AJP - Regu Fuel your research with LabChart
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Am J Physiol Regul Integr Comp Physiol 279: R769-R777, 2000;
0363-6119/00 $5.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (43)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Zou, A.-P.
Right arrow Articles by Cowley, A. W.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Zou, A.-P.
Right arrow Articles by Cowley, A. W., Jr.
Vol. 279, Issue 3, R769-R777, September 2000

alpha 2-Adrenergic receptor-mediated increase in NO production buffers renal medullary vasoconstriction

Ai-Ping Zou and Allen W. Cowley Jr.

Department of Physiology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

The present study was designed to investigate the role of nitric oxide (NO) in modulating the adrenergic vasoconstrictor response of the renal medullary circulation. In anesthetized rats, intravenous infusion of norepinephrine (NE) at a subpressor dose of 0.1 µg · kg-1 · min-1 did not alter renal cortical (CBF) and medullary (MBF) blood flows measured by laser-Doppler flowmetry nor medullary tissue PO2 (PmO2) as measured by a polarographic microelectrode. In the presence of the NO synthase inhibitor nitro-L-arginine methyl ester (L-NAME) in the renal medulla, intravenous infusion of NE significantly reduced MBF by 30% and PmO2 by 37%. With the use of an in vivo microdialysis-oxyhemoglobin NO-trapping technique, we found that intravenous infusion of NE increased interstitial NO concentrations by 43% in the renal medulla. NE-stimulated elevations of tissue NO were completely blocked either by renal medullary interstitial infusion of L-NAME or the alpha 2-antagonist rauwolscine (30 µg · kg-1 · min-1). Concurrently, intavenous infusion of NE resulted in a significant reduction of MBF in the presence of rauwolscine. The alpha 1-antagonist prazosin (10 µg · kg-1 · min-1 renal medullary interstitial infusion) did not reduce the NE-induced increase in NO production, and NE increased MBF in the presence of prazosin. Microdissection and RT-PCR analyses demonstrated that the vasa recta expressed the mRNA of alpha 2B-adrenergic receptors and that medullary thick ascending limb and collecting duct expressed the mRNA of both alpha 2A- and alpha 2B-adrenergic receptors. These subtypes of alpha 2-adrenergic receptors may mediate NE-induced NO production in the renal medulla. We conclude that the increase in medullary NO production associated with the activation of alpha 2-adrenergic receptors counteracts the vasoconstrictor effects of NE in the renal medulla and may play an important role in maintaining a constancy of MBF and medullary oxygenation.

renal hemodynamics; laser-Doppler flowmetry; kidney; rat


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

THERE IS A LARGE BODY OF EVIDENCE indicating that renal sympathetic nerves and adrenergic receptors participate in the control of total renal blood flow (RBF), glomerular filtration rate (GFR), and sodium and water excretion (2, 6, 8, 10). However, the adrenergic regulation of renal medullary blood flow (MBF) is poorly understood. In the dog kidney, the tissue norepinephrine (NE) content was found highest in the juxtamedullary area when dissected and measured along the renal corticomedullary axis. It was observed that the adrenergic nerve fibers travel with the vasa recta (VR) throughout the outer medulla and decrease in number as they approach the inner medulla (16). These morphological studies provided evidence for the rich adrenergic innervation of the medullary vasculature. In a study using the 86Rb-uptake method to measure RBF distribution, renal denervation was found to lead to a 16% increase in cortical blood flow (CBF) and 48% increase in MBF, indicating greater adrenergic tone in the medullary circulation. Renal nerve stimulation (RNS) produced a greater decrease in MBF than CBF (11), and it had little effect on cortical but markedly constricted juxtamedullary efferent arterioles, which supply blood to the renal medulla (4). In light of the high sympathetic activity, renal medullary circulation may require a strong counteracting system to buffer adrenergic vasoconstriction and to adjust the metabolic supply and demand toward a level of transport activity appropriate for the oxygen and substrate availability of the medullary tissue (6, 30).

Recent studies have indicated that nitric oxide (NO) modulates alpha -adrenergic vasoconstriction in a number of blood vessels. There is evidence that NE stimulates the NO production in isolated microvessels and endothelial cells and inhibition of the NOS activity markedly enhances the vascular reactivity to NE (13, 18). In the kidney, the high doses of NO synthase (NOS) inhibitors have been reported to accentuate the acute hemodynamic responses to renal nerve stimulation and NE infusion, suggesting that NO may play an important role in counteracting the effects of sympathetic activation and circulating NE (1, 22, 27). More specifically, there is evidence suggesting that NO may be of particular importance in buffering the vasoconstrictor actions of sympathetic stimulation within the renal medullary circulation. In isolated perfused outer medullary VR, Sai et al. (29) have shown that NE produced a concentration-dependent vasoconstriction and that Nomega -nitro-L-arginine, an NOS inhibitor, significantly enhanced NE-induced vasoconstriction. Furthermore, studies in our laboratory have indicated that the NOS protein amount and enzyme activity and NO concentrations {[NO]} are greater in the renal medulla compared with the renal cortex (15, 34). On the basis of these observations, we hypothesize that renal medullary NO counteracts the vasoconstrictor effect of adrenergic activation, thereby protecting the renal medulla from vasoconstrictive and ischemic injury. This counteracting action of NO is of particular importance in this region of the kidney, which is relatively underperfused and vulnerable to ischemic injury.

To test this hypothesis, the present study examined the effects of a moderate reduction of the medullary NOS activity on NE-induced vasoconstriction using laser-Doppler flow (LDF) techniques. A microdialysis-oxyhemoglobin NO-trapping technique was used to determine associated changes in medullary and cortical tissue [NO]. With the use of adrenergic receptor inhibitors we also determined the role of alpha 1- and alpha 2-adrenergic receptors in mediating the actions of NE on renal MBF and medullary NO production. On the basis of the results of alpha 1- and alpha 2-blockade, we went on to localize the subtypes of alpha 2-adrenergic receptors along the nephron and renal vascular tree using microdissection and RT-PCR techniques, thereby defining the possible subtypes of alpha 2-adrenergic receptors mediating medullary NO production.


    MATERIALS AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Animal preparation for medullary flowmetry and microdialysis. Male Sprague-Dawley rats (purchased from Harlan Sprague Dawley, Madison, WI) weighing between 250 and 300 g were fasted overnight but allowed free access to water. They were anesthetized with ketamine (30 mg/kg body wt im) and inactin (50 mg/kg body wt ip) and placed on a thermostatically controlled warming table to maintain body temperature at 37°C. After tracheotomy, cannulas were placed in the right femoral vein and artery for intravenous infusions and measurements of arterial pressure. An abdominal incision was made, and the left kidney was placed in a stainless steel cup to stabilize the organ for implantation of optical fibers to measure CBF and MBF or for implantation of microdialysis probes to dialyze NO from the renal interstitium as previously described (34, 35). For renal medullary interstitial infusion of drugs, a three-channel dialysis probe that contained an infusion inlet was implanted into the renal medulla. This dialysis probe (Bioanalytical Systems, West Lafayette, IN) was constructed with an inlet and outlet channel for perfusion of the microdialysis fluid as described below. After implantations, a 0.9% solution of sodium chloride was infused continuously at a rate of 0.5 ml/h to maintain the patency of interstitial infusion (35). The animals received an intravenous infusion of 2% bovine serum albumin in a 0.9% sodium chloride solution at a rate of 3 ml/h throughout the experiment to replace fluid losses and maintain a stable hematocrit of ~43 ± 3%.

LDF and polarographic measurement of medullary tissue oxygen. Experiments were performed to evaluate the effect of renal medullary interstitial infusion of the NOS inhibitor nitro-Larginine methyl ester (L-NAME) on renal cortical and medullary response to NE. The rats were anesthetized and surgically prepared as described in Animal preparation for medullary flowmetry and microdialysis. LDF (model Pf3, PERIMED KB) were used to simultaneously determine the changes in CBF and MBF. For measurement of changes in regional blood flows, we constructed one optical fiber (diameter of 500 µm), which was implanted in the renal cortex (1.5 mm depth), and another, which was in the inner medulla (5 mm depth) as described previously (35). The implanted fibers were optically connected to an external probe specifically designed for such applications using fused silica matching liquid (#50350, Cargille Laboratories, Cedar Grove, NJ) to minimize loss of light at the connection. The laser-Doppler signal, which is the product of the number of moving red blood cells and their velocity, was thereby used as an index of changes of blood flow in the different regions of the kidney.

To measure tissue PO2 in the renal medulla, a calibrated oxygen microelectrode with diameter of 50 µm was inserted into the renal medulla to a depth of 5 mm. An Ag/AgCl reference electrode (A-M System, Everett, WA) was attached to the muscle in the abdominal wall. The microelectrode was polarized at -0.7 V, and the currents were amplified, digitized, and displayed in millimeters of mercury by a two-channel polarographic amplifier (model 1900, A-M System). The amplifier was connected to a data-acquisition and processing system (DATAq Instruments, Akron, OH). We constructed PO2 microelectrodes using a platinum wire (length 5 cm, diameter 50 µm) insulated with epoxide resin and soldered to a gold pin connector (A-M System) as described previously (35).

After surgery and a 60-min equilibration period, continuous measurements of mean arterial pressure (MAP), CBF and MBF and medullary PO2 were obtained throughout the experiment using a digital online monitoring system. Saline was infused into the renal medullary interstitium for two 20-min control periods. At the end of the second control period, NE at a dose of 0.1 µg · kg-1 · min-1 was infused intravenously for 30 min and arterial pressure, flows, and PO2 were recorded. Then, L-NAME (1.4 µg · kg-1 · min-1), prazosin (10 µg · kg-1 · min-1), or rauwolscine (30 µg · kg-1 · min-1) was infused into the renal medullary interstitium. After 30 min, intravenous infusion of NE was begun, and arterial pressure, blood flows, and PO2 were continuously recorded to determine the influence of NOS inhibition on the effects of NE. The infused concentration of L-NAME was determined in preliminary studies to be one that would not reduce MBF more than 5-10% from control level (35). The objective of the NOS inhibition in this protocol was to blunt the NE-stimulated production of NO, not to totally inhibit NO production.

In vivo microdialysis. In vivo microdialysis studies in the renal medulla and cortex of rats were performed as we have described in a recent study (34). Briefly, the rats were anesthetized and surgically prepared as described in Animal preparation for medullary flowmetry and microdialysis. The microdialysis probes had 0.5 mm tip diameter and a 20-kDa transmembrane diffusion cutoff (Bioanalytical Systems, West Lafayette, IN). One was inserted into the renal cortex to a depth of 1.5 mm and another into the renal medulla (5 mm depth). The probes were perfused at a rate of 2 µl/min with 50 mM phosphate-buffered saline containing 3 µM oxyhemoglobin [Human AO hemoglobin (ferrous), Sigma Chemical, St. Louis, MO]. After a 1.5-h equilibration period, dialysate fluid was collected at 30-min intervals for a 1-h control measurement period with the medullary interstitial infusion of drug vehicle (isotonic saline). In one group of rats, NE was then intravenously infused at a dose of 0.1 µg · kg-1 · min-1. After 30 min, two 30-min dialysate samples were collected. In other groups of rats, L-NAME (1.4 µg · kg-1 · min-1), prazosin (10 µg · kg-1 · min-1), or rauwolscine (30 µg · kg-1 · min-1) was infused into the renal medulla for 1 h, and a 30-min dialysate sample was collected. Then, NE was intravenously infused, and two 30-min dialysate samples were collected.

Spectrophotometric assay of NO-induced methemoglobin formation in the dialysates was performed as described previously (34). The samples (50 µl) were added into a quartz cuvette and analyzed to record the different absorbance spectra using a wavelength-scanning mode of Du-640 Beckman spectrophotometer (Beckman Instruments, Schaumburg, IL). Methemoglobin concentration ([MetHb]) or [NO] was calculated according to the equation: c = A/epsilon b, where c is [MetHb] or [NO], A is absorbance increase at 401 nm (absorbance difference between 401 and 411 nm), epsilon  is extinction coefficient of MetHb, and b is light path in centimeters.

Microdissection of renal vascular and tubular segments. Microdissection was performed as we described previously (23, 31). Briefly, Sprague-Dawley rats weighing between 250 and 300 g were anesthetized with pentobarbital sodium (80 mg/kg body wt ip), and the aorta below left renal artery was isolated and cannulated. After ligating the aorta at a site between the origin of the left and right renal arteries, the left kidney was flushed with 20 ml ice-cold dissection solution containing (in mM) 135 NaCl, 3 KCl, 1.5 CaCl2, 1 MgSO4, 2 KH2PO4, 5.5 glucose, 5 L-alanine, and 5 HEPES (pH 7.4). Then, the kidney was perfused with 10 ml of the digestion solution, which was prepared by adding 1 mg/ml collagenase (243 U/mg) and 1 mg/ml bovine albumin in the dissection solution. After perfusion, the kidney was removed and cut into 1- to 2-mm-thick sections containing the entire corticomedullary axis. The sections were incubated at 37°C for 30 min in the same digestion solution with gentle shaking. During incubation, the samples were bubbled with 100% O2. The sections were then rinsed twice with collagenase-free dissection solution and transferred into Petri dishes filled with ice-cold dissection solution containing 0.1 mg/ml trypsin inhibitor and 20 µg/ml aprotinin. A Petri dish was mounted on the microscope stage and maintained at 4°C during dissection.

Microdissection was performed under a LEICA MZ8 stereomicroscope with dark-field illumination. The following renal vascular and tubular segments were dissected, and the length of these segments was measured with a calibrated eyepiece micrometer: arcuate artery (ArA), interlobular artery (IA), afferent arteriole (AA), VR, proximal convoluted tubule (PCT), proximal straight tubule (PST), cortical thick ascending limb (CTAL), medullary thick ascending limb (MTAL), cortical collecting duct (CCD), and medullary collecting duct (MCD). The glomeruli (Glm) were counted under microscope. The time period for dissection was limited to 1 h. In general, 20 Glm, 20 mm each of tubular segments, and 10 mm each of vascular segments were separately pooled to give one sample. For visualization of renal microvessels, 1-ml polyethylene beads with diameter of 0.2 µm were substituted for digestion solution during perfusion of the kidney.

RNA extraction and RT-PCR of alpha 2-adrenergic receptors. To extract total RNA, the microdissected segments were transferred into individual tubes containing 450 µl TRIzol reagent (GIBCO, Life Technologies), then incubated at room temperature for 5 min. Total RNA was extracted, precipitated, and washed according to the protocol described by the manufacturer. The resultant RNA was resuspended in 8 µl of RNase-free water.

A first-strand cDNA synthesis kit (Pharmacia Biotech) was used to synthesize cDNA by RT from mRNA. As described by the instruction of the manufacturer, 8 µl of total RNA were heated at 65°C for 10 min, rapidly chilled on ice, then mixed with 7 µl of the reagents supplied with the kit. The reaction mixture contained 0.2 µg random hexadeoxy nucleotides, 45 mM Tris (pH 8.3), 68 mM KCl, 15 mM dithiothreitol, 9 mM MgCl2, 0.08 mg/ml BSA, and 1.8 mM dNTPs and 100 U M-MuLV RT. The reaction mixture was incubated at 37°C for 60 min and then heated to 65°C for 10 min to inactivate the RT activity and to denature cDNA hybrids.

PCR reactions were performed in a total volume of 50 µl using a PCR Supermix kit (GIBCO) containing: 22 mM Tris · HCl (pH 8.4), 55 mM KCl, 1.65 mM MgCl2, 200 µM dNTPs, 5 µl RT reaction mixture, 22 U recombinant Taq DNA polymerase, and 400 pmol of the specific primer pairs for alpha 2A-, alpha 2B-, alpha 2C-receptor, or beta -actin. The reactions were cycled 30 times from 94°C for 1 min to 58°C for 1 min and then 72°C for 1.5 min. Samples were incubated at 72°C for an additional 5 min after last cycle was completed. alpha 2A-, alpha 2B-, and alpha 2C-adrenergic receptors and beta -actin primers spanned fragments of 276, 712, 370, and 350 bp from their respective cDNAs (14, 33). Negative control PCRs with a subsitution of dissection solution or total RNA without RT reaction were performed in parallel.

The structure of the primers was as follows: alpha 2A: sense 5'-CAT CAG CCT TGA CCG CTA CTG, antisense 5'-CGC ACG TAG ACC AGG ATC ATG; alpha 2B: sense 5'-CTC ATC ATC CCT TTC TCT CTG; antisense 5'-CCT CTT CAT CTC CCT CCT CTG; alpha 2C: sense 5'-CCT TCC CGC CTC TCG TCT CTT; antisense 5'-TCT CGC CGC CGT CCT CCT CTG; and beta -actin: sense 5'-AAC CGC GAG AAG ATG ACC CAG ATC ATG TTT; antisense 5'-AGC AGC CGT GGC CAT CTC TTG CTC GAA GTC. All primers were synthesized by Operon Technologies (Alameda, CA).

PCR products were separated by a 1.5% agarose gel electrophoresis (200 V for 1 h) in 1 × Tris-borate-EDTA (TBE) buffer, stained with ethidium bromide (0.5 µg/ml) and visualized under ultraviolet light, and a photograph made. The identity of the PCR products of alpha 2-adrenergic receptors was determined by cloning and sequencing (23).

Cloning and sequencing. The RT-PCR products of alpha 2-adrenergic receptors were ligated into pCR II vector (Invitrogen, San Diego, CA), and the subsequent plasmid DNA was purified using an ion-exchange column (QIAGEN, Chatsworth, CA). The plasmid DNA was digested with EcoR I to confirm the positive clones. The insert in these positive clones was sequenced using the dideoxynucleotide-chain termination reaction. The reaction samples were resolved on a DNA sequencer 725 (Molecular Dynamics).

Statistical analysis. Data are presented as means ± SE. The significance of differences within and between multiple groups was evaluated using one-way ANOVA followed by a post hoc test (Duncan's multiple-range test; SigmaStat, San Rafael, CA). A P value <0.05 was considered statistically significant.


    RESULTS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Effect of NE on MBF and PO2 before and during L-NAME infusion. The effects of renal medullary interstitial infusion of L-NAME on NE-induced changes in CBF and MBF are presented in Fig. 1. In control periods, the LDF signals from the fibers implanted in the renal cortex and medulla averaged 1.38 ± 0.18 and 0.58 ± 0.04 V, respectively. Medullary PO2 (PmO2) averaged 28.7 ± 3.8 mmHg. Intravenous infusion of NE (0.1 µg · kg-1 · min-1) had no significant effect on cortical LDF signal, medullary LDF signal, or PmO2 (n = 6). L-NAME infused into the renal medullary interstitial space at a dose of 1.4 µg · kg-1 · min-1 had no measurable effects on cortical and medullary LDF signals, PmO2, or arterial pressure (n = 7). However, this degree of NOS inhibition enhanced the vasoconstrictor effects of NE in the renal medulla. Medullary LDF signal was decreased significantly from 0.53 ± 0.04 to 0.37 ± 0.03 V and PmO2 from 25.4 ± 5 to 16.7 ± 4.3 mmHg (n = 6). MAP and cortical LDF signal were not significantly changed by intravenous NE infusion in the presence or absence of L-NAME in the renal medulla.


View larger version (23K):
[in this window]
[in a new window]
  		Fig. 1.   Effect of renal medullary interstitial infusion of nitro-L-arginine methyl ester (L-NAME) on norepinephrine (NE) response of cortical and medullary blood flows and medullary oxygenation. Cortical and medullary blood flows were measured by using laser-Doppler flowmetry (LDF), and medullary tissue oxygen was measured by a microelectrode polarographic technique. BP, mean arterial pressure; PmO2, medullary oxygen tension; C, control period; post, post control. * Significant difference from C.

Changes in tissue [NO] in the renal cortex and medulla. The results of these experiments are presented in Fig. 2. [NO] determined by microdialysis-oxyhemoglobin trapping assay averaged 94.8 ± 16 and 138.6 ± 16.7 nM, respectively, in the renal cortex and medulla (P < 0.05). NE intravenous infusion increased tissue [NO] by 28% in the renal cortex and 43% in the renal medulla. In the group of rats receiving the medullary interstitial infusion of L-NAME, basal tissue [NO] were decreased by 24% in the renal cortex (n = 7) and 26% in the renal medulla (n = 7). Under these conditions, the NE-induced increase in tissue [NO] was completely blocked in the renal cortex and medulla. MAP remained unchanged throughout all of the microdialysis studies.


View larger version (21K):
[in this window]
[in a new window]
  		Fig. 2.   Effect of renal medullary interstitial infusion of L-NAME on NE-induced change in NO concentrations in the renal cortex and medulla. Tissue NO concentrations were measured by microdialysis and hemoglobin-trapping technique. Oxyhemoglobin at a concentration of 3 µM was used to perfuse microdialysis probes and trap tissue NO from the renal cortex and medulla. One of the probes was implanted in the renal cortex at a depth of 1.5 mm and another in the renal medulla at a depth of 5 mm. RI, renal medullary interstitial infusion. * Significant difference from control periods. triangle  Significant difference from the values obtained before medullary interstitial infusion of L-NAME.

Effects of alpha 1- and alpha 2-receptor blockade on NE response of MBF and medullary NO. The results of these experiments are presented in Fig. 3. Renal medullary interstitial infusion of the alpha 1-receptor antagonist prazosin at a dose of 10 µg · kg-1 · min-1 slightly increased medullary LDF signal and [NO]. In the presence of prazosin in the renal medulla, intravenous infusion of NE still significantly increased medullary LDF signal and [NO] (n = 7). Renal medullary interstitial infusion of alpha 2-receptor antagonist rauwolscine at a dose of 30 µg · kg-1 · min-1 significantly decreased medullary LDF signal and [NO]. In the presence of rauwolscine in the renal medulla, intravenous infusion of NE further reduced medullary LDF signal, whereas [NO] remained unchanged (n = 8). MAP and cortical LDF signal were not altered throughout the experiment.


View larger version (25K):
[in this window]
[in a new window]
  		Fig. 3.   Effects of blockade of alpha 1- and alpha 2-adrenergic receptors on NE response of medullary blood flow and medullary NO. The alpha 1-antagonist prazosin (10 µg · kg-1 · min-1) or alpha 2-antagonist rauwolscine (30 µg · kg-1 · min-1) was infused into the renal medullary interstitium, respectively. * Significant difference from C. triangle  Significant difference from the values obtained before NE infusion.

Expression of alpha 2-adrenergic receptors in renal cortical and medullary tissues and microdissected renal segments. Figure 4 illustrates a representative photograph of ethidium bromide-stained RT-PCR products of alpha 2-adrenergic receptors and beta -actin in the renal cortex, outer medulla, and papilla. Three subtypes of alpha 2-adrenergic receptors were detected in the total RNA extracted from the renal cortex, outer medulla, and papilla, and the signal intensity for each receptor was similar in these three kidney regions, suggesting that the primers and reaction condition were optimized for the analysis of the distribution of these receptors in different regions using RT-PCR. The sizes of the RT-PCR products of alpha 2A-, alpha 2B-, and alpha 2C-receptors were 276, 712, and 370 bp, respectively, which were identical to the predicted sizes based on their cDNA sequences (14, 33). Direct addition of total RNA from the renal cortex, outer medulla, and papilla into PCR reaction without reverse transcription did not produce any signal, suggesting that PCR products are derived from cDNA synthesized by RT. Sequence analysis confirmed that the RT-PCR products detected in the present study represent corresponding receptor genes. The product of alpha 2A-receptor had 97% homology with a known rat liver alpha 2A-cDNA sequence in GeneBank, alpha 2B-receptor had 99.3% homology with a known rat kidney alpha 2B-cDNA sequence, and alpha 2C-receptor had 95.2% similarity to a known rat liver alpha 2C-cDNA (14, 33).


View larger version (41K):
[in this window]
[in a new window]
  		Fig. 4.   RT-PCR products of alpha 2-adrenergic receptors in the renal cortex and medulla. Expected sizes of the products were 276 bp for alpha 2A, 712 bp for alpha 2B, and 370 bp for alpha 2C-adrenergic receptors. (-), PCR without reverse transcription; P, papilla, M, outer medulla; C, cortex. The cDNA products were subjected to 1.5% agarose gel separation and stained by 0.5 µg/ml ethidium bromide.

Figure 5 shows a representative expression profile of three alpha 2-receptors in microdissected intrarenal vascular segments and Glm (n = 5 rats). The signal of each lane on the gel represents the RT-PCR product from 0.5 mm of dissected vessels or one glomerulus. Three subtypes of alpha 2-receptor mRNA were detected in ArA and IA, both alpha 2A- and alpha 2B-receptor mRNAs were demonstrated in afferent arterioles and Glm, and only alpha 2B-receptor mRNA was found in postglomerular VR. The sizes of RT-PCR products of three receptors were the same as those detected in cortical and medullary tissues. The RT-PCR product of beta -actin was detected with a predicted size of 350 bp in all vascular segments and Glm. When RNAs from microdissected vascular segments and Glm were directly added into the PCR reaction without reverse transcription, no signal was detected. An example of negative control without reverse transcription using RNA from ArA was shown in lane RT (-) in Fig. 5.


View larger version (58K):
[in this window]
[in a new window]
  		Fig. 5.   RT-PCR products of alpha 2-adrenergic receptors and beta -actin in the microdissected renal pre- and postglomerular vessels. Labels of molecular sizes on the right of photo are predicted sizes of the products. RT(-) indicates PCR without reverse transcription; Glm, glomerulus; AA, arcuate artery, IA, interlobular artery; Aff.A, afferent arteriole; VR, vasa recta. Gel separation and staining of the cDNA products were same as those in Fig. 4.

A representative distribution pattern of three alpha 2-receptors in isolated tubular segments is presented in Fig. 6. The signal of each lane on the gel represents the RT-PCR product from 1-mm dissected tubules. The subtypes of alpha 2A- and alpha 2B-receptors were detected in all microdissected tubular segments including PCT, PST, CTAL, CCD, MTAL, and MCD and alpha 2C-signal demonstrated in PST, CTAL, and CCD. beta -actin was detected in all the segments. RT negative control using PCT RNA was depicted in lane RT (-).


View larger version (70K):
[in this window]
[in a new window]
  		Fig. 6.   RT-PCR products of alpha 2-adrenergic receptors and beta -actin in the microdissected renal tubules. Labels of molecular sizes at right of photo are predicted sizes of the products. RT(-) indicates PCR without reverse transcription; PCT, proximal convoluted tubules; PST, proximal straight tubules; CTAL, cortical thick ascending limb; MTAL, medullary thick ascending limb; CCD, cortical collecting duct; MCD, medullary collecting duct. Gel separation and staining of the cDNA products were same as those in Fig. 4.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

The important observation of the present study is that a small subpressor dose of NE decreased MBF and tissue PO2 either when the NOS activity was blunted or alpha 2-adrenergic receptors were blocked in the renal medulla. The results show that medullary NO produced by the stimulation of alpha 2-adrenergic receptors effectively buffered the vasoconstrictor effects of NE in this region. Activation of alpha 2-adrenergic receptors may be an important mechanism to protect the renal medulla from vasoconstriction and ischemia through increase in the production of endogenous NO in response to small elevations of renal sympathetic nerve activity or circulating catecholamine. Identification of the mRNA of alpha 2-adrenergic receptors, specifically alpha 2B-receptors in renal medullary vessels and tubules, provides the evidence that alpha 2-adrenergic receptors may participate importantly in the control of renal medullary circulation and maintenance of tissue PO2 in this kidney region.

Production and action of renal medullary NO during alpha -adrenergic activation. In the present study, we examined the effect of intravenous infusion of a subpressor dose of NE (0.1 µg · kg-1 · min-1) on renal MBF and NO production in the absence and presence of the NOS inhibitor L-NAME in the renal medulla. NE was found to decrease renal MBF and PmO2 only in the presence of L-NAME at a dose that partially inhibited NO production and which had no effect on basal MBF and CBF nor PmO2. It should be noted that the dose of L-NAME chosen for this study was based on a previous study (35) and was a dose which avoided the confounding influences of reduced control levels of MBF and elevations of arterial blood pressure induced by high doses of L-NAME.

With the use of the microdialysis and oxyhemoglobin-NO trapping technique, we found that small subpressor elevations of circulating NE result in significant increases in medullary and cortical [NO] and that renal medullary interstitial infusion of L-NAME completely blocked NE-induced increase in tissue NO in the renal medulla. These results suggest that NE stimulates the production of NO in both renal cortex and medulla and that inhibition of the NO production makes the medullary circulation vulnerable to the vasoconstrictor actions of NE. In the presence of L-NAME, therefore, even a small subpressor elevation of circulating NE levels reduced renal MBF due to a reduction of NO-mediated buffering effects.

However, it remains unknown why L-NAME blocked NE-induced increase in NO but had no effect on the vasoconstrictor effect of NE in the renal cortex. It is possible that other counteracting mechanisms, such as inhibition of renin release, are also importantly involved in buffering NE-induced vasoconstriction in the renal cortex. Previous studies have shown that renin generation and release in the kidney are markedly inhibited by activation of alpha 2-adrenergic receptors (21), which may counteract the vasoconstrictor effect of NE in spite of blockade of NO production. Moreover, it appears that the sensitivity to adrenergic vasoconstriction is lower in renal cortical vessels than medullary vessels. It has been shown that renal denervation produced a 16% and 48% increase in CBF and MBF, respectively, indicating a greater adrenergic tone in the medullary circulation (11). RNS produced a larger vasoconstrictor response in juxtamedullary efferent arterioles that supply blood to the renal medulla compared with cortical efferent arterioles (4). This low intrinsic reactivity of cortical circulation to adrenergic vasoconstriction may also be related to a low response of CBF to subpressor dose of NE, irrespective of blockade of its stimulatory effect on NO production.

In previous studies, we also demonstrated that ANG II and arginine vasopressin (AVP) stimulated the production of NO in the renal medulla and that the increase in [NO] counteracted their vasoconstrictor effects in this kidney region (24, 35). The response pattern of renal MBF and medullary [NO] to NE was similar to that to ANG II. All these results indicate that NO may serve as a counteracting factor to contribute to the control of renal MBF, thereby protecting the renal medulla from vasoconstrictive or ischemic injury. The mechanism by which NO is produced in response to all these vasoconstrictors is still poorly understood. There is evidence indicating that increased NO production in the face of these vasoconstrictors may be associated with the rise in intracellular Ca2+ during vasoconstriction. Increase in intracellular Ca2+ concentration {[Ca2+]i} stimulates the activity of NOS and consequently results in the production of NO. In some vascular beds, however, NO was only increased in response to ANG II, but not to NE or vice versa. In renal medullary circulation, vasoconstriction induced by cyclooxygenase inhibitor indomethacin was not accompanied by NO production (20). These findings have challenged the view that increases in [Ca2+]i can fully account for vasoconstrictor-induced NO production. Our data also do not support the role of increase in [Ca2+]i during vasoconstriction on NO production, because the small dose of NE used in the present study did not produce vasoconstriction.

Another possible mechanism to stimulate NO production by these vasoconstrictors is the action of shear stress due to flow increase in the renal medullary vessels by vasoconstriction. Once again, however, because doses of all vasoconstrictors (NE, ANG II, and AVP) used in the present and our previous studies were chosen to not induce change in vascular tone and blood flow, it seems that alterations of blood flow or shear stress in the medullary vessels are not attributed to increased production of NO in face of these vasoconstrictors. In addition, a previous study in our laboratory demonstrated that AVP stimulated NO production through the V2 receptor, but this AVP receptor could not be detected in the renal medullary vessels (23). In contrast, vasopressin V1 receptors are abundantly expressed in VR of the outer medulla. Activation of V1 receptors produced vasoconstriction in these medullary vessels and thereby decreased renal MBF, but it had no effect on renal medullary interstitial [NO] (24). These results excluded the possibility that increased NO production in response to AVP is derived from renal medullary microvessels. Our data are, therefore, not consistent with the hypothesis that the NO responses are associated with the increase in [Ca2+]i or shear stress induced by vasoconstriction. We propose that selective receptor-mediated mechanisms account for the stimulatory effects of each of these vasoconstrictors on NO production in the renal medulla.

Role of alpha 2-adrenergic receptors in mediating medullary NO production. The present studies provide three lines of evidence, suggesting that alpha 2-adrenergic receptors may mediate NE-induced NO production in the renal medulla, which counteracts the vasoconstrictor response of renal medullary vessels to NE. First, RT-PCR analyses demonstrated that the mRNAs of alpha 2-adrenergic receptors are expressed in the renal medulla. Of particular importance is the occurrence of alpha 2-adrenergic receptors in the VR, because these medullary vessels exhibit the greatest NOS activity among the tubular and vascular segments (32). Activation of alpha 2-adrenergic receptors in these vessels may therefore stimulate the NOS to produce NO, which in turn counteracts the vasoconstrictor effect of NE. Second, by the measurement of tissue NO levels with use of microdialysis, we demonstrated that blockade of alpha 2-adrenergic receptors completely abolished the NE-induced increase in medullary NO, which was similar to the effect of L-NAME. After blockade of alpha 1-adrenergic receptors, however, NE still increased medullary [NO]. This provides direct evidence that activation of alpha 2-adrenergic receptors accounts for the NE-induced NO production. Finally, we found that in parallel to the reduction of medullary [NO], basal MBF was significantly decreased by blockade of alpha 2-adrenergic receptors, suggesting a tonic regulatory effect of alpha 2-adrenergic receptor-mediated NO production in renal medullary circulation. Moreover, the antagonism of alpha 2-adrenergic receptors substantially unmasked the vasoconstrictor effect of a subpressor dose of NE in the renal medulla. This provides functional evidence indicating that alpha 2-adrenergic receptors may mediate adrenergic vasodilation in the renal medulla through stimulation of the NO production.

Considering the importance of renal MBF in the long-term control of arterial blood pressure (5), activation of alpha 2-adrenergic receptors may be an important medullary antihypertensive mechanism by which sodium and water retention can be prevented during elevation of sympathetic tone and circulating catecholamines. However, alpha 2-adrenergic receptor-mediated effects on renal function are rather complex, as shown in previous studies. It has been reported that alpha 2-adrenergic receptor stimulation produced renal vasoconstriction and inhibited prostaglandin-induced sodium and water excretion, thereby leading to antidiuresis and antinatriuresis (25, 26). There is evidence that the density of renal alpha 2-adrenergic receptors determined by ligand binding was markedly increased in several genetically hypertensive rat strains, such as spontaneously hypertensive rats, Dahl salt-sensitive rats, and Sabra rats, even before the arterial blood pressure rose. Chronic salt loading increased renal alpha 2-receptor density in these rats (25, 26). It has been proposed that the changes in renal alpha 2-adrenergic regulation are associated with genetic hypertension. However, previous studies reported that infusion of specific alpha 2-adrenergic receptor antagonists into renal artery in genetically hypertensive rats or in dogs had no effects on renal vascular resistance and sodium excretion (7, 26). Genetic linkage analyses failed to demonstrate a cosegregation of alpha 2-adrenergic receptors with blood pressure in segregating populations involving crosses of Dahl salt-sensitive hypertensive rats with five other strains (9). These results suggest that altered renal alpha 2-adrenergic receptor regulation could not contribute to the pathogenesis of genetic hypertension. It appears that increased expression of renal alpha 2-adrenergic receptors in hypertensive rats represents an antihypertensive compensatory mechanism. The present results indicate that blockade of alpha 2-adrenergic receptors decreased the medullary NO production, and MBF would support this view.

Expression of alpha 2-adrenergic receptors in renal vessels and tubules. The present study demonstrated that the mRNAs of alpha 2B-adrenergic receptors are expressed in the microdissected VR. This suggests the presence of alpha 2B-subtype adrenergic receptors in the renal medullary microvessels. Although previous studies demonstrated the alpha 2-adrenergic ligand binding on the medullary vessels (3, 19), the subtypes of alpha 2-receptors were not dissected. The presence of alpha 2-adrenergic receptors in these medullary vessels may be involved in the stimulation of NE on NO production. Although vascular alpha 2-adrenergic receptors, especially those on the endothelium, may play an important role in mediating the NO production, as indicated by previous studies in coronary circulation (12, 13), alpha 2-adrenergic receptors found in the tubular elements may also participate in the NO production. We found alpha 2A- and alpha 2B-adrenergic receptor mRNA in all microdissected medullary tubules. The contribution of these subtypes of alpha 2-adrenergic receptors on medullary vessels and tubules to the NE-induced NO production remains to be further elucidated.

In summary, small elevations of circulating NE did not change renal MBF, but stimulated NO production. Inhibition of the NOS activity and blockade of alpha 2-adrenergic receptors suppressed the NO production and hence unmasked NE-induced medullary vasoconstriction. These results indicate that the alpha 2-adrenergic receptors may be activated to produce NO in the renal medulla in response to small elevations of sympathetic tone and circulating NE. Increased tissue [NO] appears to buffer the medullary vasoconstrictor actions of NE and prevent the associated reduction of tissue PO2. This protective mechanism may also participate in the control of renal sodium and water excretion during sympathetic stimulations or elevation of circulating catecholamine levels.

Perspectives

The present study demonstrated that alpha 2-adrenergic receptors mediate renal medullary NO production in response to small elevations of circulating NE, which represents a new function of these adrenergic receptors in the kidney. It seems that this receptor-mediated NO production is more sensitive in the renal medulla than in other kidney regions. Considering the importance of renal medullary circulation in the control of sodium and water excretion and long-term control of arterial blood pressure, alpha 2-adrenergic receptor-mediated NO production may serve as an important protecting mechanism during neural-humoral stress response, which can help maintain a circulatory homeostasis. In this regard, activation of alpha 2-adrenergic receptors in the kidney, especially in the renal medulla in response to sympathetic stimulation or elevation of circulating catecholamine would produce an antihypertensive effect through increase in NO production and sodium and water excretion in the kidney. This antihypertensive action of renal alpha 2-adrenergic receptors needs to be further established.


    ACKNOWLEDGEMENTS

The authors thank Na Su and Roxanne Allaire for technical assistance.


    FOOTNOTES

This study was supported by National Institutes of Health Grants HL-29587 and DK-54927.

Address for reprint requests and other correspondence: A.-P. Zou, Dept. of Physiology, Medical College of Wisconsin, 8701 Watertown Plank Rd., Milwaukee, WI 53226 (E-mail: azou{at}mcw.edu).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Received 28 June 1999; accepted in final form 22 March 2000.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Ajikobi, DO, and Cupples WA. Alpha 2-adrenergic mediation of the effect of angiotensin II on rat renal artery in vitro. Can J Physiol Pharmacol 72: 1019-1024, 1994[ISI][Medline].

2.   Bylund, DB. Pharmacological characteristics of alpha-2 adrenergic receptor subtypes. Ann NY Acad Sci 763: 1-7, 1995[Abstract].

3.   Calianos, T II, and Muntz KH. Autoradiographic quantification of adrenergic receptors in rat kidney. Kidney Int 38: 39-46, 1990[ISI][Medline].

4.   Chen, J, and Fleming JT. Juxtamedullary afferent and efferent arterioles constrict to renal nerve stimulation. Kidney Int 44: 684-691, 1993[ISI][Medline].

5.   Cowley, AW, Jr, Mattson DL, Lu S, and Roman RJ. The renal medulla and hypertension. Hypertension 25: 663-673, 1995[Abstract/Free Full Text].

6.   DiBona, GF, and Kopp UC. Neural control of renal function. Physiol Rev 77: 75-197, 1997[Abstract/Free Full Text].

7.   DiBona, GF, and Sawin LL. Role of renal alpha 2-adrenergic receptors in spontaneously hypertensive rats. Hypertension 9: 41-48, 1987[Abstract/Free Full Text].

8.   DiBona, GF, Sawin LL, and Jones SY. Differentiated sympathetic neural control of the kidney. Am J Physiol Regulatory Integrative Comp Physiol 271: R84-R90, 1996[Abstract/Free Full Text].

9.   Ginn, DI, Baptista CA, Alam KY, Deng AY, Dene H, Le H, Kurtz TW, and Rapp JP. Genetic analysis of alpha 2-adrenergic receptors and blood pressure using Dahl salt-sensitive rats. J Hypertens 12: 357-365, 1994[ISI][Medline].

10.   Graham, RM, Perze DM, Hwa J, and Piascik MT. alpha 1-Adrenergic receptor subtypes. Molecular structure, function and signaling. Circ Res 78: 737-749, 1996[Free Full Text].

11.   Hermansson, K, Ojte GG, and Wolgast M. The cortical and medullary blood flow at different levels of renal nerve activity. Acta Physiol Scand 120: 161-169, 1984[ISI][Medline].

12.   Indolfi, C, Piscione F, Villari B, Russolillo E, Rendina V, Golino M, and Chiariello M. Role of alpha-2 adrenoceptors in normal and atherosclerotic human coronary circulation. Circulation 86: 1116-1123, 1992[Abstract/Free Full Text].

13.   Jones, CJH, Defily DV, Patterson JL, and Chilian WM. Endothelium-dependent relaxation competes with alpha 1-and alpha 2-adrenergic constriction in the canine epicardial coronary microcirculation. Circulation 87: 1264-1274, 1993[Abstract/Free Full Text].

14.   Lanier, SM, Downing S, Duzic E, and Homcy CJ. Isolation of rat genomic clones encoding subtypes of the alpha-2-adrenergic receptor. Identification of a unique receptor subtype. J Biol Chem 266: 10470-10478, 1991[Abstract/Free Full Text].

15.   Mattson, DL, and Higgins D. Influence of dietary sodium intake on renal medullary nitric oxide synthesis. Hypertension 27: 688-692, 1996[Abstract/Free Full Text].

16.   Mckenna, OC, and Angelakos ET. Adrenergic innervation of the canine kidney. Circ Res 22: 345-353, 1968[Abstract/Free Full Text].

17.   McPherson, GA, and Summers RJ. Evidence from binding studies for alpha 2-adrenoceptors directly associated with glomeruli from rat kidney. Eur J Pharmacol 90: 333-341, 1983[ISI][Medline].

18.   Miller, VM, and Vanhoutte P. Endothelial alpha 2-adrenoceptors in canine pulmonary and systemic blood vessels. Eur J Pharmacol 118: 123-129, 1985[ISI][Medline].

19.   Muntz, KH, Meyer L, Gadol S, and Calianos TA. Alpha-2 adrenergic receptor localization in the rat heart and kidney using autoradiography and tritiated rauwolscine. J Pharmacol Exp Ther 236: 542-547, 1996[Abstract/Free Full Text].

20.   Ortiz, MC, Atucha NM, Lahera V, Vargas F, Quesada T, and Garcia-Estan J. Importance of nitric oxide and prostaglandins in the control of rat renal papillary blood flow. Hypertension 27: 377-381, 1996[Abstract/Free Full Text].

21.   Osborn, JL, Holdass JH, and Thames MC. Renal adrenoreceptor mediation of antinatriuretic and renin secretion response to low frequency renal nerve stimulation in the dog. Circ Res 53: 298-305, 1983[Free Full Text].

22.   Parekh, N, Dobrowolski L, Zou AP, and Steinhausen M. Nitric oxide modulates angiotensin II- and norepinephrine-dependent vasoconstriction in rat kidney. Am J Physiol Regulatory Integrative Comp Physiol 270: R1-R6, 1996.

23.   Park, F, Mattson DL, Skelton MM, and Cowley AW, Jr. Localization of the vasopressin V1a and V2 receptors within the renal cortical and medullary circulation. Am J Physiol Regulatory Integrative Comp Physiol 272: R243-R251, 1997[Abstract/Free Full Text].

24.   Park, F, Zou AP, and Cowley AW, Jr. Vasopressin-mediated stimulation of nitric oxide in the rat renal medulla. Hypertension 32: 896-901, 1998[Abstract/Free Full Text].

25.   Pettinger, WA. Renal alpha 2-adrenergic receptors and hypertension. Hypertension 9: 3-6, 1987[Free Full Text].

26.   Pettinger, WA, Umemura S, Smyth DD, and Jeffries W. Renal alpha 2-adrenoceptors and the adenylate cyclase-cAMP system: biochemical and physiological interactions. Am J Physiol Renal Fluid Electrolyte Physiol 252: F199-F208, 1987[Abstract/Free Full Text].

27.   Reid, JJ, and Rand MJ. Renal vasoconstriction is modulated by nitric oxide. Clin Exp Pharmacol Physiol 19: 376-379, 1992[ISI][Medline].

28.   Richard, V, Tanner FC, Tschudi M, and Luescher TF. Different activation of L-arginine pathway by bradykinin, serotonin, and clonidine in coronary arteries. Am J Physiol Heart Circ Physiol 259: H1433-H1493, 1990[Abstract/Free Full Text].

29.   Sai, Y, Silldorff EP, and Pallone TL. Effect of norepinephrine and acetylcholine on outer medullary descending vasa recta. Am J Physiol Heart Circ Physiol 269: H710-H716, 1995[Abstract/Free Full Text].

30.   Spielman, WS, and Arend LJ. Adenosine receptors and signaling in the kidney. Hypertension 17: 117-130, 1991[Abstract/Free Full Text].

31.   Wu, F, Li PL, and Zou AP. Microassay of 5'-nucleotidase and adenosine deaminase activity in microdissected nephron segments. Anal Biochem 266: 133-139, 1999[ISI][Medline].

32.   Wu, F, Park F, Cowley AW, Jr, and Mattson DL. Quantification of nitric oxide synthase activity in microdissected segments of the rat kidney. Am J Physiol Renal Physiol 276: F874-F881, 1999[Abstract/Free Full Text].

33.   Zeng, D, Harrison JD, D'Angelo DD, Barber CM, Tucker AL, Lu Z, and Lynch KR. Molecular characterization of a rat alpha-2B-adrenergic receptor. Proc Natl Acad Sci USA 87: 3102-3106, 1990[Abstract/Free Full Text].

34.   Zou, AP, and Cowley AW, Jr. Nitric oxide in renal cortex and medulla: an in vivo microdialysis study. Hypertension 29: 194-198, 1997[Abstract/Free Full Text].

35.   Zou, AP, Wu F, and Cowley AW, Jr. Protective effect of angiotensin II-induced increase in nitric oxide in the renal medullary circulation. Hypertension 31: 271-276, 1998[Abstract/Free Full Text].

Am J Physiol Regul Integr Comp Physiol 279(3):R769-R777
0363-6119/00 $5.00 Copyright © 2000 the American Physiological Society


This article has been cited by other articles:


Home page
 J. Neurosci.Home page
S. Hirano, K. Asanuma, Y. Ma, C. Tang, A. Feigin, V. Dhawan, M. Carbon, and D. Eidelberg
Dissociation of Metabolic and Neurovascular Responses to Levodopa in the Treatment of Parkinson's Disease
J. Neurosci., April 16, 2008; 28(16): 4201 - 4209.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Renal Physiol.Home page
W. Zhang and A. Edwards
A model of nitric oxide tubulovascular cross talk in a renal outer medullary cross section
Am J Physiol Renal Physiol, February 1, 2007; 292(2): F711 - F722.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Regul. Integr. Comp. Physiol.Home page
N. W. Rajapakse, A. K. Sampson, G. A. Eppel, and R. G. Evans
Angiotensin II and nitric oxide in neural control of intrarenal blood flow
Am J Physiol Regulatory Integrative Comp Physiol, September 1, 2005; 289(3): R745 - R754.
[Abstract] [Full Text] [PDF]

Home page
 Exp PhysiolHome page
K. M. McCormick, E. M. Bravo, and C. T. Kappagoda
Role of adrenergic receptors in the reflex diuresis in rabbits during pulmonary lymphatic obstruction
Exp Physiol, May 1, 2005; 90(3): 341 - 347.
[Abstract] [Full Text] [PDF]

Home page
 SEMIN CARDIOTHORAC VASC ANESTHHome page
M. Stafford-Smith
Evidence-Based Renal Protection in Cardiac Surgery
Seminars in Cardiothoracic and Vascular Anesthesia, March 1, 2005; 9(1): 65 - 76.
[Abstract] [PDF]

Home page
 Am. J. Physiol. Renal Physiol.Home page
W. Zhang, T. Pibulsonggram, and A. Edwards
Determinants of basal nitric oxide concentration in the renal medullary microcirculation
Am J Physiol Renal Physiol, December 1, 2004; 287(6): F1189 - F1203.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Regul. Integr. Comp. Physiol.Home page
A. W. Cowley Jr., T. Mori, D. Mattson, and A.-P. Zou
Role of renal NO production in the regulation of medullary blood flow
Am J Physiol Regulatory Integrative Comp Physiol, June 1, 2003; 284(6): R1355 - R1369.
[Abstract] [Full Text] [PDF]

Home page