Endothelial mitochondria regulate the intracellular Ca2+ response to fluid shear stress

Shear stress is known to stimulate an intracellular free calcium concentration ([Ca2+](i)) response in vascular endothelial cells (ECs). [Ca2+](i) is a key second messenger for signaling that leads to vasodilation and EC survival. Although it is accepted that the shear-induced [Ca2+](i) response is, in part, due to Ca2+ release from the endoplasmic reticulum (ER), the role of mitochondria (second largest Ca2+ store) is unknown. We hypothesized that the mitochondria play a role in regulating [Ca2+](i) in sheared ECs. Cultured ECs, loaded with a Ca2+-sensitive fluorophore, were exposed to physiological levels of shear stress. Shear stress elicited [Ca2+](i) transients in a percentage of cells with a fraction of them displaying oscillations. Peak magnitudes, percentage of oscillating ECs, and oscillation frequencies depended on the shear level. [Ca2+](i) transients/oscillations were present when experiments were conducted in Ca2+-free solution (plus lanthanum) but absent when ECs were treated with a phospholipase C inhibitor, suggesting that the ER inositol 1,4,5-trisphosphate receptor is responsible for the [Ca2+](i) response. Either a mitochondrial uncoupler or an electron transport chain inhibitor, but not a mitochondrial ATP synthase inhibitor, prevented the occurrence of transients and especially inhibited the oscillations. Knockdown of the mitochondrial Ca2+ uniporter also inhibited the shear-induced [Ca2+](i) transients/oscillations compared with controls. Hence, EC mitochondria, through Ca2+ uptake/release, regulate the temporal profile of shear-induced ER Ca2+ release. [Ca2+](i) oscillation frequencies detected were within the range for activation of mechanoresponsive kinases and transcription factors, suggesting that dysfunctional EC mitochondria may contribute to cardiovascular disease by deregulating the shear-induced [Ca2+](i) response.