Role of Angiotensin II in the Renal Effects Induced by Nitric Oxide and Prostaglandin Synthesis Inhibition

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1 Role of Angiotensin II in the Renal Effects Induced by Nitric Oxide and Prostaglandin Synthesis Inhibition MARIA T. LLINAS, JUAN D. GONZALEZ, EDUARDO NAVA, and F. JAVIER SALAZAR Departamento de Fisiologla, Facultad de Medicina, Murcia, Spain. Abstract. The objective of this study was to examine the renal effects of changes in intrarenal angiotensin II levels during the administration of a cyclooxygenase inhibitor, when nitric oxide synthesis is reduced. In the first group of dogs, the administration of meclofenamate and a subpressor dose of L-NAME induced an increase (P < 0.05) in arterial pressure (14 ± 2 mm Hg), a decrease (P < 0.05) in RBF ( 180 ± 1 3 to ± I 0 ml/min) and GFR (37 ± 3 to 24 ± 5 ml/min), and a reduction in the renal excretory response to a sodium load. In the second group, the administration of a converting enzyme inhibitor prevented the increase in arterial pressure, the renal vasoconstriction, and the increase in the proximal but not the distal tubular sodium reabsorption induced by the inhibition of prostaglandins and nitric oxide synthesis. In the third group, it was found that a small increase in the intrarenal angiotensin II levels, which does not produce changes in renal function in control conditions, induced a significant decrease in RBF (183 ± 14 to 71 ± 12 ml/min) and GFR (36 ± 3 to 13 ± 4 ml/min) when meclofenamate was administered and nitric oxide synthesis was slightly reduced. The results of this study suggest that renal vasoconstriction and increased proximal sodium reabsorption during the reduction of nitric oxide and prostaglandin synthesis are produced by endogenous angiotensin II levels. These results also suggest that endogenous intrarenal nitric oxide and prostaglandins may serve as homeostatic mediators of angiotensin II effects when the intrarenal levels are inappropriately elevated, as occurs in salt-sensitive hypertension. (J Am Soc Nephrol 8: , 1997) The importance of nitric oxide (NO), angiotensin II (Ang II), and prostaglandins (P0) in the regulation of renal hemodynamics and excretory function has been demonstrated in previous studies (1-7). It has been reported that small increases in intrarenal Ang II or small reductions in intrarenal NO levels do not affect renal vascular resistance but reduce the renal ability to eliminate a sodium load (2,5-7). The absence of renal hemodynamic changes during the administration of a subpressor dose of Ang II seems to be secondary to a compensatory increase in NO and/or PG levels ( 1-4). The absence of renal hemodynamic changes during the administration of a subpressor dose of a NO synthesis inhibitor appears to be the result of a compensatory increase in PG levels (7). This possible increase in PG levels is supported by the results obtained in one study, in which the administration of a cyclooxygenase inhibitor when intrarenal NO synthesis is slightly reduced induced an increase in arterial pressure (AP), a more than 40% decrease in RBF and GFR, and reduced the renal ability to eliminate a sodium load (7). With these results, it was proposed that the administration of cyclooxygenase inhibitors can produce renal vasoconstriction and an increase in sodium sensitivity in patients in whom NO synthesis is reduced (7). This hypothesis is supported by the results of Fern et al. (8), who found that cyclooxygenase inhibitors increase arterial pressure in saltsensitive but not salt-resistant hypertensive patients. On the other hand, it has been proposed that NO synthesis is diminished in salt-sensitive hypertension (9). The purpose of this study was to examine the renal effects of changes in intrarenal Ang II levels during the administration of a cyclooxygenase inhibitor when NO synthesis is partially diminished. These effects were evaluated in anesthetized dogs before and after administration of an acute sodium load. Interest in determining the effects of inhibiting Ang II synthesis or of small increases in intrarenal Ang II levels when NO and PG synthesis is reduced stems from the desire to understand the interactions between the different mechanisms involved in the regulation of renal function and from the fact that there are processes (such as aging) in which the intake of cyclooxygenase inhibitors is frequent (I 0) and NO synthesis has been suggested to be decreased (9). To evaluate the renal response to a sodium load during changes in Ang II levels when NO and PG synthesis is reduced may also have some clinical implications because it is known that sodium-sensitivity is enhanced during aging ( I I ); it has been proposed that the sodiumsensitive hypertensive patients have an inappropriate activation of the renin-angiotensin system ( I 2). Received March 18, Accepted November 1 1, Correspondence to Dr. F. Javier Salazar, Departamento de FisiologIa. Facultad de Medicina, Murcia, Spain. l / $03.00/0 Journal of the American Society of Nephrology Copyright 1997 by the American Society of Nephrology Materials and Methods Surgical Preparation Experiments were performed in mongrel dogs ( 14 to 23 kg) of either sex, which were maintained on a standard laboratory diet with free access to water. All experimental procedures were designed according to the Guiding Principles in the Care and Use of Laboratory

2 544 Journal of the American Society of Nephrology Animals approved by the Council of the American Physiological Society. The evening before the experiment, food was removed and lithium (800 mg) was given orally. Dogs were anesthetized with sodium pentobarbital (30 mg/kg, iv) and ventilated artificially. Catheters were placed in the femoral artery for measurement of AP and in the femoral vein for infusion of inulin and additional anesthetic (0.6 ml/min). The left and right kidneys were exposed through flank incisions and the ureters cannulated to allow for comparison of the renal function of both kidneys. The dogs were placed in a metal frame that mimicked their usual standing position. The renal arteries were fitted with noncannulating electromagnetic flow probes and connected to flowmeters. Distal to the flow probe, a curved 23-gauge needle attached to polyethylene tubing was inserted into the renal arteries and connected to a peristaltic pump for infusion of saline or drugs (0.6 ml/min). Finally, a 45-mm stabilization period was allowed before experimental maneuvers were begun. Experimental Groups Group 1 (N = 6). After two I 5-mm control clearances, N#{176}- nitro-l-arginine-methyl ester (L-NAME; 1 jtg/kg per mm) was continuously infused into the right renal artery. Fifteen minutes after the initiation of L-NAME infusion, meclofenamate (5 pg/kg per mm) was administered into both renal arteries for the duration of the experiment. Thirty minutes after the initiation of meclofenamate infusion, two more 15-mm clearances were obtained, and a 5% extracellular volume expansion (ECVE) with isotonic saline was then performed for 45 mm. Two clearances were obtained during the last 10 mm of volume expansion and 10 mm after cessation of the expansion. Finally, 30 mm after the end of the saline infusion, two more 15-mm clearances were obtained. Blood samples were taken during the control period and before the ECVE, during the simultaneous administration of L-NAME and meclofenamate, to determine plasma renin activity (PRA). Previous studies of our group indicate that the doses used of L-NAME and meclofenamate reduce the NO and PG synthesis, respectively (2,5,7). In meclofenamate-treated dogs, the intrarenal infusion of L-NAME (I pg/kg per mm) inhibited the vasodilatory response to the intrarenal infusion of bradykinin ( 10 ngfkg per mm) by more than 65%. A major assumption underlying the results of this study is that the dose of L-NAME used does not completely inhibit the intrarenal NO synthesis because the administration of greater doses produces a significant vasoconstriction ( 13). Group 2 (N = 6). After two 15-mm control clearances, captopril (0.8 pg/kg per mm) was infused into both renal arteries for the duration of the experiment. to inhibit the intrarenal Ang II synthesis. Fifteen minutes after initiating the captopril administration, an infusion of L-NAME and meclofenamate started, with the protocol being similar to that used in group I. In previous studies (2,5), the dose ofcaptopril used was effective in blocking 65% of the decrease in RBF induced by an intrarenal angiotensin I bolus of 0.8 jig. In pilot studies. the maximal intrarenal Ang II dose that can be infused without altering renal hemodynamic or sodium excretion and urine volume was 0.2 nglkg per mm; however, this maximal dose of Ang II is I nglkg per mm after the infusion of captopril. Group 3 (N 5). After two 15-mm control clearances, the Ang II levels were maintained constant in the right kidney by the continuous infusion of captopril (0.8 gfkg per mm) and Ang II (1 nglkg per mm) into the right renal artery. Fifteen minutes after the captopril and Ang II infusions were initiated, L-NAME ( I pg/kg per mm) and meclofenamate (5.tg/kg per mm) were continuously administered into the right renal artery to reduce NO and PG synthesis. Thirty minutes after the infusion of L-NAME and meclofenamate started, two 15-mm clearances were obtained. A 5% ECVE during 45 mm then began, with clearances performed during the last 10 mm of saline infusion and 10 mm after cessation of the expansion. Finally, 30 mm after the end of ECVE, two more 15-mm clearances were obtained. Analytical Methods Renal clearances were taken during each experimental period to determine GFR, sodium (UNaV), potassium (UKV) and lithium excretion, and urine flow rate (UV). Blood samples for hematocrit and plasma sodium, potassium, lithium, and inulin concentrations were also obtained. GFR was measured by the clearance of inulin. Inulin concentrations were analyzed by the anthrone method. Concentrations of sodium and potassium were measured by flame photometry (Corning 435; Halstead Essex, UK). PRA was measured using a commercially available RIA (Sorin Biomedica, Madrid, Spain). Proximal tubule sodium reabsorption was estimated by the lithium clearance technique. Lithium concentrations were measured by flame emission spectrophotometry (Model 5500; Perkin-Elmer Corp.. Norworth, CA). The fact that lithium is a marker for changes in proximal tubule sodium reabsorption is suggested by previous studies showing that fractional lithium excretion (FeLi) increased during administration of proximal- but not distal-acting diuretics (14). During ECVE, lithium clearance has been postulated as a qualitative index of proximal sodium reabsorption because distal reabsorption is insignificant (15). Previous studies of our group have used lithium clearance as a marker of proximal sodium reabsorption (1,2,5,7). Statistical Analyses The data for the two clearance periods for each condition were averaged for statistical comparisons because the fluid and solute excretions were in steady-state conditions. There were no differences between the results obtained during both renal clearances of each period. Data are expressed as means ± SE. Significance of differences in values of each period in the same group and kidney was evaluated using a one-way analysis of variance and the Duncan multiple range test. The significant difference between the same period of different kidneys and groups was calculated with a two-way analysis of variance and the Duncan test. Results Group 1 Table 1 shows that the simultaneous intrarenal infusion of L-NAME and meclofenamate caused an increase in AP that remained (P < 0.05) until the end of the experiment. The inhibition of NO and PG synthesis in the right kidney also induced a significant reduction of RBF (38%) and GFR (35%) before ECVE. RBF and GFR increased to control levels during the ECVE (Table 1, Figure 1) and decreased again during the postexpansion period (1 38 ± 13 ml/min and 27 ± 3 ml/min, respectively). The PG synthesis inhibition in the left kidney did not induce changes in GFR throughout the experiment, but did cause a significant decrease in RBF before the ECVE. During the ECVE and the postexpansion period, RBF increased to control levels. The simultaneous inhibition of NO and PG synthesis induced a greater reduction (P < 0.05) in RBF, UNaV, UV, and FeLi than did the inhibition of PG synthesis alone. It should be noted that RBF and GFR were lower in the

3 Control of Renal Function by Different Hormones 545 Table 1. Effects of a 5% extracellular volume expansion (ECVE) during the prostaglandin synthesis inhibition in both kidneys, with the infusion of meclofenamate (Meclo) into both renal arteries, and the administration of a low dose of a nitric oxide (NO) synthesis inhibitor (L-NAME) into the right renal artery ECVE MAP (mm Hg) 120 ± ± 2h 128 ± 2h Right Kidney Control L-NAME + Meclo L-NAME + Meclo GFR(ml/min) 37±3 245b 37±3 RBF (ml/min) 180 ± ± 10b 16 1 ± 22 UNaV (jteq/min) 47 ± 15 8 ± 3b 233 ± 74k UKV (p.eq/min) 32 ± 3 13 ± 3b 38 ± 5 UV (mi/mm) 0.33 ± ± 0.01k 3.16 ± l.2l FeLi(%) 34±6 l8±2b 30±4 Left Kidney Control Meclo Meclo GFR (ml/min) 37 ± 4 36 ± 3 36 ± 3 RBF (ml/min) 189 ± ± 5bc 187 ± 12 UNaV (p.eq/min) 64 ± ± ± 1 l4ic UKV (Eq/min) 32 ± 3 24 ± 4 59 ± 4b,c UV (ml/min) 0.44 ± ± b.c 6.05 ± I 59b.c FeLi (%) 38 ± 6 32 ± 58 ± 3b,c a Values shown are mean ± SE. MAP, mean arterial pressure: GFR, glomerular filtration rate; RBF. renal blood flow: UNaV, urinary sodium excretion; UKV, urinary potassium b p < 0.05 versus control period. excretion: UV, urine flow rate; FeLi, fractional excretion of lithium. C p < 0.05 versus contralateral kidney. right than in the left kidney before ECVE, after the infusion of L-NAME and meclofenamate started. However, during ECVE, there were no differences between the RBF and GFR of both kidneys (Table 1). During 5% ECVE, UNaV and UV increased in both kidneys, but the increments were greater (P < 0.05) in the left (474 ± 107 p.eq/min and 5.78 ± ml/min, respectively) than in the right kidney (225 ± 73 jteq/min and ± 1.14 ml/min, respectively). During the postexpansion period, UNaV and UV remained elevated (P < 0.05) in both kidneys and were greater (P < 0.05) in the left ( 183 ± 47 Eq/min and 1.55 ± 0.42 ml/min, respectively) than in the right kidney (86 ± 30.tEq/min and 0.69 ± 0.34 ml/min, respectively). Table 1 and Figure 2 (top panel) demonstrate that the ECVE-induced increase in FeLi was greater (P < 0.05) in the left than in the right kidney. In Table 1, the difference in the absolute values of FeLi in the left (58 ± 3%) and right (30 ± 4%) kidneys during ECVE is shown. FeLi remained lower (P < 0.05) in the right ( 19 ± 3%) than in the left (35 ± 5%) kidney during the recovery period of the ECVE. PRA did not change during the simultaneous reduction of NO and PG synthesis (4.2 ± 0.5 ng Ang IImL per h), compared with the control period (4.4 ± 1.4 ng Ang I/mL per h). As occurred in groups 2 and 3, the hematocrit value decreased during the ECVE (P < 0.05), and plasma sodium and potassium concentrations did not change throughout the experiment. Group 2 Table 2 and Figure 1 show that the increase in AP and the renal vasoconstriction induced by the simultaneous inhibition of NO and PG synthesis is blocked by the administration of a converting enzyme inhibitor (CEI). Table 2 also demonstrates that the decrease of UNaV and UV secondary to the NO and PG inhibition, before the ECVE, was blocked during the administration of the CEI. However, the ECVE-induced increments of UNaV, UV, and UKV were lower (P < 0.05) in the right than in the left kidney, indicating that the intrarenal Ang II is not responsible of the blunted excretory response to an ECVE when NO and PG are reduced (Table 2, Figure 2). During the postexpansion period, UNaV and UV remained elevated (P < 0.05) in both kidneys and were greater (P < 0.05) in the left (277 ± 48 peq/min and 1.59 ± 0.30 ml/min, respectively) than in the right kidney (139 ± 26 p.eq/min and 0.71 ± ml/min, respectively). During the ECVE, FeLi increased to a level (56 ± 2%) that was greater (P < 0.05) than that found in group 1 (30 ± 4%), in which a CEI was not administered. Table 2 and Figure 2 also show that FeLi was still lower (P < 0.05) in the right than in the left kidney during ECVE. These results suggest that the ECVE-induced increase in proximal sodium reabsorption in this group is secondary to an enhanced effect of Ang II and to the NO and PG synthesis inhibition. Group 3 Table 3 shows the response of both kidneys to an ECVE during the maintenance of Ang II levels and inhibition of NO and PG synthesis in the right kidney. Table 3 and Figure 1 show that the administration of a subpressor dose of Ang II in this group induced a decrease (P < 0.05) in both GFR (64%)

4 546 Journal of the American Society of Nephrology - - VEHICLE Right Kidney (L-NAME + MECLO) CE! Left Kidney (MECLO) ANGII - - C F- z ± >, z-. I }:.... Right Kidney (L-NAME + MECLO + CEI) - Left Kidney (MECLO + CE!) - F ± c-) }: 0 EXP 5 % CONTROL L-NAME L-NAME MECLO + ± MECLO Figure 1. Line graph illustrating changes in GFR and RBF in response to the simultaneous intrarenal infusion of L-NAME and meclofenamate (MECLO) before and after an 5% volume expansion (EXP 5%). These changes were evaluated in dogs in which vehicle (isotonic saline, ClNa 0.9%), a converting enzyme inhibitor (CEI), or angiotensin II (Ang II) was also infused into the renal artery. P < 0.05 versus control period; + P < 0.05 versus vehicle. and RBF (6 1 %) that remained significant throughout the experiment. The decrease in RBF induced by the simultaneous reduction of NO and PG synthesis was greater (P < 0.05) in this group than in group 1, in which Ang II was not infused. In contrast to the results obtained in group 1. GFR and RBF did not increase during the ECVE (Table 3, Figure 1) and during the postexpansion period ( 1 5 ± 3 ml/min and 73 ± 13 ml/min, respectively). As can be expected from the hemodynamic results, there was a significant decrease of UNaV (88%), UV (84%), and FeLi (76%) before the ECVE, and the ECVEinduced increases in UNaV, UV, and FeLi were almost abolished in the right kidney (Table 3). During the postexpansion period, UNaV and UV decreased (29 ± I 1 p.eq/min and ± 0.05 ml/min, respectively) with respect to the control period. Renal hemodynamics and excretory function in the Figure 2. Bar graphs showing increases in urinary sodium excretion (UNaV) and fractional lithium excretion (FeLi) in right and left kidneys after 5% extracellular volume expansion with isotonic saline. Top panel illustrates the changes during the administration of L- NAME into the right renal artery and meclofenamate (MECLO) into both renal arteries. Bottom panel illustrates the effects of the intrarenal infusion of a CEI during the administration of L-NAME into the right renal artery and meclofenamate (MECLO) into both renal arteries. contralateral kidney did not change before the ECVE. However, as expected, RBF, UNaV, UKV, UV, and FeLi increased in the left kidney during the ECVE. There were significant differences in the absolute values of UNaV, UV, and FeLi in the left and right kidneys during ECVE. The ECVE-induced increases in UNaV, UV, and FeLi tended to be greater in the left kidneys of this group than in the left kidneys of groups 1 and 2, but significant difference was not reached. Discussion This study presents evidence showing that the hypertension and renal vasoconstriction induced by the administration of a cyclooxygenase inhibitor, when NO synthesis is slightly reduced, seem to be secondary to the unrestrained vasoconstriction produced by endogenous Ang II levels. Endogenous Ang II also contributes to increased sodium reabsorption in the proximal but not in distal tubular segments during an increase in extracellular volume when the NO and PG synthesis are reduced. This study also presents new evidence that there is an

5 Control of Renal Function by Different Hormones 547 Table 2. Effects of the administration of a converting enzyme inhibitor (CE!) into both renal arteries on the renal response to a 5% extracellular volume expansion (ECVE), when prostaglandin synthesis was inhibited in both kidneys with the infusion of meclofenamate (Meclo) into both renal arteries, and a low dose of L-NAME was infused into the right renal artery MAP (mm Hg) 128 ± ± 4 ECVE 127 ± 3 Right Kidney Control L-NAME + Meclo + CEI L-NAME ± Meclo + CEI GFR (mi/mm) 31 ± 3 31 ± 4 30 ± 3 RBF (ml/min) 177 ± ± ± 12 UNaV (Eq/min) 40 ± ± ± 60 UKV (peq/min) 39 ± 8 26 ± 6 44 ± 8 UV (mi/mm) 0.23 ± ± ± 0.54 FeLi (%) 29 ± 2 25 ± 3 56 ± 2 Left Kidney Control Meclo + CE! Meclo + CE! GFR (ml/min) 31 ± 3 31 ± 3 30 ± 3 RBF (ml/min) 175 ± ± ± 14 UNV (teq/min) UKV (,teq/min) UV (ml/min) FeLi (%) ± 1 1 ± 7 ± 0.06 ± ± 23 ± 6 ± 0.17 L Values shown are mean ± SE. Abbreviations are defined in the footnote to Table 1. h p < 0.05 versus control period. C p < o.os versus contralateral kidney. ± ± 95 ± 8 ± 0.80 ± 2 important interaction between endogenous NO and PG in modulating the renal vasoconstrictor effects of Ang II. The results obtained in this study during the administration of meclofenamate and a subpressor dose of L-NAME are similar to those previously reported (7). The administration of this dose of L-NAME alone does not produce changes in renal hemodynamics, but reduces the ECVE-induced increments of sodium and water excretion (2,7). On the other hand, meclofenamate alone does not modify the renal excretory response to an ECVE (5,7). However, the administration of this PG synthesis inhibitor together with the low dose of L-NAME induces an increase in AP, renal vasoconstriction, and a decrease in the renal excretory response to an ECVE. It was previously suggested (7) that these effects could be secondary only to the simultaneous reduction of NO and PG synthesis, or also to a hypersensitivity to the vasoconstrictor effect of the endogenous Ang II. The results of this study suggest that the increase in AP and the renal vasoconstriction are produced by endogenous Ang II, because these effects were abolished by the administration of the CE!. Previous studies have suggested that the renal effects induced by the administration of a NO synthesis inhibitor could be secondary to an activation of the reninangiotensin system (16, 17). However, this is the first study showing that the hypertension and renal vasoconstriction induced by the administration of a cyclooxygenase inhibitor when NO synthesis is reduced is prevented with the inhibition of Ang II synthesis. The Ang II dependency of the renal effects secondary to the reduction of NO and PG synthesis is supported by the results obtained in a previous study by our group ( 18), which showed that these effects are only partly reduced with the administration of a calcium antagonist. The administration of the CEI prevented the vasoconstriction but not the blunted excretory response to an ECVE when NO and PG synthesis are reduced. The CE! was infused into both renal arteries to examine the possible mediation by Ang II of the antinatriuretic effects induced by the reduction of NO and PG synthesis, because the effect induced by the CE! could have resulted from the blockade of sodium reabsorption in one tubular segment, offsetting the effect of the NO and PG synthesis reduction in another tubular segment. Our results mdicate that a major component of the proximal tubular effect during ECVE is mediated by Ang II, because FeLi was a 26% greater in the group of dogs in which the CEI was infused. Taken together with results previously reported (2,5,7), these observations suggest that there is an interaction between NO and PG in the regulation of proximal sodium reabsorption during an ECVE. Besides the proximal tubule, other tubular segments could also be involved in the blunted excretory response to an ECVE when NO and PG synthesis are reduced. Previous studies have proposed that NO and PG synthesis are elevated during an ECVE (19,20), and that the renal medulla is capable of releasing large amounts of NO and PG (1 3). It might be that reduced increases in medullary blood flow and renal interstitial hydrostatic pressure (RIHP) are involved in the lowered natriuretic response to the ECVE. This hypothesis is supported by studies showing that increases in medullary blood

6 548 Journal of the American Society of Nephrology Table 3. Effects of a 5% extracellular volume expansion (ECVE) during simultaneous infusion of a CE! and Ang II into the right renal artery to maintain the Ang I! levels in the right kidney; L-NAME and meclofenamate (Meclo) were infused into right renal artery to inhibit NO and PG synthesis ECVE MAP (mm Hg) 126 ± ± ± b Right Kidney Control L-NAME + Meclo + Ang II L-NAME + Meclo + Ang II GFR (mllmin) 36 ± 3 13 ± 4b 14 ± 3b RBF (mi/mm) 183 ± ± ± 10 UNaV (jieq/min) 57 ± 12 7 ± 3b 88 ± 33 UV(p.Eq/min) 44±9 8±lb 15±3 UV (ml/min) 0.31 ± ± 0#{149}01b 0.90 ± 027b FeLi(%) 37±2 9±1 l7±4b Left Kidney Control Vehicle Vehicle GFR RBF (ml/min) (ml/min) UNaV (peq/min) UKV (Eq/min) UV (ml/min) FeLi (%) a Values shown are mean ± SE. Abbreviations are defined in the footnote to Table 1. b p < 0.05 versus control period. C p < 0.05 versus contralateral kidney. 35 ± ± ± ± ± ± 3 36 ± 4C 186 ± l3c 41 ± ± loc 0.26 ± ± 4 35 ± 4C 207 ± 14b.c 1003 ± ± 22b.c 9.45 ± 204b,c 69 ± 10b,c flow and RIHP are necessary to eliminate a sodium load (21,22) and that both NO and PG play an important role in the regulation of medullary blood flow and RIHP (1 3,22-24). On the other hand, it has been suggested that NO modulates sodium reabsorption in collecting ducts (25), and that POE2 induces a decrease of sodium reabsorption in the medullary portion of the thick ascending loop of Henle and in collecting ducts (26). The renal hemodynamic effects induced by the administration of Ang II or a NO synthesis inhibitor is dose-dependent (1, 13,27-29). The administration of a low dose of Ang II in this study during the reduction of NO and PG synthesis produced a 60% decrease in RBF and GFR, which was maintained when a sodium load was administered. This dose of Ang II does not affect renal hemodynamics when infused alone (2,5), and only causes a decrease in RBF when simultaneously infused with a PG synthesis inhibitor (5) or with a subpressor dose of L-NAME (2,7). The decrease in RBF and GFR observed in this study seems to be the result of removing intrinsic NO and PG-mediated renal vasodilation, allowing the vasoconstrictor effect of Ang II to predominate. Taken together with those results previously reported (1,2,5), the results of this study present new evidence that there is an important interaction between NO and PG in protecting the renal vasculature from the vasoconstrictor effects of Ang II. This interaction seems to be more evident in the afferent arteriole and during the administration of a low Ang I! dose. We propose that small increases in intrarenal Ang II levels induce only a decrease in RBF when NO synthesis is slightly reduced (2), because the intrarenal levels of PG protect the afferent arteriole from the vasoconstrictor effects of Ang II. It is also possible that a small increase in intrarenal Ang I! levels induces only a small decrease in RBF when PG synthesis is inhibited (5), because the endogenous intrarenal NO levels protect the renal vasculature from the vasoconstrictor effects of Ang II. The role of NO or PG in protecting the afferent and efferent arterioles from the vasoconstrictor effect of Ang II has been suggested in several studies in which different doses of Ang II were given (1,3-5,28). When a pressor dose of Ang II is infused, the administration of only a PG or a NO synthesis inhibitor induces a significant decrease of GFR (1,3,4,28). These results, together with those presented in this study, suggest that an increased level of only one vasodilator is not enough to protect the afferent arteriole from the effect of a high dose of Ang I!. The protection of the renal vasculature from the vasoconstrictor effect of Ang I! is also ineffective if the dose of this vasoconstrictor is very high, even if the synthesis of one of these vasodilators is not inhibited (27,28). The renal response to the ECVE was almost abolished during the small increase in intrarenal Ang I! levels and the administration of a cyclooxygenase inhibitor, when NO synthesis was partly diminished. This effect on sodium reabsorption could very easily be explained by the marked reduction in RBF and GFR during this sodium load. Whether there was an increased tubular sodium reabsorption independent of the renal hemodynamic effect cannot be discerned in our experiments. The effects induced by the infusion of L-NAME in this study are probably secondary to the intrarenal inhibition of NO

7 Control of Renal Function by Different Hormones 549 synthesis and not to a systemic effect of this NO synthesis inhibitor. A previous study found that the renal hemodynamic effects caused by the intravenous infusion of L-NAME seem to be partly secondary to the extrarenal effects of NO synthesis inhibition (30). Although there are contradictory results (3 1,32), it has been suggested (3 1 ) that an increase in renal sympathetic nerve activity could be partly responsible for the renal effects induced by the systemic inhibition of NO synthesis. In summary, the results of the study presented here suggest that the administration of a cyclooxygenase inhibitor when NO synthesis is slightly reduced: (1) induces an increase in arterial pressure, renal vasoconstriction, and an increase in sodium reabsorption in the proximal tubule that is Ang Il-dependent; and (2) can induce the development of a sodium-sensitive hypertension, even when Ang II synthesis is inhibited because the renal ability to eliminate a sodium load is diminished. It is also suggested that intrarenal NO and PG may serve as homeostatic mediators of Ang II effects when the intrarenal levels are inappropriately elevated, as occurs in some salt-sensitive hypertensive patients (12). Our findings could have clinical implications regarding diseases associated with an impaired modulation of intrarenal Ang II levels and a process such as aging, which seems to be associated with endothelial dysfunction (9) and in which the use of anti-inflammatory drugs is frequent (10). Acknowledgments The authors acknowledge the generous supply of meclofenamate by Parke-Davis (Warner-Lambert, Barcelona, Spain) and of captopril by Squibb Laboratories (Barcelona. Spain). This study was supported by grants from the Fondo de Investigaciones Sanitarias (FIS. 94/780) of Spain and from the European Community (ERBCHRXCT94O,645). References I. Alberola AM, Salazar FJ, Nakamura T, Granger JP: Interaction between angiotensin II and nitric oxide in control of renal hemodynamics in conscious dogs. Am J Physiol 267: Rl472- R1478, Llin#{225}sMT, Gonzalez JD, Salazar FJ: Interactions between angiotensin and nitric oxide in the renal response to volume expansion. Am J Ph% siol 269: R504-R5 10, Nasjletti A, Malik KU: Interrelations between prostaglandins and vasoconstrictor hormones: Contribution to blood pressure regulation. FedProc4l: , Olsen ME, Hall JE, Montani JP, Cornell JE: Protection of preglomerular vessels from angiotensin II vasoconstriction by renal prostaglandins. 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