Cholesterol regulates both store‐ and agonist‐induced Ca2+ entry in pulmonary arterial endothelium (1089.9)

Abstract

Endothelial Ca2+ entry is decreased in pulmonary arteries from rats exposed to chronic hypoxia (CH) (4 wk, PB = 380 torr). In addition, membrane cholesterol (Chol) is diminished in these cells and Chol supplementation restores ATP‐induced Ca2+ entry. Since both agonist‐induced and store‐operated Ca2+ entry (SOCE) are decreased after CH, we hypothesized that both require functional membrane Chol. To test this hypothesis, we administered epicholesterol (EpiCh; epimeric form of cholesterol) or vehicle to pulmonary arterial endothelial cells (PAEC) from control and CH rats to replace native Chol. The efficacy of EpiCh to diminish membrane Chol was confirmed by filipin staining in cultured ECs. EpiCh treatment greatly reduced both cyclopiazonic acid (CPA) ‐induced SOCE and ATP‐induced Ca2+ entry in PAECs from control and CH rats. We next assessed the effect of EpiCh on ATP and CPA‐induced Ca2+ currents in PAEC from control rats using patch clamp. Both EpiCh and La3+ inhibited ATP‐induced current similarly. Although T‐type voltage gated Ca2+ channels contribute to ATP responses, mibefradil (T‐type inhibitor) had no further effect in EpiCh treated cells. Likewise, CPA‐induced current was reduced in cells treated with EpiCh. We conclude that membrane Chol is required for both agonist‐induced and store‐operated Ca2+ entry in PAEC and that reduced Ca2+ seen after CH is possibly due to loss of this key regulator.Grant Funding Source: Supported by NIH grant HL95640