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(Hypertension. 1997;29:1051-1057.)
© 1997 American Heart Association, Inc.

Articles

Pressure Natriuresis and Renal Medullary Blood Flow in Dogs

Dewan S. A. Majid; Murrell Godfrey; ; L. Gabriel Navar

From the Department of Physiology, Tulane University School of Medicine, New Orleans, La.

Correspondence to Dewan S.A. Majid, PhD, Department of Physiology, Tulane University School of Medicine, 1430 Tulane Ave, New Orleans, LA 70112.

	Abstract

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Abstract In the present study, we evaluated the effects of changes in arterial pressure on regional renal blood flows and sodium excretion in anesthetized dogs during control conditions and after 5% volume expansion with isotonic saline. Medullary and cortical blood flow responses were determined with laser-Doppler needle flow probes inserted into the midmedullary and midcortical regions, and whole-kidney blood flow was assessed with an electromagnetic flow probe. Volume expansion in six dogs caused marked increases in urine flow (20.2±5.5 to 82.5±22.7 µL·min^-1·g^-1) and sodium excretion (3.2±0.5 to 11.1±2.7 µmol·min^-1·g^-1), with slight increases in glomerular filtration rate (0.92±0.03 to 1.01±0.02 mL·min^-1·g^-1) but no significant changes in total renal blood flow (4.7±0.3 to 5.2±0.6 mL·min^-1·g^-1), medullary blood flow (+6±9%), or cortical blood flow (+12±10%). During stepwise reductions in renal arterial pressure (150 to 75 mm Hg) elicited with a renal arterial occluder, both before and after volume expansion, medullary, cortical, and total renal blood flows as well as glomerular filtration rate exhibited efficient autoregulation, with slopes not significantly different from zero over this range of arterial pressure. There were marked increases in the slopes of the relationships between arterial pressure and urine flow (0.18±0.05 to 0.78±0.27 µL·min^-1·g^-1·mm Hg^-1) as well as sodium excretion (0.03±0.004 to 0.10±0.03 µmol·min^-1·g^-1·mm Hg^-1) during volume expansion. These data demonstrate that medullary blood flow is efficiently autoregulated in dogs during control and volume-expanded states and indicate that the mechanism responsible for the arterial pressure–induced changes in sodium excretion does not depend on coincident alterations in medullary blood flow.

Key Words: blood flow, renal • laser-Doppler flowmetry • calcium channel blockers • natriuresis

	Introduction

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It is well known that the kidney has the ability to alter its U_NaV in response to acute changes in RAP over a range in which filtered load and total RBF do not change.^{1 2 3 4} Such a relationship between RAP and U_NaV has remained a critical aspect of many concepts concerning the long-term regulation of blood pressure.⁵ However, the precise intrarenal mechanism responsible for arterial pressure–induced natriuretic responses has not yet been clearly defined.^{1 2 3 4} Because this phenomenon of pressure natriuresis is observed in the absence of changes in filtered load and within the autoregulatory range for RBF and GFR, it is clear that changes in tubular sodium reabsorption are primarily responsible for the changes in U_NaV. Changes in intrarenal hemodynamics^{2 6 7 8} and/or paracrine factors^{3 9 10 11 12} have been suggested to serve as the possible links between arterial pressure and the changes in tubular reabsorption. Although there is clear agreement that blood flow to the renal cortex exhibits efficient autoregulation similar to that of total RBF, the autoregulatory behavior of MBF has remained controversial.^{6 7 8 13 14}

Recent advances in LDF with single-fiber probes have provided an opportunity for the study of dynamic microvascular blood flow variations in discrete regions of the kidney.^{6 13 14 15} Recent studies using this approach have reported that autoregulatory efficiency of blood flow in the renal medulla is similar to that in the whole kidney in anesthetized dogs.^{13 14} In contrast, studies in rats^{6 7} have indicated that under volume-expanded conditions, MBF autoregulatory efficiency in response to changes in arterial pressure is significantly impaired compared with CBF.

Although it has been suggested that arterial pressure–induced changes in U_NaV are associated with coincident changes in MBF,^{6 7 8} no systematic study has been conducted to evaluate the direct correlation between changes in MBF and U_NaV during alterations in arterial pressure. We designed the present study to examine simultaneously the MBF and U_NaV responses to changes in RAP in anesthetized dogs using LDF needle probes and obtain a better understanding of the role of MBF in mediating the arterial pressure–induced changes in U_NaV. Accordingly, we evaluated renal responses to changes in RAP during control conditions and after a 5% volume expansion with an infusion of isotonic saline.^{16 17 18}

	Methods

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Experiments were conducted in 17 mongrel dogs (body weight, 20.6±0.9 kg). Eight of these dogs were used for studies validating the responsiveness of the LDF needle probes to various interventions. Pressure-natriuresis experiments were conducted in 6 dogs. An additional 3 dogs were used for examination of the autoregulatory behavior of regional RBF after calcium channel blockade. All dogs were given supplemental amounts of sodium chloride (1.5 g/kg body wt per day for 3 days) added to the normal laboratory diet so that they achieved a sodium-replete state. Dogs were anesthetized with pentobarbital sodium (30 mg/kg body wt IV) and artificially ventilated. Body temperature was maintained within the normal range (99° to 101°F) with an electric heating pad. Systemic arterial pressure was measured from a catheter placed in the abdominal aorta inserted via the right femoral artery. The catheter was connected to a pressure transducer, and mean systemic arterial pressure was recorded on a polygraph (model 7D, Grass Instruments). The left femoral artery was cannulated for collection of blood samples. The femoral and jugular veins were cannulated for administration of saline, inulin solution, and additional doses of pentobarbital sodium as necessary.

The left kidney was exposed through a flank incision and the renal artery isolated from surrounding tissue. Renal denervation was performed by cutting all the visible nerves projecting to the kidney from the aorticoadrenal ganglion. RBF was measured with an electromagnetic flow probe placed on the renal artery near its origin from the aorta and connected to a square-wave flowmeter (Carolina Medical Electronics). An adjustable plastic clamp was placed on the renal artery distal to the electromagnetic flow probe to allow acute reductions in RAP. A curved 23-gauge needle cannula was inserted into the renal artery distal to the plastic clamp and was connected to a pressure transducer for measurement of RAP. Another catheter was also connected to this needle cannula for continuous infusion of heparinized saline or drug solutions at a rate of 0.4 mL/min. Urine was collected from a catheter placed in the ureter.

Two needle probes connected to a dual-channel LDF (Periflux 4001, Perimed) were used for measurement of relative blood flows in the cortical and medullary regions of the kidney.^{6 13 14 15} These probes were inserted to approximately 5 and 15 mm in depth inside the kidney mass so the tips were positioned in the midcortical and midmedullary regions, respectively.^{13 15} As previously noted, insertion of the thin probes (500 µm diameter) did not cause any perceptible damage to the kidney.^{6 13 14 15} The positions of the tips of the needle probes were confirmed at the end of each experiment by dissecting the kidney and viewing the needle tract and regions surrounding the fiber tip. These flow probes were calibrated with a standard calibration device using a motility standard that is a colloidal suspension of latex particles (10 µm microspheres). Brownian motion of the latex particles provides the standard value of 250 perfusion units. One perfusion unit is an arbitrary value equal to an analog output of 10 mV. To avoid respiratory movement artifacts in the laser-Doppler signal, we kept the kidney in a fixed position by placing it on a plastic holder similar to that used for micropuncture experiments. Basal RBF was not reduced after the kidney was immobilized. The output signals from the LDF were recorded on a Grass polygraph. Although the changes in absolute voltage signals (ie, perfusion unit values) were continuously monitored on the digital screen of the flowmeter, the data are reported as a percentage of the basal levels at spontaneous pressures. However, it was always noted that the absolute outputs from the cortical probes were substantially greater (approximately three times) than the output from the medullary probe. An example of this difference in signal is shown in Fig 1. In 17 dogs, the absolute values of control CBF averaged 364±22 perfusion units, whereas MBF was 118±10 perfusion units, reflecting the predictable differences in CBF and MBF.

View larger version (17K): [in this window] [in a new window]   Figure 1. Example of the difference in LDF needle probe signal output from cortical and medullary regions of the kidney. At the start of the experiment, when both probes were introduced to similar depth (5 mm) into the cortical tissue, absolute signals were similar (301 vs 305 perfusion units [PU]).When one probe was advanced farther to 15 mm in depth for medullary recordings, absolute signals decreased to around 78 from 305 perfusion units, indicating a predictable difference in blood flow between the cortex and medulla. EMF indicates electromagnetic flow probe, recording total RBF; SAP, systemic arterial pressure.

After completion of surgery, a 2.5% solution of inulin in normal saline was administered into the jugular vein at least 45 minutes before the onset of the experimental protocol. An initial dose of 1.6 mL/kg body wt was followed by a continuous infusion of 0.3 mL·kg^-1·min^-1. At least 45 minutes before the start of the experimental protocol, the right common carotid artery was occluded and the left common carotid artery was partially constricted to elevate the basal level of mean arterial pressure to around 150 mm Hg.

For examination of the relationship between blood flows in the kidney and pressure natriuresis in six dogs, the experimental protocol was started with urine collections for two consecutive 10-minute periods at spontaneous RAP. At the midpoint of each collection period, an arterial blood sample (2 mL) was taken for measurement of plasma inulin, sodium, potassium, and osmolar concentrations. RAP was then reduced in steps (125, 100, and 75 mm Hg) by adjusting the clamp on the renal artery. Five minutes was allowed for stabilization at each RAP level before a 10-minute urine sample was collected. After the last reduction in RAP, the clamp was released completely to reestablish control RAP and RBF. Then the dogs were subjected to extracellular volume expansion with infusion of isotonic saline (bolus dose of 200 mL and continuous infusion of 1 mL·kg^-1·min^-1); the infusion was continued for the duration of the experiment. Forty minutes after the initiation of the saline infusion, the protocol was repeated for examination of the renal responses to reductions in RAP during volume expansion.

For examination of the sensitivity of the LDF needle probes to detect changes in MBF, bolus doses of acetylcholine (1 ng to 1 µg) and Ang II (0.05 to 5 ng) were administered intrarenally in eight dogs. It was observed that the changes in MBF recorded with the LDF needle probe in response to administration of these vasoactive agents were directly correlated with the changes in total RBF recorded with an electromagnetic flow probe. Fig 2 compares the responses from LDF needle probes with those obtained from the electromagnetic flow probe. There was a close association between the changes in MBF and changes in total RBF in response to different doses of acetylcholine (Fig 2A; r=.89, P<.001) and Ang II (Fig 2B; r=.75, P<.001). Similar sensitivity of LDF probes in detecting MBF responses to various vasoactive agents in rats has been reported by Mattson et al.⁶ Consistent with the reported observations in dogs,¹⁹ it is noted that Ang II causes greater decreases in MBF than total RBF (Fig 2B). In response to Ang II doses, the percent changes in MBF were significantly greater (P<.05) than the changes in total RBF (Fig 2B), whereas MBF and RBF responses during bolus doses of acetylcholine did not differ significantly (Fig 2A).

View larger version (22K): [in this window] [in a new window]   Figure 2. Comparison of responsiveness of LDF needle probe measuring MBF and electromagnetic flow probe (EMF) measuring total RBF in response to intrarenal administration of different doses of acetylcholine (Ach, 1 ng to 1 µg) (A) and Ang II (0.05 to 5 ng) (B).

Further validation studies were performed for examination of the MBF responses during calcium channel blockade, which has been shown to cause impairment of autoregulatory efficiency in total RBF.²⁰ Felodopine (Astra Pharmaceuticals) was administered intrarenally (1 µg·kg^-1·min^-1) in three dogs. The effects of changes in RAP on total RBF, CBF, and MBF were examined before and 10 minutes after the initiation of felodopine infusion. After control measurements at spontaneous arterial pressures, RAP was reduced in steps of 10 to 20 mm Hg for 1 to 2 minutes in each step with the adjustable clamp.¹³ After the last reduction in RAP, the clamp was released to reestablish spontaneous RAP.

At the end of each experiment, the electromagnetic flow probe was calibrated in situ by collection of timed blood samples into the graduated cylinder at different flow rates from a catheter placed in the renal artery. The kidney was then removed, stripped of all surrounding tissue, blotted dry, and weighed so that the calculated parameters could be expressed per gram of kidney weight. The localization of the fiber probes was confirmed after the kidneys were removed by direct observation of the position of the tips.

Flame photometry (Instrumentation Laboratories) was used for measurement of sodium and potassium concentrations in plasma and urine. Inulin concentrations in plasma and urine samples were determined by the anthrone colorimetric technique (Gilford Instruments). Plasma and urinary osmolality were determined by vapor pressure osmometry (Wescor Inc). Urinary concentration of nitrate/nitrite (NO₃^-/NO₂^-) was measured with the Greiss reaction technique after enzymatic reductions of nitrate to nitrite in the samples.⁹ NO₃^-/NO₂^- is the metabolic end product of NO, and changes in the urinary excretion rate of NO₃^-/NO₂^- are considered to reflect changes in the production of endogenous NO, although there is a basal excretion rate dependent on dietary nitrate intake.

Values are reported as mean±SE. Statistical comparisons of differences in the responses were made by Student's paired t test. Differences in the mean values were deemed significant at a value of P.05.

	Results

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Renal Excretory and Regional Blood Flow Responses to Changes in RAP Before Volume Expansion
Control RAP was raised from 124±3.3 to 147±1.7 mm Hg by bilateral partial carotid constriction. Hematocrit during control conditions before volume expansion averaged 0.47±0.01. Plasma protein levels and colloid osmotic pressure were 5.2±0.2 g/100 mL and 16.8±0.8 mm Hg, respectively. During the control period, RBF and GFR were 4.7±0.3 and 0.92±0.03 mL·min^-1·g^-1, respectively. Urine flow, U_NaV, FE_Na, potassium excretion, and urinary excretion of NO₃^-/NO₂^- during the control periods were 20.2±5.5 µL min^-1·g^-1, 3.2±0.5 µmol·min^-1·g^-1, 2.4±0.3%, 0.86±0.12 µmol·min^-1·g^-1, and 1.4±0.1 nmol min^-1·g^-1, respectively. Urine osmolality and osmolar excretion during the control period were 669±135 mOsm·L^-1 and 10.5±1.5 µOsm·min^-1·g^-1, respectively. As previously shown,^{9 10 11 13} RBF and GFR demonstrated efficient autoregulatory behavior in response to reductions in RAP down to 75 mm Hg (Fig 3). Both MBF and CBF maintained autoregulatory efficiency similar to that of total RBF (Figs 3 and 4). The slopes of the relationships between RAP and RBF (0.002±0.002 mL·min^-1·g^-1), MBF (-0.18±0.16 %·mm Hg^-1), CBF (-0.01±0.07 %·mm Hg^-1), and GFR (-0.001±0.001 mL·min^-1·g^-1) were not significantly different from zero. As shown in Figs 5 and 6, urine flow, U_NaV, FE_Na, and the urinary excretion rate of NO₃^-/NO₂^- decreased in response to decreases in RAP. The slopes of the relationships between RAP and urine flow (0.18±0.05 µL·min^-1·g^-1), urinary excretion rate of NO₃^-/NO₂^- (0.01±0.002 nmol·min^-1·g^-1·mm Hg^-1), U_NaV (0.03±0.004 µmol · min^-1·g^-1·mm Hg^-1), and FE_Na (0.023±0.003% · mm Hg^-1) were statistically significant. These responses in renal excretory function were similar to those previously observed in our laboratory.^{9 10 11}

View larger version (14K): [in this window] [in a new window]   Figure 3. Total RBF (A) and GFR (B) responses to changes in RAP (n=6).

View larger version (14K): [in this window] [in a new window]   Figure 4. MBF (A) and CBF (B) responses to changes in RAP (n=6).

View larger version (15K): [in this window] [in a new window]   Figure 5. Urine flow (V) (A) and urinary excretion rate of nitrate/nitrite (UNO3/NO2.V) (B) responses to changes in RAP (n=6).

View larger version (14K): [in this window] [in a new window]   Figure 6. UNaV (A) and FENa (B) responses to changes in RAP (n=6).

Renal Excretory and Regional Blood Flow Responses to Changes in RAP After Volume Expansion
After volume expansion with an intravenous infusion of isotonic saline (5% of body weight) for 45 minutes, RAP did not change significantly (142±5.8 mm Hg) from control periods. Average hematocrit was significantly decreased to 0.40±0.02 (P<.05). There were also decreases in plasma protein levels (4.4±0.3 g/100 mL, P<.001) and colloid osmotic pressure (13.3±0.7 mm Hg, P<.001) after acute saline loading. The slight increases in RBF (5.2±0.6 mL·min^-1·g^-1), MBF (+6±9%), and CBF (+12±10%) were not statistically significant, whereas GFR increased slightly (1.01±0.02 mL ·min^-1·g^-1, P<.05). In contrast, there were marked increases in urine flow (82.5±22.7 µL·min^-1·g^-1, P<.05), U_NaV (11.1±2.7 µmol·min^-1·g^-1, P<.05), and FE_Na (7.9±2.0%, P<.05) during volume expansion. The urinary excretion rates of NO₃^-/NO₂^- (1.93±0.35 nmol·min^-1·g^-1, P<.05) and potassium (1.30± 0.20 µmol·min^-1·g^-1, P<.05) also increased slightly. There were significant decreases in urine osmolality (467±98 mOsm·L^-1, P<.05) and increases in osmolar excretion rate (26.0±6.8 µOsm·min^-1·g^-1, P<.05).

During reductions in RAP down to 75 mm Hg after volume expansion, the autoregulatory efficiency of RBF and GFR remained intact (Fig 3), and the slopes of the relationships between RAP and RBF (-0.002±0.002 mL·min^-1·g^-1·mm Hg^-1) as well as between RAP and GFR (-0.0007±0.0003 mL·min^-1·g^-1· mm Hg^-1) were not significantly different from zero nor from the values obtained before volume expansion. Similar autoregulatory efficiency was observed in MBF and CBF during changes in RAP (Fig 4), without any significant change in the slopes of the relationships between RAP and MBF (-0.04±0.07 %·mm Hg^-1) and CBF (-0.14±0.06 %·mm Hg^-1) from zero or the control period. However, the volume expansion elicited marked increases in the slopes of the relationships between RAP and urine flow (0.78±0.27 µL ·min^-1·g^-1·mm Hg^-1), U_NaV (0.10±0.03 µmol · min^-1·g^-1·mm Hg^-1), and FE_Na (0.06±0.02 % ·mm Hg^-1) (Figs 5 and 6). The slope of RAP versus urinary excretion of NO₃^-/NO₂^- increased slightly (0.014±0.003 nmol·min^-1·g^-1·mm Hg^-1), but this was not statistically significant (Fig 5).

Effects of Calcium Channel Blockade on Renal Autoregulation
Infusion of felodopine (1 µg·kg^-1·min^-1) in three dogs resulted in a decrease in arterial pressure from 142±3.3 to 125±2.9 mm Hg and increases in total RBF from 4.7±0.5 to 5.8±0.3 mL·min^-1·g^-1 (P<.01). There were also increases in CBF (23±4%, P<.01) and MBF (43±10%, P<.05). As reported previously,²⁰ autoregulatory efficiency of total RBF after calcium channel blockade by felodopine was impaired (Fig 7). The slope of the pressure-flow relationship within the autoregulatory range of RAP (140 to 70 mm Hg) was significantly altered (-0.0025±0.0008 to 0.023±0.007 mL ·min^-1·g^-1·mm Hg^-1) during felodopine infusion. Similar impairment of autoregulatory efficiency was also observed in CBF and MBF (Fig 7). During the control period, the slopes of RAP versus MBF as well as RAP versus CBF within the autoregulatory range were not significantly different from zero. However, during felodopine infusion, these slopes were significantly altered (0.61±0.08 and 0.60±0.12 %·mm Hg^-1, respectively) from the control period.

View larger version (21K): [in this window] [in a new window]   Figure 7. Responses to changes in RAP on total RBF (A), MBF (B), and CBF (C) during calcium channel blockade by intrarenal administration of felodopine (1 µg·kg-1·min-1, n=3). Mean curves for RBF, CBF, and MBF autoregulation were generated by extrapolating blood flow values at different RAP levels (at 20–mm Hg intervals ranging from 10 mm Hg to highest spontaneous pressure) from each individual curve obtained in each dog. RBF was recorded by electromagnetic flow probe; CBF and MBF, by LDF.

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
In the present study, arterial pressure–induced changes in urine flow and U_NaV were observed under conditions of highly efficient autoregulation of blood flow to the renal cortex and medulla, measured simultaneously with LDF needle probes. The changes in U_NaV observed in the present study were similar to those reported previously by our laboratory.^{9 10 11} The MBF responses are consistent with recent studies that used LDF optical fiber probes.^{13 14} In further experiments, we also demonstrated that the autoregulatory efficiency of MBF could be altered (Fig 7) when whole-kidney blood flow autoregulation was impaired by calcium channel blockade in dogs.²⁰ Thus, this technique is able to detect changes in MBF in response to changes in arterial pressure when they occur. Although another recent study in anesthetized dogs⁸ indicated that MBF was not autoregulated with high efficiency, it should be considered that MBF measurements in that study were made with a multifiber LDF probe placed on the surface of the medulla exposed after excision of renal tissue. With the needle probes used in the present study, surgical excision was not required to provide access to the medulla, and it was thus possible to observe simultaneously the MBF and urinary excretory responses to changes in RAP.

Although LDF has provided the means to study dynamic regional blood flow responses within specific sites in an organ, some potential limitations of this technique remain. One concern is that measurements with the LDF needle probe do not provide an absolute measure of blood flow. However, the relative changes in blood flow signals from the LDF needle probes in response to various vasoactive agents were found to be closely correlated (Fig 2 and References 6, 13, and 15^{6 13 15} ) to the changes in total RBF. Moreover, it was observed that the LDF signals obtained from the cortical tissue were substantially greater (approximately three times) than the signals from medullary tissue (Fig 1), in general agreement with the well-recognized differences in CBF and MBF when expressed per gram of tissue (for review, see Reference 21²¹ ). Thus, the available data indicate that LDF methodology can provide a good indication of changes in regional blood flow within renal tissue. However, signals from the LDF needle probe also depend on a number of factors, such as hematocrit, tissue morphology, and vessel alignment.²² It is possible that changes in hematocrit affect the magnitude of the LDF signals, as this depends on red blood cell flux (numberxvelocity of red blood cells) in the volume of tissue sampled by the LDF probe. In the present study, hematocrit clearly decreased during acute saline loading, and this could have diminished the relative changes observed after volume expansion. However, hematocrit values did not change during stabilized periods when responses to stepwise reductions in RAP were examined both before and after volume expansion. It is also possible that alterations in the vascular component of the unit volume sampled by LDF needle probes could lead to a change in the magnitude of LDF signals. However, total RBF remained unchanged within the autoregulatory range, and net renal volume did not change perceptibly during alterations in RAP. Thus, a change in unit volume sampled by the LDF probe may be insignificant in producing a discernible change in LDF signals. However, direct assessment of the implications of various physical factors in the renal tissue on the absolute LDF signals may not be possible in the context of the present study.

Studies in rats^{6 7 23} reported that during volume expansion with isotonic saline, the autoregulatory efficiency in renal MBF was reduced, as recorded by LDF. As these volume-expanded rats exhibited an exaggerated pressure-natriuresis response, it was postulated that the changes in MBF may mediate the enhanced natriuretic responses to changes in arterial pressure. However, direct correlations between the changes in MBF and U_NaV in response to alterations in RAP were not shown.^{6 7 23} Little information is available related to MBF responses to volume expansion in dogs. In the present study, we did not observe a significant increase in MBF during volume expansion in dogs. In addition, MBF exhibited efficient autoregulation similar to that seen during control measurements. At present, we have no satisfactory explanation for the differences between the results obtained in dogs and rats. Species differences in medullary morphology could help explain these contrasting findings. The long papilla of the rat, which is distinct from that in the dog, may exhibit different hemodynamic responses.^{6 7 23} It can be argued that differences in methodological approaches may be partially responsible for the differences between the findings in the dog and rat. In the present study, needle probes were inserted directly into the tissue, whereas the previous studies in rats relied on implanted optical fibers.⁶ However, it seems unlikely that this factor is responsible for the differences observed because both the needle probes and the optical fibers should be able to respond to the same volume of tissue in front of the tip. Moreover, MBF measured with optical fiber probes in anesthetized dogs¹⁴ also showed efficient autoregulatory behavior in agreement with the present study. We also demonstrated that the changes in MBF recorded by LDF needle probes in response to various vasoactive agents were closely correlated with the changes in total RBF recorded with electromagnetic flow probes (Fig 2). Mattson et al⁶ reported similar sensitivity with LDF optical fiber probes in rat MBF. The needle probes were clearly able to reflect changes in MBF autoregulatory behavior when they occurred during calcium channel blockade (Fig 7) or when RAP was reduced below the autoregulatory range.¹³ Finally, studies conducted in our laboratory have also demonstrated that intrarenal administration of other vasodilator agents—bradykinin, angiotensin-converting enzyme inhibitors,²⁴ and adrenomedullin¹⁵ —elicit increases in MBF.

Another factor that should be considered is that possible elevations of circulating levels of catecholamines, vasopressin, plasma renin activity, or Ang II in these dogs subjected to partial occlusion of the common carotid arteries may have influenced the MBF responses in the present study. Although we did not measure the circulating levels of these hormones in this study, it should be noted that we performed partial carotid constriction to raise systemic arterial pressure at least 45 minutes before the start of the experimental protocol and maintained this occlusion throughout the experimental period. During this period, systemic arterial pressure remained stable. Thus, it seems unlikely that subtle fluctuations in the release of catecholamines and vasopressin might influence the MBF response to selective changes in renal perfusion pressure elicited with a renal arterial occluder. Previous studies in rats⁷ have demonstrated that arterial pressure–dependent changes in MBF can be observed in animals maintained at fixed high levels of these hormones achieved by their continuous infusion. It is also unlikely that a vasoconstrictor effect of increases in intrarenal Ang II levels on MBF in these dogs subjected to partial carotid occlusion influenced the MBF responses because such effects would have led to progressive decreases in MBF with decreases in renal perfusion pressure and would have counteracted rather than been responsible for the high degree of autoregulatory efficiency observed. In addition, the dogs were given a high sodium diet before the experiment and then subsequently were infused with a large isotonic saline load. Clearly, the dogs used in this study were not hypovolemic, as they exhibited high FE_Na, confirming that they were in a sodium-replete state. The marked reductions in plasma protein concentrations (15%) and colloid osmotic pressure in response to saline loading, which were comparable to the changes observed in a previous study,¹⁸ clearly demonstrated the adequacy of the volume expansion in this group of dogs.

Several mechanisms have been considered to explain the loss of MBF autoregulatory behavior in volume-expanded rats.^{6 7 18} It has been reported that the loss of MBF autoregulatory efficiency in rats was restored by inhibition of NO synthase.²⁵ A possible role for prostaglandins was also indicated, as it was shown that indomethacin treatment also restored MBF autoregulatory efficiency in rats.²⁶ Thus, both of these paracrine factors may play a role in determining the autoregulatory ability of MBF in rats. In studies by Salazar et al¹⁷ and Alberola et al¹⁶ in anesthetized dogs, it was reported that both prostaglandins and NO were involved in mediating sodium excretory responses to acute volume expansion with isotonic saline. In the present study, volume expansion also caused enhancement of endogenous NO production, as evidenced by slight but significant increases in the urinary excretion rate of NO₃^-/NO₂^-. This slight increase in NO production could have contributed partly to the increases in urine flow and U_NaV. Although some studies in rats²⁷ indicate that intrarenal production of NO may be greater in the renal medulla than in the cortex, this has not been examined in dogs. Recently, we have reported¹⁵ that inhibition of NO synthesis in anesthetized dogs resulted in similar reductions in both CBF (28%) and MBF (27%) in the kidney. Of interest, a recent study in rats by Mohaupt et al²⁸ has shown that the inducible NO synthase isoform is constitutively expressed in abundance in the medullary thick ascending limb of the loop of Henle. Thus, the medullary ascending limb of the loop of Henle could be a source of the high NO levels, which may be trapped more efficiently in the medulla of the rat.

Although the mechanism of pressure natriuresis is not yet clearly defined, several recent studies^{9 10 11 12} have suggested that changes in intrarenal NO activity during changes in arterial pressure may be involved in mediating the pressure-natriuresis response. This is supported further by the present findings showing a significant correlation between RAP and urinary excretion rate of NO₃^-/NO₂^- during both control and volume-expanded conditions. Increases in NO release during increases in RAP could occur as a consequence of increases in shear stress in the vessel wall during autoregulatory adjustments in arteriolar resistance.^{29 30} The increased NO production would raise the tissue levels³¹ and could thus serve as an important mediator of the natriuretic responses by influencing tubular transport directly^{32 33} and/or altering the intrarenal hemodynamic environment.^{2 27 34} Changes in renal interstitial hydrostatic pressure in response to changes in RAP have been suggested to be involved in mediating the change in tubular transport.^{2 28} As we did not measure the changes in renal interstitial hydrostatic pressure in this study, however, we cannot provide data regarding changes in this parameter under conditions in which there is highly efficient autoregulation of total RBF, CBF, and MBF.

In conclusion, the results of the present investigation demonstrate that the changes in U_NaV in response to alterations in arterial pressure both during a control sodium-replete state and after acute extracellular volume expansion occur in the presence of efficient autoregulation of renal MBF as well as CBF in dogs. Thus, the data indicate that the mechanism of pressure natriuresis does not depend on coincident changes in MBF.

	Selected Abbreviations and Acronyms

 

Ang II = angiotensin II CBF = cortical blood flow FENa = fractional excretion of sodium GFR = glomerular filtration rate LDF = laser-Doppler flowmetry MBF = medullary blood flow NO = nitric oxide RAP = renal arterial pressure RBF = renal blood flow UNaV = urinary sodium excretion

	Acknowledgments

 
This study was supported by a Young Investigator Grant from the National Kidney Foundation, a grant from the Louisiana Education Quality Support Fund (LEQSF), and grants HL-51306 and HL-18426 from the National Heart, Lung, and Blood Institute, National Institutes of Health. The authors are grateful to George Prophet for technical assistance and Agnes C. Buffone for preparing the manuscript.

Received September 19, 1995; first decision December 4, 1995; accepted July 15, 1996.

	References

Top Abstract Introduction Methods Results Discussion References

 
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