title: Membrane cholesterol depletion with beta-cyclodextrin impairs pressure-induced contraction and calcium signalling in isolated skeletal muscle arterioles.
abstract: OBJECTIVE
  Given evidence for clustering of signalling molecules and ion channels in cholesterol-rich membrane domains, the involvement of such structures in arteriolar smooth muscle mechanotransduction was examined.
  
  
  METHODS
  To determine the contribution of smooth muscle cholesterol-rich membrane domains to the myogenic response, isolated arterioles were exposed to the cholesterol-depleting agent beta-cyclodextrin (1-10 mM) in the absence and presence of excess exogenous cholesterol.
  
  
  RESULTS
  beta-Cyclodextrin significantly impaired pressure-induced vasoconstriction, while excess cholesterol attenuated this effect. Impaired myogenic constriction was evident in de-endothelialized vessels, indicating an action at the level of smooth muscle. beta-Cyclodextrin treatment uncoupled increases in intracellular Ca(2+) from myogenic constriction and depleted intracellular Ca(2+) stores consistent with a loss of connectivity between plasma membrane and sarcoplasmic reticulum signalling. However, beta-cyclodextrin-treated arterioles showed unaltered constrictor responses to KCl and phenylephrine. Electron microscopy verified that beta-cyclodextrin caused a decrease in caveolae, while confirmation of smooth muscle containing caveolae was obtained by immunostaining for caveolin-1. Viability of beta-cyclodextrin-treated arterioles was confirmed by agonist sensitivity and propidium iodide nuclear staining.
  
  
  CONCLUSIONS
  The data suggest that smooth muscle cholesterol-rich membrane domains contribute to the myogenic response. Further studies are required to determine whether this relates to specific mechanosensory events or generalized alterations in membrane function.
authors.affiliation: Microvascular Biology Group, School of Medical Sciences, RMIT University, Bundoora, Australia. Potocnik@rmit.edu.au
authors.forename: Simon J
authors.initials: SJ
authors.lastname: Potocnik
authors.forename: Nicole
authors.initials: N
authors.lastname: Jenkins
authors.forename: Timothy V
authors.initials: TV
authors.lastname: Murphy
authors.forename: Michael A
authors.initials: MA
authors.lastname: Hill
mesh_terms: D000818:Animals; D001160:Arterioles / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002273:Carcinogens / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D002784:Cholesterol / Q000378:metabolism*; D066298:In Vitro Techniques; D008297:Male; D008854:Microscopy, Electron; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D018482:Muscle, Skeletal / Q000098:blood supply; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D011312:Pressure; D051381:Rats; D017207:Rats, Sprague-Dawley; D014661:Vasoconstriction / Q000187:drug effects / Q000502:physiology; D047392:beta-Cyclodextrins / Q000494:pharmacology
chemical_list: D002273:Carcinogens; D047392:beta-Cyclodextrins; D002784:Cholesterol; C031215:betadex; D002118:Calcium