Joaquín García-Estañ

Pressure-diuresis in volume-expanded rats. Cortical and medullary hemodynamics.

This study evaluated whether pressure-diuretic and pressure-natriuretlc responses are associated with alterations in vasa recta hemodynamics. Autoregulation of cortical and papillary blood flow was studied using a laser-DoppIer flowmeter in volume-expanded and hydropenk rats. Superficial cortical flow and whole kidney renal blood flow were antoregulated in volume-expanded rats and decreased by less than 10% after renal perfusion pressure was lowered from 150 to 100 mm Hg. In contrast, papillary blood flow was not autoregulated and fell by 24 ± 2%. The failure of papillary blood flow to autoregulate was due to changes in the number of perfused vessels as well as to alterations In blood flow in individual ascending and descending vasa recta. Pressure hi vasa recta capillaries increased from 6.8 ± 0.8 to 13.8 ± 1.2 mm Hg after renal perfusion pressure was elevated from 100 to 150 mm Hg, and renal Interstitial pressure rose from 7.4 ± 0.8 to 12.3 ± 1.4 mm Hg. In hydropenic rats, papillary blood flow was autoregulated to a significant extent, but it still decreased by 19% after renal perfusion pressure was lowered from 150 to 100 mm Hg. The pressure-diuretic and presure-natriuretic responses in hydropenic rats were blunted in comparison to those observed in volume-expanded rats. These findings indicate that the pressure-diuretic and pressure-natriuretic responses are associated with changes in vasa recta hemodynamics and renal interstitial pressure.

Nitric Oxide, the Kidney, and Hypertension

The acute administration of nitric oxide (NO) synthesis inhibitors reduces the renal capacity to excrete sodium under normal or volume expanded conditions and increases renovascular resistances in the absence of changes in systemic blood pressure (BP). This indicates a sensitivity of renal vasculature higher than that of systemic vessels to NO synthesis inhibition. Medullary circulation is the renovascular territory most dependent on NO availability. Thus, alterations in medullary blood flow seems to account for the blunted pressure-natriuresis and sodium retention during acute NO synthesis inhibition. By contrast, during chronic administration of L-arginine analogs, systemic BP rises and overrides initial sodium retention by a resetting of the pressure-natriuresis relationship. This BP increase appears to be dependent on an overexpression of the actions of vasoconstrictor systems due to an imbalance created by the diminished NO production. Prolonged NO synthesis inhibition not only elevates BP, but also produces renal vascular and parenchymal damage. Antihypertensive therapy impedes BP elevation and ameliorates kidney deterioration. Finally, there is evidence of the possibility that a certain alteration in the L-arginine-NO pathway exists in genetic models and in human essential hypertension. In conclusion, according to the data contained in the literature, NO plays a significant role in the regulation of systemic and renal hemodynamics and excretory function, and could participate in the development of hypertension.

Effects of chronic increased salt intake on nitric oxide synthesis inhibition-induced hypertension

Objective and method Experimental evidence suggests that endogenous nitric oxide plays an important role in the homeostatic response to an increase in sodium intake. In the present study we evaluated the influence of a high sodium intake (1% NaCl as drinking water) on arterial hypertension induced by long-term (6–7 weeks) inhibition of nitric oxide synthesis [NG-nitro-L-arginine methyl ester (L-NAME), 75 mg/100 ml in the drinking fluid] in rats. Results Treatment with L-NAME induced progressive elevations in tail-cuff systolic blood pressure, but there were no differences between rats drinking tap water and rats drinking 1% NaCl. Direct measurement of blood pressure at the end of the treatment confirmed the hypertension and the lack of differences between the two groups treated with L-NAME. Metabolic studies performed at the end of L-NAME treatment showed a reduced glomerular filtration rate and elevated urinary excretion of immunoreactive endothelin in the two hypertensive groups treated with L-NAME. Drinking intake, diuresis and natriuresis were significantly higher only in the L-NAME group drinking 1% NaCI. Both groups treated with L-NAME showed an accelerated and increased diuretic and natriuretic response to an isotonic 0.9% NaCl load (2.5 ml/100 g body weight, intraperitoneally). At the end of the study ventricular hypertrophy was observed in both L-NAME groups. Conclusion The present results indicate that the time-dependent elevation in blood pressure produced by long-term inhibition of nitric oxide production is not affected by an increased sodium intake. However, salt supplementation induced the development of a polyuria and polydipsia syndrome in rats treated with L-NAME. The elevated excretion of endothelin in both groups treated with L-NAME suggests the possible participation of endothelin in the development of L-NAME hypertension.

Role of AT1 receptors in the renal papillary effects of acute and chronic nitric oxide inhibition

Nitric oxide (NO) is a vasodilator substance controlling renal papillary blood flow (PBF) in the rat. In this study we have evaluated the role of AT1 angiotensin II receptors as modulators of the whole kidney and papillary vasoconstrictor effects induced by the acute or chronic inhibition of NO synthesis. Experiments have been performed in anesthetized, euvolemic Munich-Wistar rats prepared for the study of renal blood flow (RBF) and PBF. In normal rats, acute administration of the NO synthesis inhibitor N ω-nitro-l-arginine methyl ester (l-NAME) increased mean arterial pressure (MAP) and decreased RBF and PBF. Either acute or chronic treatment with the AT1 receptor blocker losartan did not modify the decreases in RBF or PBF secondary to l-NAME. In animals made hypertensive by chronic inhibition of NO, basal MAP was higher, whereas RBF and PBF were lower than in the controls. In these animals, acute or chronic administration of losartan decreased MAP and increased both RBF and PBF significantly. These results indicate that, under normal conditions, the decreases in RBF or PBF induced by the acute inhibition of NO synthesis are not modulated by AT1-receptor stimulation. However, the arterial hypertension, renal vasoconstriction, and reduced PBF present in chronic NO-deficient hypertensive rats is partially due to the effects of angiotensin II, via stimulation of AT1-receptors.Nitric oxide (NO) is a vasodilator substance controlling renal papillary blood flow (PBF) in the rat. In this study we have evaluated the role of AT1 angiotensin II receptors as modulators of the whole kidney and papillary vasoconstrictor effects induced by the acute or chronic inhibition of NO synthesis. Experiments have been performed in anesthetized, euvolemic Munich-Wistar rats prepared for the study of renal blood flow (RBF) and PBF. In normal rats, acute administration of the NO synthesis inhibitor N omega-nitro-L-arginine methyl ester (L-NAME) increased mean arterial pressure (MAP) and decreased RBF and PBF. Either acute or chronic treatment with the AT1 receptor blocker losartan did not modify the decreases in RBF or PBF secondary to L-NAME. In animals made hypertensive by chronic inhibition of NO, basal MAP was higher, whereas RBF and PBF were lower than in the controls. In these animals, acute or chronic administration of losartan decreased MAP and increased both RBF and PBF significantly. These results indicate that, under normal conditions, the decreases in RBF or PBF induced by the acute inhibition of NO synthesis are not modulated by AT1-receptor stimulation. However, the arterial hypertension, renal vasoconstriction, and reduced PBF present in chronic NO-deficient hypertensive rats is partially due to the effects of angiotensin II, via stimulation of AT1-receptors.

Role of nitric oxide in the early renal hemodynamic response after unilateral nephrectomy

We evaluated the involvement of nitric oxide (NO) in the early hemodynamic response to uninephrectomy (UNX) in rats. Animals were uninephrectomized, and 48 h after removal of the kidney, the effect of infusing N G-nitro-l-arginine methyl ester (l-NAME) on renal function was studied. Glomeruli were isolated, and glomerular nitrite and cGMP productions were measured. In addition, endothelial constitutive NO synthase (NOS III) and inducible NO synthase (iNOS) were assessed by Western blot and by measuring the conversion of arginine to citrulline. UNX animals showed an increase in renal plasma flow that was inhibited by l-NAME in a higher proportion than in sham-operated (SO) animals. No differences were observed in systemic NO-dependent vascular tone, since mean arterial pressure showed similar increments in SO and UNX rats. Glomeruli from UNX animals showed an increase in glomerular nitrite production that was blunted byl-NAME addition. Also, cGMP levels were increased in glomeruli from UNX animals, and this increase was inhibited byl-NAME. Western blot analysis showed no differences in NOS III but a higher iNOS amount in glomeruli from UNX than in those from SO rats. No significant differences between UNX and SO rats were found in calcium-dependent NOS enzymatic activity in the renal cortex. However, calcium-independent enzymatic activity was markedly higher in the renal cortex of UNX than in those from SO animals. In conclusion, glomeruli from rats 48 h after UNX had a greater production of NO than those from SO animals. This increased glomerular NO production is based on an increase in the iNOS isoform. Increased glomerular NO synthesis seems to play a role in the decreased renal vascular resistance observed after unilateral nephrectomy in rats.

Effect of homocysteine on calcium mobilization and platelet function in type 2 diabetes mellitus

Type 2 diabetes mellitus induces a characteristic platelet hyperactivity that might be due to several factors including oxidativ stress and abnormal intracellular Ca2+ homeostasis. Hyperhomocysteinaemia is considered a risk factor in the development of thrombosis although its effect on platelet function and the mechanisms involved are still poorly understood. Here we show tha homocysteine induce a concentration‐dependent increase in endogenous production of reactive oxygen species (ROS), which was significantly greater in platelets from diabetic patients than in controls. Platelet treatment with homocysteine resulted in Ca2+ release from the dense tubular system and the acidic stores. Ca2+ mobilization‐induced by homocysteine consisted in two components, an initial slow increase in intracellular free Ca + concentration ([Ca +]i) and a rapid and marked increase in [Ca2+]i, th second leading to the activation of platelet aggregation. As well as ROS generation, Ca2+ mobilization and platelet aggregation were significantly greater in platelets from diabetic donors than in controls, which indicate that platelets from diabetic donors are more sensitive to homocysteine. These findings, together with the hyperhomocysteinaemia reported in diabetic patients, strongly suggest that homocysteine might be considered a risk factor in the development of cardiovascular complications associated to type 2 diabetes mellitus.

Mechanisms of the increased pressor response to vasopressors in the mesenteric bed of nitric oxide-deficient hypertensive rats.

In the present study we analyzed mesenteric vascular reactivity of chronic nitric oxide (NO)-deficient hypertensive rats (NW-nitro-L-Arginine Methyl Ester, L-NAME, 50 mg/kg/day, oral, 3 weeks). Perfusion pressure changes in response to cumulative additions of methoxamine and KCl were significantly increased in the mesenteric vessels of the L-NAME-treated as compared with vessels of the controls. Verapamil reduced the responses to methoxamine, but those of the hypertensive rats were still enhanced. In contrast, responses to KCl were almost completely abolished by verapamil. In mesenteric vessels perfused with zero calcium and high-potassium Krebs, pressor responses to the re-addition of calcium were also significantly enhanced in the hypertensive rats compared to the controls. Vasodilator responses to acetylcholine in KCl-preconstricted vessels, while still significant, were reduced in the L-NAME-treated rats. In this case, acute inhibition of NO blocked the vasodilator responses to acetylcholine and abolished the differences between the two groups. In methoxamine-preconstricted vessels and in the presence of acute inhibition of NO and prostaglandins, vasodilator responses to acetylcholine were significantly greater in the hypertensive vessels than in controls. In conclusion, the mesenteric vessels of L-NAME hypertensive rats show an enhanced response to vasopressors which is related to calcium entry. These data also reveal the existence of an enhanced role of a NO and prostaglandin-independent vasodilator factor, probably endothelium-derived hyperpolarizing factor that may play a compensatory role in the deficiency of NO.

Hemodynamic effects of hypertonic saline in the conscious rat.

The present study examines the role of vasopressin and the sympathetic nervous system on the hemodynamic effects of an infusion of hypertonic saline (NaCl 1.5 M) in conscious rats. The cardiovascular response to hypertonic saline was similar in both untreated and hexamethonium-pretreated rats. Mean arterial pressure increased by 15 mmHg as a consequence of the elevation of total peripheral resistance, while cardiac index was decreased. The administration of an antagonist to the pressor activity of vasopressin in rats with intact reflexes, partially decreased mean arterial pressure and total peripheral resistance and increased cardiac index toward basal values. In contrast, the hemodynamic response to hypertonic saline was totally reverted when the vasopressin antagonist was injected in the hexamethonium-pretreated rats. The results of the present study indicate that the hypertensive response induced by hypertonic saline in conscious rats is due to the vasoconstrictor effects of both vasopressin and the sympathetic nervous system.

Importance of Nitric Oxide and Prostaglandins in the Control of Rat Renal Papillary Blood Flow

The role of nitric oxide and prostaglandins in the control of rat renal papillary blood flow has been studied in anesthetized Munich-Wistar rats by use of laser Doppler flowmeter. Acute administration of N omega-nitro-L-arginine methyl ester (L-NAME) 10 mg/kg IV (n=8) increased mean arterial pressure by 27.8 +/- 3.6%, decreased papillary blood flow by 39.4 +/- 3.8%, and decreased renal blood flow by 47.4 +/- 1.9%. The subsequent administration of indomethacin (7.5 mg/kg IV) further decreased papillary blood flow (35.2 +/- 2.5%) without significant changes in mean arterial pressure or renal blood flow. In a second group (n = 6), administration of indomethacin before L-NAME decreased papillary blood flow by 39.6 +/- 2.1% without significantly altering mean arterial ressure or renal blood flow. The subsequent injection of L-NAME further decreased papillary blood flow (32.9 +/- 1.8%) and renal blood flow (49.8 +/- 6.6%) while increasing mean arterial pressure to a level not significantly different from that found in the first group. Autoregulation studies showed that L-NAME but not indomethacin reduced the renal perfusion pressure-renal blood flow relationship without altering autoregulation. However, both nitric oxide and prostaglandins importantly affected the renal perfusion pressure-papillary blood flow relationship because L-NAME and indomethacin significantly decreased this relationship in an additive fashion. Although both drugs reduced the sensitivity of the pressure-papillary flow relationship, only L-NAME affected autoregulation so that papillary blood flow was autoregulated at higher renal perfusion pressures. Thus, the present results indicate that both nitric oxide and prostaglandins control a similar percentage of rat renal papillary blood flow, but nitric oxide seems to be more important than prostaglandins as a mediator of the pressure-blood flow relationship. In contrast, only nitric oxide modifies the renal blood flow level, although it does not disturb whole-kidney blood flow autoregulation.

Role of protein kinase C in mesenteric pressor responses of rats with portal hypertension.

1 Hyporesponsiveness to vasoconstrictors is a characteristic abnormality of liver diseases of uncertain origin. In the present study, we have evaluated the involvement of protein kinase C (PKC) in the reduced pressor response to methoxamine (MTX) of a rat model of portal hypertension induced by partial portal vein ligation (PVL). Experiments were performed in the isolated and perfused mesentery. 2 The pressor response to MTX was reduced in PVL compared to that of control animals (Sham) and pretreatment with NG‐nitro‐L‐arginine (L‐NOARG, 10−4 m) or removal of the endothelium potentiated the response of both groups. However, only removal of the endothelium completely eliminated the reduced pressor response to MTX of the PVL vessels. 3 Pretreatment of the mesentric vessels with calphostin C (10−6 m), a PKC inhibitor, reduced the response to MTX of Sham to a level similar to that of untreated PVL vessels, but did not change that of PVL animals. 4 Mesenteric pressor responses to a PKC activator, phorbol 12,13‐dibutyrate (PDBu), were similar in vessels from both PVL and Sham rats and pretreatment with L‐NOARG or removal of the endothelium enhanced those responses while indomethacin (10−5 m) decreased them. In all cases, the responses to PDBU were similar in PVL vessels compared to Sham. 5 These results indicate that the reduced pressor response to MTX of the mesenteric vascular bed of PVL rats is due to an endothelial alteration, compatible with an enhanced production of nitric oxide. The lack of response to calphostin C in PVL vessels suggests an impairment in agonist‐induced PKC activation. Since direct activation of PKC induces a normal pressor response, it is concluded that the endothelial alteration interacts with the mechanism producing PKC activation, which results in a lower pressor response of the PVL mesenteric vaculature.