Abstract
Endothelial injury is the primary event that leads to a variety of severe vascular disorders. The signal transduction pathway which drives the subsequent healing process is far from being fully elucidated. Mechanical injury elicits a Ca2+ response in the endothelium of intact rat aorta, which comprises an initial Ca2+ release from inositol-1,4,5-trisphosphate-sensitive (IP3Rs) stores followed by a long-lasting decay phase due to Ca2+ entry through uncoupled connexons. In a minor fraction of cells, the Ca2+ signal adopts an oscillatory pattern whose molecular underpinnings are yet to be elucidated. In the light of the role played by repetitive Ca2+ spikes in regulating tissue regeneration, the present study aims at elucidating the mechanisms underlying injury-induced Ca2+ oscillations. The repetitive Ca2+ signal reversibly ceased upon removal of extracellular Ca2+ or addition of the inorganic cations, La3+ and Ni2+. Moreover, the spiking response was abolished by the gap-junction blockers, heptanol and 18 beta-glycyrrhetinic acid and by interfering with the Ca2+ entry-mode mode of the Na+/Ca2+ exchanger (NCX). The InsP3-producing agonist, ATP, resumed Ca2+ oscillations in silent cells, while the phospholipase C inhibitor, U73122, inhibited the oscillatory signal. The latter was also prevented by the SERCA inhibitors, thapsigargin and cyclopiazonic acid acid. These data show that injury-induced [Ca2+]i oscillations require the coordinated interplay between NCX-mediated Ca2+ entry and InsP3-dependent Ca2+ release. Besides directly gating Ca2+ inflow, uncoupled connexons might let Na+ into the cells and stimulate Ca2+ entry through NCX by increasing submembranal Na+ levels.
