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. 2005 Oct 20;569(Pt 3):849–856. doi: 10.1113/jphysiol.2005.097709

Nitric oxide inhibition and the impact on renal nerve-mediated antinatriuresis and antidiuresis in the anaesthetized rat

NM Bagnall 1, PC Dent 1, A Walkowska 2, J Sadowski 2, EJ Johns 1
PMCID: PMC1464264  PMID: 16239274

Abstract

The contribution of nitric oxide (NO) to the antinatriuresis and antidiuresis caused by low-level electrical stimulation of the renal sympathetic nerves (RNS) was investigated in rats anaesthetized with chloralose–urethane. Groups of rats, n = 6, were given i.v. infusions of vehicle, l-NAME (10 μg kg−1 min−1), 1400W (20 μg kg−1 min−1), or S-methyl-thiocitrulline (SMTC) (20 μg kg−1 min−1) to inhibit NO synthesis non-selectively or selectively to block the inducible or neuronal NOS isoforms (iNOS and nNOS, respectively). Following baseline measurements of blood pressure (BP), renal blood flow (RBF), glomerular filtration rate (GFR), urine flow (UV) and sodium excretion (UNaV), RNS was performed at 15 V, 2 ms duration with a frequency between 0.5 and 1.0 Hz. RNS did not cause measurable changes in BP, RBF or GFR in any of the groups. In untreated rats, RNS decreased UV and UNaV by 40–50% (both P < 0.01), but these excretory responses were prevented in l-NAME-treated rats. In the presence of 1400W i.v., RNS caused reversible reductions in both UV and UNaV of 40–50% (both P < 0.01), while in SMTC-treated rats, RNS caused an inconsistent fall in UV, but a significant reduction (P < 0.05) in UNaV of 21%. These data demonstrated that the renal nerve-mediated antinatriuresis and antidiuresis was dependent on the presence of NO, generated in part by nNOS. The findings suggest that NO importantly modulates the neural control of fluid reabsorption; the control may be facilitatory at a presynaptic level but inhibitory on tubular reabsorptive processes.

The kidney is extensively innervated by the sympathetic nervous system, and neuroeffector junctions have been described along the vasculature as well as the tubules (Liu et al. 1996). Activation of the sympathetic nervous system regulates all aspects of kidney function; thus, at low levels of activity there is a primary influence to stimulate renin release (Hesse & Johns, 1984), while at higher degrees of activation there is a concomitant increase in sodium reabsorption along the proximal tubule (Bello-Reuss et al. 1976) and thick ascending loop of Henle (DiBona & Sawin, 1982). It is only when the renal nerves are stimulated at high rates that there are short-term decreases in both renal blood flow and glomerular filtration rate (DiBona & Kopp, 1997). There are a number of autocrine and paracrine factors within the kidney that also impact on vascular and tubular function, and may buffer or enhance the influence of extrinsic factors such as activity of the renal sympathetic nerves. One of these is nitric oxide (NO), which is generated by the enzyme nitric oxide synthase (NOS), and which exists in three isoforms. Endothelial NOS (eNOS) has been shown to occur along the renal vessels and glomerular capillaries (Bachmann et al. 1995; Mattson & Wu, 2000), while the neuronal isoform (nNOS) has been found in nitrergic nerve fibres and terminals (Liu et al. 1996), at low concentrations along the tubules but at high concentrations at the macula densa (Bosse et al. 1995). The inducible form (iNOS) has been reported to be expressed constitutively in the medullary regions of the kidney (Kone & Baylis, 1997).

At the neuro-effector junction, NO has the potential of exerting both pre- and postsynaptic actions, which complicate studies of its role in neuro-transmission. There is evidence in the anaesthetized dog that in response to low-level renal nerve stimulation, noradrenaline output is enhanced following NOS blockade, and suppressed by exogenous NO, suggesting that NO exerts a tonic inhibitory action at the presynaptic level (Egi et al. 1994; Maekawa et al. 1996). In contrast, in the rat, it has been reported that sympathetic nerve-induced noradrenaline release from the heart is enhanced (Schwartz et al. 1995) but is suppressed from the mesenteric vasculature (Yamamoto et al. 1994, 1997) as a consequence of NOS inhibition. Tanioka et al. (2002), using the isolated pump perfused rat kidney, reported that NOS blockade suppressed sympathetic nerve-mediated noradrenaline release and the vasoconstrictor responses to high levels of renal nerve stimulation consistent with a presynaptic facilitatory action of NO. Taken together, these divergent findings mean that there is a lack of clarity as to the exact action of NO at neuroeffector junctions and would suggest that the role played by NO is dependent upon the tissue, organ and species studied.

Most reports have focused on the role of NO in modulating neurotransmission at the renal vasculature, but another major site of neural action is at the tubular epithelia to stimulate sodium reabsorption. Recent reports from this laboratory have shown that administration of l-NAME into the proximal tubules increased, and an NO donor decreased, fluid reabsorption, indicative of NO having a direct action to inhibit fluid reabsorption (Wu et al. 1999). Moreover, low-level RNS, which increased fluid reabsorption in a frequency-related way, was prevented following intraluminal administration of l-NAME and 7-nitroindazole (7-NI), which is a relatively selective nNOS blocker (Wu & Johns 2002, 2004). However, whether this local application of the NOS blockers was sufficient to penetrate into the neuro-effector junction was unclear. These observations were, in part, compatible with NO having a facilitatory action at the neuro-effector junction.

The present study aimed to investigate the importance of NO in mediating neurally induced sodium reabsorption in a whole-kidney setting and to evaluate the isoform of NOS involved. This was done by directly stimulating the renal nerves at low frequencies that had no effect on renal haemodynamics, and determining how the antinatriuretic and antidiuretic responses were affected following relatively selective blockade of different NOS isoforms.

Methods

The experimental procedures all conformed to European, National and local Biomedical Ethical Committee guidelines, with approval from the University Local Ethical Committee. Male Wistar rats, 275–360 g, were maintained on a regular rat diet (SDS, RMI; Lillico, Surrey, UK) and had restricted food availability, but water ad libitum, over the night prior to use. The animals were prepared as previously described (Huang & Johns, 2001). Briefly, anaesthesia was induced using a mixture of halothane–O2–NO2, and the right femoral vein was cannulated and 0.8 ml of chloralose–urethane (12 and 180 mg ml−1) was given over 35 min as the gaseous anaesthesia was gradually reduced. Supplementary doses of the chloralose–urethane, 0.05 ml, were given i.v. every 30 min. A saline (150 mm NaCl) infusion i.v. was begun at 3 ml h−1 and continued throughout the experiment. A further cannula was placed in the right femoral artery to measure arterial blood pressure. Another cannula was inserted into the trachea to facilitate spontaneous respiration. The right ureter was exposed through a flank incision and cannulated to allow urine collection. The left kidney was similarly exposed, its ureter cannulated, its artery cleared in order to fit an electromagnetic flow probe (Carolina Medical Electronic, King, NC, USA), and the renal nerves were exposed and placed on stimulating electrodes. After completion of surgery, a 2 ml bolus of inulin (15 mg ml−1) in saline was given i.v. as a primer, which was followed by an infusion of saline containing 15 mg ml−1 inulin at 3 ml h−1 for the remainder of the experiment. The animals were allowed 2 h to stabilize before the experimental protocols were begun.

The experimental protocol comprised five 15 min clearance periods, two before and two following a period in which the renal sympathetic nerves were stimulated at 15 V, 2 ms duration and at a frequency which was just subthreshold for causing a reduction in renal blood flow (0.5–1.0 Hz). The nerves were stimulated for a total of 20 min, but urine collections were not begun until 5 min after the start of stimulation in order to clear preformed urine from the ureteral cannula. The rats were killed humanely with a 1 ml overdose of anaesthetic at the end of the experiment.

Four groups of rats were studied. Group I received an infusion of saline throughout, and acted as a control group. Group II received a continuous i.v. infusion of Nω-nitro-l-arginine methyl ester (l-NAME) at 10 μg kg−1 min−1 for 30 min prior to the start of the clearance protocol. This infusion rate of l-NAME has been reported to block all NOS isoforms, but without increasing blood pressure or reducing renal haemodynamics (Lahera et al. 1991). Group III was given 1400W at 20 μg kg−1 min−1i.v. for 30 min before beginning the sequence of clearance measurements. This compound has a 500-fold selectivity for blocking iNOS compared with eNOS, and at this infusion rate it has been demonstrated to prevent iNOS-mediated events (Garvey et al. 1997). Group IV was infused with S-methyl thiocitrulline (SMTC) at 20 μg kg−1 min−1i.v. for 30 min before the start of the clearance collections. SMTC is a relatively selective nNOS blocker having a 17-fold greater inhibitory action for nNOS than eNOS (Furfine et al. 1994), and at this dose level it has been reported to depress renal levels of NO (Walkowska et al. 2005). All drugs were obtained from ‘Sigma, (Dorset, UK)’.

All data are the average values calculated from individual animals and are given as means ± s.e.m. The renal responses to stimulation were calculated by averaging the two clearances before and the two following renal nerve stimulation and comparing the values to those obtained whilst the nerves were stimulated. Comparisons were undertaken using a two-way ANOVA (SuperANOVA Software, Abacus, CA, USA). Significance was taken when P < 0.05.

Results

Table 1 shows the basal blood pressure and renal haemodynamic and excretory variables in animals that were either untreated or treated with one of the three NOS inhibitors. Administration of l-NAME had no effect on blood pressure or left renal blood flow, and although the left glomerular filtration rate was significantly (P < 0.001) lower than that in the saline-treated rats, the lower right glomerular filtration rats was not, while fluid excretions from both kidneys were similar to those of the rats infused with saline. Infusion of 1400W had no effect on blood pressure, or renal blood flow, glomerular filtration rate or fluid excretions from either kidney. The infusion of SMTC resulted in a blood pressure significantly (P < 0.05) higher, by some 14 mmHg, than that of the saline-infused group, but all other variables were comparable to the saline-infused rats.

Table 1.

Basal levels of blood pressure, blood flow, left and right glomerular filtration rate, left and right urine flow and left and right absolute sodium excretion

Saline (n = 6) l-NAME (n = 6) 1400W (n = 6) SMTC (n = 6)
BP (mmHg) 105 ± 2 108 ± 3 108 ± 3 119 ± 4*
Left RBF (ml min−1 kg−1) 24.1 ± 2.8 21.8 ± 3.3 21.3 ± 4.6 17.0 ± 4.2
Left GFR (ml min−1 kg−1) 4.88 ± 0.55 2.80 ± 0.55** 7.93 ± 2.53 4.89 ± 0.47
Left UV (μl min−1 kg−1) 87.7 ± 11.6 63.7 ± 8.4 101.9 ± 16.6 53.2 ± 12.3
Left UNaV (μmol min−1 kg−1) 11.7 ± 1.7 10.1 ± 1.3 22.1 ± 16.6 8.35 ± 2.28
Right GFR (ml min−1 kg−1) 5.57 ± 0.81 3.35 ± 0.82 6.92 ± 1.03 5.15 ± 0.32
Right UV (μl min−1 kg−1) 99.2 ± 17.0 64.1 ± 11.2 92.5 ± 16.6 61.4 ± 14.2
Right UNaV (μmol min−1 kg−1) 13.6 ± 2.6 10.4 ± 0.7 19.4 ± 3.9 8.9 ± 1.7
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BP, blood pressure; RBF, renal blood flow; GFR, glomerular filtration rate; UV, urine flow; UNaV, absolute sodium excretion. n = number of animals.

*

P < 0.05

**

P < 0.01 compared with the saline group.

Stimulation of the left renal sympathetic nerves at low levels in the saline-infused group had no effect on blood pressure or left renal blood flow (Table 2), and although there was a small inconsistent fall in left glomerular filtration rate, right glomerular filtration rate was unchanged (Fig. 1). Under these conditions of RNS, there were significant (P < 0.01) reversible decreases in left urine flow and sodium excretion of some 49 and 39%, respectively (Fig. 1). In contrast, right kidney urine flow and sodium excretion did not change during this period of left renal nerve stimulation (Fig. 1).

Table 2.

Blood pressure and renal blood flow under basal conditions, during renal nerve stimulation and over the recovery period in the groups of rats (n = 6 in all groups) receiving saline, L-NAME, 1400W or SMTC

Basal Stimulation Recovery
Saline control
 BP (mmHg) 105 ± 2 109 ± 2 106 ± 2
 RBF (ml min−1 kg−1) 24.1 ± 2.8 23.9 ± 2.9 23.1 ± 2.3
l-NAME
 Blood pressure (mmHg) 108 ± 3 111 ± 4 108 ± 3
 RBF (ml min−1 kg−1) 21.8 ± 3.3 20.2 ± 3.4 18.6 ± 3.2
1400W
 BP (mmHg) 108 ± 3 110 ± 3 107 ± 3
 RBF (ml min−1 kg−1) 21.3 ± 4.6 17.2 ± 2.8 18.7 ± 3.4
SMTC
 Blood pressure (mmHg) 119 ± 4 121 ± 4 116 ± 4
 Renal blood flow (ml min−1 kg−1) 17.0 ± 4.2 14.0 ± 3.5 14.1 ± 3.1
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BP, blood pressure; RBF, renal blood flow; SMTC, S-methyl-thiocitrulline.

Figure 1. Renal haemodynamic and excretory responses to nerve stimulation.

Figure 1

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Effect of left renal nerve stimulation on glomerular filtration rate, urine flow and sodium excretion in the left (open bars) and right kidneys (filled bars) (n = 6) before, during renal nerve stimulation (Stim) and in the recovery (Rec) period. *P < 0.05.

In the group of rats receiving l-NAME, left renal nerve stimulation had no effect on blood pressure and did not change left renal blood flow (Table 2). Moreover, glomerular filtration rate, urine flow and sodium excretion of the left kidney were not altered during the period of renal nerve stimulation (Fig. 2); these results were different from the pattern observed in the control rats receiving saline only. Over the time of the experiment, glomerular filtration rate, urine flow and sodium excretion of the right kidney remained at a stable level (Fig. 2).

Figure 2. Effect of L-NAME on the renal haemodynamic and excretory responses to nerve stimulation.

Figure 2

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Effect of left renal nerve stimulation on glomerular filtration rate, urine flow and sodium excretion in left (open bars) and right kidneys (filled bars) in the presence of l-NAME (n = 6) before, during renal nerve stimulation (Stim) and in the recovery (Rec) period.

During infusion of 1400W, stimulation of the left renal nerves was without effect on either blood pressure or renal blood flow (Table 2). Renal nerve stimulation had no effect on left glomerular filtration rate but reversibly decreased both left urine flow and sodium excretion (both P < 0.01) by 46 and 39%, respectively (Fig. 3). The magnitude and pattern of these responses were comparable with those observed in the rats infused with saline. There was no change in right glomerular filtration rate, but there was a gradual decline in right urine flow and sodium excretion over the period when the left renal nerves were stimulated (Fig. 3), but none of these changes reached statistical significance.

Figure 3. The impact of 1400 W on the renal haemodynamic and excretory responses to nerve stimulation.

Figure 3

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Effect of left renal nerve stimulation on glomerular filtration rate, urine flow and sodium excretion in left (open bars) and right (filled bars) kidneys in the presence of 1400W (n = 6) before, during renal nerve stimulation (Stim) and in the recovery (Rec) period. *P < 0.05.

In the group of rats receiving SMTC (Fig. 4), stimulation of the left renal nerves had no effect on blood pressure or left renal blood flow (Table 2). Under these conditions (Fig. 4), left glomerular filtration rate did not change and, while there were non-significant falls in left urine flow, left sodium excretion was reduced significantly (P < 0.05) by 22%, which was significantly (P < 0.05) smaller than the magnitude of the reduction observed in the group of rats receiving saline. Over this period, right glomerular filtration rate, urine flow and sodium excretion showed small gradual increases that did not reach statistical significance (Fig. 4).

Figure 4. The influence of SMTC on the renal haemodynamic and excretory responses to nerve stimulation.

Figure 4

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The effect of renal nerve stimulation on glomerular filtration rate, urine flow and sodium excretion in the left (open bars) and right (filled bars) kidneys in the presence of S-methyl-thiocitrulline (SMTC; n = 6) before, during renal nerve stimulation (Stim) and in the recovery (Rec) period. *P < 0.05.

Discussion

This investigation was focused on outlining the contribution that NO might make to the neural control of urinary sodium excretion in the anaesthetized rat, and identifying the isoform of NOS that might be responsible. To this end, the renal sympathetic nerves to the left kidney were exposed and stimulated at rates which were subthreshold for reducing renal blood flow and glomerular filtration rate, but which caused an antinatriuresis and an antidiuresis. Three compounds were used to block NOS, one non-selective and two with relative selectivity. Thus, l-NAME was chosen to block all isoforms of NOS and it was given as a slow i.v. infusion. At the dose and method of administration chosen, l-NAME causes minimal changes in blood pressure, and it is one compound which has been shown to block NOS activity (Lahera et al. 1991) when given in this way. It was considered important that large bolus doses of l-NAME were avoided since these result in large increases in blood pressure and reductions in renal blood flow and glomerular filtration rate which would have impacted on the ability to demonstrate renal nerve-induced changes. Under these conditions, l-NAME had no measurable effect on basal renal haemodynamics or sodium and water excretion from either kidney compared with the group of rats given saline only, and these observations were comparable with those of Lahera et al. (1991). Moreover, this dosing regime has been used previously in this laboratory to study the role of NOS in the control of renal function (Wongmekiat & Johns 2001a, b, c).

The 1400W compound is one that has a 500-fold greater selectivity of inhibition for iNOS as against eNOS (Garvey et al. 1997), and the dose chosen for the present study has been used previously to prevent iNOS-mediated activities (Rocha et al. 2002). There was no evidence that 1400W given in this way had any effect on blood pressure or kidney function. The SMTC was used to cause a relatively selective blockade of nNOS and the dose used in the present study was aimed at causing an inhibition of nNOS with minimal action on eNOS. Furfine et al. (1994) have demonstrated that SMTC has a 17-fold inhibitory selectivity for nNOS as against eNOS, and at these doses. This compound has been reported to have differential actions on the renal vasculature (Ichihara et al. 1998), to selectively increase sodium reabsorption-dependent oxygen consumption (Deng et al. 2005) and to cause a decrease in the renal content of NO (Walkowska et al. 2005) at concentrations where it is probably acting preferentially on nNOS. This compound was utilized in preference to 7-NI used previously (Wu et al. 1999; Wu & Johns, 2002), which has only a 5-fold greater inhibitory action on nNOS compared with eNOS, and has to be dissolved in oil and given i.p. to obtain sufficient systemic concentrations (Wongmekiat & Johns 2001a, b, c). Again, it was apparent from the basal values that the compound did not result in any major differences in renal haemodynamic or excretory function compared with the control group of rats receiving saline, although blood pressure was higher. There have been several reports of SMTC being used in a range of studies, and all appear to report small but variable effects on blood pressure which may be dependent on the dose of drug used, type of anaesthesia and the experimental conditions (Walkowska et al. 2005).

Electrical stimulation of the left renal nerves had no consistent effect on renal blood flow or glomerular filtration rate, but resulted in a 40–50% reduction in urine flow and sodium excretion. These observations are very comparable with those we previously reported in the anaesthetized rat and rabbit (Hesse & Johns, 1984; Manitius & Johns, 1987), and those reported by others in the dog (Slick et al. 1975). Micropuncture studies have demonstrated that such low-level activation of the renal sympathetic nerves, which has no haemodynamic action, directly stimulates fluid reabsorption at the proximal tubular epithelial cells (Bello-Reuss et al. 1976; Wu & Johns 2002, 2004) and at the thick ascending loop of Henle (DiBona & Sawin, 1982). It is very likely that the whole-kidney antinatriuresis and antidiuresis in response to renal sympathetic nerve stimulation, reported herein, reflects this tubular action of the renal nerves. It is also important to point out that the action of the renal nerve stimulation on sodium and water excretion was limited to the left kidney, since over this time neither water nor sodium excretion from the right kidney was changed.

Infusion of l-NAME resulted in blood pressures which were comparable with those of control rats receiving saline, and this was important as it meant that there were no confounding effects of pressure on the level of sodium and water excretion (Granger, 1992). It was clear that following the infusion of l-NAME, the antinatriuresis and antidiuresis associated with the low-level renal nerve stimulation was prevented. This observation would suggest that the presence of NO was necessary for the nerves to exert their effect at the epithelial cells. The question arises as to whether this action represents a pre- or postsynaptic action of the NO. Interestingly, in previous reports from this laboratory, a role for NO in adrenergically mediated proximal tubular fluid reabsorption became apparent. The first indication of an interaction was that local infusion of l-NAME into the proximal tubular lumen for a few seconds resulted in a significant increase in proximal tubular fluid reabsorption (Wu et al. 1999). This suggested that the basal level of proximal tubular fluid reabsorption was under the influence of NO which exerted a tonic inhibitory action (Ortiz & Garvin, 2002). Interestingly, this action of l-NAME to increase tubular fluid reabsorption occurred only if the renal sympathetic nerves were intact. It was unclear how NO might interact with the renal nerves, but in a subsequent study it was reported that the effect of low-level renal nerve stimulation, which caused a frequency-dependent increase in proximal tubule fluid reabsorption, was also blocked by l-NAME (Wu & Johns, 2002). This finding was interpreted as indicating a potential second site of action, i.e. at the neuro-effector junction, where NO might be important in facilitating the release of noradrenaline. This latter concept was supported by the observations in the rat mesenteric bed where l-NAME blunted the noradrenaline release in response to stimulation of the sympathetic nerves (Yamamoto et al. 1994, 1997). Moreover, Tanioka et al. (2002) demonstrated not only that noradrenaline release from the isolated pump-perfused kidney was suppressed following NOS inhibition with l-NAME, but also that the nerve-mediated vasoconstrictions were attenuated. However, there is a body of evidence indicating that NO has a negative action to inhibit noradrenaline release at the neuro-effector junction and to depress adrenergically mediated actions (Egi et al. 1994; Schwartz et al. 1995; Maekawa et al. 1996; Chowdhary & Townend, 1999). Exactly why there should be such divergent reports in the literature is unclear at the present time, but it does indicate that the situation at each neuro-effector junction may be very different.

Together, the micropuncture studies (Wu et al. 1999; Wu & Johns, 2001) suggested two sites at which NO could have been exerting an action. However, there were concerns that in these micropuncture studies, because the NOS blockers were given into the tubular lumen and were present for a relatively short period, uncertainty arose as to whether there was sufficient time for the NOS enzymes to be blocked in sites beyond the epithelial cell and in the neuro-effector junction. Based on these considerations, it was hypothesized that the NO could have been acting not only on basal rates of fluid reabsorption where it had a tonic inhibitory action, but also on the noradrenaline-stimulated fluid reabsorption, where it had a facilitatory action. The question remained as to whether there might have been a presynaptic site where the NO could have been acting.

The present study was an attempt to provide further evidence for this interaction. In a whole-kidney setting, the NOS blocker l-NAME was given i.v., with the aim of providing blockade of NOS, including that involved in noradrenaline release and transmission at the neuro-effector junction. Clearly in this whole-kidney setting the presence of NO was necessary if the low-level renal nerve stimulation was to cause an antinatriuresis and an antidiuresis. These observations reinforce the earlier findings and provide supportive evidence for NO being importantly involved in allowing adrenergically mediated sodium reabsorption to occur within the kidney.

A further objective concerned the identification of the NOS isoform involved in generating the NO necessary to ensure effective adrenergic regulation of fluid reabsorption. It was evident from the results that it was unlikely to be a consequence of iNOS activity, since in the presence of 1400W, at dose levels which have been shown to inhibit iNOS activity in vivo (Garvey et al. 1997; Rocha et al. 2002), the magnitudes of the antinatriuresis and antidiuresis were the same as in the control rats infused with saline. Thus, in spite of reports that iNOS was constitutively active in the medullary regions of the kidney (Kone & Baylis, 1997), any NO generated by iNOS was unlikely to be involved in the neural control of fluid handling. The study with SMTC aimed to investigate whether nNOS provided a source for the NO. It was clear that following administration of the SMTC the magnitude of the reductions in sodium and water excretion were markedly blunted, although not totally prevented. This would suggest that the nNOS isoform was, to a degree, responsible for the generation of NO required to allow an effective action of the renal nerves on fluid reabsorption. However, the renal nerve-mediated antinatriuretic and antidiuretic responses were not completely blocked and it remains uncertain as to whether this could have been due to an insufficient dose of SMTC used or whether a component of the NO did indeed arise from activity of eNOS. This is a limitation consequent on the relative lack of selectivity of the available compounds, as even if a higher dose of SMTC were used, the possibility would still arise that the compound was at a concentration at which it would begin to block eNOS

This study investigated the interaction between NO and the renal nerves to cause a retention of sodium and water. Using the anaesthetized rat, low-level stimulation of the renal nerves decreased urine flow and sodium excretion by almost one-half. In the presence of l-NAME, the renal nerve-induced antinatriuresis and antidiuresis was blocked but this was not the case in the presence of the relatively selective iNOS blocker 1400W. In the presence of SMTC, the magnitudes of the renal nerve-mediated excretory responses were markedly reduced. Together these findings show that NO is importantly involved in allowing the renal nerves to stimulate tubular fluid reabsorption. This action of NO may be at a presynaptic as well as a postsynaptic site at the neuro-effector junction, and may involve both nNOS and eNOS isoforms.

Acknowledgments

This work was funded by a collaborative joint project grant from the Royal Society, and a project grant from the Wellcome Trust to Edward J. Johns.

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