EXPOSURE OF ENDOTHELIAL-CELLS TO CYCLIC STRAIN INDUCES ELEVATIONS OF CYTOSOLIC CA2+ CONCENTRATION THROUGH MOBILIZATION OF INTRACELLULAR ANDEXTRACELLULAR POOLS

We have previously reported that exposure of endothelial cells to cycl ic strain elicited a rapid but transient generation of inositol 1,4,5- trisphosphate (IP3), which reached a peak 10 s after the initiation of cyclic deformation. To address the effect of cyclic strain on intrace llular Ca2+ concentration ([Ca2+](i)) and its temporal relationship to IP3 generation, confluent bovine aortic endothelial cells were grown on flexible membranes, loaded with aequorin and the membranes placed i n a custom-designed flow-through chamber. The chamber was housed insid e photomultiplier tube, and vacuum was utilized to deform the membrane s. Our results indicate that the initiation of 10% average strain indu ced a rapid increase in [Ca2+](i) which contained two distinct compone nts: a large initial peak 12 s after the initiation of stretch which c losely followed the IF, peak, and a subsequent lower but sustained pha se. Pretreatment with 5 mu M GdCl3 for 10 min or nominally Ca2+-free m edium (CFM) for 3 min reduced the magnitude of the initial rise and ab olished the sustained phase. Repetitive 10% average strain at a freque ncy of 60 cycles/min also elicited a single IP3 peak at 10 s. However, there was also a large initial [Ca2+](i) peak followed by multiple sm aller transient [Ca2+](i) elevations. Preincubation with 5 mu M GdCl3 or CFM diminished the initial [Ca2+](i), transient and markedly inhibi ted the late-phase component. Preincubation with 25 mu M 2,5-di-(t-but yl)-1,4-benzohydroquinone (BHQ) attenuated the initial [Ca2+](i) trans ient. Cyclic-strain-mediated IF, formation in confluent endothelial ce lls at 10 s, however, was not modified by pretreatment with 25 mu M BH Q, 500 mu M NiCl2, 10 nM charybdotoxin, 5 mu M GdCl3 or CFM. We conclu de that in endothelial cells exposed to cyclic strain, Ca2+ enters the cytosol from intracellular and extracellular pools but IF, formation is not dependent on Ca2+ entry via the plasma membrane.