Characterization of Single Isolated Vascular Smooth Muscle Cells from Bovine Coronary Artery

Dijk, Adele; Laird, J. D.

doi:10.1159/000158529

Cited by 12 publications

(10 citation statements)

References 10 publications

(10 reference statements)

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“…Fresh dispersion of smooth muscle cells from the coronary artery Single cells from the bovine left circumflex and the proximal 1/3 of the left anterior descending coronary artery were enzymatically dispersed (Van Dijk & Laird, 1984; Wagner-Mann, Bowman & Sturek, 1991). At a local abbatoir cows were killed by exsanguination according to US Department of Agriculture guidelines.…”

Section: Methodsmentioning

confidence: 99%

Sarcoplasmic reticulum buffering of myoplasmic calcium in bovine coronary artery smooth muscle.

Sturek¹,

Kunda²,

Hu³

1992

The Journal of Physiology

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Section: Methodsmentioning

confidence: 99%

Sarcoplasmic reticulum buffering of myoplasmic calcium in bovine coronary artery smooth muscle.

Sturek¹,

Kunda²,

Hu³

1992

The Journal of Physiology

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“…First, we were unable to relax the vascular smooth muscle cells con stricted by KC1 in spite of repeated washing to remove KC1. This observation, however, is the usual finding reported in the literature [14][15][16][29][30][31] ; but see 11,32], It may be due partly to the fact that in blood vessels the vas cular smooth muscle cells in situ may be under strain by attachment to the elastic connective tissues. This inter pretation, although not proven yet.…”

Section: Functional Considerationsmentioning

confidence: 67%

Morphological and Functional Characterization of Vascular Muscle Cells Enzymatically Dispersed from Dog Mesenteric Arteries

Kwan

Gaspar²,

Berezin³

et al. 1992

J Vasc Res

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The objective of this study was to compare the properties of single smooth muscle cells enzymatically dispersed from the dog mesenteric arteries to the properties of similar cells functioning in tissue strips. The isolated cells remained relaxed in nominally Ca²⁺-free medium for about 1 -2 h after exposure to 1 mM Ca²⁺ and like intact mesenteric artery rings did not contract spontaneously. Enzymatically dispersed cells maintained all the characteristic morphological features observed in strips of muscle prior to isolation except that the amorphous materials covering the smooth muscle cell surfaces (basal lamina) were absent after enzymatic dispersion. Addition of 100 mM KCl to these vascular muscle cells elicited maximal shortening in the presence but not in the absence of extracellular Ca²⁺ and KCl-induced cell shortening was prevented by 10^–7M nifedipine indicating the presence of functional voltage-operated Ca²⁺ channels. However, in contrast to the vascular muscle strips, in which graded contractile responses were observed with increasing KCl concentrations, isolated vascular muscle cells underwent nearly maximal contraction at concentrations as low as 15 mM KCl. Both intact tissue and isolated cell preparations responded similarly to phenylephrine in a concentration-dependent manner and the responses were blocked by prazosin. In contrast to muscle strips, the isolated cells did not shorten in response to phenylephrine in Ca²⁺-free medium. Isolated muscle shortened in the presence of sarcoplasmic reticulum selective Ca²⁺ transport ATPase inhibitors, cyclopiazonic acid or thapsigargin. Ryanodine also caused contraction. We conclude that enzymatically dispersed smooth muscle cells from dog mesenteric arteries are potentially useful for studies of the regulation of smooth muscle contractility, but have significantly increased sensitivity to external K⁺, implying an altered membrane potential or voltage dependence of ion channels. Their impaired ability to contract to phenylephrine in Ca²⁺-free medium implies some alteration in intracellular Ca²⁺ stores of their coupling to cellular activation. These differences will affect how the data obtained from freshly isolated enzymatically dispersed vascular muscle cells may be extrapolated to cell studies in intact tissues.

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“…In contrast, nonmigratory dVSM cells have no need for complete FA turnover. However, it is known that dVSM cells display remarkable cytoskeletal plasticity, with the ability, when unrestrained and activated by an agonist, to shorten to 50% of their initial length (57). Recycling/relocation of a subset of dVSM FA proteins may allow maintenance of plasticity of the connection to the contractile filaments while maintaining a stable FA protein core and a linkage to the matrix in the vessel wall.…”

Section: Discussionmentioning

confidence: 99%

Vasoconstrictor-induced endocytic recycling regulates focal adhesion protein localization and function in vascular smooth muscle

Poythress

Gallant

Vetterkind

et al. 2013

American Journal of Physiology-Cell Physiology

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Turnover of focal adhesions (FAs) is known to be critical for cell migration and adhesion of proliferative vascular smooth muscle (VSM) cells. However, it is often assumed that FAs in nonmigratory, differentiated VSM (dVSM) cells embedded in the wall of healthy blood vessels are stable structures. Recent work has demonstrated agonist-induced actin polymerization and Src-dependent FA phosphorylation in dVSM cells, suggesting that agonist-induced FA remodeling occurs. However, the mechanisms and extent of FA remodeling are largely unknown in dVSM. Here we show, for the first time, that a distinct subpopulation of dVSM FA proteins, but not the entire FA, remodels in response to the ␣-agonist phenylephrine. Vasodilator-stimulated phosphoprotein and zyxin displayed the largest redistributions, while ␤-integrin and FA kinase showed undetectable redistribution. Vinculin, metavinculin, Src, Crk-associated substrate, and paxillin displayed intermediate degrees of redistribution. Redistributions into membrane fractions were especially prominent, suggesting endosomal mechanisms. Deconvolution microscopy, quantitative colocalization analysis, and Duolink proximity ligation assays revealed that phenylephrine increases the association of FA proteins with early endosomal markers Rab5 and early endosomal antigen 1. Endosomal disruption with the small-molecule inhibitor primaquine inhibits agonist-induced redistribution of FA proteins, confirming endosomal recycling. FA recycling was also inhibited by cytochalasin D, latrunculin B, and colchicine, indicating that the redistribution is actin-and microtubule-dependent. Furthermore, inhibition of endosomal recycling causes a significant inhibition of the rate of development of agonist-induced dVSM contractions. Thus these studies are consistent with the concept that FAs in dVSM cells, embedded in the wall of the aorta, remodel during the action of a vasoconstrictor. endosomes; microtubules; zyxin; Src DYNAMIC REMODELING of focal adhesions (FAs) and the associated actin cytoskeleton is known to be critical for cell function in proliferative and migratory cells. FA formation at the leading edge, maturation of FAs, and eventual turnover have been extensively studied (4, 61). However, the degree to which FAs in nonmigratory, fully differentiated cellular phenotypes are dynamic and the functions of those FAs embedded in tissue have yet to be fully clarified.Recent studies have demonstrated that the actin cytoskeleton in nonmigrating, contractile smooth muscle is not a totally static structure (for reviews see Refs. 24,32,60,62). In fully differentiated vascular smooth muscle (dVSM) cells, actin polymerization increases in response to the vasoconstrictor phenylephrine (PE) (33). Furthermore, it has been shown for the aorta that the actin elongation factor vasodilator-stimulated phosphoprotein (VASP) is necessary for ␣-agonist-induced actin polymerization (34). Interestingly, in airway smooth muscle, neural Wiscott-Aldrich syndrome protein activation of the actin-related protein 2/3 (Arp2...

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Characterization of Single Isolated Vascular Smooth Muscle Cells from Bovine Coronary Artery

Cited by 12 publications

References 10 publications

Sarcoplasmic reticulum buffering of myoplasmic calcium in bovine coronary artery smooth muscle.

Sarcoplasmic reticulum buffering of myoplasmic calcium in bovine coronary artery smooth muscle.

Morphological and Functional Characterization of Vascular Muscle Cells Enzymatically Dispersed from Dog Mesenteric Arteries

Vasoconstrictor-induced endocytic recycling regulates focal adhesion protein localization and function in vascular smooth muscle

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