Abstract
Substance P (SP) is a potent endothelium-dependent vasodilator. This study examined the ionic mechanisms involved in SP action on porcine coronary artery endothelial cells (PCAECs), using a combination of patch-clamp techniques and fura-2 microfluorimetry. Two major hypotheses were tested: (1) SP induces hyperpolarization of PCAECs by activating Ca$sp{2+}$-activated K$sp+$ (K$sb{rm Ca}$) channels; (2) SP-induced Ca$sp{2+}$ influx occurs due to activation of a Ca$sp{2+}$-permeable influx pathway coupled to depletion of intracellular stores. SP (10 nM) induced an elevation in cytosolic Ca$sp{2+}$ ($rmlbrack Casp{2+}rbracksb{i}$) due to Ca$sp{2+}$ release from intracellular stores and influx from extracellular space. Under voltage-clamp using the perforated patch technique, SP stimulated outward whole-cell currents, correlating with changes in $rmlbrack Casp{2+}rbracksb{i}$ and were eliminated by buffering $rmlbrack Casp{2+}rbracksb{i}$ using intracellular BAPTA. The reversal potentials of these currents were strongly dependent on the extracellular K$sp+$ concentration. In 62% of patches tested, inside-out recordings revealed a small-conductance K$sb{rm Ca}$ channel. SP induced a 5-fold increase in Ca$sp{2+}$ membrane conductance. When K$sb{rm Ca}$ channels were blocked, SP activated inward currents that reversed at 2.5$sppm$1 mV, a value not significantly different from the reversal potential for inward current induced in response to store-depletion by 1,4-benzohydroquinone (BHQ, 10 $mu$M) (2$sppm$1 mV). Increasing bath Ca$sp{2+}$ or reducing bath Na$sp+$ induced similar shifts in reversal potentials for SP- and BHQ-induced current. IP$sb3$ (20 $mu$M) applied through the patch pipette activated an inward current with reversal potential not significantly different from the reversal potentials for SP- and BHQ-activated currents. Dialysis of the cells with heparin (5 mg/ml) completely blocked SP-induced inward current but not BHQ-induced current. Thus we conclude that in PCAECs, SP-induced $rmlbrack Casp{2+}rbracksb{i}$ elevation activates a K$sb{rm Ca}$ channel resulting in hyperpolarization which promotes Ca$sp{2+}$ influx. Ca$sp{2+}$ entry occurs through an influx pathway coupled to depletion of intracellular stores.
Sharma, Neeraj Roshanlal (1994). Ionic mechanisms in the action of substance P on coronary artery endotheliel cells. Texas A&M University. Texas A&M University. Libraries. Available electronically from
https : / /hdl .handle .net /1969 .1 /DISSERTATIONS -1554814.