2ϩ channel blockers and, thus, presumably not associated with decreased activity of Ca 2ϩ sparks and spontaneous transient outward currents, as reported for m␤cd-treated cells and after caveolin-1 knockout. 23,31 Our results suggest that the signaling step impaired by cholesterol depletion is located either to ligand-receptor or to receptor-second-messenger interaction. Both steps could poten- tially be influenced by local variations in membrane fluidity 3 imposed by caveolin-cholesterol interactions.…”

Section: Discussionmentioning

confidence: 80%

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Cholesterol Depletion Disrupts Caveolae and Differentially Impairs Agonist-Induced Arterial Contraction

Dreja

Voldstedlund

Vinten

et al. 2002

ATVB

165

158

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Objective-This study assessed the role of cholesterol-rich membrane regions, including caveolae, in the regulation of arterial contractility. Methods and Results-Rat tail artery devoid of endothelium was treated with the cholesterol acceptor methyl-␤-cyclodextrin, and the effects on force and Ca 2ϩ handling were evaluated. In cholesterol-depleted preparations, the force responses to ␣ 1 -adrenergic receptors, membrane depolarization, inhibition of myosin light chain phosphatase, and activation of G proteins with a mixture of 20 mmol/L NaF and 60 mol/L AlCl 3 were unaffected. In contrast, responses to 5-hydroxytryptamine (5-HT), vasopressin, and endothelin were reduced by Ͼ50%. The rise in global intracellular free Ca 2ϩ concentration in response to 5-HT was attenuated, as was the generation of Ca 2ϩ waves at the cellular level. By electron microscopy, cholesterol depletion was found to disrupt caveolae. The 5-HT response could be restored by exogenous cholesterol, which also restored caveolae. Western blots showed that the levels of 5-HT 2A receptor and of caveolin-1 were unaffected by cholesterol extraction. Sucrose gradient centrifugation showed enrichment of 5-HT 2A receptors, but not ␣ 1 -adrenergic receptors, in the caveolin-1-containing fractions, suggesting localization of the former to caveolae. Conclusions-These results show that a subset of signaling pathways that regulate smooth muscle contraction depends specifically on cholesterol. Furthermore, the cholesterol-dependent step in serotonergic signaling occurs early in the pathway and depends on the integrity of caveolae. Key Words: smooth muscle Ⅲ caveolae Ⅲ 5-hydroxytryptamine Ⅲ endothelin Ⅲ intracellular calcium C ellular cholesterol, of which most (up to 90%) 1 resides in the plasma membrane, is crucial for normal membrane permeability and fluidity and also plays a role in cellular signaling, via several proposed mechanisms that fall into at least 4 categories. First, cellular cholesterol may influence gene transcription in the nucleus through sterol regulatory element binding proteins. 2 Second, the activity of membrane receptors, ion channels, and transporters may depend on the membrane fluidity, per se. 3 Third, membrane protein function may be regulated through specific cholesterol-protein interactions. 3,4 Fourth, cholesterol stabilizes the structure of caveolae and lipid rafts.Caveolae, which are 50-to 100-nm membrane invaginations that are abundant in vascular endothelium and smooth muscle cells, are defined by their characteristic morphology and contents of caveolin and cav-p60. 5,6 No definitive definition of rafts has appeared because they do not exhibit a characteristic structure, but the term is used for planar aggregations of specific lipids and proteins. Caveolae and lipid rafts are envisaged to serve as platforms for a dynamic association of signaling proteins and for the initiation or modulation of signaling. 5,7,8 Some agonists causing contraction of vascular smooth muscle act on receptors that are believed to be located in caveo...

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Section: Discussionsupporting

confidence: 80%

Section: Discussionmentioning

confidence: 80%

Cholesterol Depletion Disrupts Caveolae and Differentially Impairs Agonist-Induced Arterial Contraction

Dreja

Voldstedlund

Vinten

et al. 2002

ATVB

165

158

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“…This result was expected for the adenosine A 1 receptor as this receptor is a known resident of caveolae in cardiomyocytes [16]. The presence of DHPRs in caveolae has previously been inferred from the effects of methyl-β-cycoldextrin (a reagent that sequesters cholesterol and disrupts lipid rafts) on Ca 2+ sparks [22]. We now provide direct biochemical data that DHPRs are components of lipid rafts.…”

Section: Discussionsupporting

confidence: 70%

Localization of sarcolemmal proteins to lipid rafts in the myocardium

et al. 2007

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The localization of sarcolemmal proteins within the membrane can have a dramatic effect on excitation-contraction coupling. We examine the localization of the Na + -Ca 2+ exchanger, the dihydropyridine receptor, and other proteins involved in excitation-contraction coupling in rat heart using biochemical and immunolocalization techniques. Specifically, we assess the distribution of proteins within the lipid raft fraction of the sarcolemma. We find that the distribution of proteins in lipid raft fractions is very dependent on the solubilization technique. A common technique using sodium carbonate/pH 11 to solubilize non-lipid raft proteins was inappropriate for use with sarcolemmal membranes. Use of Triton X-100 was more efficacious as a solubilization agent. A large majority of the Na + -Ca 2+ exchanger, Na + /K + -ATPase, and plasma membrane Ca 2+ pump are not present in lipid rafts. In contrast, most adenosine A 1 receptors and dihydropyridine receptors were in lipid raft fractions. Most of the adenosine A 1 receptors could be co-immunoprecipitated with caveolin indicating a localization to caveolae (a subclass of lipid rafts). In contrast, the dihydropyridine receptors could not be co-immunoprecipitated with caveolin. Most biochemical data were confirmed by high resolution immunolocalization studies. Using correlation analysis, only a small fraction of the Na + -Ca 2+ exchangers colocalized with caveolin whereas a substantial fraction of dihydropyridine and adenosine A 1 receptors did colocalize with caveolin. The most pertinent findings are that the Na + -Ca 2+ exchanger and the dihydropyridine receptor are in separate sarcolemmal subcompartments. These spatial relationships may be relevant for understanding excitation-contraction coupling.

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“…For example, caveolae have been implicated as sites of Ca 2ϩ sparks (28) in cardiomyocytes and the location of capacitive Ca 2ϩ entry in other cells (19). If Ca 2ϩ entry is localized in caveolae, expression of AC6 (a Ca 2ϩ -inhibitable isoform) in this same microdomain should result in cAMP levels that are highly sensitive to Ca 2ϩ entry from outside the cell.…”

Section: Discussionmentioning

confidence: 99%

Receptor Number and Caveolar Co-localization Determine Receptor Coupling Efficiency to Adenylyl Cyclase

Ostrom

Gregorian

Drenan

et al. 2001

Journal of Biological Chemistry

232

213

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Recent evidence suggests that many signaling molecules localize in microdomains of the plasma membrane, particularly caveolae. In this study, overexpression of adenylyl cyclase was used as a functional probe of G protein-coupled receptor (GPCR) compartmentation. We found that three endogenous receptors in neonatal rat cardiomyocytes couple with different levels of efficiency to the activation of adenylyl cyclase type 6 (AC6), which localizes to caveolin-rich membrane fractions. Overexpression of AC6 enhanced the maximal cAMP response to ␤ 1 -adrenergic receptor (␤ 1 AR)-selective activation 3.7-fold, to ␤ 2 AR-selective activation only 1.6-fold and to prostaglandin E 2 (PGE 2 ) not at all. Therefore, the rank order of efficacy in coupling to AC6 is ␤ 1 AR > ␤ 2 AR > prostaglandin E 2 receptor (EP 2 R). ␤ 2 AR coupling efficiency was greater when we overexpressed the receptor or blocked its desensitization by expressing ␤ARKct, an inhibitor of G protein-coupled receptor kinase activation, but was not significantly greater when cells were treated with pertussis toxin. Assessment of receptor and AC expression indicated co-localization of AC5/6, ␤ 1 AR, and ␤ 2 AR, but not EP 2 R, in caveolin-rich membranes and caveolin-3 immunoprecipitates, likely explaining the observed activation of AC6 by ␤AR subtypes but lack thereof by PGE 2 . When cardiomyocytes were stimulated with a ␤AR agonist, ␤ 2 AR were no longer found in caveolin-3 immunoprecipitates; an effect that was blocked by expression of ␤ARKct. Thus, agonist-induced translocation of ␤ 2 AR out of caveolae causes a sequestration of receptor from effector and likely contributes to the lower efficacy of ␤ 2 AR coupling to AC6 as compared with ␤ 1 AR, which do not similarly translocate. Therefore, spatial co-localization is a key determinant of efficiency of coupling by particular extracellular signals to activation of GPCR-linked effectors.

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Ignition of Calcium Sparks in Arterial and Cardiac Muscle Through Caveolae

Cited by 156 publications

References 31 publications

Cholesterol Depletion Disrupts Caveolae and Differentially Impairs Agonist-Induced Arterial Contraction

Cholesterol Depletion Disrupts Caveolae and Differentially Impairs Agonist-Induced Arterial Contraction

Localization of sarcolemmal proteins to lipid rafts in the myocardium

Receptor Number and Caveolar Co-localization Determine Receptor Coupling Efficiency to Adenylyl Cyclase

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