Membrane cholesterol depletion with beta-cyclodextrin impairs pressure-induced contraction and calcium signalling in isolated skeletal muscle arterioles

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. METHOD - 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 m M) 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 Ca2+ from myogenic constriction and depleted intracellular Ca2+ 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. CONCLUSION - 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