The effect of prostaglandin synthetase inhibitors on renal function: studies in normal rats during sodium or water diuresis and in rats with chronic renal insufficiency

The effect of acute infusion of the prostaglandin synthetase inhibitors - meclofenamate or indomethacin - was examined in awake rats. Studies were performed in normal rats undergoing either sodium or water diuresis and in salt-replete rats with chronic renal insufficiency. Prostaglandin synthetase inhibitors had no effect on renal plasma flow, glomerular filtration rate or fractional excretion of sodium in any of the groups. Absolute urinary excretion rates for sodium and potassium decreased only in the normal, salt-replete rats. In contrast, prostaglandin synthetase inhibitors consistently decreased urinary flow and osmolar clearance under all experimental conditions studied. In the normal, salt-replete rats the fall in urine flow was preceded by an increase in urinary excretion of cyclic AMP. These results show that inhibitors of prostaglandin synthesis enhance the ability of the kidney to reabsorb water. This effect may be secondary to increased cyclic AMP generation and to increased urea recirculation resulting in higher urea accumulation in the renal medulla.     

The effect of prostaglandin synthetase inhibitors on renal function: Studies in normal rats during sodium or water diuresis and in rats with chronic renal insufficiency

The effect of acute infusion of the prostaglandin synthetase inhibitors — meclofenamate or indomethacin — was examined in awake rats. Studies were performed in normal rats undergoing either sodium or water diuresis and in salt-replete rats with chronic renal insufficiency. Prostaglandin synthetase inhibitors had no effect on renal plasma flow, glomerular filtration rate or fractional excretion of sodium in any of the groups. Absolute urinary excretion rates for sodium and potassium decreased only in the normal, salt-replete rats. In contrast, prostaglandin synthetase inhibitors consistently decreased urinary flow and osmolar clearance under all experimental conditions studied. In the normal, salt-replete rats the fall in urine flow was preceded by an increase in urinary excretion of cyclic AMP. These results show that inhibitors of prostaglandin synthesis enhance the ability of the kidney to reabsorb water. This effect may be secondary to increased cyclic AMP generation and to increased urea recirculation resulting in higher urea accumulation in the renal medulla.     

Plasma atrial natriuretic peptide in conscious rats with reduced renal mass

The effect of salt intake and reduction of renal mass (RRM) on plasma immunoreactive atrial natriuretic peptide (iANP) levels in conscious rats was studied. Rats were divided into RRM and sham-operated groups, and then further subdivided into groups infused with 1 or 6 mEq of sodium per day. Plasma urea nitrogen increased in the groups with RRM. Plasma sodium, sodium balance, and heart rate did not differ between the sham and RRM groups. Rats with RRM maintained on 1 mEq of sodium per day did not have an elevation of water intake, arterial pressure, or plasma iANP. Rats with RRM maintained on 6 mEq of sodium per day had significantly (P less than 0.05) elevated water intake, arterial pressure, and plasma iANP. Arterial pressure and plasma iANP were correlated (r = 0.800) for rats with RRM on either 1 or 6 mEq of sodium per day. Increased plasma iANP in the RRM group on 6 mEq per day was not caused by either RRM or high sodium alone; it was an effect of RRM plus high salt intake. The increase in plasma iANP in the RRM group may be caused by the increase in arterial pressure, possibly due to an increase in extracellular fluid volume. ANP may not be responsible for the sustained increase in fractional sodium excretion observed in RRM.     

Prostaglandins and renal function II. The effect of prostaglandin inhibition on autoregulation of blood flow in the intact kidney of the dog

In order to evaluate the effect of prostaglandin release on renal autoregulation in the intact kidney of the dog, pressure-flow curves were obtained before and after the administration of either indomethacin or meclofenamate, two potent prostaglandin synthetase inhibitors. After drug administration renal venous prostaglandin E decreased in each of eight studies with a mean change from 286 to 141 pg/ml (p < .001). In addition, prostaglandin inhibition was associated with a 31 percent decrease in renal blood flow and a 58 percent increase in renal resistance. Yet, as renal perfusion pressure was decreased by aortic constriction, the change in flow per pressure reduction and the percent change in renal resistance were not significantly different after prostaglandin inhibition when compared to control values in the same animals. The magnitude of the pressure range over which autoregulation was maintained was also similar in the two groups although both the initial and lowest level of autoregulation were slightly higher after prostaglandin inhibition. It is concluded that the administration of these prostaglandin synthetase inhibitors does not significantly impair renal autoregulation in the intact dog kidney.     

Estradiol is responsible for reduced renal prostaglandin dehydrogenase activity in female rats

The contribution of sex steroids to sex-related differences in renal prostaglandin dehydrogenase activity and urinary prostaglandin excretion was examined in 7-8-week-old male and female rats subjected to sham-operation or gonadectomy at 3 weeks of age. Rats were injected subcutaneously twice over a 6-day interval with vehicle (peanut oil, 0.5 mg/kg) or with depot forms of testosterone (10 mg/kg), estradiol (0.1 mg/kg), progesterone (5 mg/kg), or with estradiol and progesterone combined (0.1 and 5 mg/kg). After the second injection, 24-h urine samples were collected for prostaglandin measurement by radioimmunoassay; the rats were killed, and renal and pulmonary prostaglandin dehydrogenase activities were determined by radiochemical assay. Renal prostaglandin dehydrogenase activity was 10-times higher in intact male rats than in intact females. Gonadectomy increased renal prostaglandin dehydrogenase activity 4-fold in females, but had no effect in males; estradiol, alone or combined with progesterone, markedly suppressed renal prostaglandin dehydrogenase activity in both sexes, while testosterone or progesterone alone had no effect. Pulmonary prostaglandin dehydrogenase did not differ between the sexes and was unaffected by gonadectomy or sex-steroid treatment. Intact female sham-operated rats excreted 70-100% more prostaglandin E2, prostaglandin F2 alpha, and 6-keto-prostaglandin F1 alpha in urine than did males; gonadectomy abolished the difference in urinary prostaglandin E2 excretion. Estradiol decreased urinary prostaglandin E2 in females but not in males; treatment with other sex steroids did not alter urinary prostaglandin excretion.     

Increased severity of the acute renal failure induced by HgCl2 on rats with reduced renal mass

In rats unilaterally nephrectomized 2 days before and sham operated controls, an acute fenal failure (ARF) has been induced by subcutaneous HgCl2 injection. The uninephrectomized animals showed a more severe ARF than the sham operated, 60% of the former and 10% of the controls became anuric 48 hours after ARF induction. The increased diuresis and natriuresis produced by acute saline overload did not improve the severity of the ARF. The marked difference in the evolution of this model of ARF with respect to the glycerol induced ARF, which is ameliorated by reduction of renal mass, emphasizes the different pathogenetic mechanism of these two experimental models.     

The effect of sodium intake on renal prostaglandin production

The effect of chronic alterations in dietary sodium intake on renal arachidonic acid (AA) metabolism was studied in male Wistar rats who were maintained for 14 days on a diet consisting of sodium-deficient food and either deionized water (low salt intake, LSI), 1% saline (normal salt intake, NSI), or 2% saline (high salt intake, HSI). 24 h Urinary Sodium (UNaV) and plasma renin activity (PRA) measurements were shown to validate the dietary protocol. Microsomal preparations from the cortices and medullae were incubated with radiolabeled exogenous AA, and endogenous urinary prostaglandin (PG) levels were assayed by RIA to quantify renal PG synthesis. Cortical PGF2 alpha and PGE2 synthesis was found to be the greatest following LSI. In contrast, medullary PGF2 alpha was shown to be the least following LSI and to increase with increased sodium intake. Likewise, urinary PGF2 alpha levels significantly increased with increasing sodium intake. Changes in urinary PGE2 levels showed the same trend as PGF2 alpha but did not achieve statistical significance. These data show that dietary sodium differentially affects renal cortical and medullary PG synthesis and may reflect physiological differences in the regulation of cyclooxygenase in these zones. These data further suggest that the major source of urinary PGs is the renal medulla since the relationship of urinary levels to sodium intake mimics that described for the synthesis of PGs by the medullary tissue.     

Influence of NSAID-induced inhibition of renal prostaglandin synthesis on inorganic sulfate clearance in rats.

The objective of the present investigation was to examine the influence of inhibition of renal prostaglandin synthesis on the renal clearance of inorganic sulfate, an electrolyte involved in the biotransformation of both exogenous and endogenous substrates. Homeostasis of inorganic sulfate is maintained predominantly by renal reabsorption in the proximal tubule. Using a crossover study design, the renal clearance of sulfate was assessed in conscious female Lewis rats during control periods and following the infusion of two structurally dissimilar nonsteroidal anti-inflammatory drugs, ibuprofen (IBU) and indomethacin (INDO). Animals were infused with IBU or INDO to achieve steady state concentrations of 59 +/- 8 micrograms/ml (mean +/- SD) of IBU and 22 +/- 3 micrograms/ml of INDO. At these serum concentrations, IBU and INDO produced greater than 80% decrease in the urinary excretion of prostaglandin (PG) E2. Treatment with either IBU or INDO significantly increased the renal clearance of sulfate, but did not alter the glomerular filtration rate as assessed by creatinine clearance. The role of prostaglandins in the effects of IBU and INDO on sulfate homeostasis was investigated by examining the influence of concomitant intraarterial PGE2 administration (infusion of 0.1 micrograms/min) on nonsteroidal anti-inflammatory drug-induced alterations in sulfate renal clearance. Although PGE2 alone did not significantly alter the renal clearance of inorganic sulfate or that of creatinine, the PGE2 infusion abolished the effects of IBU on sulfate renal clearance. Concomitant PGE2 administration also significantly increased the sulfate reabsorption rate in INDO-treated animals; other parameters were not significantly changed, although the fractional reabsorption of sulfate tended to increase (P = 0.17). The reason for the less pronounced effect on PGE2 on the INDO-sulfate interaction is as yet unknown, but may be partly due to additional mechanisms involved in the INDO-induced alterations in sulfate clearance. The results of these studies suggest that prostaglandin inhibition represents one mechanism whereby IBU can alter the renal clearance of inorganic sulfate.     

Kallidin effect on renal tubular function in meclofenamate- and vehicle-pretreated rats

The effect of kallidin (lysyl-bradykinin) on the urinary recovery of sodium-22 was examined in anesthetized, volume-expanded rats. Sodium-22 was microinfused into the lumen of late proximal convoluted tubules with and without kallidin (100 pg/ml). Kallidin enhanced mean sodium-22 recovery from a control of 2.24 +/- 0.29% to 6.22 +/- 1.30% (delta = 3.98 +/- 1.31%, P less than 0.005). The urinary recovery of simultaneously microinfused inulin, mean blood pressure, urine flow, and the rate of tubular infusion were similar during control and kallidin microinfusions. Pretreatment of rats with meclofenamate (3.0 mg/kg) to inhibit renal prostaglandin synthesis blunted, but did not abolish, the effect of kallidin to promote sodium-22 recovery. The changes in sodium recovery induced by kallidin represent a 175 +/- 47% and a 58 +/- 11% increase from control values in vehicle- and meclofenamate-pretreated rats, respectively. The results indicate that kallidin, microinfused in high doses into the lumen of late proximal tubules, may lower sodium efflux in that nephron. Inhibition of prostaglandin synthesis reduced the tubular effect of kallidin, suggesting that enhanced prostaglandin synthesis may contribute to the natriuretic effects of kallidin. Alternatively, meclofenamate may directly oppose the tubular effect of kallidin.     

Effects of the antioxidant drug tempol on renal oxygenation in mice with reduced renal mass

We tested the hypothesis that reactive oxygen species (ROS) contributed to renal hypoxia in C57BL/6 mice with ⅚ surgical reduction of renal mass (RRM). ROS can activate the mitochondrial uncoupling protein 2 (UCP-2) and increase O(2) usage. However, UCP-2 can be inactivated by glutathionylation. Mice were fed normal (NS)- or high-salt (HS) diets, and HS mice received the antioxidant drug tempol or vehicle for 3 mo. Since salt intake did not affect the tubular Na(+) transport per O(2) consumed (T(Na/)Q(O2)), further studies were confined to HS mice. RRM mice had increased excretion of 8-isoprostane F(2α) and H(2)O(2), renal expression of UCP-2 and renal O(2) extraction, and reduced T(Na/)Q(O2) (sham: 20 ± 2 vs. RRM: 10 ± 1 μmol/μmol; P < 0.05) and cortical Po(2) (sham: 43 ± 2, RRM: 29 ± 2 mmHg; P < 0.02). Tempol normalized all these parameters while further increasing compensatory renal growth and glomerular volume. RRM mice had preserved blood pressure, glomeruli, and patchy tubulointerstitial fibrosis. The patterns of protein expression in the renal cortex suggested that RRM kidneys had increased ROS from upregulated p22(phox), NOX-2, and -4 and that ROS-dependent increases in UCP-2 led to hypoxia that activated transforming growth factor-β whereas erythroid-related factor 2 (Nrf-2), glutathione peroxidase-1, and glutathione-S-transferase mu-1 were upregulated independently of ROS. We conclude that RRM activated distinct processes: a ROS-dependent activation of UCP-2 leading to inefficient renal O(2) usage and cortical hypoxia that was offset by Nrf-2-dependent glutathionylation. Thus hypoxia in RRM may be the outcome of NADPH oxidase-initiated ROS generation, leading to mitochondrial uncoupling counteracted by defense pathways coordinated by Nrf-2.     

The effect of dehydroepiandrosterone (DHEA) on renal function and metabolism in diabetic rats

Dehydroepiandrosterone (DHEA) is an endogenous steroid hormone involved in a number of biological actions in humans and rodents, but its effects on renal tissue have not yet been fully understood. The aim of this study is to assess the effect of DHEA treatment on diabetic rats, mainly in relation to renal function and metabolism. Diabetic rats were treated with subcutaneous injections of a 10 mg/kg dose of DHEA diluted in oil. Plasma glucose and creatinine, in addition to urine creatinine, were quantified espectophotometrically. Glucose uptake and oxidation were quantified using radioactive glucose, the urinary Transforming Growth Factor β₁ (TGF-β₁) was assessed by enzyme immunoassay, and the total glutathione in the renal tissue was also measured. The diabetic rats displayed higher levels of glycemia, and DHEA treatment reduced hyperglycemia. Plasmatic creatinine levels were higher in the diabetic rats treated with DHEA, while creatinine clearance was lower. Glucose uptake and oxidation were lower in the renal medulla of the diabetic rats treated with DHEA, and urinary TGF-β₁, as well as total gluthatione levels, were higher in the diabetic rats treated with DHEA. DHEA treatment was not beneficial to renal tissue, since it reduced the glomerular filtration rate and renal medulla metabolism, while increasing the urinary excretion of TGF-β₁ and the compensatory response by the glutathione system, probably due to a mechanism involving a pro-oxidant action or a pro-fibrotic effect of this androgen or its derivatives. In conclusion, this study reports that DHEA treatment may be harmful to renal tissue, but the mechanisms of this action have not yet been fully understood.     

Localization of exaggerated prostaglandin synthesis associated with renal damage

Regional localization of the exaggerated prostaglandin E₂ (PGE₂) synthesis caused by hydronephrosis was studied in unilateral ureteral ligated rabbits. The renal distribution of PGE₂ production was compared in the hydronephrotic and contralateral kidneys. Basal and bradykinin-stimulated PGE₂ synthesis were increased in cortical and medullary slices of the hydronephrotic kidneys. Contralateral (control) cortical slices produced very low levels of PGE₂ and were insensitive to stimulation by bradykinin (BK). The hydronephrotic cortex produced 10 times more PGE₂ than the contralateral cortex and responded to BK stimulation with increased PGE₂ synthesis. Cortical slices from the hydronephrotic kidney exhibited a time-dependent increase in PGE₂ release, presumably as a result of new protein synthesis. The division of the hydronephrotic cortex into outer and inner regions revealed that the inner cortex produced more PGE₂ than the outer cortex. A similar division of the hydronephrotic medulla showed that the inner medulla produced slightly greater amounts of PGE₂ than the outer medulla. The present study demonstrates that hydronephrosis causes increases in prostaglandin synthesis throughout the kidney. We suggest from these results and other studies that a possible explanation for this finding is the involvement of the collecting duct system in this response. The gradient of PGE₂ production detected in the cortex may have a very significant role in the control of renal hemodynamics and could provide an explanation for the large decrease in blood flow to the inner cortex caused by indomethacin treatment.     

The effect of vasopressin on renal function in young rats a clearance and micropuncture study.

Using micropuncture techniques, the author studied the effect of vasopressin on renal function in young rats at three stages of development -- in the middle of the weaning period (22 days), after weaning was over (30 days) and at the beginning of the sexual maturation period (42 days). In the presence of a hypotonic load, a small dose of vasopressin (12 muU/100 g b.w., i.v.) was most effective in the youngest age group, where it reduced the urine flow by 82% both by increasing water reabsorption and by reducing the GFR. In this group, vasopressin lowered the TF/P Na+ ratio and raised the TF/P K+ ratio in the initial part of the distal tubules of the superficial nephrons, but raised water absorption only beyond the initial part of the distal tubules. Vasopressin reduced the urine flow by 72% in 30-day-old rats by raising water reabsorption beyond the initial part of the distal tubules. The only ion to be affected was K+, whose concentration rose in the final urine. In 42-day-old rats the effect of vasopressin was manifested in only mild depression of the GFR. In this age group, as distinct from younger animals, anaesthesia and surgery evidently led to endogenous vasopressin release, so that the small dose of exogenous vasopressin did not significantly influence the test parameters. This is also underlined by the significant difference between the control urine flow of the 42-day-old and the younger rats.     

The effect of prostaglandin synthesis inhibition on the direct stimulation of renin release from rabbit renal cortical slices

Prostaglandins have been hypothesized to have several mechanistic functions in sympathetically mediated release of renin. The rabbit renal cortical slice system was chosen to examine the prostaglandin dependency of renin release directly stimulated by either a direct adenylate cyclase activator, forskolin, or a β-agonist, isoproterenol. In this study, we demonstrate that with forskolin (1 × 10⁻⁵M) or isoproterenol (1 × 10⁻⁶M), renin release was elevated 2–3 fold above control, and that this increase was shown to accompany a substantial increase in the tissue levels f cAMP (19.5 fold and 3.5 fold respectively). We also demonstrate that the increase in renin release produced by these compounds was not inhibited by cyclooxygenase inhibitors, indomethacin (25 uM) or eicosatetraynoic acid (30 ug/ml), nor was it inhibited by the selective prostacyclin synthesis inhibitor, U-51605 (30 ug/ml). Each of these inhibitors was demonstrated to block the synthesis of prostaglandins in the cortical slices at the concentrations used. Thus we propose that prostaglandins do not play a role in the induction of renin release resulting from elevated cyclic nucleotide levels or β-adrenergic stimulation.     

The effect of prostaglandin synthesis inhibition on the direct stimulation of renin release from rabbit renal cortical slices

Prostaglandins have been hypothesized to have several mechanistic functions in sympathetically mediated release of renin. The rabbit renal cortical slice system was chosen to examine the prostaglandin dependency of renin release directly stimulated by either a direct adenylate cyclase activator, forskolin, or a beta-agonist, isoproterenol. In this study, we demonstrate that with forskolin (1 X 10(-5) M) or isoproterenol (1 X 10(-6) M), renin release was elevated 2-3 fold above control, and that this increase was shown to accompany a substantial increase in the tissue levels of cAMP (19.5 fold and 3.5 fold respectively). We also demonstrate that the increase in renin release produced by these compounds was not inhibited by cyclooxygenase inhibitors, indomethacin (25 microM) or eicosatetraynoic acid (30 micrograms/ml), nor was it inhibited by the selective prostacyclin synthesis inhibitor, U-51605 (30 micrograms/ml). Each of these inhibitors was demonstrated to block the synthesis of prostaglandins in the cortical slices at the concentrations used. Thus we propose that prostaglandins do not play a role in the induction of renin release resulting from elevated cyclic nucleotide levels or beta-adrenergic stimulation.     

The effect of inhibitors of prostaglandin synthesis on hematocrits of mice

Four experiments were performed to evaluate the effects of nonsteroidal anti-inflammatory drugs (NSAID) on hematocrit of mice. Indomethacin (experiment I) and meclofenamic acid (experiment II) reduced (P<.01) the hematocrit by the fourth day of treatment. Following withdrawal of the drug, hematocrit values rose (P<.01) but the degree of recovery four days after withdrawal was dependent on the dose of drug administered. Decreased hematocrit values were accompanied by a decrease (P<.01) in the number of red blood cells per unit volume of blood. However, there were no significant changes in white blood cell numbers (experiment III). Replacement therapy (experiment IV) with PGE₂ partially reversed (P<.01) the effect of indomethacin on hematocrit, but PGF₂α was without effect. The results of these studies indicate that inhibitors of prostaglandin synthesis decreased hematocrit by increasing plasma volume.