The hypothesis that vascular protection in females and its absence in males reflects gender differences in [Ca(2+)](i) and Ca(2+) mobilization mechanisms of vascular smooth muscle contraction was tested in fura 2-loaded aortic smooth muscle cells isolated from intact and gonadectomized male and female Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) rats. In WKY cells incubated in Hanks' solution (1 mM Ca(2+)), the resting length and [Ca(2+)](i) were significantly different in intact males (64.5 +/- 1.2 microm and 83 +/- 3 nM) than in intact females (76.5 +/- 1.5 microm and 64 +/- 7 nM). In intact male WKY, phenylephrine (Phe, 10(-5) M) caused transient increase in [Ca(2+)](i) to 428 +/- 13 nM followed by maintained increase to 201 +/- 8 nM and 32% cell contraction. In intact female WKY, the Phe-induced [Ca(2+)](i) transient was not significantly different, but the maintained [Ca(2+)](i) (159 +/- 7 nM) and cell contraction (26%) were significantly less than in intact male WKY. In Ca(2+)-free (2 mM EGTA) Hanks', Phe and caffeine (10 mM) caused transient increases in [Ca(2+)](i) and contraction that were not significantly different between males and females. Membrane depolarization by 51 mM KCl caused 31% cell contraction and increased [Ca(2+)](i) to 259 +/- 9 nM in intact male WKY, which were significantly greater than a 24% contraction and 214 +/- 8 nM [Ca(2+)](i) in intact female WKY. Maintained Phe- and KCl-stimulated cell contraction and [Ca(2+)](i) were significantly greater in SHR than WKY in all groups of rats. Reduction in cell contraction and [Ca(2+)](i) in intact females compared with intact males was significantly greater in SHR ( approximately 30%) than WKY ( approximately 20%). No significant differences in cell contraction or [Ca(2+)](i) were observed between castrated males, ovariectomized (OVX) females, and intact males, or between OVX females with 17beta-estradiol implants and intact females. Exogenous application of 17beta-estradiol (10(-8) M) to cells from OVX females caused greater reduction in Phe- and KCl-induced contraction and [Ca(2+)](i) in SHR than WKY. Thus the basal, maintained Phe- and depolarization-induced [Ca(2+)](i) and contraction of vascular smooth muscle triggered by Ca(2+) entry from the extracellular space exhibit differences depending on gender and the presence or absence of female gonads. Cell contraction and [Ca(2+)](i) due to Ca(2+) release from the intracellular stores are not affected by gender or gonadectomy. Gender-specific reduction in contractility and [Ca(2+)](i) in vascular smooth muscle of female rats is greater in SHR than WKY rats.

Abstract-We investigated whether putative vascular protection against hypertension in females reflects differences in the Ca 2ϩ mobilization mechanisms of vasoconstriction depending on the gender and the status of the gonads. Active stress and 45 Ca 2ϩ influx were measured in aortic strips isolated from intact and gonadectomized male and female Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR

Reduction of uterine perfusion pressure (RUPP) during late pregnancy has been suggested to trigger increases in renal vascular resistance and lead to hypertension of pregnancy. We investigated whether the increased renal vascular resistance associated with RUPP in late pregnancy reflects increases in intracellular Ca(2+) concentration ([Ca(2+)](i)) and contraction of renal arterial smooth muscle. Single smooth muscle cells were isolated from renal interlobular arteries of normal pregnant Sprague-Dawley rats and a rat model of RUPP during late pregnancy. The cells were loaded with fura 2 and both cell length and [Ca(2+)](i) were measured. In cells of normal pregnant rats incubated in Hanks' solution (1 mM Ca(2+)), ANG II (10(-7) M) caused an initial increase in [Ca(2+)](i) to 414 +/- 13 nM, a maintained increase to 149 +/- 8 nM, and 21 +/- 1% cell contraction. In RUPP rats, the initial ANG II-induced [Ca(2+)](i) (431 +/- 18 nM) was not different from pregnant rats, but both the maintained [Ca(2+)](i) (225 +/- 9 nM) and cell contraction (48 +/- 2%) were increased. Membrane depolarization by 51 mM KCl and the Ca(2+) channel agonist BAY K 8644 (10(-6) M), which stimulate Ca(2+) entry from the extracellular space, caused maintained increases in [Ca(2+)](i) and cell contraction that were greater in RUPP rats than control pregnant rats. In Ca(2+)-free (2 mM EGTA) Hanks' solution, the ANG II- and caffeine (10 mM)-induced [Ca(2+)](i) transient and cell contraction were not different between normal pregnant and RUPP rats, suggesting no difference in Ca(2+) release from the intracellular stores. The enhanced maintained ANG II-, KCl- and BAY K 8644-induced [Ca(2+)](i) and cell contraction in RUPP rats compared with normal pregnant rats suggest enhanced Ca(2+) entry mechanisms of smooth muscle contraction in resistance renal arteries and may explain the increased renal vascular resistance associated with hypertension of pregnancy.

Vascular resistance and arterial pressure are reduced during normal pregnancy, but dangerously elevated during pregnancy-induced hypertension (PIH), and changes in nitric oxide (NO) synthesis have been hypothesized as one potential cause. In support of this hypothesis, chronic inhibition of NO synthesis in pregnant rats has been shown to cause significant increases in renal vascular resistance and hypertension; however, the cellular mechanisms involved are unclear. We tested the hypothesis that the pregnancy-associated changes in renal vascular resistance reflect changes in contractility and intracellular Ca(2+) concentration ([Ca(2+)](i)) of renal arterial smooth muscle. Smooth muscle cells were isolated from renal interlobular arteries of virgin and pregnant Sprague-Dawley rats untreated or treated with the NO synthase inhibitor nitro-L-arginine methyl ester (L-NAME; 4 mg. kg(-1). day(-1) for 5 days), then loaded with fura 2. In cells of virgin rats incubated in Hanks' solution (1 mM Ca(2+)), the basal [Ca(2+)](i) was 86 +/- 6 nM. Phenylephrine (Phe, 10(-5) M) caused a transient increase in [Ca(2+)](i) to 417 +/- 11 nM and maintained an increase to 183 +/- 8 nM and 32 +/- 3% cell contraction. Membrane depolarization by 51 mM KCl, which stimulates Ca(2+) entry from the extracellular space, caused maintained increase in [Ca(2+)](i) to 292 +/- 12 nM and 31 +/- 2% contraction. The maintained Phe- and KCl-induced [Ca(2+)](i) and contractions were reduced in pregnant rats but significantly enhanced in pregnant rats treated with L-NAME. Phe- and KCl-induced contraction and [Ca(2+)](i) were not significantly different between untreated and L-NAME-treated virgin rats or between untreated and L-NAME + L-arginine treated pregnant rats. In Ca(2+)-free Hanks', application of Phe or caffeine (10 mM), to stimulate Ca(2+) release from the intracellular stores, caused a transient increase in [Ca(2+)](i) and a small cell contraction that were not significantly different among the different groups. Thus renal interlobular smooth muscle of normal pregnant rats exhibits reduction in [Ca(2+)](i) signaling that involves Ca(2+) entry from the extracellular space but not Ca(2+) release from the intracellular stores. The reduced renal smooth muscle cell contraction and [Ca(2+)](i) in pregnant rats may explain the decreased renal vascular resistance associated with normal pregnancy, whereas the enhanced cell contraction and [Ca(2+)](i) during inhibition of NO synthesis in pregnant rats may, in part, explain the increased renal vascular resistance associated with PIH.