Abstract 1925 Pulmonary Vascular Biology Platform, Monday, 5/3

In fetal lambs, pulmonary vasodilation in response to increased oxygen, tension, shear stress and nitric oxide (NO) are blocked after pharmacologic inhibition of calcium-sensitive potassium (KCa) channels. While pulmonary artery endothelial cells (PAEC) play an essential role in modulating perinatal pulmonary vascular tone, little is known about the role of K+ channel activity in the control of cytosolic Ca2+ ([Ca2+]i) and nitric oxide synthase (NOS) activity in fetal PAEC. Since KCa channel activation mediates perinatal pulmonary vasodilation in response to several critical birth-related stimuli, we hypothesized that KCa channel activity determines (i) basal [Ca2+]i; and (ii) NOS activity in PAEC. It follows that pharmacologic inhibition of the KCa channel would result in a decrease of both basal [Ca2+]i and NOS activity in PAEC. To test these hypotheses, PAEC were obtained from late gestation ovine fetuses and maintained in cell culture. Low passaged, sub-confluent monolayers of PAEC were loaded with the Ca2+-sensitive fluorophore, fura-2, to measure apparent [Ca2+]i, and treated with either iberiotoxin, a specific KCa channel antagonist, or glibenclamide (GLI), and ATP-sensitive K+ channel antagonist. Iberiotoxin at 10-10 M caused a 43% decrease in [Ca2+]i from 175±16nM to 102±9nM (p<0.001; n=101 cells). Glibenclamide at 10-5M had no effect on [Ca2+]i (n=37 cells). To test the second hypothesis, NOS activity was determined in intact PAEC by measuring 3H-L-arginine conversion to 3H-L-citrulline in whole cells during a 15 min incubation. Tetraethylammonium, a KCa channel blocker in mM concentrations, at 0.5 mM caused a decrease in NOS activity to 76±9% of control levels (p<0.05, n=4). GLI at 1 uM caused no change in NOS activity (116±23% of control levels, NS, n=4). We conclude that under normoxic conditions, basal [Ca2+]i and NOS activity in fetal PAEC is determined primarily by KCa channel activity. We speculate that physiologic stimuli at birth cause PAEC KCa channel activation, resulting in an elevation of PAEC [Ca2+]i that may provide the signal for the production of endothelial derived NO.