Chronic hypoxia (CH)-induced pulmonary hypertension is associated with diminished production of endothelium-derived Ca2+-dependent vasodilators such as nitric oxide. Interestingly, ATP-induced endothelial Ca2+ entry as well as membrane cholesterol (Chol) are decreased in pulmonary arteries from CH rats (4 wk, PB = 380 Torr) compared to normoxic controls. Store-operated Ca2+ entry (SOCE) and depolarization-induced Ca2+ entry are major components of the response to ATP and are similarly decreased after CH. We hypothesized that membrane Chol facilitates both SOCE and depolarization-induced pulmonary endothelial Ca2+ entry, and that CH attenuates these responses by decreasing membrane Chol. To test these hypotheses, we administered Chol or epicholesterol (Epichol) to acutely isolated pulmonary arterial endothelial cells (PAEC) from control and CH rats to either supplement or replace native Chol, respectively. The efficacy of membrane Chol manipulation was confirmed by filipin staining. Epichol greatly reduced ATP-induced Ca2+ influx in PAEC from control rats. Whereas Epichol similarly blunted endothelial SOCE in PAEC from both groups, Chol supplementation restored diminished SOCE in PAEC from CH rats while having no effect in controls. Similar effects of Chol manipulation on PAEC Ca2+ influx were observed in response to a depolarizing stimulus of KCl. Furthermore, KCl-induced Ca2+ entry was inhibited by the T-type Ca2+ channel antagonist, mibefradil, but not the L-channel inhibitor, diltiazem. We conclude that PAEC membrane Chol is required for ATP -induced Ca2+ entry and its two components, SOCE and depolarization-induced Ca2+ entry, and that reduced Ca2+ entry after CH may be due to loss of this key regulator.
- pulmonary hypertension
- T-type calcium channels
- store-operated Ca2+ entry
- depolarization-induced Ca2+ entry
- Copyright © 2017, American Journal of Physiology-Heart and Circulatory Physiology