Cholesterol depletion disrupts caveolae and differentially impairs agonist-induced arterial contraction.
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-beta-cyclodextrin, and the effects on force and Ca2+ handling were evaluated. In cholesterol-depleted preparations, the force responses to alpha1-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 micro mol/L AlCl3 were unaffected. In contrast, responses to 5-hydroxytryptamine (5-HT), vasopressin, and endothelin were reduced by >50%. The rise in global intracellular free Ca2+ concentration in response to 5-HT was attenuated, as was the generation of Ca2+ 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-HT2A receptor and of caveolin-1 were unaffected by cholesterol extraction. Sucrose gradient centrifugation showed enrichment of 5-HT2A receptors, but not alpha1-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.
Dreja
Karl
K
Department of Physiological Sciences, Lund University, Lund, Sweden.
Voldstedlund
Marianne
M
Vinten
Jørgen
J
Tranum-Jensen
Jørgen
J
Hellstrand
Per
P
Swärd
Karl
K
D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000818:Animals; D001158:Arteries / Q000502:physiology; D021941:Caveolae / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism*; D003505:Cyclodextrins / Q000494:pharmacology; D005260:Female; D007093:Imidazoles / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011985:Receptors, Serotonin / Q000378:metabolism; D013623:Tail / Q000098:blood supply; D014661:Vasoconstriction / Q000187:drug effects / Q000502:physiology*
D000316:Adrenergic alpha-Agonists; D003505:Cyclodextrins; D007093:Imidazoles; D011985:Receptors, Serotonin; D002784:Cholesterol; C014282:cirazoline