J. Phyaiol. (1975), 245, pp. 137-162 With 5 text-ftgure8 Printed in Creat Britain

137

RENAL FUNCTION IN SHEEP DURING INFUSION OF ALKALI METAL IONS INTO THE RENAL ARTERY

BY A. M. BEAL AND F. A. HARRISON From the Agricultural Research Council Institute of Animal Physiology, Babraham, Cambridge CB2 4AT

(Received 22 April 1974) SUMMARY

1. The effect on renal function of 1 M solutions of LiCl, NaCi, KC1, RbCl and CsCl and 3 M-NaCl infused close-arterially to the kidney for 10 min at 0*7 ml./min has been studied in nine experiments on four unilaterally nephrectomized sheep. The levels of flow, electrolyte concentration and electrolyte excretion in the urine were measured before, during and for 50 min after the infusions. 2. The infusion of 1 M-NaCl produced little change in urine flow and composition whereas 3 m-NaCl resulted in relatively small increases in urine flow and sodium excretion. 3. The infusion of lithium, potassium, rubidium and caesium resulted in marked increases in urine flow, urinary sodium concentration and excretion, urinary potassium excretion and osmolal clearance while the urinary potassium concentration decreased. 4. Changes in urine flow and urinary pH during the infusions of all the alkali ions except sodium were consistent with increased urinary bicarbonate excretion. 5. The osmolal clearance was increased by the infusion of lithium, potassium, rubidium and caesium, but equivalent increases in the rate of solutefree water reabsorption did not occur. 6. The infusion of caesium resulted in a depression of the glomerular filtration rate (G.F.R.) which was not observed when the other alkali ions were infused. 7. The effects of lithium, potassium and rubidium on urine flow and composition were rapid in onset and the residual effects of these ions, on cessation of infusion, were relatively short. The effects of caesium were slow in onset and prolonged in duration. 8. It was concluded that lithium, potassium, rubidium and caesium altered urine flow and electrolyte excretion by acting upon common mechanisms which were predominantly intra-renal and located in the proximal segment of the nephron.

138

A. M. BEAL AND F. A. HARRISON INTRODUCTION

Elevation of the plasma potassium concentration, by ingestion or by infusion of potassium salts has been found to alter renal electrolyte excretion in a number of species including the sheep. Typically, hyperkalaemia is associated with increased excretion of sodium, potassium, chloride, bicarbonate and water and decreased excretion of hydrion (Vogel, 1959; Dewhurst, Harrison & Keynes, 1968; Beal, Budtz-Olsen, Clark, Cross & French, 1973). Other members of the alkali metal series have been substituted for sodium or potassium in various biological situations and their administration has resulted in changes in urinary composition which bear striking similarities to those produced by potassium administration. The ingestion of rubidium inhibits the renal excretion of acid thus preventing the metabolic alkalosis of potassium depletion and producing acidosis in both normal and potassium-depleted rats (Relman, Roy & Schwartz, 1953). To a lesser extent caesium ingestion also blocks the renal excretion of acid during potassium deficiency (Hall & Relman, 1959). In the dog the infusion of acidified lithium solutions suppressed the rate of hydrion secretion so that renal excretion of bicarbonate was increased (Orloff & Kennedy, 1952) and the infusion of isotonic lithium solutions produced concurrent increases in urine flow and in the renal excretion of sodium and potassium (Foulks, Mudge & Gilman, 1952). The ingestion of lithium by man has been associated with an increase in renal elimination of sodium approximating the quantity of lithium administered (Trautner, Morris, Noack & Gershon, 1955). Despite the similarities in the renal response to the administration of the alkali ions there has been no attempt to compare their actions on the kidney under uniform conditions. The following series of experiments was designed to compare the effects on renal function of equivalent increases in the concentration of the alkali metal ions in the plasma perfusing the kidney of sheep and to ascertain whether the effects on renal function were primarily of intra-renal or extra-renal origin. A preliminary report of this work has been presented to the Physiological Society (Beal & Harrison, 1973). METHODS

Experimental procedure The four Clun Forest x Merino x Welsh Mountain ewes (body weight 49-65 kg) used in the nine experiments reported in this paper were surgically prepared over several years to enable other studies to be made on renal function (see Harrison, Keynes & Pearson, 1969). Each sheep had the left adrenal gland transplanted to a neck loop (McDonald, Goding & Wright, 1958) and, as a consequence of this operation, was left nephrectomized. To make the animal dependent on the transplanted

ALKALI METALS ON RENAL FUNCTION

139

gland for adrenal steroids, the right adrenal gland was removed and to eliminate any ovarian steroid influence both ovaries were removed. Each animal had the right carotid artery exteriorized in a skin loop. At a final operation permanent clear vinyl catheters were implanted into the right renal artery (0.45 mm i.d., 0-75 mm o.d. expanded at one end to 0-9 mm i.d.; Dural Plastics Ltd, Dural, Australia) and right renal vein (1-4 mm i.d., 2-0 mm o.d.; Portex Ltd, Hythe) using the technique of Herd & Barger (1964) at least 4 weeks before the experiments and these catheters were maintained as described by Harrison, McDonald & Olsson (1970). Six days before the infusion experiments the sheep were transferred into metabolism cages and maintained on 1000 g hay chaff and 200 g crushed oats with distilled water for drinking and no salt supplementation. A 14 FG Foley catheter with a 5 ml. balloon was inserted into the urinary bladder 16-18 hr before the commencement of the experiment. This procedure was assisted by the application of a topical anaesthetic solution ('Xylocaine' spray; Astra Chemicals Ltd, Watford) to the vulva and urethral orifice and by lubricating the tip of the catheter with sterile anaesthetic jelly ('Xylocaine' Gel; Astra Chemicals Ltd, Watford). Any uneaten food was removed at this time. On the morning of the experiment, a nylon catheter (2-1 mm o.d., Portex Ltd, Hythe) was inserted into the right jugular vein and the right carotid artery was catheterized with a disposable plastic catheter (Braunula, size 1; Armour Pharmaceutical Company Ltd, Eastbourne). Both catheterizations were done under local anaesthesia (Xylocaine 2 % with adrenaline 1: 80,000; Astra Chemicals Ltd,Watford). Following an injection into the jugular catheter of 40 ml. of a solution containing 8% inulin (Thomas Kerfoot & Co. Ltd, Vale of Bardsley, Lancashire) and 1 % para-aminohippurate (Merck, Sharpe & Dohme Ltd. Hoddesdon, Hertfordshire) in pyrogen-free sterile water the same solution was infused at approximately 0-35 ml./ min for the remainder of the experiment. After a 90 min period of stabilization, the close-arterial infusions through the renal artery catheter began witb a 30 min infusion of isotonic saline containing potassium, 5 m-mole/l. Thereafter, each hour of the experiment was divided into an initial 10 min interval during which a test solution was infused followed by a 50 min recovery period when the isotonic saline solution was again infused. The infusion rate into the renal artery was approximately 0-7 ml./ min throughout the experiments. The first test solution was invariably 1 M-NaCl and subsequent test infusions were any of the following solutions, 1 M-NaCl, 1 m-KCl, 1 m-LiCl, 1 m-RbCl, 1 M-CsCl or 3 m-NaCl. These solutions were given in a number of different sequences but all possible combinations were not attempted. All infusates were not necessarily given on each occasion and no solution was given more than once in each experiment with the following exceptions. 1 M-NaCl infusion was given twice during most experiments. In four experiments 1 M-CsCl was infused immediately after the 50 min recovery period of the previous caesium infusion and was followed by a recovery period of 2 hr. Urine samples were taken at 10 min intervals except during the period of test infusion when the sampling periods were either 2, 3 and 5 min or two 5 min periods if the urine volume was low. Blood samples were taken from the carotid artery and renal vein to coincide with the mid-point of the 10 min urinary collection immediately preceding the first test infusion (5 min before the hypertonic infusion), with the midpoint of the second 5 min urinary collection of the hypertonic test infusion and with the mid-point of the last urinary collection of the recovery period following the test infusion (45 min after the hypertonic infusion). Thereafter, the same time sequence for blood sampling was used for each hourly period of the experiment and because of the serial nature of the experiments the sample at 45 min of the recovery period following one test infusion was also the sample at 5 mi before the next hypertonic test infusion. During the 2 hr recovery periods allowed after the repeat caesium infusions blood samples were taken at 45 mi and at the mid-point of the last urinary

140

A. M. BEAL AND F. A. HARRISON

collection (1 15 min after the hypertonic infusion). In the first two experiments another blood sample was taken from both vessels 5 min after the hypertonic test infusions but as the plasma inulin and para-aminohippurate (PAH) concentrations did not vary greatly this sampling was omitted for the sake of reducing the blood loss suffered by the animals. Analytical methods Blood samples were withdrawn into glass syringes heparinized with one drop of heparin (5000 i.u./ml.) and centrifuged in plastic tubes at 4500 rev/min for 20 min to obtain plasma for analysis. Urinary pH was estimated at 390 C immediately after collection using Radiometer micro-electrodes. Micro-haematocrit determinations were made in triplicate within 30 min of blood sampling using a micro-haematocrit centrifuge spinning at 11,000 revlmin for 10 min. Inulin in the urine and plasma was estimated in duplicate by the colorimetric method of Heyrovsky (1956) adapted to the Technicon autoanalyser by Dawborn (1965). The inulin clearance was calculated as follows. Inulin clearance (Cl.)

=

urinary inulin excretion rate plasma inulin concentration (mid-period)

Variations in plasma inulin concentration during each experiment were small and did not appear to be associated with any type of infusion. As the concentration of inulin rose slightly with the increasing duration of the experiment (r = 0-96; P < 0.001) the plasma inulin concentrations for periods between blood samples were calculated as a linear function of time from sample to sample. PAH was estimated in duplicate by the method of Bratton & Marshall (1939) as adapted for the Technicon autoanalyser by Harvey & Brothers (1962). The PAH clearance was calculated as follows:

PAH clearance (CPAH) =

urinary PAH excretion rate plasma PAH concentration (mid-period)

The PAH concentration of plasma showed small variations which were not related to the type of infusion or to the elasped time in the experiment. The plasma PAH concentrations for periods between blood samples were calculated as for inulin. Sodium and potassium were estimated simultaneously by emission flame photometry in an oxygen-acetylene flame (Autotechnicon) using mixed standards to correct for the mutual interference. The reference background provided by lithium, 15 m-mole/l. in the diluent prevented variations in lithium concentration in the urine and plasma from affecting the estimation of sodium or potassium. The accuracy of the estimation of sodium was not affected by the levels of rubidium or caesium in the samples but these ions enhanced the estimated concentration of potassium. Calibration curves were produced for varying concentrations of potassium estimated in the presence of varying levels of either rubidium or caesium. Having estimated the concentration of rubidium and caesium by other means, the amount of enhancement of each potassium estimation in the presence of rubidium or caesium could then be calculated and subtracted to correct the potassium levels. Lithium was estimated by atomic absorption in an air-acetylene flame (PyeUnicam SP-90). Only sodium interfered with the accuracy of estimation at the concentrations of sample presented to the instrument and the inclusion of sodium 140 m-mole/l. in the diluting fluid and in the standards corrected this problem. Rubidium was estimated by atomic absorption in an air-propane flame (Pye-

ALKALI METALS ON RENAL FUNCTION

141

Unicam SP-90). Lithium and caesium did not affect the estimation and interference due to sodium and potassium was overcome by the inclusion of sodium, 100 m-mole/l. and potassium 100 m-mole/l. in the diluting fluid and standards. Caesium was estimated by emission flame photometry in an air-propane flame (Pye-Unicam SP-90). Only lithium affected the accuracy of this estimation and the addition of lithium 100 m-mole/I. to the diluting fluid and standards was sufficient to correct for the levels of lithium in the samples. The renal clearances of lithium, rubidium and caesium were calculated for the last urine samples of the individual recovery periods following their infusion (40-50 min after test infusion) using the alkali metal excretion rate for this period and the concentration of the ion in the plasma of a blood sample taken at the mid-point of this urine collection. During the long recovery periods which followed some caesium infusions this procedure was repeated for the blood sample taken at 115 min. Alkali metal clearance = urinary alkali metal excretion rate plasma alkali metal concentration The distribution volumes of the above ions were also calculated at the same intervals as the clearances using the same plasma values amount infused - total renal excretion from start of infusion to blood sample Volume = plasma concentration at 45 min (or 115 min) Osmolality was estimated in duplicate by freezing-point depression using a Fiske Osmometer. The osmolal clearance (Co.) and the solute free water reabsorption (T0H2o) were calculated as follows: urinary solute excretion rate COSM ~~ plasma osmolality TCH20 = CO-m - V During the hyperosmotic test infusions the osmolality of the plasma perfusing kidney was calculated from the known osmolal infusion rate, the arterial plasma osmolality and the renal plasma flow which was taken as the mean of the two PAH clearances prior to each test infusion. Although the PAH clearance values were elevated during some test infusions, for reasons discussed later, the true renal plasma flow was believed to be relatively unaltered by the treatments and the use of the lower renal plasma flow values gave the maximum plasma osmolality likely to occur. The osmolal clearance for all other periods was calculated using the mean plasma osmolality of all arterial samples taken during any one experiment. =

Statistical procedures Although attempts were made to achieve similar states of electrolyte balance for each experiment, considerable variation was found in the urinary excretion of electrolytes from experiment to experiment both between sheep and between experiments within the same animal. To reduce the effects of differences in electrolyte status, analysis of covariance was applied to the data for each corresponding period of the close-arterial infusions. The two covariates used for each parameter analysed were (a) the value obtained for the initial 10 min urine sample of each of the nine experiments and (b) the value obtained for the penultimate 10 min sample before each test renal infusion (filled circles in Figs. 1, 2 and 5). When the analyses of covariance produced significant variance ratios, the differences between individual treatments were found by t test.

142

A. M. BEAL AND F. A. HARRISON RESULTS

The mean values for the haematocrit, plasma osmolality and the concentrations of sodium and potassium in the plasma of carotid arterial blood collected before, during and after the infusion of each of the alkali metals are given in Table 1. TABLE 1. The haematocrit, osmolal concentration (m-osmolefl.) and the concentration of ions (m-mole/l.) in the plasma of carotid arterial blood before, during and after the close-arterial infusion of alkali metal solutions into the renal artery (mean ± s.E. of mean) 5 min 45 min 7-5 min before infusion into infusion after infusion Infusate Parameter 1 M-NaCl 31-74 + 1-58 32-90 + 1-61 Haematocrit 31-73 ± 1-58 297 + 4 294+ 2 298 + 4 Osmolality 142-7 +0-9 141-9 +0-8 142-5 +0-9 Na 443+ 0-12 4-50+ 0-11 4-36 + 0-11 K 1 M-KC1 31-20 + 2-36 29-96 + 1-92 31-05+ 1-80 Haematocrit 296 ± 5 295+ 4 297 + 5 Osmolality 142-8 +0-8 142-2 +1-2 142-7 +1-3 Na 4-41 + 0-12 4-49 0-12 K 5-04 + 0-13 29-03 + 1-53 29-08 + 1-49 29-47 + 1-41 1 M-LiCl Haematocrit 296 + 4 296 3 298+ 4 Osmolality 142-4 ±0-6 142-3 ±0-6 Na 143-2 + 0-7 4-34 + 0-12 4-44 + 0-11 K 4-36 ± 0-09 0 Li 0-46 + 0-01 1-05 0-04 1 M-RbCl 29-37 + 0-89 28-64± 1-40 28-55 1-19 Haematocrit 296 4 293 + 2 295 3 Osmolality 141-2 + 1-0 141-3 ±1-1 141-5 ±1-1 Na 4-28+ 0-15 K 4-19±0-12 4-15+0-13 0 Rb 0-08 ± 0-01 0-58 ± 0-06 28-29+ 1-00 28-43 + 0-99 1 M-CsCl 29-31+ 0-97 Haematocrit 295 + 3 293 + 3 293 + 3 Osmolality 142-1 +0-7 142-0 +0-6 140-6 + 1-0 Na 4-17+ 0-10 4-14 + 0-12 4-16±0-10 K 0 0-15 + 0-01 0-79 + 0-05 Cs ± ± 0-57 1-01 27-78 29-01 0-97 30-43 Haematocrit M-NaCl 3 295+ 3 297 3 297+ 3 Osmolality 141-3 + 1-3 142-4 +1-2 140-6 1-1 Na 4-35± 0-11 K 4-10 0-12 4-22+ 0-13

Close-arterial infusion of 1 m-NaCi (nine replications) A molar solution of sodium chloride was infused as the first test solution in all experiments and the infusion of this solution was repeated later in most experiments as an internal control. As no differences were observed in the results of the two periods of infusion, either in the raw data or after

143 ALKALI METALS ON RENAL FUNCTION statistical analysis, the data for the repeat infusion have not been presented in this paper. The infusion of 1 m-NaCl into the renal artery resulted in an immediate increase in the sodium concentration of the plasma perfusing the kidney which was estimated to increase by 2-13 m-mole/l. + 017 m-mole/l. (S.E. of mean). By the mid-point of the second 5 min clearance period of the arterial infusion, the mean sodium concentration was raised by 303+ 0 37 mmole/l. as a result of a small increase in the sodium concentration of the plasma being re-circulated to the kidney. These values were calculated by assuming that, during the sodium chloride infusion, renal plasma flow (RPF) did not vary greatly from the mean of the PAH clearance values for the two clearance periods immediately before the 1 M-NaCl infusion. The urine flow fell slightly after the commencement of the sodium infusion and this was accompanied by a small increase in solute-free water reabsorption and by increases in the sodium and potassium concentrations of the urine sufficient to maintain the excretion rates of these ions at pre-infusion levels (Fig. 1). Urinary pH rose before this first test infusion in some experiments but this trend was curtailed at the commencement of the NaCI infusion. The inulin and PAH clearances showed little change.

Close-arterial infusion of 3 m-NaCl (six replications) The infusion of 3 M-NaCl into the renal artery increased the mean sodium concentration of the plasma perfusing the kidney by 6-39 + 070 mmole/i. at the start of the infusion and by 8-15 + 091 m-mole/l. after 7-5 min of infusion. Simultaneously the urine flow, C0Hm, To20 and the excretions of sodium and potassium rose slightly. After the infusion the urine flow fell below the pre-infusion levels but the rate of excretion of sodium and potassium tended to be maintained by compensating increases in the urinary concentrations of these ions. Urinary pH was little affected during the infusion but subsequently fell to levels significantly below those of the 1 M-NaCl infusion. The inulin and PAH clearances were increased during the first 5 min of the 3 M-NaCl infusion and decreased during the first 10 min of the recovery period (Fig. 2). Statistically the results of this infusion were not different from those of the 1 m-NaCl infusion with the exception of urinary pH.

Close-arterial infusion of 1 m-KCI (seven replications) The infusion of potassium chloride into the renal artery increased the concentration of potassium in the plasma perfusing the kidney by an estimated 2-04 + 0-17 m-mole/l. at the start of infusion and by 2'67 + 0-21 m-mole/l. at the mid-point of the second 5 min clearance period of the potassium infusion. The infusion of potassium was accompanied by

144

A. M. BEAL AND P. A. HARRISON

marked increases in urine flow rate, osmolal clearance and in the excretion rates of sodium and potassium to levels above those observed during the infusions of 1 M or 3 M-NaCl (Figs. 1 and 2). The differences between the three treatments were statistically significant (Table 2). Concurrent with these changes the urine sodium concentration rose and the potassium concentration fell. The concentrations of these ions at this time were significantly different from those of the 1 m-NaCl infusion but not the 3 M-NaCl infusion (Table 2). The mean urine flow rate during the first 2 min TABLE 2. Summary of the final t test analyses of the effects of infusion into the renal artery of 1 M-KC1 compared with the effects of similar infusions of 1 M-NaCl and 3 m-NaCl. Every parameter shown in Figs. 1, 2 and 5 was tested and omission from this Table means that no significant differences were found. Differences are indicated aslevelsofsignificance(* = P < 005;** = P < 0-01;*** = P < 0001; - = not significant). When the response to KC1 was less than that of another treatment t replaces * Infusion Recovery Al Parameter KCI , Degrees of freedom = 42 vS. 0 1 2 1 2 3 4 5 Urine flow ** NaCl *** *** * 3 m-NaCl - ** *** *** Osmolal clearance NaCl * *** ** * -

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._

145 ALKALI METALS ON RENAL FUNCTION rose during the potassium infusion so that the maximum fall in hydrogen on concentration was 51P3 n-mole/i., the greatest decrease occurring when the initial pH was lowest. The latter effect predominates in the combined K

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Fig. 1. Mean values for the urine flow, osmolal clearance (CoQm), solute free water reabsorption (TH20), urinary sodium concentration and excretion rate, urinary potassium concentration and excretion rate, urinary pH, inulin clearance (CI.) and PAH clearance (CpA) during and after the infusion of 1 m-NaCl, 1 M-KCI and 1 m-LiCl into the renal artery. The filled circles indicate the values of the covariates used in the statistical analyses of each of the renal parameters shown on the Figure.

60

146 A. M. BEAL AND F. A. HARRISON results given in Fig. 1. Both the inulin and PAH clearances gave high values during the first 5 min of potassium infusion and low values during the first sample of the recovery period. TABLE 3. Summary of the final t test analyses of the effects of infusion into the renal artery of 1 M-LiCl compared with the effects of similar infusions of 1 M-NaCl, 3 M-NaC1 and 1 M-KC1. Every parameter shown in Figs. 1, 2 and 5 was tested and omission from this Table means that no significant differences were found. Differences are indicated as levels of significance (* = P < 005; ** = P < 001; *** = P < 0001; - = not significant). When the response to LiCl was less than that of another treatment t replaces* Recovery Infusion Parameter LiCl Degrees of freedom = 42 VS. 0 1 2 1 2 3 4 5 ** Urine flow NaCl *** _-. 3 m-NaCl KCI ttt * * Osmolal clearance NaCl * 3 M-NaCl KCI Ittt * Solute-free water reabsorption NaCl * ** Urine Na excretion NaCl KCI - fI * *** ** Urine Na concentration NaCl *** _ * * Urine K excretion NaCl * ** 3 m-NaCl KCI - Ittt Urine K concentration NaCl - - ltt tt 3 m-NaCl - t t t t _ * ** Osmolal clearance/C1. NaCl KCI - ttt * * Urine Na excretion/CG. NaCl KCI - ttt ** Urine K excretion/C1. NaCl * 3 M-NaCl KCI - ttt t *

Close-arterial infusion of 1 M-LiCl (eight replications) The infusion of lithium chloride into the renal artery increased the lithium concentration of the plasma perfusing the kidney from zero to an estimated 2*10 + 0-17 m-mole/l. at the commencement of the infusion. The plasma concentration rose further to 3 14 + 0418 m-molefl. by the mid-point of the second 5 min of lithium infusion as a result of re-circulation of lithium. During the infusion, the urine flow,Co0m and Tc20 were increased as were the excretion rates of sodium and potassium (Fig. 1). With the exception

ALKALI METALS ON RENAL FUNCTION 147 of the Tc,0, these urinary parameters were increased to levels which were significantly higher than during the 1 M and 3 M-NaCl infusions but less than those of the potassium chloride infusion (Table 3). The mean urine flow for the 2 min immediately following the commencement of lithium infusion was 2- 18 + 0 18 times the flow prior to the infusion. The increased excretion of sodium was the result of simultaneous increases in urine flow and sodium concentration whereas the increased potassium excretion was dependent on the increase in urine flow since the potassium concentration in the urine fell (Fig. 1; Table 3). The mean urinary pH rose to levels similar to those observed during the potassium infusion and, in a similar way, the experiments could be divided into 3 responses. The maximum increase in hydrion concentration in the high urinary pH group was 1-43 n-mole/l. while the maximum decrease in concentration in the low pH group was 1406 n-mole/l. Both the inulin and PAH clearances were elevated during the first 5 min of lithium infusion and depressed during the first 10 min of the post-infusion recovery period (Fig. 1). The urinary concentration and excretion rate of lithium resulting from the infusion are given in Fig. 3. At the end of the 50 min recovery period the mean distribution volume of lithium in the sheep was 13-71 + 0-55 1. (s.E. of mean) which was 24-91 + 1'05 % of the body weight of the animals. The mean lithium clearance at this time was 12-12 + 0-99 ml./min or 25-90 + 1-55 % of the inulin clearance. Close-arterial infusion of 1 M-RbCl (seven replications) The infusion of rubidium chloride into the renal artery increased the rubidium concentration of the plasma perfusing the kidney from zero to an estimated concentration of 1-94 + 0-28 m-mole/i. at the start of the infusion. By the mid-point of the second 5 min of rubidium infusion this concentration of rubidium had risen to 2-51 + 0'30 m-mole/l. as a result of recirculation. During the infusion of rubidium, marked increases were observed in the urine flow, C0smand the rate of excretion of sodium and potassium (Fig. 2). With the exception of potassium excretion the levels of these parameters were significantly higher than those for the infusions of molar NaCl, KCl, LiCl and for 3 M-NaCl. The rate of excretion of potassium was significantly higher than that during 1 m-NaCl infusion but less than that during 1 M-KCl (Table 4). The mean urine flow for the first 2 min of the rubidium infusion was 1-84 + 0-24 times the flow rate before the infusion. Urinary sodium concentration was higher than during the infusion of sodium chloride at either concentration and the potassium concentration was lower than during any of the previously described infusions (Table 4). The mean urinary pH rose as a result of the rubidium infusion (Fig. 2), and the maximum increase in hydrogen ion concentration in those sheep with a high urinary pH was 4-5 n-mole/l. while the maximum decrease in hydrion

A. M. BEAL AND F. A. HARRISON

148

TABLE 4. Summary of the final t test analyses of the effects of infusion into the renal artery of 1 m-RbCl compared with the effects of similar infusions of 1 M-NaCl, 3 m-NaCl, 1 M-KC1 and 1 M-LiCl. Every parameter shown in Figs. 1, 2 and 5 was tested and omission from this Table means that no significant differences were found. Differences are indicated as levels of significance (* = P < 0-05; ** = P < 0-01; *** = P < 0-001; - = not significant). When the response to RbC1 was less than that of another treatment t replaces * Infusion Recovery Parameter RbCl 1 O 1 2 2 3 4 5 Degrees of freedom = 42 V8. *** *** Urine flow NaCl ** **$ ** 3 m-NaCl * KC1 LiCl ** *** ** * Osmolal clearance NaCl *** *** 3 m-NaCl ** *** *_ _KCl *** ** LiCl Solute-free water reabsorption NaCl *** KC1 LiCl * Urine Na excretion NaCl 3 M-NaCl ** KC1 * LiCl *** ** * Urine Na concentration NaCl ttt 3 M-NaCl Urine K excretion NaCl 3 m-NaCl KC1 ttt LiCl ** t Urine K concentration NaCl t ** ttt 3 m-NaCl ttt t KC1 tt LiCl tt* ** Osmolal clearance/CQ. NaCl ** * 3 M-NaCl *** KC1 *** ** *** LiCl *** Solute-free water *** reabsorption/CQ, NaCl *** ** 3 M-NaCl *** KC1 ** LiCl *

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149 ALKALI METALS ON RENAL FUNCTION concentration in the sheep with a low urinary pH was 2430 n-mole/I. Both the inulin and PAH clearances were elevated during the first 5 min of rubidium infusion and were less than pre-infusion values during the first 10 min of the period of recovery after the infusion (Fig. 2). The mean Rb 5,. -

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-

n---

*

(ml./min)

' ' ' | | ' ' ' '

0

Fig. 2. Mean values for the urine flow, osmolal clearance (CO,'m) solute free water reabsorption (T120), urinary sodium concentration and excretion rate, urinary potassium concentration and excretion rate, urinary pH, inulin clearance (Cl.) and PAH clearance (CPAH) during and after the infusion of 1 M-RbCl, 1 M-CsCl and 3 M-NaCl into the renal artery. The filled circles indicate the values of the covariates used in the statistical analyses of each of the renal parameters shown on the Figure.

60

150 A. M. BEAL AND F. A. HARRISON urinary concentration and excretion rate of rubidium are given in Fig. 3. By the end of the 50 min recovery period the distribution volume of rubidium was greater than 571., the upper limit being impossible to calculate as the rubidium concentration of the plasma had fallen to immeasurably small values. The mean rubidium clearance at the end of the recovery period for the four experiments in which plasma rubidium could be measured was 15-69 + 404 ml./min or 33-75 + 10-33 % of the inulin clearance.

Close-arterial infusion of 1 M-CsCl (eight replications) The infusion of caesium chloride into the renal artery produced an estimated plasma level of caesium 1P94 + 0-20 m-mole/l. initially and 2-72 + 0-21 m-mole/l. by the mid-point of the second 5 min of infusion. During the infusion urine flow, Cosm, TO 0 and the rates of excretion of sodium and potassium changed very little so that the levels of these parameters, while being less than those of the corresponding periods of the potassium, rubidium and lithium infusions, were not statistically different from those of the 1 M and 3 m-NaCl treatments (Fig. 2; Table 5). After the infusion prolonged rises in urine flow, Cosm and sodium excretion were observed and during this time, the levels of these parameters exceeded those of all other treatments (Fig. 2; Table 5). The urinary sodium concentration exceeded that following the 1 M-NaCl infusion during the first post-infusion period only and the potassium concentration of the urine, while not falling during the caesium infusion, fell to levels below that of the other treatments during the recovery period (Table 5). Urinary pH rose in all experiments but one (Fig. 2), the maximum decrease in hydrogen ion concentration being 762 n-mole/l. and the increase in hydrion concentration in the experiment where pH fell, being 1-52 n-mole/l. Both the inulin and PAH clearances underwent small changes which appeared to be related to the rate of urine flow during the caesium infusion. The inulin clearance was lower after the infusion than before and significantly less than at the corresponding stage of the other treatments (Fig. 2; Table 5). The PAH clearance was not depressed significantly during this period of recovery. The urinary concentration and excretion rate of caesium resulting from the infusion are given in Fig. 3. At the end of the 50 min recovery period following the caesium infusion the mean distribution volume of caesium was 47*74 + 5-16 1. which was 84-69 + 7-67 % of the body weight of the sheep. The mean caesium clearance by the kidney at this time was 17-69 + 2-61 ml./min or 41-66 + 5-63 % of the inulin clearance.

ALKALI METALS ON RENAL FUNCTION

151

TABLE 5. Summary of the final t test analyses of the effects of infusion into the renal artery of 1 m-CsCl compared with the effects of similar infuisiois of 1 M-NaCl, 1 M-KC1, 1 M-LiCl and 1 M-RbCl. Every parameter shown in Figs. 1, 2 and 5 was tested and omission from this Table means that no significant differences were found. Differences are indicated as levels of significance (* = P < 0-05; ** = P < 0-01; *** = P < 0-001; - = not significant). When the response to CsCl was less than that of another treatment t replaces * Recovery

Infusion Parameter Degrees of freedom Urine flow

CsCl =

42

Vs.

NaCI 3 m-NaCl KC1 LiCl RbCl NaCl Osmolal clearance 3 N-NaCl KCI LiCl RkCI Solute-free water reabsorption KC1

-

-

Urine Na concentration Urine K excretion

Urine K concentration

Inulin clearance (CIn)

Osmolal clearance/CQ.

Solute-free water reabsorption/Cfn Urine Na excretion/Cf.

Urine

K

excretionfC1.

RbCl NaCl 3 Ai-NaCl KCI LiCI RbCl NaCl KC1 RLCI KCI LiCl RbCl NaCl 3 m -NaCl KC1 LiCl RbCl NaCl 3 At-NaCl KC1 LiCI RbCl NaCl 3 M NaCi KCI LiCl RbCl LiCl RbCl NaCl 3 Mi-NaCl KC1 LiCl RbCl NaCl 3 N-NaCl KC1 LiCl RbCl

4

a

*** ***

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** *

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** *** **

-

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***

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-

** *** **

** *** ***

*** *** ***

*** *** ***

_**

-

t

- ttt tit - ti-i t-t -

tt

-

t

LiCl

Urine Na excretion

3

*** ***

1

tit ttt ttt ttt ttt ttt

-

2

* *

2

1

0

-

-

*

-_

ttt tt

-

ttt ttt tt t ttt ttt

t tt ttt ttt - ttt tt - ttt tt -- *** *** - -* ***

**

_

-

-

t -

t t **

t - - - - --

-

ttt ttt ttt ttt tt

t ttt t t t tt tt t tt tt

tt tt ttt ttt tt tt t it -

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ttt tt tt tt t

it

t t t t tt ttt t tt

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A. M. BEAL AND F. A. HARRISON

152

Li

20 Li concentration (m-mole/l.)

20 Llt ' p

1 ]

Li

140r-1 Liexcretionn'

,,(#momle/min)

G

E

o

40 r

Rb concentration

(m-mole/l.)

20

Rb excretion

(#mole/min)

II 1

400

0L

Cs

:

Cs concentration

10 _

(m-mole/I.)

20 _

Cs excretion (4umole/min) 0

LI

,

5I

o _

0

60

Time (min)

Fig. 3. Urinary concentration and excretion rates of lithium, rubidium and caesium during and after the infusion of each of the ions into the renal artery (means ± s.E. of mean).

Repeated close-arterial infusion of 1 M-CsCl (four replications) In the four experiments which had two caesium infusions, the estimated concentration of caesium resulting from the first infusion into the renal artery was 1P88 + 0-28 m-molefl. initially and 2-67 + 0-32 m-mole/i. by the mid-point of the second 5 min of caesium infusion. Repeating the caesium infusion 50 min later raised the caesium concentration of renal plasma from the residual level of 0416 + 0 01 m-mole/l. to 2-04 + 0-23 m-mole/l. at the beginning of infusion and to 3-01 + 0-27 m-mole/l. by the mid-point of the second 5 min of infusion. During the subsequent 120 min recovery period,

ALKALI METALS ON RENAL FUNCTION 153 the plasma caesium concentration fell to 0-27 + 0-02 m-molefl. after approximately 45 min and 0-18 + 0'02 m-molefl. at 115 min. The mean values for the haematocrit, plasma osmolality and the concentrations of sodium and potassium in the plasma of carotid arterial blood collected before, during and after the repeated infusions of caesium are given in Table 6. The first .Cs concentration

.15 _ _

(flmolefmin)

Cs

L

(m-mole/l.)

Cs excretion

Cs .:

15 0

a

70 _

(mi./min)

2 L

20', 500 _

. .

.

CPAHTT

(mPi./min)to Urine flow

200

'

0*075

_

(ml./mi.) Com and Tc,0

(Ml./Ml.)H0

F L

Naexcretion

(Fmole/ml.)TTT

K excretion

(,umolelml.)

I~~~~~ 0

T

v*

.

o

I

I

Ij

I

60

I

11 I I 120

1111 I 180

Time (min) Fig. 4. Mean values for urinary caesium concentration and excretion, inulin clearance (CI), PAH clearance (CPAH) and for the ratios of urine flow/Cj., osmolal clearance/C1,, solute free water reabsorption/C_%, urinary sodium excretion/C,, and urinary potassium excretion/C,, during and after repeated infusions of 1 m-CsCl into the renal artery (means ± s.E. of mean).

154 A. M. BEAL AND F. A. HARRISON infusion of caesium chloride increased urine flow,Cosm and urinary sodium excretion as previously described and the repeat infusion caused further increases in these parameters. After the first caesium infusion the inulin clearance was depressed to an extent that was statistically significant while the PAH clearance was relatively unaffected. The repeat infusion of caesium further lowered the inulin clearance and the PAH clearance was depressed during the second 5 min of the infusion and the first 10 min of the recovery period. Thereafter, the PAH clearance returned immediately to preinfusion levels while the inulin clearance increased slowly and approached the rates observed before the caesium infusions by the end of the 2 hr recovery period. The latency in response of the kidney to caesium infusion was shortened in only one of the 4 experiments by the previous priming of the animal with the ion. By the end of the 2 hr recovery period following the second caesium infusion the renal excretion of water and electrolytes had not returned to the levels observed before any caesium was infused. As a depression of the G.F.R. by the caesium treatment would affect urine flow and electrolyte excretion these parameters have been expressed as a ration of the inulin clearance in Fig. 4. The mean caesium clearances calculated for the three plasma samples collected during the recovery periods were 14*76 + 1-83, 13*10 + 2*20 and 12 20 + 2-35 ml./min and as a percentage of the inulin clearance these were 34'95 + 434, 3034 + 3'55 and 26*32 + 2*43 % respectively. TABLE 6. The haematocrit, osmolal concentration (m-osmole/l.) and the concentrations of ions (m-mole/l.) in the plasma of carotid arterial blood before, during and after the repeated infusion 1 m-CsCl into the renal artery (mean+ S.E. of mean)

Infusion CsCl (1) CsCl(2) CsCl (1) CsCl (2) CsCl (1) CsCl (2) CsCl (1) CsCl (2) CsCl (1) CsCl (2)

Parameter

Haematocrit Osmolality Na K

5 min before infusion

7-5 min into infusion

30*2 + 1*3 30.6+1-2

300 + 1.5

30-6 ± 1-2

305+2-2

303+1-7

296±4 294 + 4 141-7±0-8 140-7 + 1.0

295±5 297 + 5 141-4+ 1-1 141-8 + 1-5

294+4 296 + 4 140*7+ 1*0 142.7 + 1.3

4*17+0-09 4-15+ 0-05

Cs

0

0*16± 0-01

4*17+±006 4-14+0-05 0-80+ 0-05 0-98 ± 0-06

45 min after infusion

115 min after infusion

28-4+1±7 295 + 5 1430 + 1-2

4*15±0-05 4-11+ 0-05

4.08+ 002

0-16+0-01 0-27 + 0*02

0-18 +0-02

DISCUSSION

The sheep were bilaterally ovariectomized to prevent variations in the levels of ovarian steroids from influencing water and mineral balance between experiments. The right adrenalectomy and the transplantation to

155 ALKALI METALS ON RENAL FUNCTION the neck of the left adrenal gland removed all adrenal tissue from the site of close-arterial infusion in the abdomen and reduced the possibility that elevated concentrations of any of the alkali metals might directly alter the

rate of mineralocorticoid secretion during each experiment. The period of hypertonic infusion was limited to 10 min so that any increase in vasopressin release would not affect renal water and electrolyte excretion during the test infusion. Despite these precautions and the use of identical husbandry techniques between experiments considerable variations in the urinary excretion of electrolytes still occurred between experiments even in the same animal. Renal function, particularly sodium excretion, tended to alter during the experiment possibly because the residual effects of some of the alkali metal cations lasted more than the 50 min recovery period between the test infusions. -To ensure that these factors did not bias the interpretation of the results, the data for corresponding time intervals of each cation infusion were subjected to analysis of covariance. The presentation of the results of these experiments in terms of statistically determined differences between each of the alkali metals examined should not obscure the clear similarities which have been found in the responses to all ions except sodium. Of the 2352 t test analyses performed, 11 % showed significant differences between the effects of infusion of lithium, potassium, rubidium or caesium compared to the effects of sodium infusion and 8 % differences in the levels of response to the infusion of any pair of the ions lithium, potassium, rubidium and caesium. Because the urinary tract has a relatively large 'dead-space' volume, the concentration of solutes in urine collected from a bladder catheter does not change as soon as the urine flow alters, so that during short periods of collection, rapid increases in flow tend to augment the rates of excretion of urinary solutes whereas decreases in flow have the opposite effect. This phenomenon was clearly demonstrated during the infusions of lithium, potassium and rubidium by the excretion rates and clearances of inulin and PAH which were markedly augmented during the first 5 min of infusion by the rapidly rising urine flow, were either equal to or only slightly elevated above pre-infusion levels during the second 5 min of infusion when the rate of increase in urine flow had lessened and were depressed during the first 10 min of the recovery period when the urine flow was falling rapidly. Presumably the rates of excretion of ions such as sodium and potassium were similarly affected. By dividing the rates of excretion of these ions by the inulin clearance, the plasma concentrations of inulin being reasonably stable, the effect of the alkali metals on renal excretion could be observed unbiased by the effects of dead space and flow change. The modified values are given in Fig. 5 and the statistical analyses of these results are appended to the bottom of each of the appropriate

156 A. M. BEAL AND F. A. HARRISON Tables 2-5. From these modified results and their statistical analysis, it is clear that there was a marked and significant increase in osmolal clearance and sodium excretion rate during the first 5 min of the infusions of lithium, potassium and rubidium. During the entire 10 min of close-arterial infusion of potassium into the Na

0 1 5_

Li

K

,.

a

I

CosmandT2O_ I 'L 0*1

6r Na excretion

(4umole/ml.) 6F 2L

K excretion

(,umole/ml.)

2L_*Cs

Cosm andTH%

1

,___,___,______I .

-

(mI./mI.) =

010 L

Na excretion

0

(Fumole/ml.)

*K

.iiit. t i

1

5r 2 2

]

0 60 06 Tim(m.

(pumole/ml.)

K excretion

3M-Na

:0 0

60

Fig. 5. Mean values for the ratios of osmolal clearance/CI, solute free water

reabsorption/CO, urinary sodium excretion/lk and urinary potassium excretion/C1. during and after the infusion of 1 M-NaCl, 1 M-KCl, 1 m-LiCl,

1 M-RbCl, 1 M-CsCl and 3 M-NaCl into the renal artery. The filled circles indicate the values of the covariates used in the statistical analyses of each of the renal parameters shown on the Figure.

157 ALKALI METALS ON RENAL FUNCTION renal artery the urine flow and osmolal clearance were markedly increased with relatively little change in the rate of solute-free water reabsorption. Sodium excretion was always increased as the result of simultaneous increases in urine flow and urinary sodium concentration. Urinary potassium excretion was also increased but, in contrast to sodium, the potassium concentration of the urine fell. Although urinary bicarbonate concentration was not estimated directly it can be shown that the excretion of this ion must have increased regardless of whether the urinary pH rose or fell. In those experiments where urinary pH tended to be less than that of blood before the potassium infusion, the rapid increase in urine pH and urine flow during the infusion must have been associated with increased urinary bicarbonate concentration and excretion. In those experiments where the urinary pH was high initially, the small fall in urinary pH (maximum fall during KCl infusion being 0418 units) would lead to a fall in urinary bicarbonate concentration. However, it can be seen from the data of Nave, Beal, Budtz-Olsen, Clark, Cross, French, Ward & Wilson (1969) and of Scott (1969) that the pH of sheep's urine would have to fall by approximately 0*3 of a pH unit to produce 50 % reduction in urinary bicarbonate concentration. Any fall in bicarbonate concentration was therefore more than compensated by the increase in urine flow rate in these experiments. The changes in urine electrolyte concentration and excretion during closearterial infusion of potassium were consistent with those reported for more general potassium loading of sheep and other animals. Of the four other species of alkali metal cation investigated in these experiments, lithium, rubidium and caesium were found to produce changes in urinary electrolyte composition and excretion similar to those of potassium. The infusion into the renal artery of these three ions invariably resulted in increased urine flow, increased urinary sodium concentration and excretion and increased urinary potassium excretion associated with decreased urinary potassium concentration. The urinary pH was elevated by these infusions except when the pH was already very high and the magnitude of the changes in urinary pH and urine flow meant that the bicarbonate concentration of the urine would usually rise and that the rate of bicarbonate excretion would always be increased by the infusions. Therefore, the four ions, lithium, potassium, rubidium and caesium differed in their effects on urinary electrolyte composition and excretion only in the rate of onset of action and in the degree and duration of effect. The infusion of sodium at the same concentration as the other alkali metals had little effect on urinary composition and excretion while sodium infusion at 3 times this concentration produced relatively small effects on urine flow and sodium excretion compared to those of the other alkali metals. The sodium chloride infusions eliminate the possibility that changes in osmolality or

158 A. M. BEAL AND F. A. HARRISON chloride concentration of the plasma perfusing the kidney contribute significantly to the renal response to alkali metal infusion.The obvious similarity of the effects of the infusions of lithium, potassium, rubidium and caesium on renal excretion provides evidence that these four ions act by similar

mechanisms. In the dog, potassium loading suppresses the Peo2 dependent reabsorption of bicarbonate (Buttram, Rector & Seldin, 1961) which occurs in the proximal tubule (Rector, Seldin, Roberts & Smith, 1960). Malnic, Klose & Giebisch (1966) found that potassium loading increased the sodium/inulin clearance ratio of the proximal segment of the rat kidney, while Brandis, Keyes & Winghager (1970) have shown that increased potassium concentration in the peritubular fluid depresses the reabsorption of water and sodium by the proximal tubule of the rat. If the increase in osmolal clearance during the infusion of potassium is predominantly due to a reduction in reabsorption in the proximal tubule the ratio of the increase in solute-free water reabsorption to the increase in osmolal clearance (ATe 0/IlCOsm) would tend to approach zero whereas increased osmolal clearance resulting from the addition of solute in the distal segments of the nephron would cause the ratio to approach unity. The mean value for ATci0I/.COsm was calculated for each cation using the results for the period just before alkali metal infusion and those for the period of peak increase in renal electrolyte excretion resulting from each infusion. All data used to estimate the AT'0/IACO.m were expressed as ratios of the inulin clearance to make allowance for any errors in the measurement of solute excretion caused by the 'dead-space' effect and for real changes in G.F.R. The values of ATci0/AGCOsm obtained for the infusion of lithium, potassium, rubidium and caesium were 0-20, 0-17, 0-28 and 0-22 respectively while that for 3 m-NaCl was 0 57. The low value for the ratio during potassium infusion was interpreted as indicating that the proximal tubule was the primary site of action of potassium for increasing electrolyte excretion by the sheep's kidney and that any subsequent effects of the ion on more distal sites in the nephron were of much less importance. Although opinions differ as to the value and interpretation of changes in Te 0 and C0osm the above interpretation is consistent with the evidence of other workers that the proximal tubule is the site at which potassium reduces the reabsorption of sodium and bicarbonate in the nephron. If a lowvalue of ATH,0I/\COsm is evidence that potassium depresses reabsorption by the proximal tubule the similarly low values of this ratio for all cations except sodium, coinciding invariably with marked increases in sodium and bicarbonate excretion, support the view that lithium, potassium, rubidium and caesium alter electrolyte excretion through the same mechanisms which are primarily located in the proximal segment of the nephron. An increase in G.F.R. would also augment

ALKALI METALS ON RENAL FUNCTION 159 electrolyte excretion but, as the inulin clearances during the second 5 min of the infusions of lithium, potassium or rubidium were either equal to or only slightly elevated above the pre-infusion levels, any real increase in filtration rate must have been small and have had a correspondingly small effect on electrolyte excretion. Caesium infusion resulted in a depression of the inulin clearance coincident with the increase in electrolyte excretion. The infusion of potassium into the renal artery of sheep with divided bladders resulted in ipsilateral increases in sodium excretion as well as potassium excretion (Harrison et al. 1970) indicating that the effect of potassium loading under these conditions was intrarenal rather than extrarenal. The depression of proximal water and sodium reabsorption during the microperfusion of capillaries adjacent to the proximal tubules of rats with high-potassium Ringer solution (Brandis et al. 1970), supports the concept that the natriuretic effect of potassium loading is basically intrarenal. Three of the cations infused, lithium, potassium and rubidium, produced marked increases in urine flow during the first 2 min of infusion showing that the action of these ions was very rapid in onset and that, initially at least, it must be intrarenal rather than as a result of some extrarenal mechanism. It was intended to investigate changes in PAH extraction ratio during these infusions. However, the results obtained in one animal indicated that the renal venous catheter was sampling both renal venous and vena caval blood and this was subsequently confirmed by laparotomy. In the other animals, withdrawal of blood from the renal venous catheter became increasingly position dependent and unreliable during the life of the preparations so that the data for PAH extraction ratio were incomplete. Following the peak response during the infusion of lithium the urinary excretion ofsodium returned to and stabilized at levels which were obviously higher than those prior to the infusion. This effect was probably due to the slow removal of lithium from the plasma and hence a significant level of recirculation of the ion to the kidney. The slow movement of lithium into the intracellular space which has been reported in several other species (Radomski, Fuyat, Nelson & Smith, 1950; Foulks et al. 1952; Trautner et al. 1955) results in the distribution volume of the ion, 50 min after the infusion, being approximately the theoretical extracellular volume of the sheep. The distribution volumes of rubidium and caesium, calculated on a one compartment basis, were much higher than those of lithium since rubidium and caesium, like potassium, distribute themselves unequally in the intracellular and extracellular compartments. The slow response of the kidney to the infusion of caesium suggests that this ion has difficulty either in reaching the site of action in the nephron or in producing the appropriate changes in cellular function once there. 6

P H Y 245

A. M. BEAL AND F. A. HARRISON 160 The slow onset of action may mean that the maximum renal response to caesium observed in these experiments was that appropriate to the level of caesium in the plasma during the recovery phase rather than that perfusing the kidney during the infusion. The possibility that there was a threshold level of caesium necessary for the effect and that once primed the response would be more rapid was not supported by the data obtained during the repeated infusions. The response of the kidney to the two sequential infusions of caesium lasted for more than 2 hr and judging by the similar flat excretion curves the rate of recovery after one 10 min infusion would have been of similar duration. Caesium infusion resulted in a prolonged reduction in the G.F.R. which was not observed during or after the infusion of the other alkali metal ions. The large increases in inulin clearance resulting from the infusion of lithium, potassium and rubidium were believed to be erroneous since the combination of the 'dead space' volume of the urinary tract with rapid increases in urine flow would tend to augment these clearance values. The observation of low values for the inulin clearance following caesium infusion must be considered to be highly significant since they coincide with a period of rising urine flow and therefore the true G.F.R. should be even lower than that given by the inulin clearance. The most characteristic effect of the infusion of the alkali metals was the marked natriuretic diuresis. The order of effectiveness of the cations ranked according to their ability to cause the greatest natriuresis was Rb > K > Li > Cs. The rate at which each treatment reached its maximum response fell into the order Li > Ke Rb > Cs. However, arranging the elements into hierarchies of relative effect is not necessarily reasonable since the relative changes in plasma concentration for the sodium and potassium were much less than for the other three ions. The results of these experiments indicate that, although extrarenal factors may contribute in the long term, the primary mechanisms for the increases in renal sodium and bicarbonate excretion during loading with salts of the alkali metals, lithium, potassium, rubidium and caesium are

situated within the kidney. We are indebted to Miss G. Needham, Mr R. C. Saunders and Mr P. Burrow for their technical assistance and to Mr J. Christie, S.R.N. for assistance at operations.

ALKALI METALS ON RENAL FUNCTION1161 REFERENCES BEAL, A. M., BUDTZ-OLSEN, 0. E., CLARK, R. C., CROSS, R. B. & FRENCH, T. J. (1973). Renal and salivary responses to infusion of potassium chloride, bicarbonate and phosphate in Merino sheep. Q. JI exp. Physiol. 58, 251-265. BEAL, A. M. & HARRISON, F. A. (1973). The effect of renal arterial infusions of alkali metal cations on electrolyte excretionby the sheep kidney. J. Physiol. 234,105-106 P. BRANDIS, M., KEYES, J. & WINDHAGER, E. E. (1970). Potassium-induced inhibition of proximal tubular fluid reabsorption. Physiologist 13, 154. BRATTON, A. C. & MARSHALL, E. K. (1939). A new coupling component for sulphanilamide determination. J. biol. Chem. 128, 537-550. BUTTRAM, H. M., RECTOR, JR. F. C. & SELDIN, D. W. (1961). Effect of potassium on renal bicarbonate reabsorption. Clin. Res. 9, 56. DAWBORN, J. K. (1965). Application of Heyrovsky's inulin method to automatic analysis. Clin. Chim. Acta 12, 63-66. DEWHURST, J. K., HARRISON, F. A. & KIEYNEs, R. D. (1968). Renal excretion of potassium in the sheep. J. Physiol. 195, 609-621. FouiXs, J., MUDGE, G. H. & GmImAN, A. (1952). Renal excretion of cation in the dog during infusion of isotonic solutions of lithium chloride. Am. J. Physiol. 168, 642-649. HALL, P. W. & RELMAN, A. S. (1959). Acid excretion in rubidium- and caesiumsubstituted rats. Clin. Re8. 7, 30-31. HARRISON, F. A., KEYNES, R. D. & PEARSON, JENNIFER L. (1969). Occlusion of the autotransplanted adrenal gland in the sheep and its effect on renal excretion of potassium. J. Physiol. 200, 31-32P. HARRISON, F. A., MCDONALD, I. R. & OLSSON, K. (1970). Unilateral renal excretory responses to close arterial infusions of potassium chloride in conscious sheep. J. Physiol. 210, 125-127P. HARVEY, R. D. & BROTHERS, A. J. (1962). Renal extraction of para-aminohippurate and creatinine measured by continuous in vivo sampling of arterial and renal vein blood. Ann. N.Y. Acad. Sci. 102, 46-54. HERD, J. A. & BARGER, A. C. (1964). Simplified technique for chronic catheterization of blood vessels. J. appl. Physiol. 19, 791-792. HEYROVSKY, A. (1956). A new method for the determination of inulin in plasma and urine. Clin chim. Acta 1, 470-474. MCDONALD, I. R., GODING, J. R. & WRIGHT, R. D. (1958). Transplantation of the adrenal gland of the sheep to provide access to its blood supply. Aust. J. exp. Biol. med. Sci. 36, 83-96. MALNIc, G., KLOSE, R. M. & GIEBISCH, G. (1966). Micropuncture study of distal tubular potassium and sodium transport in the rat kidney. Am. J. Physiol. 211, 529-547. NAVE, J. M., BEAL, A. M., BUDTZ-OLSEN, 0. E., CLARK, R. C., CROSS, R. B.,FRENCH, T. J., WARD, G. M. & WILSON, K. (1969). Urinary bicarbonate excretion in the sheep. Aust. J. exp. Biol. med. Sci. 47, 20-21. ORLOFF, J. & ]KENNEDY, T. J. (1952). Effect of lithium on acidification of urine. Fedn Proc. 11, 115.'4 RADOMSKI, J. L., FUYAT, H. N., NELSON, A. A. & SMITH, P. K. (1950). The toxic effects, excretion and distribution of lithium chloride. J. Pharmac. exp. Ther. 100, 429-444. RECTOR, F. C., SELDIN, D. W., ROBERTS, A. D. & SMITH, J. S. (1960). The role of plasma CO2 tension and carbonic anhydrase activity in the renal reabsorption of bicarbonate. J. clin. Invest. 39, 1706-1721.

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RELMAN, A. S., Roy, M. A. & SCHWARTZ, W. B. (1953). Effect of rubidium on acidbase balance in potassium-deficient and normal rats. J. clin. Invest. 32, 597. SCOTT, D. (1969). Renal excretion of potassium and acid by sheep. Q. Ji exp. Physiol. 54, 412-422. TRAUJTNER, E. M., MoRRIs, R., NOACK, C. H. & GERSHON, S. (1955). The excretion and retention of ingested lithium and its effect on the ionic balance of man. Med. J. Aust. 42, 280-291. VOGEL, G. (1959). Vergleichende Untersuchungen zur Kalium-Exkretion der Nieren verschieden Haussaugetiere. Pflugers Arch. ges. Physiol. 269, 339-343.

Renal function in sheep during infusion of alkali metal ions into the renal artery.

J. Phyaiol. (1975), 245, pp. 137-162 With 5 text-ftgure8 Printed in Creat Britain 137 RENAL FUNCTION IN SHEEP DURING INFUSION OF ALKALI METAL IONS I...
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