This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Plato, C. F. Articles by Garvin, J. L. Search for Related Content PubMed PubMed Citation Articles by Plato, C. F. Articles by Garvin, J. L. Pubmed/NCBI databases Compound via MeSH Substance via MeSH Hazardous Substances DB (L)-ARGININE NITRIC OXIDE SODIUM Related Collections Genetically altered mice

(Hypertension. 2000;35:319.)
© 2000 American Heart Association, Inc.

Scientific Contributions

eNOS Mediates L-Arginine–Induced Inhibition of Thick Ascending Limb Chloride Flux

Craig F. Plato; Edward G. Shesely; Jeffrey L. Garvin

From the Division of Hypertension and Vascular Research, Henry Ford Hospital, Detroit, Mich.

Correspondence to Jeffrey L. Garvin, PhD, Henry Ford Hospital, Hypertension and Vascular Research Division, 2799 W Grand Blvd, Detroit, MI 48202.

	Abstract

Top Abstract Introduction Methods Results Discussion References

 
Abstract—We recently reported that the rat thick ascending limb (THAL) possesses an active isoform of nitric oxide synthase (NOS) that is substrate-limited in vitro. NO produced by THAL NOS inhibits chloride flux. Protein and transcript for each of the primary NOS isoforms-endothelial (eNOS), inducible (iNOS), and neuronal (nNOS)-have been demonstrated in THALs. However, the NOS isoform that mediates NO-induced inhibition of chloride flux is unknown. We hypothesized that NO produced from eNOS in the THAL inhibits NaCl transport. THALs from male eNOS, iNOS, and nNOS knockout mice and C57BL/6J wild-type controls were perfused in vitro and the response of transepithelial chloride flux (J_Cl) to L-arginine (L-Arg), the substrate for NOS, and spermine NONOate (SPM), an NO donor was measured. We first tested whether isolated mouse THALs could synthesize NO and whether this NO inhibits transport. Addition of 0.5 mmol/L L-Arg to the bath decreased J_Cl from 105.8±17.5 to 79.2±15.8 pmol/mm per minute (P<0.01) in C57BL/6J wild-type mice, whereas addition of D-Arginine had no effects on J_Cl. In contrast, addition of 0.5 mmol/L L-Arg to the bath did not alter J_Cl of THALs from eNOS knockout mice. When 10 µmol/L SPM was added to the bath of eNOS knockout THALs, J_Cl decreased from 89.1±8.6 to 74.8±7.5 pmol/mm/min (P<0.05). Thus the lack of responsiveness of eNOS knockout THALs to L-Arg was not due to an inability to respond to NO. We next evaluated the role of iNOS and nNOS in the response to L-Arg. Addition of 0.5 mmol/L L-Arg to the bath decreased J_Cl in THALs from iNOS and nNOS knockout mice by 37.7±6.4% (P<0.05) and 31.8±8.3% (P<0.01), respectively. We conclude that eNOS is the active isoform of NOS in the THAL under basal conditions. Mouse THAL eNOS responds to exogenous L-Arg by increasing NO production, which, in turn, inhibits J_Cl.

Key Words: nitric oxide • kidney • mice

	Introduction

Top Abstract Introduction Methods Results Discussion References

 
Renal nitric oxide (NO) is an important controller of urinary sodium excretion. NO may enhance natriuresis by inhibiting transport along the nephron as well as altering renal hemodynamics and glomerular filtration rate.¹ Several in vivo studies support the hypothesis that NO directly inhibits tubular sodium reabsorption. Urinary sodium excretion increases when NO production is stimulated² and decreases when NO synthase (NOS) is inhibited³ in the absence of altered renal hemodynamics.

In vitro studies indicate that NO directly affects tubular absorption.^{4 5 6} We recently reported that L-Arginine (L-Arg) inhibits chloride flux through the thick ascending limb of the loop of Henle (THAL) of rats via an L-NAME–sensitive mechanism. These data indicate that endogenously produced NO inhibits THAL transport.⁷ However, these studies did not address the NOS isoform(s) mediating tubular NO synthesis.

mRNA for each of the NOS isoforms has been detected in THALs. With the use of reverse transcription polymerase chain reaction (RT-PCR) of microdissected nephrons, Ujiie et al⁸ detected endothelial (eNOS) mRNA in rat THALs, whereas Mohaupt et al⁹ used competitive RT-PCR to demonstrate high expression of inducible (iNOS) mRNA in medullary THALs of rats. Finally, nNOS transcript was detected by in situ hybridization in THALs and macula densa.¹⁰ NOS protein expression has also been demonstrated in the THAL. Tojo et al¹¹ described positive immunolabeling of constitutive NOS in THALs. More recently, Mattson and Higgins,¹² with the use of Western blots, showed that the rat outer medulla expresses all 3 NOS isoforms.

Currently, the isoform(s) of NOS mediating the effects of L-Arg on THAL transport have not been extensively studied. We hypothesized that L-Arg inhibits mouse THAL chloride absorption through stimulation of eNOS rather than either iNOS or the neuronal (nNOS) isoform. We studied genetically mutated mice to directly evaluate the effects of L-Arg on THAL transport in the absence of specific isoforms of NOS. Our findings indicate that L-Arg inhibits THAL chloride absorption through activation of eNOS.

	Methods

Top Abstract Introduction Methods Results Discussion References

 
Preparation of Isolated Nephron Segments
eNOS- and iNOS-homozygous knockout mice on a C57BL/6J background and their wild-type controls were bred in the Henry Ford Hospital animal facility. Homozygous nNOS knockout mice on a hybrid B6129S background¹³ and B6129S hybrid wild-type controls were obtained commercially (Jackson Laboratory). Thick ascending limbs were obtained from 6-week-old male mice (19 to 23.5 g) maintained on a diet containing 0.22% sodium and 1.1% potassium (Purina) with water ad libitum for at least 5 days. On the day of the experiment, mice were anesthetized with ketamine (150 mg/kg body wt IP) and xylazine (30 mg/kg body wt IP), and the abdominal cavity was opened. The left kidney was bathed in ice-cold saline and removed. Coronal slices were placed in oxygenated physiological saline at 12°C. Thick ascending limbs were dissected in the same solution under a stereomicroscope.

Thick Ascending Limb Perfusion
Thick ascending limbs (0.5 to 0.9 mm in length) were transferred to a temperature-regulated chamber and perfused between concentric glass pipettes at 37°C as described previously.⁷ The composition of the basolateral bath and perfusate (in mmol/L) was NaCl, 114; NaHCO₃, 25; NaH₂PO₄, 2.5; KCL, 4; MgSO₄, 1.2; alanine, 6; Na₃ citrate, 1; glucose, 5.5; Ca-lactate₂, 2; raffinose, 5. The solution was bubbled with 5% CO₂-95%O₂ before and during the experiment, and the pH of the bath was 7.4. The osmolality of the bath solution was 290±3 mosmol/kg H₂O as measured by freezing-point depression. The basolateral bath was exchanged at a rate of 0.5 mL/min, and tubules were perfused at 5 to 10 nL/mm per minute. Time-control experiments were conducted for each protocol to determine the stability of tubular transport.

An NO donor, 1,3-propanediamine, N-[4-[1-(3-aminopropyl)-2-hydroxy-2-nitrosohydrazino]-butyl]C₁₀H₂₆N₆O₂ (spermine NONOate) was purchased from Cayman Chemical. The substrate for NOS, L-Arg, and its stereoisomer D-arginine (D-Arg) were purchased from Sigma Chemical Co.

Net Chloride Flux
Chloride concentrations were determined in samples of perfusate and collected fluid with the use of a previously described fluorometric technique.¹⁴ Because chloride reabsorption was not accompanied by significant fluid reabsorption, net chloride flux (J_Cl) was calculated according to the formula

where PR is the perfusion rate normalized for tubule length, Cl_o is the chloride concentration in the perfusion fluid, and Cl_l is the chloride concentration in the collected tubular fluid.

The typical experimental protocol was as follows. After a 20-minute equilibration period, 3 basal measurements were performed (control period). One of the compounds was then added to the bath, and 20 minutes later 3 additional collections were made (experimental period). Spermine NONOate (SPM), L-Arg, and D-Arg were added to the bath as indicated in the text.

Blood Pressure Measurement
Arterial blood pressure was measured in separate groups of knockout and corresponding control mice under inactin (25 mg/kg SC) anesthesia. Briefly, after a stable plane of anesthesia was obtained, a cervical midline incision was made and a catheter (PE-10) was inserted into the carotid artery. The catheter was advanced to the aortic arch. Mean arterial pressure values for the eNOS knockout mice (n=12) was 102±2 mm Hg, whereas the C57Bl6J controls (n=6) averaged 86±2 mm Hg. The iNOS knockout mice (n=3) averaged 99±5 mm Hg, whereas their C57Bl6J controls (n=4) averaged 101±3 mm Hg. Last, the nNOS knockout mice (n=5) averaged 83±5 mm Hg, whereas their B6129S controls (n=5) averaged 93±4 mm Hg.

Statistics
Experimental results are expressed as mean±SEM. Data were evaluated with Student’s paired t test. The criterion for statistical significance was P<0.05 in all experiments.

	Results

Top Abstract Introduction Methods Results Discussion References

 
We have previously reported that the rat thick ascending limb contains active NOS and that locally produced NO inhibits thick ascending limb transport.⁷ In addition, others have reported detection of both transcript and protein for all 3 isoforms of NOS in the thick ascending limb.^{8 10 12 15} Accordingly, we evaluated the response to L-Arg in isolated perfused thick ascending limbs from control and selective NOS isoform knockout mice to L-Arg.

We first determined whether the mouse thick ascending limb contains active NOS and whether endogenously produced NO inhibits transport. Figure 1 illustrates the effect of the substrate for NOS, L-Arg (0.5 mmol/L), on chloride flux in 6 isolated thick ascending limbs from C57BL/6J controls. During the control period, tubules absorbed chloride at a rate of 105.8±17.5 pmol/mm per minute. After 0.5 mmol/L L-Arg was added to the bath, tubules absorbed chloride at a rate of 79.2±15.8 pmol/mm per minute. Perfusion rates did not differ between the 2 periods. To determine whether the inhibitory effects were specific to the L-isomer, we next evaluated the effects of D-Arg on thick ascending limb J_Cl. During the control period, tubules absorbed chloride at a rate of 152.5±21.5 pmol/mm per minute. After the tubules were treated with 0.5 mmol/L D-Arg, they absorbed chloride at a rate of 166.8±30.3 pmol/mm per minute (n=4). Thus 0.5 mmol/L L-Arg inhibited chloride flux by 26.9±5.5% (P<0.01), indicating that the transport-inhibiting effects of arginine are specific for the L-isomer and that the mouse thick ascending limb possesses a constitutively active isoform of NOS.

View larger version (17K): [in this window] [in a new window]   Figure 1. Exogenous L-Arg inhibits JCl in isolated perfused THALs from C57Bl6J control mice. Addition of 0.5 mmol/L L-Arg to the basolateral bath resulted in a 27% reduction in THAL JCl, from 105.8±17.5 to 79.2±15.8 pmol/mm per minute (n=6; *P<0.01).

We next evaluated the role of endothelial NOS in the inhibitory effects of L-Arg on THAL chloride flux. Figure 2 illustrates the effect of the substrate for NOS, L-Arg (0.5 mmol/L), on chloride flux in 6 isolated thick ascending limbs from eNOS knockout mice on a C57BL/6J background. During the control period, tubules absorbed chloride at a rate of 102.0±26.8 pmol/mm per minute. After 0.5 mmol/L L-Arg was added to the bath, tubules absorbed chloride at a rate of 111.1±19.9 pmol/mm per minute. Perfusion rates did not differ between the 2 periods. These data indicate that selective genetic ablation of the endothelial isoform of NOS prevents the inhibitory effects of L-Arg on THAL J_Cl.

View larger version (18K): [in this window] [in a new window]   Figure 2. Exogenous L-Arg does not inhibit JCl in THALs from eNOS knockout mice. Addition of 0.5 mmol/L L-Arg to the basolateral bath did not alter JCl (102.0±26.8 vs 111.1±19.9 pmol/mm per minute, n=7).

To determine whether the absence of a reaction to L-Arg was due to a defect in responsiveness of eNOS knockout mice to NO, we next evaluated the effect of an NO donor on eNOS thick ascending limb chloride flux. Figure 3 illustrates the effect of the NO donor SPM (10 µmol/L) on chloride flux in 6 isolated thick ascending limbs from eNOS knockout mice. During the control period, tubules absorbed chloride at a rate of 111.5±14.7 pmol/mm per minute. After the tubules were treated with 10 µmol/L SPM, they absorbed chloride at a rate of 74.2±4.7 pmol/mm per minute. Perfusion rates did not differ between the 2 periods. Thus 10 µmol/L SPM inhibited chloride flux by 29.0±8.1% (P<0.05). The inhibition of eNOS knockout THAL chloride flux by exogenous NO indicates that the lack of responsiveness of eNOS knockouts to L-Arg was not due to an inability to respond to NO.

View larger version (17K): [in this window] [in a new window]   Figure 3. Exogenous NO inhibits JCl in THALs from eNOS knockout mice. Addition of an NO donor (10 µmol/L SPM) to the basolateral bath resulted in a 29% reduction in THAL JCl, from 111.5±14.7 to 74.2±4.7 pmol/mm per minute (n=6; *P<0.05).

THALs also have been reported to express iNOS.¹⁵ Therefore we next evaluated the effects of L-Arg on THAL chloride flux in iNOS knockout mice on a C57BL/6J background. Figure 4 illustrates the effect of 0.5 mmol/L L-Arg on chloride flux in 5 isolated thick ascending limbs from iNOS knockout mice. During the control period, tubules absorbed chloride at a rate of 126.9±28.2 pmol/mm per minute. After 0.5 mmol/L L-Arg was added to the bath, tubules absorbed chloride at a rate of 72.4±9.7 pmol/mm per minute. Perfusion rates did not differ between the 2 periods. Thus 0.5 mmol/L L-Arg inhibited chloride flux in iNOS knockout THALs by 37.7±6.4% (P<0.05). These data indicate that selective genetic ablation of the inducible isoform of NOS does not alter the inhibitory effects of L-Arg on THAL J_Cl. In a separate group of iNOS knockout mice, pretreatment with the NOS inhibitor L-NAME (5 mmol/L) blocked the inhibitory effects of L-Arg on THAL chloride flux. These data indicate that the inhibition of THAL transport by exogenous L-Arg is not likely to be caused by a nonspecific effect but by the catabolism of L-Arg to NO by NOS.

View larger version (17K): [in this window] [in a new window]   Figure 4. Exogenous L-Arg inhibits JCl in THALs from iNOS knockout mice. Addition of 0.5 mmol/L L-Arg to the basolateral bath resulted in a 38% reduction in THAL JCl, from 126.9±28.2 to 72.4±9.7 pmol/mm per minute (n=5; *P<0.05).

Expression of nNOS has also been reported in the thick ascending limb.¹⁰ Therefore we next evaluated the effects of L-Arg on chloride flux by THALs from hybrid nNOS knockout mice on a B6129S background. Figure 5 illustrates the effect of 0.5 mmol/L L-Arg on chloride flux in 6 isolated thick ascending limbs from nNOS knockout mice. During the control period, tubules absorbed chloride at a rate of 162.7±27.3 pmol/mm per minute. After 0.5 mmol/L L-Arg was added to the bath, tubules absorbed chloride at a rate of 114.9±23.2 pmol/mm per minute. Perfusion rates did not differ between the 2 periods. Thus 0.5 mmol/L L-Arg inhibited chloride flux in nNOS knockout THALs by 31.8±8.3% (P<0.05). These data indicate that selective genetic ablation of the neuronal isoform of NOS does not alter the inhibitory effects of L-Arg on THAL J_Cl.

View larger version (17K): [in this window] [in a new window]   Figure 5. Exogenous L-Arg inhibits JCl in THALs from nNOS knockout mice. Addition of 0.5 mmol/L L-Arg to the basolateral bath resulted in a 32% reduction in THAL JCl, from 162.7±27.3 to 114.9±23.2 pmol/mm per minute (n=6; *P<0.01).

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
Our data show that (1) L-Arg inhibits chloride flux by isolated mouse thick ascending limbs, whereas D-Arg does not; (2) inhibition of chloride flux is dependent on the presence of the endothelial isoform of NOS and not iNOS or nNOS; and (3) the inability of eNOS knockout mice to respond to L-Arg is not associated with an inability to respond to NO. Taken together, these findings suggest that L-Arg–induced inhibition of thick ascending limb transport is mediated by the action of eNOS. Thus the current data indicate that eNOS may be the primary effector of NO production in response to basolaterally delivered L-Arg. We believe these are the first data showing that L-Arg acts specifically through eNOS to inhibit transport in any nephron segment.

Our findings suggest that L-Arg inhibits thick ascending limb transport, supporting in vivo data¹⁶ that suggest that renal L-Arg exerts a direct effect on urinary sodium excretion. When NOS inhibitors are administered intrarenally, they lower urinary sodium excretion,¹⁷ whereas intrarenal infusion of L-Arg induces natriuresis.¹⁸ These data suggest that L-Arg affects urinary sodium excretion by a direct tubular effect. Our own data indicate that at least part of this effect may reside in the thick ascending limb, where L-Arg inhibits NaCl absorption.

Regulation of thick ascending limb function is critical in the control of urinary sodium excretion because this nephron segment absorbs 25% of the filtered sodium load. Because it is impermeable to water, its absorption of salt both establishes and maintains a hypertonic medullary solute gradient as well as generating dilute tubular fluid.^{19 20} Therefore the ability of L-Arg to directly alter thick ascending limb absorption could have potent effects on urinary NaCl excretion and concentrating ability.

Numerous studies have demonstrated the expression of endothelial, inducible, and neuronal NOS in the thick ascending limb.^{8 9 10} Our current data demonstrating inhibition of chloride absorption with L-Arg and the absence of this response in eNOS knockout mice suggest that the endothelial isoform of NOS is responsible for L-Arg–induced inhibition in chloride flux. These data appear to conflict with other reports that eNOS requires increased intracellular calcium to become activated.²¹ In contrast, the inducible isoform does not have that requirement²¹ but is dependent on substrate availability in vivo²² and in vitro.²³ However, normal thick ascending limb intracellular calcium concentrations are 100 nmol/L,²⁴ whereas the K_1/2 of NOS for calcium is 200 nmol/L.²⁵ Thus, based on Michelis-Menten kinetics, addition of excess substrate (0.5 mmol/L) in the presence of basal intracellular calcium concentrations should be sufficient for 33% of maximal NOS activity. Therefore elevation of intracellular calcium may be unnecessary for activation of thick ascending limb NOS, provided that adequate substrate is available. Further study into the mechanism of L-Arg–induced activation of tubular NOS is needed.

The mechanism by which L-Arg ultimately inhibits chloride absorption in the thick ascending limb is unknown, although the abolition of this response in eNOS knockout mice suggests it is secondary to NO production. NO has been shown to act through a variety of second-messenger cascades, although most of its effects are mediated by cGMP.²⁶ In particular, NO-induced natriuresis is linked to increased cGMP production in the kidney.²⁷ Our laboratory has previously shown that NO increases cGMP in collecting duct cells by activating soluble guanylate cyclase²⁸ and that NO increases cGMP in the thick ascending limb.²⁹ Thus it is possible that L-Arg–mediated NO inhibits transport in the thick ascending limb through stimulation of soluble guanylate cyclase, resulting in an increase in cGMP. We have previously demonstrated that NO stimulates activation of cGMP-dependent protein kinase in cortical collecting ducts.³⁰ Because thick ascending limb sodium chloride absorption depends on the Na-K-2Cl cotransporter, Na-K-ATPase, apical K channel, and basolateral Cl channels, it could be affected by a change in cGMP concentration and in turn decrease chloride transport.

In conclusion, we found that L-Arg–induced inhibition of chloride absorption by isolated mouse thick ascending limbs is stereospecific and dependent on the presence of eNOS. Such inhibition of chloride transport is not altered by the absence of either the inducible or neuronal isoform of NOS and is not secondary to an inability to respond to NO. These findings indicate that eNOS is involved in L-Arg–induced inhibition of thick ascending limb transport under basal conditions. Thus L-Arg may be a physiological regulator of thick ascending limb NO production through activation of eNOS, and the inhibitory effects of L-Arg on thick ascending limb chloride absorption may partially explain the ability of L-Arg to increase urinary sodium excretion in vivo.

	Acknowledgments

 
This work was conducted during the tenure of an American Heart Association Fellowship Grant awarded to C.F. Plato. This work was supported by National Heart, Lung, and Blood Institute grants HL28-982 and HL02-891 awarded to J.L. Garvin.

Received September 14, 1999; first decision October 11, 1999; accepted October 19, 1999.

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Raij L, Baylis C. Glomerular actions of nitric oxide. Kidney Int. 1995;48:20–32.[Medline] [Order article via Infotrieve]

2. Lahera V, Salom MG, Fiksen-Olsen MJ, Raij L, Romero JC. Effects of N^G-monomethyl-L-arginine and L-arginine on the acetylcholine response. Hypertension. 1990;15:659–663.[Abstract/Free Full Text]

3. Lahera V, Salom MG, Fiksen-Olsen MJ, Romero JC. Mediatory role of endothelium-derived nitric oxide in renal vasodilatory and excretory effects of bradykinin. Am J Hypertens. 1991;4:260–262.[Medline] [Order article via Infotrieve]

4. Garcia NH, Pomposiello SI, Garvin JL. Nitric oxide inhibits ADH-stimulated osmotic water permeability in cortical collecting ducts. Am J Physiol. 1996;270:F206–F210.[Abstract/Free Full Text]

5. Roczniak A, Burns KD. Nitric oxide stimulates guanylate cyclase and regulates sodium transport in rabbit proximal tubule. Am J Physiol. 1996;270:F106–F115.[Abstract/Free Full Text]

6. Stoos BA, Garcia NH, Garvin JL. Nitric oxide inhibits sodium reabsorption in the isolated perfused cortical collecting duct. J Am Soc Nephrol. 1995;6:89–94.[Abstract]

7. Plato CF, Stoos BA, Wang D, Garvin JL. Endogenous nitric oxide inhibits chloride transport in the thick ascending limb. Am J Physiol. 1999;276:F159–F163.[Abstract/Free Full Text]

8. Ujiie K, Yuen J, Hogarth L, Danziger R, Star RA. Localization and regulation of endothelial NO synthase mRNA expression in rat kidney. Am J Physiol. 1994;267:F296–F302.[Abstract/Free Full Text]

9. Mohaupt MG, Elzie JL, Ahn KY, Clapp WL, Wilcox CS, Kone BC. Differential expression and induction of mRNAs encoding two inducible nitric oxide synthases in rat kidney. Kidney Int. 1994;46:653–665.[Medline] [Order article via Infotrieve]

10. Bachmann S, Bosse HM, Mundel P. Topography of nitric oxide synthase by localizing constitutive NO synthases in mammalian kidney. Am J Physiol. 1995;268:F885–F898.[Abstract/Free Full Text]

11. Tojo A, Gross SS, Zhang L, Tisher CC, Schmidt HHHW, Wilcox CS, Madsen KM. Immunocytochemical localization of distinct isoforms of nitric oxide synthase in the juxtaglomerular apparatus of normal rat kidney. J Am Soc Nephrol. 1994;4:1438–1447.[Abstract]

12. Mattson DL, Higgins D. Influence of dietary sodium intake on renal medullary nitric oxide synthase. Hypertension. 1996;27:688–692.[Abstract/Free Full Text]

13. Huang PL, Dawson TM, Bredt DS, Snyder SH, Fishman MC. Targeted disruption of the neuronal nitric oxide synthase gene. Cell. 1993;75:1273–1286.[Medline] [Order article via Infotrieve]

14. Garcia NH, Plato CF, Garvin JL. Fluorescent determination of chloride in nanoliter samples. Kidney Int. 1999;55:321–325.[Medline] [Order article via Infotrieve]

15. Morrisey JJ, McCracken R, Kaneto H, Vehaskari M, Montani D, Klahr S. Location of an inducible nitric oxide synthase mRNA in the normal kidney. Kidney Int. 1994;45:998–1005.[Medline] [Order article via Infotrieve]

16. Kanno K, Hirata Y, Emori T, Ohta K, Eguchi S, Imai T, Muramo F. L-Arginine infusion induces hypotension and diuresis/natriuresis with concomitant increased urinary excretion of nitrite/nitrate and cGMP in humans. Clin Exp Pharmacol Physiol. 1992;10:619–625.

17. Mattson DL, Lu S, Nakanishi K, Papanek PE, Cowley AW Jr. Effect of chronic renal medullary nitric inhibition on blood pressure. Am J Physiol. 1994;266:H1918–H1926.[Abstract/Free Full Text]

18. Baylis C, Harton P, Engels K. Endothelial-derived relaxing factor controls renal hemodynamics in normal rat kidney. J Am Soc Nephrol. 1990;1:875–881.[Abstract]

19. Greger R, Velazquez H. The cortical thick ascending limb and early distal convoluted tubule in the urinary concentrating mechanism. Kidney Int. 1987;31:590–596.[Medline] [Order article via Infotrieve]

20. Molony DA, Reeves WR, Andreoli TA. Na⁺:K⁺:2Cl^- cotransport and the thick ascending limb. Kidney Int. 1989;36:418–426.[Medline] [Order article via Infotrieve]

21. White KA, Pufahl RA, Olken NM, Hevel JM, Richards MK, Marletta MA. Nitric oxide synthase: mechanisms and relationship to cytochrome P450. In: Lechner MC, ed. Cytochrome P450. 8th International Congress. Paris: Libbey Eurotest; 1994:43–48.

22. Schott CA, Gray GA, Stoclet JC. Dependence of endotoxin-induced vascular hyporeactivity on extracellular L-arginine. Br J Pharmacol. 1993;108:38–43.[Medline] [Order article via Infotrieve]

23. Beasley D, Schwartz JH, Brenner BM. Interleukin 1 induces prolonged L-arginine-dependent cyclic guanosine monophosphate and nitrite production in rat vascular smooth muscle cells. J Clin Invest. 1991;87:602–608.

24. Naruse M, Uchida S, Ogata E, Kurokawa K. Endothelin 1 increases cell calcium in mouse collecting duct cells. Am J Physiol. 1991;261:F720–F725.[Abstract/Free Full Text]

25. Griffith OW, Steuhr DJ. Nitric oxide synthases: properties and catalytic mechanism. Ann Rev Physiol. 1995;57:707–736.[Medline] [Order article via Infotrieve]

26. Biondi ML, Romero JC. Nitric oxide-mediated reactions stimulate cGMP in the dog kidney. J Vasc Med Biol. 1990;2:294–298.

27. Grandes S, Gallego MJ, Riesco A, Lopez-Farre A, Millas I, Casado S, Hernando L, Caramelo C. Mechanisms of renal effects of different agents stimulating production of cGMP. Am J Physiol. 1991;261:H1109–H1114.[Abstract/Free Full Text]

28. Stoos BA, Carretero OA, Farhy RD, Scicli G, Garvin JL. Endothelium-derived relaxing factor inhibits transport and increases cGMP content in cultured mouse cortical collecting duct. J Clin Invest. 1992;89:761–765.

29. Garcia NH, Plato CF, Stoos BA, Garvin JL. NO-induced inhibition of transport by thick ascending limbs from Dahl S rats. Hypertension. 1999;34:508–513.[Abstract/Free Full Text]

30. Garcia NH, Stoos BA, Carretero OA, Garvin JL. Mechanism of the nitric oxide–induced blockade of collecting duct water permeability. Hypertension. 1996;27:679–683.[Abstract/Free Full Text]

This article has been cited by other articles:

				N. J. Hong and J. L. Garvin Nitric oxide reduces flow-induced superoxide production via cGMP-dependent protein kinase in thick ascending limbs Am J Physiol Renal Physiol, May 1, 2009; 296(5): F1061 - F1066. [Abstract] [Full Text] [PDF]

				E. T. B. Olesen, S. de Seigneux, G. Wang, S. C. Lutken, J. Frokiaer, T.-H. Kwon, and S. Nielsen Rapid and segmental specific dysregulation of AQP2, S256-pAQP2 and renal sodium transporters in rats with LPS-induced endotoxaemia Nephrol. Dial. Transplant., February 17, 2009; (2009) gfp011v2. [Abstract] [Full Text] [PDF]

				V. D. Ramseyer and J. L. Garvin Angiotensin II Decreases Nitric Oxide Synthase 3 Expression via Nitric Oxide and Superoxide in the Thick Ascending Limb Hypertension, February 1, 2009; 53(2): 313 - 318. [Abstract] [Full Text] [PDF]

				M. Herrera, N. J. Hong, P. A. Ortiz, and J. L. Garvin Endothelin-1 Inhibits Thick Ascending Limb Transport via Akt-stimulated Nitric Oxide Production J. Biol. Chem., January 16, 2009; 284(3): 1454 - 1460. [Abstract] [Full Text] [PDF]

				D. Nakano, J. S. Pollock, and D. M. Pollock Renal medullary ETB receptors produce diuresis and natriuresis via NOS1 Am J Physiol Renal Physiol, May 1, 2008; 294(5): F1205 - F1211. [Abstract] [Full Text] [PDF]

				M. Herrera, P. A. Ortiz, and J. L. Garvin Regulation of thick ascending limb transport: role of nitric oxide Am J Physiol Renal Physiol, June 1, 2006; 290(6): F1279 - F1284. [Abstract] [Full Text] [PDF]

				G. Silva, W. H. Beierwaltes, and J. L. Garvin Extracellular ATP Stimulates NO Production in Rat Thick Ascending Limb Hypertension, March 1, 2006; 47(3): 563 - 567. [Abstract] [Full Text] [PDF]

				M. Herrera, G. Silva, and J. L. Garvin A High-Salt Diet Dissociates NO Synthase-3 Expression and NO Production by the Thick Ascending Limb Hypertension, January 1, 2006; 47(1): 95 - 101. [Abstract] [Full Text] [PDF]

				C. S. Wilcox Oxidative stress and nitric oxide deficiency in the kidney: a critical link to hypertension? Am J Physiol Regulatory Integrative Comp Physiol, October 1, 2005; 289(4): R913 - R935. [Abstract] [Full Text] [PDF]

				M. Herrera and J. L. Garvin Recent Advances in the Regulation of Nitric Oxide in the Kidney Hypertension, June 1, 2005; 45(6): 1062 - 1067. [Abstract] [Full Text] [PDF]

				M. Herrera and J. L. Garvin A high-salt diet stimulates thick ascending limb eNOS expression by raising medullary osmolality and increasing release of endothelin-1 Am J Physiol Renal Physiol, January 1, 2005; 288(1): F58 - F64. [Abstract] [Full Text] [PDF]

				M. Varela, M. Herrera, and J. L. Garvin Inhibition of Na-K-ATPase in thick ascending limbs by NO depends on O2- and is diminished by a high-salt diet Am J Physiol Renal Physiol, August 1, 2004; 287(2): F224 - F230. [Abstract] [Full Text] [PDF]

				M. Herrera and J. L. Garvin Endothelin stimulates endothelial nitric oxide synthase expression in the thick ascending limb Am J Physiol Renal Physiol, August 1, 2004; 287(2): F231 - F235. [Abstract] [Full Text] [PDF]

				P. A. Ortiz, N. J. Hong, and J. L. Garvin Luminal flow induces eNOS activation and translocation in the rat thick ascending limb Am J Physiol Renal Physiol, August 1, 2004; 287(2): F274 - F280. [Abstract] [Full Text] [PDF]

				P. A. Ortiz, N. J. Hong, and J. L. Garvin Luminal flow induces eNOS activation and translocation in the rat thick ascending limb. II. Role of PI3-kinase and Hsp90 Am J Physiol Renal Physiol, August 1, 2004; 287(2): F281 - F288. [Abstract] [Full Text] [PDF]

				M. Varela and J. L. Garvin Acute and Chronic Regulation of Thick Ascending Limb Endothelial Nitric Oxide Synthase by Statins J. Am. Soc. Nephrol., February 1, 2004; 15(2): 269 - 275. [Abstract] [Full Text] [PDF]

				P. A. Ortiz, N. J. Hong, D. Wang, and J. L. Garvin Gene Transfer of eNOS to the Thick Ascending Limb of eNOS-KO Mice Restores the Effects of L-Arginine on NaCl Absorption Hypertension, October 1, 2003; 42(4): 674 - 679. [Abstract] [Full Text] [PDF]

				P. A. Ortiz and J. L. Garvin Cardiovascular and renal control in NOS-deficient mouse models Am J Physiol Regulatory Integrative Comp Physiol, March 1, 2003; 284(3): R628 - R638. [Abstract] [Full Text] [PDF]

				P. Ortiz, B. A. Stoos, N. J. Hong, D. M. Boesch, C. F. Plato, and J. L. Garvin High-Salt Diet Increases Sensitivity to NO and eNOS Expression But Not NO Production in THALs Hypertension, March 1, 2003; 41(3): 682 - 687. [Abstract] [Full Text] [PDF]

				T. L. Pallone, Z. Zhang, and K. Rhinehart Physiology of the renal medullary microcirculation Am J Physiol Renal Physiol, February 1, 2003; 284(2): F253 - F266. [Abstract] [Full Text] [PDF]

				P. A. Ortiz and J. L. Garvin Role of nitric oxide in the regulation of nephron transport Am J Physiol Renal Physiol, May 1, 2002; 282(5): F777 - F784. [Abstract] [Full Text] [PDF]

				P. A. Ortiz and J. L. Garvin Interaction of O2- and NO in the Thick Ascending Limb Hypertension, February 1, 2002; 39(2): 591 - 596. [Abstract] [Full Text] [PDF]

				H. Wang, O. A. Carretero, and J. L. Garvin Nitric Oxide Produced by THAL Nitric Oxide Synthase Inhibits TGF Hypertension, February 1, 2002; 39(2): 662 - 666. [Abstract] [Full Text] [PDF]

				S. Adler, H. Huang, J. N. Trochu, X. Xu, S. Gupta, and T. H. Hintze Simvastatin reverses impaired regulation of renal oxygen consumption in congestive heart failure Am J Physiol Renal Physiol, November 1, 2001; 281(5): F802 - F809. [Abstract] [Full Text] [PDF]

				C. F. Plato and J. L. Garvin alpha 2-Adrenergic-mediated tubular NO production inhibits thick ascending limb chloride absorption Am J Physiol Renal Physiol, October 1, 2001; 281(4): F679 - F686. [Abstract] [Full Text] [PDF]

				Y. Kotelevtsev and D. J Webb Endothelin as a natriuretic hormone: the case for a paracrine action mediated by nitric oxide Cardiovasc Res, August 15, 2001; 51(3): 481 - 488. [Full Text] [PDF]

				V. VALLON, T. TRAYNOR, L. BARAJAS, Y. G. HUANG, J. P. BRIGGS, and J. SCHNERMANN Feedback Control of Glomerular Vascular Tone in Neuronal Nitric Oxide Synthase Knockout Mice J. Am. Soc. Nephrol., August 1, 2001; 12(8): 1599 - 1606. [Abstract] [Full Text] [PDF]

				R. Govers and T. J. Rabelink Cellular regulation of endothelial nitric oxide synthase Am J Physiol Renal Physiol, February 1, 2001; 280(2): F193 - F206. [Abstract] [Full Text] [PDF]

				C. F. Plato, D. M. Pollock, and J. L. Garvin Endothelin inhibits thick ascending limb chloride flux via ETB receptor-mediated NO release Am J Physiol Renal Physiol, August 1, 2000; 279(2): F326 - F333. [Abstract] [Full Text] [PDF]

				G. Kovacs, J. Peti-Peterdi, L. Rosivall, and P. D. Bell Angiotensin II directly stimulates macula densa Na-2Cl-K cotransport via apical AT1 receptors Am J Physiol Renal Physiol, February 1, 2002; 282(2): F301 - F306. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Plato, C. F. Articles by Garvin, J. L. Search for Related Content PubMed PubMed Citation Articles by Plato, C. F. Articles by Garvin, J. L. Pubmed/NCBI databases Compound via MeSH Substance via MeSH Hazardous Substances DB (L)-ARGININE NITRIC OXIDE SODIUM Related Collections Genetically altered mice