The effect of cholesterol enrichment on vascular smooth muscle cell (VSMC) calcium homeostasis was studied by evaluating calcium uptake, efflux, and intracellular content in cultured VSMC derived from the rat pulmonary artery. Incubation of VSMC with liposomes consisting of free cholesterol (FC) and phospholipid (2:1 molar ratio, 1 mg FC/ml medium) for 24 h resulted in a 69 +/- 19% increase (P less than 0.01; n = 10) in FC which was associated with a 73 +/- 11% increase (P less than 0.005; n = 10) in intracellular calcium content as assessed by isotopic equilibrium with 45Ca2+ and a 65 +/- 11% increase (P less than 0.024; n = 3) as assessed by atomic absorption spectroscopy. Cholesterol enrichment caused a marked increase in the unidirectional calcium uptake rate from 0.026 +/- 0.03 to 0.158 +/- 0.022 nmol calcium/s per mg protein (P less than 0.01; n = 3), but had no effect on calcium efflux. Nifedipine (1 microM) reduced (P less than 0.05; n = 6) the effect of cholesterol enrichment on unidirectional calcium uptake by 78 +/- 16%; and verapamil (10 microM), diltiazem (1 microM), and nifedipine (1 microM) each significantly inhibited the effect of cholesterol enrichment on intracellular calcium accumulation. Exposure of cholesterol-enriched VSMC to cholesterol-poor liposomes for 24 h returned both FC and calcium contents to control levels. Serum- and serotonin-stimulated calcium uptakes were potentiated 3.7- and 1.7-fold, respectively, in cholesterol-enriched VSMC, whereas endothelin, vasopressin, and thrombin-stimulated calcium uptakes were not affected. We conclude that VSMC FC content plays a role in regulating cellular calcium homeostasis, both under basal conditions and in response to selected agonists.
R A Bialecki, T N Tulenko, W S Colucci