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While parathyroid hormone type 1 receptor (PTH1R)-mediated vasodilatory, cardiac stimulatory, and renin-activating effects of exogenous PTH/PTH-related protein (PTHrP) are acknowledged, interactions of endogenous PTHrP with these systems remain unclear, mainly because the unavailability of viable PTHrP/PTH1R knockout mice. Transgenic mice overexpressing PTH1R in smooth muscle strongly have supported the PTHrP/PTH1R system as a cardiovascular system (CVS) regulator, but the consequences on renovascular (RVS) and renin-angiotensin systems (RAS) have not been explored in these studies. The aim was to develop a model in which one could study the consequences on CVS, RVS, and RAS of generalized PTH1R overexpression. Systemic PTH1R cDNA plasmid delivery was used in adult rats, a system that is amenable to studies in isolated perfused kidneys and that minimizes development-induced compensatory mechanisms. Intravenous administration of hPTH1R or green fluorescence protein-tagged hPTH1R in pcDNA3 resulted 3 wk later, in generalized expression of hPTH1R (mRNA and protein), especially in vessels, liver, heart, kidney, and central nervous system, where it is expressed physiologically. As expected, PTH1R overexpression decreased BP and renal tone. Unexpected, however, PTH1R overexpression decreased heart rate. These studies also revealed that endogenous PTHrP actually inhibits renin release and that hPTH1R overexpression tends to increase that effect. Striking, liver production and circulatory level of angiotensinogen and hence plasma renin activity were markedly reduced. Thus, abrupt PTH1R overexpression in adult rats profoundly alters the CVS, RVS, and RAS, strongly supporting the PTH/PTHrP/PTH1R system as crucial for heart and vascular tone regulation. In addition, these results revealed that PTH1R-mediated mechanisms might have protective effects against cardiovascular stress-induced responses, including stimulations in heart rate and RAS.

These studies examine whether PTHrP(1-36), a vasodilator, modulates BP and renal vascular resistance (RVR) in spontaneously hypertensive rat (SHR). Within the kidney of normotensive rats, PTHrP(1-36) was enriched in vessels. In vessels of SHR, PTHrP was upregulated by 40% and type 1 PTH receptor (PTH1R) was downregulated by 65% compared with normotensive rats. To investigate the role of endogenous PTHrP in the regulation of BP and RVR, SHR were subjected to somatic human (h)PTH1R gene delivery. Three weeks after a single intravenous injection of pcDNA1.1 plasmid containing the hPTH1R gene under the control of the cytomegalovirus promoter, hPTH1R mRNA was detected in all of the main organs. Within the kidney, the transgene was enriched in vessels. In the isolated perfused kidney, RVR was reduced by 23% and PTHrP(1-36)-induced vasodilation, which is depressed in SHR, was restored and a vasoconstrictory response to PTH(3-34), a PTH1R antagonist, was revealed. These effects were not observed in control SHR treated with empty plasmid. BP remained unchanged, and plasma renin activity increased by 60%. Thus, in SHR renal vessels, a reduced number of PTH1R contributes to the high RVR, despite the higher expression of vasodilatory PTHrP. Moreover, these studies provide evidence for a direct link between the density of PTH1R and plasma renin activity, which might be responsible for the absence of effect of PTH1R gene delivery on BP in SHR. Overall, PTHrP significantly contributes to the homeostasis of renal and systemic hemodynamics in SHR.

We recently showed that brain mineralocorticoid receptors (MRs) are involved in blood pressure and kidney function control in normotensive Wistar rats. We now assessed the involvement of brain MRs in spontaneously hypertensive rats (SHR), in which the presence of adrenocorticoids has been shown to be required for the development of hypertension. The effect of a single intracerebroventricular (ICV) injection of an MR antagonist (RU28318) on systolic blood pressure (SBP) and renal function was examined in conscious adult SHR and Wistar-Kyoto rats (WKY) maintained on a standard-sodium diet (0.4% Na + ). In WKY, a long-lasting decrease in SBP was caused by the ICV injection of 10 ng RU28318 as previously reported in Wistar rats, associated with increased urinary excretion of water and electrolytes. In SHR maintained on the standard diet, the ICV injection of RU28318 (10 or 100 ng) had no effect on cardiovascular and renal functions. However, the ICV injection of 10 ng RU28318 in SHR after 3 weeks of high sodium intake (8% Na + ) caused a long-lasting decrease in SBP. The effect was present at 8 hours (ΔSBP 34±2 mm Hg), persisted at 24 hours (ΔSBP 29±1 mm Hg), and disappeared at 48 hours after the injection. The hypotension was not associated with changes in heart rate, urinary excretion of water and electrolytes, and plasma renin activity, whereas renal denervation did not affect the decrease in SBP. A more pronounced decrease in SBP (49±3 mm Hg at 8 hours) was observed with 100 ng RU28318. This dose of the antagonist was without effect after subcutaneous administration. Thus, brain MRs appear to participate in the maintenance of hypertension in conscious adult SHR sensitized by sodium loading.

Abstract —Nitric oxide attenuates both vasopressin-induced vasoconstriction and vasopressin release. We tested whether hypertension and renal dysfunction elicited by chronic inhibition of nitric oxide (NO) synthesis using N G -nitro- l -arginine (L-NNA) could be mediated in part by vasopressin V 1A receptors. Male rats were treated orally for 6 weeks with L-NNA (15 mg/kg per day), a nonpeptide V 1A receptor antagonist (2 S )-1-[(2 R ,3 S )-5-chloro-3-(2-chlorophenyl)-1-(3,4-dimethoxybenzene-sulfonyl)-3-hydroxy-2,3-dihydro-1 H -indole-2-carbonyl]-pyrrolidine-2-carboxamide (SR 49059, 30 mg/kg per day), or a combination of SR 49059 and L-NNA (same doses), or they received no treatment. Both drugs were added to the food. Measurements were performed in conscious rats (urine collection in metabolic cages, tail-cuff arterial pressure) and at the end of the study in anesthetized rats (clearance measurements). L-NNA produced sustained hypertension, decreased glomerular filtration rate, and increased renal vascular resistance, plasma renin activity, and urinary albumin excretion. SR 49059 had no effect per se on these parameters and also did not attenuate the hypertension and renal dysfunction induced by L-NNA. Surprisingly, SR 49059 potentiated L-NNA–induced hypertension at the end of the 6-week treatment. However, the blood pressure response and the renal and mesenteric vasoconstriction elicited by exogenous vasopressin were attenuated in rats treated with SR 49059. L-NNA did not change plasma vasopressin concentration or 24-hour urinary vasopressin excretion. Our findings suggest that activation of vasopressin V 1A receptors does not contribute to the hypertension and renal dysfunction induced by chronic NO synthesis inhibition. They also document unchanged plasma vasopressin concentration in NO-deficient hypertension.

Recent data have revealed biological and genetic variability in normotensive Wistar-Kyoto rats, which are considered to be the most appropriate control strain for spontaneously hypertensive rats. To investigate the possibility that angiotensin converting enzyme activity could be affected by this variability, we measured plasma and tissue (lung, heart, renal cortex, renal medulla, and adrenal gland) angiotensin converting enzyme activity in spontaneously hypertensive rats and normotensive Wistar-Kyoto rats from three commercial suppliers in France: Iffa-Credo, Janvier, and Charles River Laboratories. Angiotensin converting enzyme activity was measured in vitro with a fluorometric assay using carbobenzoxy-Phe-His-Leu as substrate. Angiotensin converting enzyme activity in both rat strains varied considerably from one supplier to another, and therefore, comparisons of spontaneously hypertensive rats and Wistar-Kyoto rats from the different suppliers produced conflicting results. For Wistar-Kyoto rats, angiotensin converting enzyme activity in the plasma, heart, kidney, and adrenal glands was highest in rats from Iffa-Credo and lowest in rats from Charles River. For spontaneously hypertensive rats, angiotensin converting enzyme activity in the plasma and tissues was highest in rats from Janvier, whereas no difference could be observed between rats from Iffa-Credo and Charles River. These data confirm the problem of how to interpret and compare studies that use spontaneously hypertensive and Wistar-Kyoto rat strains.

The inhibition of angiotensin converting enzyme by ramipril, ramiprilat, enalapril, enalaprilat, and captopril was studied in the plasma and various tissues (lung, heart, renal cortex, renal medulla) of normotensive rats and spontaneously hypertensive rats. Displacement curves for [3H]ramiprilat were established on each tissue with the converting enzyme inhibitors, and their potencies were expressed as the concentration that inhibited 50% of the specific [3H]ramiprilat binding. In the plasma, lung, and heart, the order of activities was: ramiprilat greater than enalaprilat greater than captopril greater than ramipril greater than enalapril. This order was different in the kidney (cortex and medulla): ramiprilat greater than enalaprilat greater than ramipril greater than captopril greater than enalapril. For ramiprilat, enalaprilat, and captopril, there were no differences in their respective potencies between tissues or between rat strains. However, the two prodrugs ramipril and enalapril were 10-30 times more active in the kidney than in the other tissues in both groups of rats. This was due to the deesterification of the prodrugs: in the presence of an esterase inhibitor (diethyl nitrophenyl phosphate, 10 microM), the potencies of ramipril in the kidney were not different from that obtained in the lung, which was not affected by the presence of the esterase inhibitor. These results suggest that the variations in the tissue activities of an angiotensin converting enzyme inhibitor are probably not due to differences in tissue affinities of the angiotensin converting enzyme inhibitor but depend on the concentration of this angiotensin converting enzyme inhibitor in each tissue.

The distinction between a circulating renin-angiotensin system and a tissue renin-angiotensin system led us to determine tissue angiotensin converting enzyme (ACE) activity. This study establishes the experimental conditions for a good reproducibility of the fluorimetric assay of ACE and describes the use of [3H]ramiprilat to characterize ACE. Angiotensin converting enzyme activity was determined in rat lung, heart, aorta, and kidney (cortex and medulla) and in rabbit kidney (cortex, medulla, tubules, and glomeruli). ACE activity and [3H]ramiprilat binding does not increase in a linear fashion with the protein content of tissue extracts. Linearity limits varied from 1.0 to 2.0 mg of protein/ml (fluorimetry) and from 0.4 to 1.0 mg of protein/ml [( 3H]ramiprilat binding). Comparing ACE activity, measured by fluorimetry, with the amount of [3H]ramiprilat bound shows that the two techniques yield similar results.