Touyz R M, Larivière R, Schiffrin E L
Clinical Research Institute of Montréal, University of Montréal, QC, Canada.
Can J Physiol Pharmacol. 1995 Sep;73(9):1262-73. doi: 10.1139/y95-178.
Calcium responses to the endothelin B receptor (ETB) selective agonist IRL-1620 (IRL) and to endothelin 1 (ET-1) in the absence and presence of the endothelin A receptor (ETA) antagonist BQ-123 were determined in primary cultured unpassaged vascular smooth muscle cells derived from mesenteric resistance vessels of 3-, 9-, and 17-week-old spontaneously hypertensive rats (SHR), Wistar rats, and Wistar-Kyoto (WKY) rats. Intracellular free calcium concentration ([Ca2+]i) was measured microphotometrically and by digital imaging using fura 2 methodology. ET receptor affinity and density were determined in membrane preparations from the mesenteric vascular bed from these rats. Binding studies were performed using [125I]ET-1 in the presence of increasing concentrations of ET-1, BQ-123, or IRL. Basal [Ca2+]i was significantly greater (p < 0.01) in adult SHR compared with age-matched normotensive groups. IRL (10(-7) mol/L) and ET-1 (10(-9) mol/L) increased [Ca2+]i in cells from all strains of all age groups. Responses were significantly lower (p < 0.05) in vascular smooth muscle cells from 9- and 17-week SHR compared with cells from age-matched Wistar and WKY rats. Concentration-response curves for IRL were blunted in cells from 17-week SHR versus cells from WKY rats. The selective ETA receptor antagonist BQ-123 (10(-7)-10(-5) mol/L) had no effect on IRL-induced [Ca2+]i but significantly reduced ET-1-induced [Ca2+]i by 135 +/- 4, 80 +/- 6, and 91 +/- 4 nmol/L in cells from 17-week SHR, WKY, and Wistar groups, respectively. The selective ETB receptor antagonist BQ-788 (10(-6) mol/L) significantly reduced ET-1-induced [Ca2+]i responses in 17-week rats. The Bmax for ET-1 binding was significantly lower in mesenteric artery membrane preparations from 17-week SHR (627 +/- 163 fmol/mg protein) compared with WKY and Wistar rats (1190 +/- 43 and 1059 +/- 55 fmol/mg protein, respectively). These data demonstrate the presence of both ETA and ETB receptors in rat mesenteric vascular smooth muscle cells. In adult SHR, reduced responses of vascular smooth muscle cells to IRL, which represents ETB-mediated effects, and to ET-1, may be partly due, as demonstrated in the binding studies, to lower ET receptor density than in normotensive rats.
在无和有内皮素A受体（ETA）拮抗剂BQ - 123的情况下，测定了来自3周龄、9周龄和17周龄自发性高血压大鼠（SHR）、Wistar大鼠和Wistar - Kyoto（WKY）大鼠肠系膜阻力血管的原代培养未传代血管平滑肌细胞对内皮素B受体（ETB）选择性激动剂IRL - 1620（IRL）和内皮素1（ET - 1）的钙反应。使用fura 2方法通过显微光度法和数字成像测量细胞内游离钙浓度（[Ca2 + ]i）。在这些大鼠肠系膜血管床的膜制剂中测定ET受体亲和力和密度。在存在递增浓度的ET - 1、BQ - 123或IRL的情况下，使用[125I]ET - 1进行结合研究。与年龄匹配的正常血压组相比，成年SHR的基础[Ca2 + ]i显著更高（p < 0.01）。IRL（10（ - 7）mol/L）和ET - 1（10（ - 9）mol/L）使所有年龄组所有品系细胞中的[Ca2 + ]i升高。与年龄匹配的Wistar和WKY大鼠的细胞相比，9周龄和17周龄SHR的血管平滑肌细胞中的反应显著更低（p < 0.05）。与WKY大鼠的细胞相比，17周龄SHR的细胞中IRL的浓度 - 反应曲线变钝。选择性ETA受体拮抗剂BQ - 123（10（ - 7） - 10（ - 5）mol/L）对IRL诱导的[Ca2 + ]i无影响，但在17周龄SHR、WKY和Wistar组的细胞中，分别使ET - 1诱导的[Ca2 + ]i显著降低135±4、80±6和91±4 nmol/L。选择性ETB受体拮抗剂BQ - 788（10（ - 6）mol/L）显著降低17周龄大鼠中ET - 1诱导的[Ca2 + ]i反应。与WKY和Wistar大鼠（分别为1190±43和1059±55 fmol/mg蛋白）相比，17周龄SHR的肠系膜动脉膜制剂中ET - 1结合的Bmax显著更低（627±163 fmol/mg蛋白）。这些数据表明大鼠肠系膜血管平滑肌细胞中同时存在ETA和ETB受体。如结合研究所示，在成年SHR中，血管平滑肌细胞对代表ETB介导作用的IRL和对ET - 1的反应降低，可能部分归因于ET受体密度低于正常血压大鼠。
