J. Physiol. (1976), 254, pp. 183-202 With 10 text-figures Printed in Great Britain

183

PLASMA SODIUM CONCENTRATION AND SODIUM EXCRETION IN THE ANAESTHETIZED DOG

BY F. S. NASHAT, J. W. TAPPIN AND C. S. WILCOX From the Departments of Physiology, Physics, Pharmacology and Therapeutics, Middlesex Hospital Medical School, London, W1

(Received 6 May 1975) SUMMARY

1. The effect of acute alterations of plasma sodium concentration (PNa) on renal sodium excretion (UNaV) was investigated by three types of experiments on anaesthetized dogs: (a) A local increase in PNa at one kidney was produced by infusion of hypertonic saline directly into its artery while systemic levels of PNa were stabilized by haemodialysis. (b) Systemic levels of PNa were lowered by exchange transfusion of blood for an equal volume of salt-free dextran-in-dextrose solution. The results were contrasted with those observed after similar exchanges, but using dextran-in-saline solution. (c) The level of PN. was altered by varying the sodium concentration of a saline solution infused at a fixed rate either intravenously or into one renal artery. 2. All three types of experiment suggest a dependence of UNa V on PNa. Analysis demonstrated that this relationship was not due to contemporary changes in: packed cell volume; plasma solids concentration; plasma potassium concentration; blood pressure or plasma hydrogen ion concentration. The distribution of these variables did not change with PNaexcept for plasma hydrogen ion concentration. Moreover, the relationship persisted when data were selected to exclude clearance periods in which the value for any variable had shifted past the group mean obtained before PNa was altered. 3. The fall in UNaV at low levels of PNa could be attributed to a fall in glomerular filtration rate (GFR), but the progressive rise in UNaV seen as PNa exceeded 150 m-mole l-l occurred despite a fall in GFR and no apparent change in the mean filtered load of sodium. These results suggest that the increased sodium excretion accompanying raised levels of PNa is due to reduced tubular re-absorption of sodium.

184

F. S. NASHAT, J. W. TAPPIN AND C. S. WILCOX INTRODUCTION

An investigation of the mechanisms relating sodium excretion (UNaV) to the plasma sodium concentration (PN.) would normally demand that plasma sodium be altered while all other variables are held constant. Yet changing plasma sodium concentration has such widespread effects in the organism, that to contain all the variables would be well nigh impossible if any semblance of the resulting preparation to physiological reality were to be maintained. So, despite an enormous literature, an unequivocal definition of the relationship between PNa and UN, V has not yet been formulated. Green & Farah (1949) and Selkurt & Post (1950) noted that when PNa was increased by rapid intravenous infusions of strongly hypertonic saline solutions in the dog there was, besides an increase in sodium excretion, an alteration in blood pressure and a fall in plasma protein concentration, haematocrit and arterial pH. Green & Farah (1949) argued that sodium excretion correlated with the volume of the intracellular water transferred to the extracellular fluid compartment; they concluded that PNa influences sodium excretion by the extracellular volume expansion it produces. In experiments on conscious dogs, O'Connor (1962) studied the sodium excretion that followed the administration of small intragastric loads of either isotonic or hypertonic saline solutions. In both instances, the increased rate of sodium excretion was closely related to the fall in plasma protein concentration produced. He found 'no compelling evidence' that a small rise in PNa per se caused a rise in sodium excretion. In other experiments, the injection of hypertonic saline directly into the blood flowing into one kidney increased its sodium excretion considerably above that of the contralateral kidney (Green & Farah, 1949; Goodyer & Glenn, 1952; Selkurt, 1954; Kamm & Levinsky, 1965; Nash, Rostorfer, Bailie, Wathen & Schneider, 1968; Forgacs, Chatel & Visy, 1969; Puschett, Goldstein, Godshall, Staum & Goldberg, 1971). This could not be wholly attributed to systemic effects which must be represented at both kidneys. But changes in the renal vasculature or in the concentration of plasma proteins or other blood constituents produced locally by the infusion could not be excluded. Thus O'Connor (1962) argued that osmotic abstraction of fluid from red blood cells within the renal artery may lead to a sufficient local haemodilution to contribute to that kidney's response. Moreover, in these experiments the PNa of the general circulation must eventually rise and thereby deny the contralateral kidney its function as a true control. The experiments of Nizet, Godon & Mahieu (1968) using the isolated, perfused dog's kidney provided more convincing evidence that PNa can determine UNaV directly.

PNaAND SODIUM EXCRETION 185 In experiments on seventy-nine dogs, three different procedures were used to explore the relationship between PNa and UNa V in the whole animal. In the first, the PNa at one kidney was raised by infusion of hypertonic saline into its artery, while the PNa of the animal's blood which perfused the other kidney was stabilized by haemodialysis. In the second, the PNa was reduced by exchange transfusions of blood for dextran-indextrose and the effects of this on sodium excretion compared to that produced by similar exchanges with dextran-in-saline. In the third series, the PNa was altered by infusion of hypo- or hypertonic saline without attempting to stabilize systemic levels of PNa. Contemporary fluctuations in a number of variables were recorded and their contribution to the renal response was analysed later from the pooled results. Effects due to variations in any condition could be excluded by simply selecting, for analysis, the results in which that condition had not changed. This procedure obviated considerable interference with the preparation that the experimental control of these conditions would have entailed. This latter series of experiments is, therefore, taken to represent the closest approximation to 'physiological reality' and the results obtained therefrom

emphasized accordingly. METHODS Experiments were performed on greyhounds of either sex weighing between 18 and 35 kg (mean ± S.D., 26-5 ± 3-8). Anaesthesia was induced by the i.v. administration of 30 mg kg-' body weight pf pentobarbitone sodium. A light surgical anaesthesia, sufficient to abolish spontaneous movements, reflex reactions to painful stimuli and the corneal reflex, was maintained by further injections of 30-60 mg of the anaesthetic as required. The trachea was either intubated or cannulated through a mid-line incision in the neck. A quarter of the animals breathed spontaneously, 65% were maintained on artificial ventilation throughout. The ventilation was adjusted to give a Pco, of between 30 and 40 mmHg at the outset, but was not subsequently changed. In the remaining 10 % of the experiments the animals had to be artificially ventilated during the experiment to counteract signs of respiratory depression. The blood pressure was measured by a mercury manometer connected to a cannulated femoral artery. Glomerular filtration rate was estimated as the clearance of [125I]sodium diatrizoate, or [51Cr]ethylene diamine tetra-acetic acid ([51Cr]EDTA). The markers were infused continuously after a priming dose and the plasma level measured at regular intervals (ca. 30 min). Radioactivity was measured in 2 ml. samples of urine or plasma in a well type scintillation counter. Throughout the experiment the animals received two infusions.

Infusion A This contained 2 parts of isotonic (0.154 M) saline and 1 part of 20% mannitol solution. It was infused i.v. at a rate of about 1 ml. min' and contained the maintenance doses of the markers used to measure glomerular filtration rate (GFR).

186

F. S. NASHAT, J. W. TAPPIN AND C. S. WILCOX

Infusion B This consisted initially of isotonic (0.154 M) saline which was given either intravenously or into a renal artery at a rate of 0-1 ml. kg-' min-'. The concentration of sodium in this infusion was changed either to hypotonic (0-077 M) or hypertonic (0-616 or 1-232 M). The ureters were approached through a low abdominal incision and cannulated with soft pliable polyethylene tubing. Urine from the two kidneys was collected separately over 4-30 min periods (average 15 min). The left kidney was exposed retroperitoneally through a flank incision and its artery dissected. Particular care was taken to keep the renal nerves and lymphatics intact. An 18-gauge hooked needle whose tip was blocked but a new hole bored in its shaft was used doubly to transfix the renal artery. The needle was used to deliver infusion B into the renal artery. Sodium and potassium levels in urine and plasma were measured using a flame photometer. When hypertonic saline was infused into the left renal artery, the plasma sodium concentration (PN.) at the left kidney was calculated assuming simple admixture of saline with the plasma. The formula used was: PW&*(RPF- + RPF where PNx* is the systemic level of plasma sodium, C and V are the concentration and flow rate of the infusate and RPF is the renal plasma flow measured as the clearance of ['25I]Hippuran corrected for extraction or calculated from the renal blood flow recorded by an electromagnetic flow meter (Biotronix BL-610). Osmolarity was estimated cryoscopically using an 'Advanced Osmometer'. Haematocrit (packed cell volume) was measured in a 'Hawkesley Microhaematocrit centrifuge'. Plasma solids were measured by desiccating measured volumes of plasma to constant weight; the value was corrected by subtracting the weight of the measured NaCl and KC1. The pH and Pco, of arterial blood and pH of urine were estimated with a Radiometer (model BMS 3). After completion of surgery, 60 min were allowed before measurements were made. The following additional procedures were undertaken. In series I either a Kolff double layer (eleven experiments) or a Travenol coil (three experiments) haemodialysis unit was connected between two femoral veins. Blood was pumped through the dialyser by a peristaltic (Sigmamotor) pump at 200 ml. min-. It was continuously dialysed against a 100 1. solution of bath fluid, kept at 370 C, and made up from a modified Fulham dialysis concentrate ('Renalyte', no. S/124; Macarthy's Laboratories, Romford, Essex) to yield the following concentrations: 124 m-mole I.sodium calcium 1-6 m-mole 1.-' potassium 3-5 m-mole 1.magnesium 1-0 m-mole 1.-' chloride 100 m-mole l.acetate 35 m-mole .-' 12 m-mole l.-1 dextrose Two Palmer slow injection pumps were used to deliver heparin (33 i.u. min-' of 'Heparin injections, mucous' B.P., Paines and Byrne Ltd) into the blood coming from the animal and protamine (0-30 mg min-' of 'Protamine sulphate injection' B.P., I %. Weddel Pharmaceuticals) into the blood going to the animal. The 'dead

VI) U1(V)

PNa

AND SODIUM EXCRETION

187 space' in the dialyser (200-400 ml.) was primed with a solution of 6 g 100 ml.-' dextran-in-isotonic (0. 154 M) saline ('Dextraven 110 ', Fisons Ltd). During the first 40-60 min of haemodialysis, dextran-in-saline was infused i.v. at a rate sufficient to maintain the central venous pressure within + 2 cm H20 of the predialysis value. Central venous pressure was measured, by a saline manometer, from the right atrium through catheters inserted via the jugular vein. The volume of the dextran solution required ranged between 0 and 1000 ml., and averaged about 300 ml. A period of 30 min was then allowed before any data were collected. In these experiments, the dog first received isotonic (0.154 M) saline delivered into the left renal artery. After 30-60 min the saline sodium concentration was increased to 1*232 M for 50-150 min after which it was reduced again to isotonic concentration until the end of the experiment, 30-90 min later. In series 2 500 ml. of the animal's blood was exchanged for an equal volume of iso-oncotic dextran (6 g 100 ml.-'; 'Dextraven 110', Fisons Ltd) in either isotonic (0-154 M) saline solution (eighteen experiments) or isotonic (0.28 M) dextrose solution (six experiments) as described previously (Nashat, Scholefield, Tappin & Wilcox, 1969). In this series, infusion B was of isotonic saline and was delivered throughout via the needle in the left renal artery. Measurements were made - from the two kidneys - for 30 min before the exchange transfusion, and for two 30 min periods (at 15-45 and 45-75 min) following the completion of the procedure. In 8er'e8 3 infusion B was delivered into a femoral vein (ten dogs) or into the left renal artery (twenty-six dogs); initially isotonic saline was infused. After 60-120 min the sodium concentration of the infusate was abruptly changed to either hypotonic (0-077) M) or hypertonic (0*616 or 1*232 M) for 15-240 min (usually about 45 min). There followed 40-120 min during which the animal received isotonic (0 154 M) saline again. With the shorter time intervals this sequence was repeated up to five times, thereby inducing fluctuating levels of PNa during the course of an experiment. In ten experiments from this series the renal vein was cannulated through a gonadal tributary. The tip of the cannula lay pointing towards the kidney and within 2 cm of its hilum. Data from the third series of experiments were analysed by computer. Results obtained from the left and right kidneys were entered separately. Data were excluded when GFR was below 15 ml. min-' or when urine flow rates were changing rapidly. Data obtained from the left kidney within 30 min of starting a hypertonic saline infusion into its artery were also excluded unless the Pa in the renal venous blood had been measured. For each clearance period, the following data were entered in the computer: animal's body weight, plasma sodium concentration, blood pressure, plasma pH, concentration of plasma solids, haematocrit, plasma potassium concentration, plasma osmolarity, urine flow rate, urine sodium excretion, urine potassium excretion, urine pH, urine osmolarity and* glomerular filtration rate. A computer programme was written which allowed the calculation from these variables (or their derivatives) of mean values, standard deviations, standard errors of the mean, simple linear regression constants and correlation coefficients. The programmes were based on statistical methods described by Armitage (1971). They were also adapted to select data corresponding to spec fied values of any of these variables.

F. S. NASHAT, J. W. TAPPIN AND C. S. WILCOX

188

RESULTS

Series 1: experiments in which a 1-232 M solution of NaCI was infused into one renal artery while haemodialysis was used in an attempt to stabilize systemic PNa

Two examples from this series are shown in Fig. 1. In both, the infusion of hypertonic saline into one renal artery increased sodium excretion from that kidney. Sodium excretion from the contralateral kidney increased, after a delay, only when haemodialysis was ineffective in checking PNa (Fig. 1 A); it was unchanged and remained low when PNa was contained below 155 m-mole 1.-i (Fig. 1 B). When the infusion of the hypertonic saline was discontinued, UNaV from the experimental kidney 190 A

B

E 150/

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0 0

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100

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50 100 Time (min)

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200

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Fig. 1. Graphic representation of results from two experiments. In both cases the animal's blood was continuously dialysed. During the periods marked by solid bars the animals received hypertonic (1232 m-mole 1.-i) saline through a needle placed in the left renal artery. This raised the calculated plasma sodium concentration (PN,) at that kidney to the levels indicated by open circles and discontinuous lines. The measured systemic PK. is shown by filled circles and continuous lines. Sodium excretion (UNAV) from the left kidney is indicated by discontinuous lines and from the right by continuous lines. Note that in panel B when systemic PNa is maintained below 155 m-mole I.-' sodium excretion from the right kidney does not rise. In both experiments the hypertonic infusion increases UN.V from the left kidney.

PNa AND SODIUM EXCRETION

189 reverted towards its initial level. In all fourteen experiments in this series sodium excretion from the kidney receiving the infusion of hypertonic saline into its artery rose above that from its contralateral pair. +50

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Fig. 2. Composite graph showing mean ( +.E. of mean) values for changes in GFR (panel A), plasma sodium concentration (panel B), urine volume (panel C) and sodium excretion (panel D) 15-45 and 45-75 min after exchange transfusions: with dextran-in-saline in eighteen. experiments (filled circles and continuous lines); with dextran-in-dextrose in six experiments (open circles and interrupted lines). The changes are expressed as percentages of the pre-exchange values.

Series 2: experiments in which PNa was reduced by 'exchange transfu8ion8' with dextran-in-dextrose The results from twenty-four experiments are summarized in Fig. 2. In these experiments 500 ml. blood was exchanged, over 10-15 min, for an equal volume of either dextran-in-isotonic (0-154 M) saline or dextran-inisotonic (0-28 M) dextrose. Both exchanges are seen to produce a diuresis. Dextran-in-saline did not change PNa or GFR significantly but increased sodium excretion. Dextran-in-dextrose, however, reduced PN., GFR and sodium excretion. On two occasions, one dog had exchange transfusions with both dextran solutions separated by a period of 90 min. The responses to either exchange were similar to those noted above.

I

F. S. NASHAT, J. W. TAPPIN AND C. S. WILCOX 190 A glycosuria lasting not more than 15 min was observed after the exchange with dextran-in-dextrose.

Series 3: experiments in which variations in PNa were produced by altering the sodium concentration of a saline solution infused i.v. or into a renal artery During infusion of hyper-tonic saline into one renal artery a simple mixing of the saline with plasma would alter the plasma sodium concentration at the kidney to a new level given by the formula described in A

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Time (min) Fig. 3. Graphic representation of results from two experiments. In each panel clearance of hypaque (GFR), plasma sodium concentration of renal venous blood and sodium excretion are plotted against time. Both dogs received isotonic (154 m-mole I.-') saline i.v. at 2-7 and 2 ml. min- respectively. During the periods marked by the rectangles the composition of the infusion was changed to 77 m-mole L.-i in A or 1230 m-mole 1.-i in B. Measurements refer to the left kidney.

Methods. In forty-nine instances plasma sodium concentration of blood leaving the kidney was measured during the infusion, and compared with the calculated values. The calculated values significantly exceeded the values simultaneously measured from the renal vein (P < 0-01), during

191

PNa AND SODIUM EXCRETION

the first 30 min of the infusion, but were not significantly different thereafter. In the same experiments the arteriovenous differences for haematocrit and plasma solids across the kidney were also measured. This showed that the mean fall throughout the period of infusion was 3* 1 % for haematocrit and 3-6 % for plasma solids. Fig. 3 shows representative results from two experiments in this series. In Fig. 3A the PNa measured in the left renal vein was reduced from 145 to 137 m-mole L.-i by the infusion of hypotonic (0.077 M) saline into the left renal artery. This was associated with a fall in UNaV and GFR from that kidney; in Fig. 3B raising PNa from 144 to 169 m-mole l.-' was accompanied by a fall in GFR but a marked rise in UiaV. T 200 4'~~~~~~~~~~~~~~~~~~~~~~%

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Plasma sodium concentration and sodium excretion in the anaesthetized dog.

J. Physiol. (1976), 254, pp. 183-202 With 10 text-figures Printed in Great Britain 183 PLASMA SODIUM CONCENTRATION AND SODIUM EXCRETION IN THE ANAES...
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