Rat arterial smooth muscle devoid of ryanodine receptor function: effects on cellular Ca<sup>2+</sup> handling

Dreja, Karl; Nordström, Ina; Hellstrand, Per

doi:10.1038/sj.bjp.0703986

Cited by 19 publications

(18 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Agonist-stimulated Ca 2ϩ wave activity, which depends on recurring inositol trisphosphate (InsP 3 )-mediated Ca 2ϩ release from the SR, 16,20 was selectively affected by cholesterol extraction, with a larger decrease for 5-HT than for cirazoline stimulation, consistent with fura 2 measurements of global [Ca 2ϩ ] i . This suggests that InsP 3 formation by 5-HT stimulation may have been compromised by cholesterol depletion, as may also be inferred from a comparison of agonist-induced Ca 2ϩ release responses in Ca 2ϩ -free medium.…”

Section: Discussionsupporting

confidence: 56%

“…Proteins were extracted and separated on 12% SDS-PAGE gels, and Western blotting on nitrocellulose membranes was then performed essentially as described, 16 with the use of antibodies to ␣ 1A -adrenergic receptor (1:50), caveolin-1 (6B6, 1 g/mL), 5-hydroxytryptamine (5-HT) 2A receptor (1:500), or Cav-p60 (1 g/mL). Peroxidase-conjugated secondary antibodies were used, and peroxidase activity was detected by chemiluminescence (Pierce).…”

Section: Western Blottingmentioning

confidence: 99%

See 1 more Smart Citation

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

Dreja

Voldstedlund

Vinten

et al. 2002

ATVB

Self Cite

165

158

View full text Add to dashboard Cite

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...

show abstract

Section: Discussionsupporting

confidence: 56%

Section: Western Blottingmentioning

confidence: 99%

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

Dreja

Voldstedlund

Vinten

et al. 2002

ATVB

Self Cite

165

158

View full text Add to dashboard Cite

show abstract

“…In the rat tail artery, normal wave activity was found in preparations where the ryanodine receptors had been functionally inactivated, 19 and thus InsP 3 receptor activity seems to be the major cause of waves in this vessel. A crucial factor in the generation of waves is the Ca 2ϩ sensitivity of the InsP 3 receptor, 24 implying a positive feedback control to increase the rate of Ca 2ϩ release.…”

Section: Discussionmentioning

confidence: 99%

“…They were subsequently mounted in a Zeiss LSM 510 laser scanning confocal microscope as described. 19 Fluo-4 was excited at 488 nm, and emission recorded at Ͼ505 nm with a ϫ63 oil immersion objective (NAϭ1.25). Pinhole settings were adjusted to give confocal planes of 1 m. Image series were recorded at 0.3 to 0.7 s/image.…”

Section: Confocal Imaging Of Local [Ca 2؉ ] Imentioning

confidence: 99%

Influence of Mitochondrial Inhibition on Global and Local [Ca ²⁺ ] _i in Rat Tail Artery

Swärd

Dreja

Lindqvist

et al. 2002

Circulation Research

Self Cite

View full text Add to dashboard Cite

Abstract-Inhibition of oxidative metabolism is often found to decrease contractility of systemic vascular smooth muscle, but not to reduce global [Ca 2ϩ ] i . In the present study, we probe the hypothesis that it is associated with an altered pattern of intracellular Ca 2ϩ oscillations (waves) influencing force development. In the rat tail artery, mitochondrial inhibitors (rotenone, antimycin A, and cyanide) reduced ␣ 1 -adrenoceptor-stimulated force by 50% to 80%, but did not reduce global [Ca 2ϩ ] i . Less relaxation (about 30%) was observed after inhibition of myosin phosphatase activity with calyculin A, suggesting that part of the metabolic sensitivity involves the regulation of myosin 20-kDa light chain phosphorylation, although no decrease in phosphorylation was found in freeze-clamped tissue. Confocal imaging revealed that the mitochondrial inhibitors increased the frequency but reduced the amplitude of asynchronous cellular Ca 2ϩ waves elicited by ␣ 1 stimulation. The altered wave pattern, in association with increased basal [Ca 2ϩ ] i , accounted for the unchanged global [Ca 2ϩ ] i . Inhibition of glycolytic ATP production by arsenate caused similar effects on Ca 2ϩ waves and global [Ca 2ϩ ] i , developing gradually in parallel with decreased contractility. Inhibition of wave activity by the InsP 3 receptor antagonist 2-APB correlated closely with relaxation. Furthermore, abolition of waves with thapsigargin in the presence of verapamil reduced force by about 50%, despite unaltered global [Ca 2ϩ ] i , suggesting that contraction may at least partly depend on Ca 2ϩ wave activity. This study therefore indicates that mitochondrial inhibition influences Ca 2ϩ wave activity, possibly due to a close spatial relationship of mitochondria and the sarcoplasmic reticulum and that this contributes to metabolic vascular relaxation. Key Words: arterial smooth muscle Ⅲ metabolic inhibition Ⅲ myosin phosphorylation Ⅲ calcium waves Ⅲ confocal imaging T he distribution of blood flow in tissue depends on the sensitivity of arterial tone to the local oxygen tension, but the nature of the mediating signal is yet unclear. Proposed mechanisms include reduced Ca 2ϩ inflow across the cell membrane, 1-4 decreased myosin phosphorylation, 5 or inhibition of cross-bridge cycling. 6 In some cases of vascular hypoxia or mitochondrial inhibition, the intracellular free Ca 2ϩ concentration ([Ca 2ϩ ] i ), measured as an average from multiple cells, is unchanged or even increased at high stimulus levels while force is reduced. [7][8][9][10] Thus mechanisms based on decreased global [Ca 2ϩ ] i are insufficient to explain hypoxic relaxation. It is puzzling also that in many studies myosin light chain phosphorylation does not decrease, 6,9 even though exceptions are seen. 5 Finally, insufficient MgATP for cross-bridge interaction, although clearly a possibility with severe metabolic inhibition, does not seem to be a general explanation for hypoxic relaxation. 10 Within the vessel wall, individual cells may exhibit asynchron...

show abstract

“…The elevated [Ca 2ϩ ] that accompanies MT could possibly have many effects, including activation of PLC (leading to production of InsP 3 ), activation of conventional PKCs (e.g., PKC␣), and activation and inactivation of InsP 3 Rs (depending on subtype). InsP 3 Rs are particularly important in propagating asynchronous Ca 2ϩ waves in smooth muscle, and therefore, any action of elevated Ca 2ϩ to inactivate them might be particularly important in agonist-induced Ca 2ϩ signaling. To determine whether the differences in ␣ 1 -adrenoceptorinduced Ca 2ϩ signaling in arteries with and without MT might be related to the elevation of cytoplasmic [Ca 2ϩ ] in the presence of MT, we examined ␣ 1 -adrenoceptor-induced Ca (Fig.…”

Section: Resultsmentioning

confidence: 99%

Ca²⁺ signaling in mouse mesenteric small arteries: myogenic tone and adrenergic vasoconstriction

Zacharia

Zhang²,

Wier³

2007

American Journal of Physiology-Heart and Circulatory Physiology

View full text Add to dashboard Cite

show abstract

Rat arterial smooth muscle devoid of ryanodine receptor function: effects on cellular Ca²⁺ handling

Cited by 19 publications

References 33 publications

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

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

Influence of Mitochondrial Inhibition on Global and Local [Ca ²⁺ ] _i in Rat Tail Artery

Ca²⁺ signaling in mouse mesenteric small arteries: myogenic tone and adrenergic vasoconstriction

Contact Info

Product

Resources

About

Rat arterial smooth muscle devoid of ryanodine receptor function: effects on cellular Ca2+ handling

Cited by 19 publications

References 33 publications

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

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

Influence of Mitochondrial Inhibition on Global and Local [Ca 2+ ] i in Rat Tail Artery

Ca2+ signaling in mouse mesenteric small arteries: myogenic tone and adrenergic vasoconstriction

Contact Info

Product

Resources

About

Rat arterial smooth muscle devoid of ryanodine receptor function: effects on cellular Ca²⁺ handling

Influence of Mitochondrial Inhibition on Global and Local [Ca ²⁺ ] _i in Rat Tail Artery

Ca²⁺ signaling in mouse mesenteric small arteries: myogenic tone and adrenergic vasoconstriction