IN a previous paper [O'Connor, 1950] comparison was made between. hormone from the neurohypophysis when hypertonic solutions were

Transcription

1 THE EXCRETION OF ADMINISTERED SODIUM CHLORIDE BY THE CONSCIOUS DOG, AND THE EFFECT OF OCCLUSION OF THE CAROTID ARTERIES. By W. J. O'CONNOR. From the Department of Physiology, School of Medicine, University of Leeds. (Received for publication 9th March 1955) CONTENTS INTRODUCTION..... METHODS..... RESULTS..... I. Excretion of Administered Sodium Chloride Effect of Denervation of the Kidneys Effect of Desoxycorticosterone Acetate Changes in the Composition of the Plasma Relationship between the Solid Content of Excretion Renal Response to Sodium Bicarbonate Renal Response to Ammonium Chloride Renal Response to Water II. Effect of Occlusion of the Carotid Arteries Description Effect of Denervation of the Kidneys Changes in Arterial Blood Pressure DIscussIoN SUMMARY..... ACKNOWLEDGMENTS REFERENCES..... the PAGE Plasma and Sodium * INTRODUCTION IN a previous paper [O'Connor, 1950] comparison was made between the renal response in the conscious dog to the administration by stomach tube of 2*5-3-5 g. of sodium chloride in hypertonic or isotonic solution. The rate of excretion of chloride rose equally; but in the first case the urine was of small volume and high concentration, while in the second the same amount of chloride was excreted in a larger volume of more dilute urine. The difference was shown to be due to the release of hormone from the neurohypophysis when hypertonic solutions were given, and from this and other evidence it was concluded that the hormone of the neurohypophysis promoted the tubular reabsorption 237

2 238 O'Connor of water without affecting the excretion of sodium chloride. As the excretion of administered sodium chloride is not determined by the release of hormone from the neurohypophysis, the first object in continuing the work was to study what factors do determine the excretion of sodium chloride. During the work it was observed that occlusion of both carotid arteries in the dog caused an increased excretion of sodium and chloride, and this has also been investigated. The results show that increased excretion of sodium in the one case may be attributed to dilution of the plasma proteins, and in the other to increase in arterial blood pressure, both of which may be expected to increase the glomerular filtration rate. Glomerular filtration rate, among the many factors affecting sodium excretion (see recent reviews by Berliner [1950], Smith [1952] and Selkurt [1954]), is agreed to be of great importance; but as Berliner [1950] has pointed out, the quantitative relationship between glomerular filtration rate and excretion of sodium is not known, and hence accurate assessment of other factors is impossible. It is to a consideration of this important relationship that the experimental work leads, the discussion in this paper being largely concerned with the importance of glomerular factors in determining high rates of excretion of sodium. METHODS Bitches (wt kg.), fed g. of dog biscuit and 120 g. of meat each evening, were used. The perineum was split to expose the urethra for easy catheterization. Also the carotid arteries were enclosed in van Leersum [1911] skin loops, and in some of the animals one femoral artery was enclosed in a loop for the measurement of the arterial blood pressure [O'Connor, 1955]. On days when the response to administered sodium chloride was to be tested, a preliminary dose of 200 ml. of 0 9 per cent sodium chloride was given by stomach tube at about a.m. At approximately 1.30 p.m. a self-retaining rubber catheter was passed into the bladder and the urine collected in measuring cylinders in periods of 7-20 min. A T-piece in the tubing near the vulva allowed the connecting tubing to be drained at the end of each period, after the bladder had been emptied by gentle movements of the catheter. The test dose of sodium chloride was given at about 2.00 p.m., usually by stomach tube but in a few experiments by intravenous infusion. In some experiments the animal stood in a Pavlov stand; in others, where intravenous infusion, collection of blood samples or measurement of the blood pressure was required, the animal lay quietly on its side on a warmed table with light restraint. On any day when no experiment was performed ml. of 0 9 per cent sodium chloride was given by stomach tube during the afternoon.

3 Excretion of Sodium Chloride 239 Occlusion of the Carotid Arteries contained in the skin loops was obtained by connecting compression cuffs to a reservoir of air at 240 mm. Hg pressure. Blood Samples were obtained by puncture of the carotid loop with a No. 19 hypodermic needle. Samples of 2-4 ml. were drawn into a syringe containing 20 units of heparin in 0-02 ml. of 0 9 per cent sodium chloride, and centrifuged in air at 900 g for 30 min. to obtain plasma. Intravenous Infusion. In those experiments where saline was given by intravenous infusion a length of polythene tubing was inserted into the cephalic or malleolar vein under local anaesthesia. Saline entered from a funnel through a warming bottle which kept the infusion at a temperature of C. Arterial Blood Pressure was determined indirectly by the method of O'Connor [1955]. Analytical Methods. Solid content of plasma was determined by weighing, drying and reweighing about 0-3 ml. of plasma. The samples were dried at C. for 3 hrs., after which there was no further weight loss. Chloride in plasma was determined by applying the method of Smirk [1927] to the dried plasma used for the estimation of solid content. Sodium in plasma was determined by a flame photometer (external standard). Duplicates of these determinations on plasma agreed to within 1-5 per cent. Sodium in urine was determined by the flame photometer, chloride by the method of Wilson and Ball [Peters and van Slyke, 1932]. RESULTS I. The Excretion of Administered Sodium Chloride The Effect of Denervation of the Kidneys.-Berne [1952] has shown equal excretion of sodium by the denervated left and the normal right kidney of conscious dogs at rates of sodium excretion from 0-02 to 0-5 m.equiv./min. The experiments shown in fig. LA and B confirm the work of Berne in the present conditions. The normal excretion of chloride in this animal following the administration of sodium chloride was established by 7 experiments (fig. IA). Then, under ether anaesthesia and with full aseptic precautions, all connexions of the kidneys were severed except the ureter, renal artery and vein, and these were carefully denuded of visible nervous or connective tissue; and the splanchnic nerves were cut as they emerged from under the crura of the diaphragm. Fig. 1B shows the unaltered response when the same dose of sodium chloride was given during the 3rd to 5th week after operation. Similarly in two other animals the excretion of sodium and chloride after the administration of the test dose was not altered by denervation of the kidneys. Clearly the renal nerves play no immediate part in determining the excretion of administered sodium chloride.

4 240 O'Connor The Effect of Desoxycorticosterone Acetate.-DCA in daily doses of 1-2 mg. in oil is known to decrease the excretion of sodium by adrenalectomized dogs and to decrease low rates of sodium excretion in normal dogs [Thorn, Engel and Eisenberg, 1938]. Fig. 1C shows that the excretion of administered sodium chloride was not decreased by 5 mg. DCA in oil injected 4 hrs. and again 15 min. before the test dose. In other experiments 20 mg. DCA in oil 3 hrs., or 10 mg. 6 and 24 hrs., before the test dose failed to modify the excretion of administered sodium chloride; and a normal response was also obtained 4 hrs. after A B... C CHLORIDE ++ m. equivj/min. O TIME (min.) FIG. 1."Jeeps", 8-5 kg. The excretion of chloride following the administration of 250 ml. of 0-9 per cent sodium chloride by stomach tube at zero time. Abscissae: time in minutes; ordinates: rate of excretion of chloride in m.equiv./min. A, Mean of 7 experiments before operation; B, mean of 6 experiments in the 3rd, 4th and 5th weeks after denervation of the kidneys and section of the splanchnic nerves; C, mean of 4 experiments before operation but with the subcutaneous injection of 5 mg. DCA in oil 240 and 15 min. before the test dose of salt. The dotted lines are placed one standard error on either side of the mean. the last of 6 daily injections of 10 mg. DCA in oil. The intravenous injection of 40 mg. desoxycorticosterone glucoside ("Percorten watersoluble", Ciba) during the response to sodium chloride had no effect on excretion of chloride. Similarly, Dorfman, Potts and Fail [1947] found that doses of DCA adequate to decrease low rates of excretion of sodium in rats had no effect at higher rates. If the increased renal elimination of sodium and chloride following salt administration were determined by decreased production of a salt-retaining hormone of the suprarenal cortex, DCA would be expected to decrease the excretion of sodium chloride. Changes in the Composition of the Plasma.-In the experiments shown in fig. 2A and B, the same dose of sodium chloride was given by stomach

5 Excretion of Sodium Chloride 241 tube either as hypertonic (3.5 per cent) or as isotonic (0.9 per cent) solution. In the lower part of each graph the rate of excretion of both sodium and chloride is recorded, whereas in the earlier experiments of O'Connor [1950] chloride only was measured. In all experiments where sodium chloride was administered the rate of excretion of sodium ran TIME (min.) FIG. 2.-"Skewbald", 15 kg. The excretion of sodium and chloride, and the changes in the solid content, sodium and chloride of plasma following the administration by stomach tube at zero time of (A) 90 ml. of 3.5 per cent sodium chloride and (B) 350 ml. of 0.9 per cent sodium chloride. In (C) 400 ml. of a solution of per cent. sodium chloride and 0-32 per cent sodium bicarbonate was infused in 33 min during the period marked by the black rectangle. Abscissea: time in min.; ordinates: from above down, plasma sodium and chloride, Molar; plasma solids, g./100 g.; urinary sodium (continuous lines) and chloride (broken lines), m.equiv./min. The figures on the graphs give the urinary sodium concentration (Molar) at the times indicated. parallel with that of chloride but about 25 per cent higher, so that changes in the excretion of the salt can be followed by determination of either ion. In confirmation of the previous experiments [O'Connor, 1950], the increase in excretion of sodium and chloride was similar whether the salt was given in hypertonic solution (e.g. fig. 2A) or as isotonic solution (e.g. fig. 2B). However, the volume and concentration of the urine differed, as the figures show. The upper frames show the changes in sodium, chloride and total

6 242 O'Connor solid content of plasma from blood samples drawn from a carotid artery, The determinations in plasma were made to show which, if any, of these values could be correlated with changes in the urinary excretion of sodium and chloride. Clearly there was no correlation between urinary excretion and changes in the sodium or chloride content of the plasma. For example, in both fig. 2A and B there was an increase in sodium excretion, but plasma sodium was increased in fig. 2A and not in fig. 2B; and in fig. 2C, where the infusion fluid had approximately the same sodium and chloride concentration (sodium, M; chloride, 0x115 M) as plasma, a large excretion occurred without significant change in the plasma concentration of sodium or chloride. These are examples from 17 experiments in which sodium chloride was ingested or infused, and in which excretion of sodium and chloride was quite independent of changes in plasma sodium or chloride content due to hypertonicity of the administered solution (e.g. fig. 2A) or its high chloride content as compared to plasma (e.g. fig. 2B). On the other hand, in each of the experiments of fig. 2 there appears to be a correlation between the rate of excretion of sodium chloride and the solid content of the plasma, and these are typical of 17 experiments on 4 dogs. Firstly, similar dilution of the plasma solids occurred in 4 experiments where the sodium chloride was given as hypertonic solution (e.g. fig. 2A) as in 10 experiments when the same dose of salt was given in isotonic solution (e.g. fig. 2B), and similar increases in the excretion of sodium and chloride occurred under these two conditions. Secondly, the time course of the urinary response followed closely the time course of the fall in plasma solid content. This is best seen in fig. 2C, where the infusion at 12 ml. per min. caused a rapid fall in the plasma solid content, which rose again when the infusion was ended; the excretion of sodium and chloride rose and fell about 5 min. after the changes in plasma solid content. A time lag of about 5 min. was found also in two other experiments like that of fig. 2C. Relationship between the Solid Content of the Plasma and Sodium Excretion.-With the purpose of expressing in graphic form the apparent relationship of the rate of excretion of sodium to solid content of the plasma, a value for the plasma solids during each of the urine collection periods in fig. 2 was obtained by interpolation from the observed values, allowance being made for the delay of 5 min. mentioned above. In each experiment about 8 points were thus obtained for plotting as in fig. 3A, B, C, which are derived from the experiments of fig. 2A, B, C respectively. In each case the apparent correlation between rate of sodium excretion and the solid content of plasma is better expressed by a curved line of the type of those drawn in fig. 3 than by a straight line, and this was true of all of 17 experiments of this type which have been performed in 4 dogs. The data in each experiment individually was not sufficient to define

7 Excretion of Sodium Chloride 243 URINE Na m.equiv./min. / ~ ~ ~ ~ X /~~~~~~~~ 00~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~J PLASMA SOLIDS g./100 g. FIG. 3.-Plotting of results derived from fig. 2A, B, C. The rate of excretion of sodium in each urine collection period (ordinates: m.equiv. per min.) is plotted against the solid content of plasma (abscissoe: g. per 100 g.) estimated by interpolation from the observations of fig. 2A, B, C. the relationship more accurately, and so in fig. 4 the data from 6 experiments on " Skewbald " have been plotted on the same graph. To do so, allowance had to be made for the variation in the initial values for plasma solids from 7-6 to 8-35 g./100 g. on the six days. In preparing fig. 4 the following procedure was adopted. Individual graphs like those of fig. 3 were plotted on transparent paper and superimposed, each 05 URINE Na t/. m.equiv./min x0~~~~...t 0~~~~~ V w~~ 0o.X *O. 7 X 7' PLASMA SOLIDS g./100 g. FIG. 4.-"Skewbald", 15 kg. The relationship between the solid content of the plasma and the rate of sodium excretion in 6 experiments in which sodium chloride was given by stomach tube or intravenous infusion. Method of plotting is described in the text. Points from each individual experiment are plotted by a different symbol.

8 244 O'Connor graph being moved along the abscissa scale until the best fit was obtained. The amount which was thus added to all values of plasma solids in any one experiment varied from to This method of fitting together the results from different experiments was adopted on the assumption that there was a daily variation in the amount of solids other than proteins in the plasma, but that this amount would not change in the course of an experiment. Plotted in this way in fig. 4, the points from the 6 experiments lie quite close together, and the line was drawn freehand to express the relationship in this dog between changes in the plasma solid content and the rate of excretion of sodium. It was this line, transposed to the appropriate abscissa scale, which was drawn in each of the graphs of fig. 3; of the 6 experiments on " Skewbald " the points lay close to this line in 4 (e.g. fig. 3A and C), while in 2 the fit was less satisfactory (e.g. fig. 3B). Sufficient experiments were performed with two other animals to allow graphs like that of fig. 4 to be constructed. In each case the curve had the same general characteristics; as the plasma solid values fell, the rate of excretion of sodium increased slowly, until at rates above m.equiv./min. the excretion of sodium increased rapidly with further dilution of the plasma solids. With "Skewbald" excretion of sodium occurred at 0-15 m.equiv./min. when the plasma solid value fell to 7-4 g./100 g. (mean of 6 experiments), and to 7-2 with "Bess" and "Black" (mean of 6 and 3 experiments). In fig. 4 ("Skewbald") the steeply rising part of the curve was drawn as a straight line such that each decrease of 0.1 g./100 g. in the plasma solid content was accompanied by an increase of 0O085 m.equiv./min. in the rate of exeretion of sodium; with "Bess" and "Black" and 0075 respectively. Sodium bicarbonate, ammonium chloride, or water were also given by stomach tube, with the object of testing whether the subsequent changes in sodium and chloride excretion could be explained by changes in plasma solid concentration, in accordance with the findings after the administration of sodium chloride (fig. 4). Renal Response to Sodium Bicarbonate.-On several occasions the dose of sodium given by stomach tube as bicarbonate was identical with that usually given as chloride. The rate of excretion of sodium increased whether the sodium was given as the chloride (fig. 2A and B) or as the bicarbonate (fig. 5A); but when bicarbonate was administered (as in fig. 5A) there was no increased excretion of chloride, the increased excretion of sodium being presumably accompanied by increased excretion of bicarbonate. In two experiments, of which fig. 5A is an example, arterial blood samples were drawn for plasma determinations. The plasma sodium was unchanged, as might be expected from the ingestion of an isotonic solution of sodium bicarbonate, and the plasma solid content fell. In

9 Excretion of Sodium Chloride each experiment the fall was rather smaller than that produced by sodium chloride, and its time course approximated to that of the increased excretion of sodium shown in the lower part of fig. 5A. Clearly dilution of the plasma protein may be an important factor in determining the rapid excretion of sodium following ingestion of sodium bicarbonate. Other factors must then operate to determine that the excreted sodium PLASMA Na (M) 245 CI (M) SOLIDS g./ioog URINE Na. Cl m.equiv./min TIME (min.) FIG. 5.-" Skewbald ", 15 kg. Changes in the plasma and urine following the administration by stomach tube of (A) 4-5 g. sodium bicarbonate in 350 ml. of water, (B) 2 9 g. ammonium chloride in 120 ml. of water. Plotting as in fig. 2. is accompanied in the one instance by bicarbonate, in the other by chloride, but this has not been investigated in the present experiments. The question still remains whether the nature of the urinary anions (bicarbonate or chloride) or the production of alkalosis can affect sodium excretion, and so modify the effects of dilution of the plasma protein. A first attempt to answer this question is shown in fig. 6. Here the data from the two experiments with sodium bicarbonate have been plotted in the same way as the data from experiments with sodium chloride were plotted in fig. 4. The points from each of the experiments with sodium bicarbonate was brought as close as possible to the line drawn in fig. 4 to represent the relation between plasma solid and rate of excretion of sodium after the administration of sodium chloride. The

10 246 O'Connor data from the two bicarbonate experiments can be fitted to this line, but the experiments are too few to justify a firm conclusion that sodium excretion after the administration of sodium bicarbonate is solely determined by dilution of the plasma proteins, and is not influenced by substitution of bicarbonate for chloride as the urinary anion, or by alkalosis. Renal Response to Ammonium Chloride.-In other experiments, one of which is illustrated in fig. 5C, ammonium chloride was given by stomach tube in a dose containing the same amount of chloride as E the usual dose of sodium 0 3 chloride; this produced URINE / * in the next 21 hrs. no Na 0 increase in the excretion of sodium and a small m.equiv./min. - increase in chloride excretion. The experiments o0 _.. 0 were not continued to include the rise in sodium excretion to about 0-15 l I ai m.equiv./min. which was found by Cort and PLASMA SOLIDS g./100 g. McCance [1954] 3-4 hrs. FIG. 6.-"Skewbald", 15 kg. The relationship be- after the intraperitoneal tween the solid content of the plasma and the rate of administration of ammonexcretion of sodium in 2 experiments in which sodium bicarbonate was given by stomach tube. Plotting as ium chloride or sulphate. in fig. 4. The curve drawn in this graph is that of fig. 4. The absence of significant change in plasma sodium and the increase in plasma chloride resemble the findings of Cort and McCance [1954], and there was no large dilution of the plasma solids. In this instance, absence of an increase in the rate of excretion of sodium was associated with very small dilution of the plasma solids. Renal Response to Water.-The changes in plasma solid content were followed during the two hours after the administration of water to two dogs and the results in one are given in fig. 7. In 4 experiments collected in fig. 7A, the administration of ml. of per cent sodium chloride resulted in the usual excretion of chloride in the urine and in each case a fall of approximately 0-4 g./100 g. in the plasma solids. Administration of ml. of water produced the typical water diuresis with a fall rather than a rise in the excretion of chloride (fig. 7B). The changes in the plasma solid concentration were far less consistent than when saline was given, and in several experiments on the two animals considerable and persistent dilution of the plasma solids did occur (e.g. A, +, fig. 7B) without increase in the excretion of sodium or

11 Excretion of Sodium Chloride 247 chloride. Reference to graphs like that of fig. 4 showed, however, that even in these instances the dilution of the plasma solids was not sufficient to reach values at which rapid excretion of sodium or chloride would 0 TIME (min.) FIG. 7.-"Bessie", 19 kg. Changes in the solid content of the plasma (increase or decrease, g./100 g.) and mean rate of excretion of chloride (m.equiv./min.) (A) in 4 experiments in which ml. of 0 8-0*9 per cent sodium chloride and (B) in 4 experiments in which ml. of water was given by stomach tube at zero time. Abscissee: time in min. Each experiment is indicated by a separate symbol. occur. To this extent water diuresis conforms to the general conclusion that rapid excretion of sodium and chloride (i.e. at rate above m.equiv./min.) is dependent upon dilution of the plasma proteins below a critical value. II. Effect of Occlusion of the Carotid Arteries Description.-Fig. 8A and B are examples of the changes which occurred in urinary volume, sodium and chloride when both carotid arteries were occluded during the course of the urinary response to the administration of sodium chloride by stomach tube. During the period of occlusion the urinary volume (curve V) was increased, without great change in the concentration of the urine (curve U), so that the rate of excretion (curve UV) of sodium (fig. 8A) or chloride (fig. 8B) was also increased. The experiment has been performed 36 times on 3 dogs, always with the same result, the rate of urine flow and excretion of 60

12 ~~~~~~~~~~~~~~~~~~~~~~~~~~~~I'-,...v, I 248 O'Connor sodium and chloride being increased by per cent. Fig. 9A provides a further example. In fig. 8B the urine was collected in periods of 3-min. duration in order to determine as accurately as possible the time course of the change. In this, as in other similar experiments, the increase in the volume of the urine was practically complete in the first 3-min. period, and when the compression was released, the urine volume fell equally quickly. The speed of the response is such as to make it unlikely that A URINE B URINE VOL. URINE Na mli /min CI 03 U UV m equiv./min. -r m-equiv /min (M) -2 M UV TIME (min.) FIG. 8.-"Skewbald", 15 kg. Changes in the urine during occlusion of the carotid arteries. 350 ml. of 0 9 per cent sodium chloride was given by stomach tube at zero time. Abscissoe: time in min. Curve V shows the volume of the urine (ml./min., scale in centre); U shows in experiment (A) sodium and in (B) chloride concentration of the urine (Molar, scale to left and right). Curve UV shows the rate of excretion of (A) sodium and (B) chloride (m.equiv./min., scale to left and right). Both carotid arteries were occluded during the periods indicated by the black rectangles. the changes on carotid occlusion can be due to changing concentration in the blood of a circulating hormone, and it has not been possible to imitate the effect of carotid occlusion by infusion of hormones believed to affect renal function. The response to occlusion is entirely different from that produced by infusion of extract of the posterior lobe of the pituitary [O'Connor, 1950], and it has already been shown that saline diuresis is not influenced by DCA. Adrenaline and nor-adrenaline may be released during carotid occlusion, but it has been found that under the conditions of these experiments their infusion caused decreased excretion of salt (unpublished observations). Hormonal mediation being thus unlikely, the possibility was examined that the urinary changes during carotid occlusion might be produced via the renal nerves.

13 Excretion of Sodium Chloride 249 Effect of Denervation of the Kidneys.-In two animals, after the response to carotid occlusion had been established, the kidneys were denervated by severing all renal connexions except the artery, vein and ureter, which were carefully denuded of all visible nervous and connective tissue. Fig. 9B shows that the renal response to occlusion of the carotid arteries was not altered by this procedure, and in a second I "* E el0 * (A) 04- o 0 00~~~~ L ( inmi.afe030ml0f 60 -bsci0 00 o- 00t 0pi z FBo.cao ar-ttyer we.e occlusion of both catid arther (A) before operation, (B) days u after denervation of the kidneys, and (C) 4 months later,18 days after section of the splanchnic nerves on both sides. Abscissy: time in mm. after 300 ml. of 09 per cent sodium chloride by stomach tube. Ordinates: lower, rate of excretion of sodium (m.equiv./min.); upper, systolic and diastolic blood pressure (sm. Hg). Both carotid arteries were occluded during the periods indicated by the black rectangles. animal also denervation did not diminish the response. Sixteen weeks after denervation of the kidneys the splanchnic nerves on both sides were cut as they emerged from under the crura of the diaphragm, and fig. 9C shows that 18 days later carotid occlusion still increased sodium excretion. Clearly the response to carotid occlusion is not mediated by the renal nerves. Changes in Arterial Blood Pressure.-In the experiments of fig. 9 the arterial blood pressure was recorded by the method of O'Connor [1955]. As described in more detail in that paper, the blood pressure rose by mm. Hg on occlusion of both carotid arteries, and had reached its maximum at the first reading one minute after application of the occluding pressure, and fell within one minute of release of the carotid occlusion. The fact that the changes in the urine also occurred quickly,

14 250 O'Connor and the absence of evidence for hormonal or nervous mediation, suggests that the increased excretion of water and salt during carotid occlusion was the direct effect on the kidney of increased arterial blood pressure. A few experiments in which one carotid artery was occluded agree with this explanation: the increase in blood pressure was about one-third of that produced by occlusion of both carotid arteries and the excretion of salt and water increased by about 25 per cent, a third of the increase which resulted from occlusion of both vessels. In attempts to increase the blood pressure by means other than carotid occlusion, adrenaline, nor-adrenaline and 5-hydroxy-tryptamine have been infused intravenously. All had very little effect on the blood pressure of the conscious dog, and any effect on the urine flow was a decrease, presumably due to their constrictor action on the renal vessels (unpublished observations). With one of the animals, spontaneous increases in blood pressure occurred in some experiments associated with mild restlessness [O'Connor, 1955], and then the rate of excretion of salt and water also increased. So unpredictable a circumstance could not be investigated further, and so no procedure has been available to decide whether increased blood pressure from causes other than carotid occlusion would produce increased excretion of sodium. DIscuSSION The experiments have disclosed two factors which may act on the kidney to increase the rate of excretion of sodium: dilution of plasma solids (i.e. protein) and increased arterial blood pressure. Each may be expected to increase glomerular filtration, and so the conclusions are in accordance with the accepted view that glomerular filtration rate is one determinant of sodium excretion. As has been emphasized by Berliner [1950], accurate appraisal of other factors which may influence sodium excretion is impossible without knowledge of the quantitative relationship between the rate of glomerular filtration and the rate of excretion of sodium, and this cannot be determined directly for reasons which will be stated later in this discussion. Indirect evidence however allows estimates to be made of the changes in glomerular filtration rate which are involved in these experiments. In fig. 10, the curve of fig. 4 has been redrawn with calculated abscissa scales to show the changes in plasma protein concentration and in glomerular filtration presumably associated with the changes in plasma solid concentration actually recorded. The scale showing plasma protein was calculated on the assumptions (1) that the plasma contained 0*9 g. per 100 g. of sodium salts, the concentration of which was not changed significantly by the ingestion of sodium chloride; and (2) that the plasma contained 1 g. per 100 g. of solids other than protein (fats, lecithin, etc.) which were diluted equally with the protein, but did not

15 Excretion of Sodium Chloride 251 otherwise change in the course of any one experiment. The scale showing glomerular filtration rate was based on data of Wesson, Anslow, Riasz, Bolomey and Ladd [1950] from experiments in which large quantities of saline solution were infused. In two experiments of which protocols are presented in that paper, plasma protein fell from 5-7 to 4-1-4*5 g./100 g., and from 6-6 to 4-5 g./100 g., and glomerular filtration rate increased from 47 to and from 40 to ml./min. E z z 0.5 E ~~~~~~~~~~~~~/ E ~~~~~~~~~~~~~~/ v03 0.I....~~~~~~~~~.. PLASMA PROTEIN g./i00 g. G.F.R ml./min. FIG. 10.-Replotting of fig. 4, with calculated abscissa scales showing plasma protein concentration (g./100 g.) and glomerular filtration rate (ml./min.). Ordinates: rate of excretion of sodium (m.equiv./min.). The assumptions used to derive the scales are explained in the text. -i.e. for a fall of 0-1 g./100 g. in plasma protein there was an increase of 1.15 ml./min. in glomerular filtration rate. In the scale of fig. 10 the initial glomerular filtration rate was placed as 45 ml./min., and 1.15 ml./min. added for each fall of 0-1 g./100 g. in the plasma protein concentration. In the absence of direct measurements of the changes in glomerular filtration rate, fig. 10 depicts the probable relationship between plasma protein concentration, glomerular ifitration rate and sodium excretion when the plasma is diluted by the ingestion of sodium chloride. In fig. 11, comparison is made between dilution of plasma protein and increased arterial blood pressure as causes of increased sodium excretion. Curve A, fig. 11, is replotted from fig. 10, and so expresses the relationship between plasma protein concentration and sodium excretion in experiments on "Skewbald" without carotid occlusion, when the mean arterial pressure was 83 mm. Hg. The results of 19 experiments in which the carotid arteries were occluded have been VOL. XL, NO

16 252 O'Connor plotted as in the following example. In fig. 8a the rate of excretion of sodium in the absence of carotid occlusion would have been 0*15 m.equiv./min. but, as the result of the occlusion, was increased to 0-31 m.equiv./min. In plotting this experiment on fig. 11, it was first found from curve A that excretion of 0-15 m.equiv./min. corresponds to plasma protein content of _ ' I _ 5-5 g./100 g., and the occlusion experiment is then plotted as 5, The 19 points from 05s/_X.* _ occlusion experiments on this dog fell close to a line (B) parallel to (A), but moved. t z _ 2 abscissa divisions (i.e. E S g./100 g. plasma protein) >0 / /..to the left, and this new line *@ ;g.. thus represents the relation 0-3 _ / between plasma solids and to X..sodium excretion when the z~~~~ LU o o...mean arterial blood pressure z _,.*._ was raised to 133 mm. Hg by v v... carotid occlusion. Thus in 0oI... a I 56 5 PLAsMA PROTEIN 5g./109g.) this animal an increase of arterial pressure by 50 mm. Hg was equal in its effect to a fall of 0417 g./100 g. in plasma protein, which, according to 509 the assumptions on which the PLASMA POscale of fig. 10 is based, would FIG. 11.-"Skewbald", 15 kg. Comparison of be accompanied by an increase increased blood pressure from carotid occlusion of n. with dilution of plasma in protein as means glomerular of increasing sodium excretion. Curve A shows filtration rate. In two other the relation between sodium excretion and plasma protein concentration at the normal anmals, risesofbloodpressure mean blood pressure of 83 mm. Hg (replotting of 41 and 39 mm. Hg from of fig. 10). Curve B shows the relation with g mean blood pressure of 133 mm. Hg during carotid occlusion caused incarotid occlusion, the points from individual creased sodium excretion equal experiments being plotted as described in the text. to that from falls of 0 21 and 0415 g./100 g. in plasma protein with estimated increases of glomerular filtration rate of 2-5 and 1-7 ml./min. These estimates agree in many respects with the findings of other workers. Taking the mean of the 3 animals, a rise of 43 mm. Hg is thus equated in its effects with a fall of 0.18 g./100 g. in plasma protein, causing an increase of 2-1 ml./min. in glomerular filtration rate. Winton [1951] states that an increase of 10 mm. Hg in arterial blood pressure causes the glomerular filtration rate to increase by 0 33 ml./min. in

17 Excretion of Sodium Chloride 253 anaesthetized dogs, by 1-12 ml./min. in anesthetized dogs with kidneys denervated, and by 0-6 ml./min. in perfused kidneys. The increase of 2-1 ml./min. associated above with an increase of 43 mm. Hg in blood pressure is within this range. Eggleton, Pappenheimer and Winton [1940] compared the effect of changes in arterial pressure and of plasma dilution in the perfused kidney and in anaesthetized dogs. They assumed that the glomerular blood pressure was 60 per cent of the mean arterial pressure, and that a fall of 1 g./100 g. in plasma protein concentration meant a fall of 4 mm. Hg in the osmotic pressure of the plasma proteins. Using these factors, an increase of 43 mm. Hg in mean arterial blood pressure and a fall of 0-18 g./100 g. in plasma protein would cause increase of effective filtration pressure by 26 mm. Hg and 0-72 mm. Hg respectively; i.e. dilution of plasma protein appears to be 36 times more effective than increase in blood pressure in promoting sodium excretion. Eggleton, Pappenheimer and Winton [1940] found that the ratio was 6: 1 in perfused kidneys and 15: 1 in innervated kidneys of anmesthetized animals. The discrepancy is presumably due to the same factors as those which limit changes in blood flow in the kidneys when the arterial pressure is changed. These factors have been discussed by Winton [1951], and may well operate even more effectively in the conscious dog than in anaesthetized animals or in the perfused kidney. It appears from figs. 10 and 11 that measurements of inulin or creatinine cannot attain the accuracy needed to plot the relationships more directly. Under the most favourable conditions of stable plasma levels and high, stable rates of urine flow, the standard deviation of successive estimations of inulin clearance is 7-9 per cent [Smith, Goldring and Chasis, 1938; Ferguson, Olbrich, Robson and Stewart, 1950; Mandell, Jones, Willis and Cargill, 1953]. In experiments testing the effect of different rates of infusion of saline, such stable conditions cannot be obtained and still larger errors may be expected, chiefly due to the effect of the renal dead space. Even if an accuracy of 8 per cent were obtained, it would not be possible in individual experiments to distinguish glomerular filtration rates of 52 and 56 ml./min., which in fig. 10 are associated with change of sodium excretion from 0-14 to 0 47 m.equiv./min. The absence of demonstrable changes in inulin or creatinine clearance during changes in the rate of excretion of sodium has in recent years often been cited as proof of variations in the tubular reabsorption of sodium, but fig. 10 suggests that the errors inherent in clearance measurements are such as to make this argument invalid in most instances. It may be added that this difficulty persists despite the calculation of rates of reabsorption of sodium or more complex mathematical manipulation of the two variables, glomerular filtration rate and rate of excretion of sodium. Brief consideration will now be given to the shape of the curves of

18 254 O'Connor figs. 4 and 10. On the steeply rising part of the curve the slope was such that, using the mean of 3 animals, an increased excretion of sodium of m.equiv./min. was obtained for each increase of 1 ml./min. in glomerular filtration rate. In the main, these curves were constructed from experiments in which isotonic solutions had been administered by stomach tube or intravenously, so that the plasma sodium concentration remained unchanged throughout each experiment, the mean being M. Thus an increase of 1 ml./min. in glomerular filtration rate meant an increase of m.equiv./min. in the load of sodium filtered through the glomerulus, and of this increase m.equiv./min. or 60 per cent appeared in the urine. This may be interpreted as meaning that the effect of increasing glomerular filtration rate is to saturate a part of the mechanism of tubular reabsorption of sodium, with the corollary that there is a tubular maximum for this part. However, care must be taken in identifying this example with the excretion of substances such as glucose where tubular maxima have been clearly defined, because of the results of those experiments in which sodium chloride was given by stomach tube as hypertonic solution. Then, in addition to dilution of the plasma solids nearly equal to that produced by isotonic salt solutions, the plasma sodium concentration increased from 0x153 to 0x158 M but the rate of excretion of sodium was no higher. Thus in the conscious dog with small changes in filtration rate or plasma sodium concentration, increase in glomerular filtration rate appears more effective in causing urinary excretion of sodium than increased load due to a higher sodium concentration in the glomerular filtrate. Selkurt [1954] has come to a different conclusion when reviewing experiments in which large increases in plasma sodium were produced in anmesthetized dogs by infusion of hypertonic solutions. Whatever intrinsic renal processes may be involved, the relationship of figs. 4 and 10 must be of importance in the regulation of the body content of sodium. According to the graphs, dilution of plasma protein results in increased excretion of sodium; retention of sodium in the body involves expansion of the extracellular fluid compartments of the body with dilution of the plasma protein. In the renal excretion of sodium, dilution of the plasma protein is thus a stimulus which increases when the body content of sodium increases. No other factor causing increased excretion of sodium has been definitely shown to be increased by body retention of sodium, and so the plasma protein concentration could play a role in regulation of the sodium content of the body similar to that of the CO2 tension of the plasma in the regulation of respiration. In a more quantitative consideration, at the end of the two experiments with " Skewbald ", in which sodium chloride was given by intravenous infusion, 35 and 36 m.equiv. of sodium had been retained and the plasma solids were 0-62 and 0 54 g./100 g. below the initial levels. Thus retention of 6-5 m.equiv. of sodium was associated with each fall of 0.1 g./100 g. in

19 Excretion of Sodium Chloride 255 the plasma solid; which on the steeply rising part of the curve of fig. 4 will determine an increase of m.equiv./min. in the rate of excretion of sodium. Clearly the mechanism represented in figs. 4 and 10 provides a process by which a large excess of sodium is rapidly excreted and the plasma protein concentration returned to near the critical level. The efficiency of this mechanism in the dog is indicated by Ladd and Raisz [1949], who fed a meal, containing 60 g. of sodium chloride. Sodium balance was maintained on the high intake without a large increase in body weight. Twenty hours after such a meal the rate of excretion of sodium in the urine was 0-08 m.equiv./min., which was very little larger than the rate of 0 05 m.equiv./min. on those days when the meal contained only 5 g. of sodium chloride. Fig. 4 indicates that this difference would be determined by a difference of 0-25 g./100 g. in plasma solids, representing the retention of 16 m.equiv. of sodium (0.94 g. of sodium chloride) and an increase of only 110 ml. in the volume of the extracellular fluids. In ordinary life intake of such large amounts of sodium does not occur, whereas periods of low sodium intake or sodium depletion are more common, when the urinary sodium falls to very low levels. The mechanism of salt conservation cannot be deduced from the present experiments, which were concerned only with high salt intakes. The experiments in themselves give no justification for extrapolation to the left of the curves of figs. 4 and 10; the shape of the curves make it unsafe to assume that factors important at high rates of excretion of sodium will be of equal importance at lower rates: the two circumstances require separate investigation. However, the work of Mueller, Surtskin, Carlin and White [1951] on the effects in the dog of partial constriction of the renal artery suggests that, at low rates also, the excretion of sodium can be related to changes in glomerular filtration rate Ṫhe results in this paper are insufficient to decide to what extent the relationship between sodium excretion and the plasma protein concentration may vary from time to time in the same dog or may be altered by experimental procedures. The most obvious difference between experiments on the same dog was the wide range of values of plasma solids, but, as mentioned already, this may have been due to differences in the plasma content of solids other than protein; and the relationship between plasma protein concentration and the rate of excretion of sodium could be quite stable for each animal. In general, the curve relating plasma protein concentration to the rate of excretion of sodium would be moved to the left by factors which increase the glomerular filtration rate (e.g. increased arterial pressure, fig. 11), or by factors which decrease the tubular reabsorption of sodium. Plotting the relationship between plasma protein concentration and sodium excretion under controlled conditions is likely to prove a more accurate

20 256 O'Connor way of assessing factors influencing the renal excretion of sodium, than has hitherto been achieved by procedures taking no account of the large changes in sodium excretion which may result from small changes in glomerular filtration rate. The effect on sodium excretion of the hormones of the suprarenal cortex, or of associated urinary anions, could be so investigated, to mention two circumstances which have been incompletely considered in this paper. SUMMARY 1. The excretion of sodium chloride administered by stomach tube to the conscious dog was not altered by previous denervation of the kidneys or by injections of desoxycorticosterone acetate. 2. The excretion of administered sodium chloride was not determined by changes in the sodium or chloride concentration of the plasma, but there was a close relationship between the rate of excretion of sodium and total solid content of the plasma, which was used to follow changes in the plasma protein concentration. Ingestion of ammonium chloride or water caused no increase in sodium excretion, and there was no large fall in plasma solid. Ingestion of sodium bicarbonate caused an increased excretion of sodium associated with dilution of the plasma solids, but there was no corresponding increase in the excretion of chloride. 3. Occlusion of both carotid arteries caused immediately an increased output of urine and of both sodium and chloride. This persisted after denervation of the kidneys, and was attributed to the increased arterial pressure produced by the occlusion. 4. The relationship between plasma solid content and rate of excretion of sodium has been plotted, and from this an estimate made of the probable relationship between plasma protein concentration, glomerular filtration rate and sodium excretion. The effects of dilution of plasma protein and of increased arterial blood pressure in increasing sodium excretion have been compared quantitatively. 5. Measurements of inulin or creatinine clearance are not sufficiently accurate to elucidate problems of sodium excretion. 6. The relationship between plasma solid content and rate of excretion of sodium is discussed with reference to the sodium balance of the body. ACKNOWLEDGMENTS The expenses of this work were met in part by the Government Grants Committee of the Royal Society. My thanks are due to Dr. W. J. Allen and other members of the Department who assisted at the operations, and especially to Mr. K. A. Pearce for his technical assistance.

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