capillaries, and a consequent increased transudation, without necessarily altering to any marked extent the total circulation of blood

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1 THE CONTROL OF THE GLOMERULAR PRESSURE BY VASCULAR CHANGES WITHIN THE ISOLATED MAMMALIAN KIDNEY, DEMONSTRATED BY THE ACTIONS OF ADRENALINE. BY F. R. WINT0N (Beit Memorial Research Fellow). (Depaortment of Pharmacology, University College, London.) INTRODUCTION. THERE are two ways in which the glomerular pressure of the kidney may be affected, (1) by variation of the blood-pressure external to the kidney in the renal artery or vein, and (2) by an internal redistribution of the resistance to the blood flow through the kidney, as between the vasa afferentia and efferentia. The second of these possibilities was first recognized by Ludwig [1856] and clearly enunciated by Starling [1912] in the following terms: "A dilatation of the afferent vessels and a slight constriction of the efferent vessels would cause a considerable rise of pressure in the glomerular capillaries, and a consequent increased transudation, without necessarily altering to any marked extent the total circulation of blood through the whole organ. The changes in the afferent and efferent vessels are, however, beyond our control or powers of observation, so that it is impossible to devise at the present time any crucial experiment which might decide the nature of the process occurring in the glomeruli." That small doses of adrenaline act on this mechanism was suggested by the observations of Richards and Plant [1922] which showed that when a rabbit's kidney was perfused with blood by a pump which maintained the rate of flow constant and independent of vaso-constrictor or dilator changes in the organ, adrenaline induced a rise of pressure in the renal artery (vaso-constriction), diuresis, and increase in kidney volume. They attributed the diuresis and kidney swelling to an increase in the glomerular pressure due to constriction of the vas efferens. It has been objected to their interpretation, wrongly as I hope to show, that the diuresis may have resulted entirely from the associated increase of arterial pressure, and that the significance of kidney swelling is

2 152 F. R. WINTON. ambiguous. Although these authors put forward the hypothesis that adrenaline acts differentially on the vas afferens and vas efferens somewhat tentatively, their experiments provide the best evidence of the existence of an internal control of the glomerular pressure which has been available. An increase of glomerular pressure due to adrenaline should be accompanied by diuresis even if the pressure in the renal artery remains unchanged. The following experiments were designed to discover whether such a diuretic response occurred, and if so, to analyse the vascular changes in the kidney to which it might be attributed, with a view to measuring the magnitude of the internal control of the glomerular pressure in the kidney. THE DIURETIC AND VASO-CONSTRICTOR RESPONSES TO ADRENALINE. In experiments performed on the isolated kidney of the dog perfused with defibrinated blood from a heart-lung circulation [Starling and Verney, 1925], there is no difficulty in controlling the arterial pressure, so that it is unaffected by the action of adrenaline on the heart and kidney; this preparation was therefore employed. At constant arterial pressure, concentrations of adrenaline of about one in twenty million elicited a well-marked diuretic response in seventeen kidneys out of the twenty-five to which this stimulus was applied. Of the rest, six kidneys increased their rates of secretion by amounts which were within the range of error, and two kidneys responded with a reduced rate of secretion. The blood flow was measured in most of the kidneys and in all of these it was diminished by adrenaline. This greater regularity in the vascular responses than in the urinary responses of isolated kidneys to drugs has previously been described, in connection with the effects of caffeine, by Verney and Winton [1930]. Larger concentrations of adrenaline invariably retard or abolish the secretion of urine, and further reduce the blood flow. These effects are antagonized by ergotoxine, in the presence of which even large concentrations of adrenaline often produce an increase of blood flow and only a small change, usually anti-diuretic, in the urine flow. Fig. 1 illustrates the diuretic and vaso-constrictor effects of small doses of adrenaline on the isolated kidney. The changes are relatively short-lived owing to the rapidity of the destruction of the drug in the blood. That the diuretic action is not an artefact due to the expression of urine preformed in the kidney, can be seen by the facts (1) that the excess urine excreted is too great to be accounted for in this way, and

3 VARIATION OF GLOMERULAR PRESSURE. (2) that if successive doses of adrenaline be added to the circulation at suitable intervals so as to maintain an appropriate concentration for a prolonged period, the diuretic and vascular effects persist. A suspicion that an increase of arterial pressure, so small as to escape notice, might contribute materially to the diuresis was shown to be Blood fiow (cc. pe nn.) 1 0C t_. 10 Q=2 QJrine flo WT(cc. per(minutes O 01 Time (l iinutes)l I u vu ou 40 Du 60 Fig. 1. The influence of adrenaline on the urine flow and blood flow of the isolated kidney of the dog. The vertical line A represents the time at which adrenaline was added to the circulation, the concentration being about 1 in 20 million. The effects are transient, presumably owing to the rapid decomposition of the drug in the blood stream. unfounded in an experiment in which a deliberate lowering of the arterial pressure from 120 mm. to 105 mm. Hg was insufficient to reduce the urine flow to the value obtaining before adding the adrenaline. This is confirmed by the glomerular pressure measurements described below. An analogous test of the authenticity of the diuresis is shown in Fig. 2,

4 154 F. R. WINTON. '3 X f140 0 Urine flow 2~~~~~~~~~~~~~~~~~ Ureter pressure 10 mm. S mm. Time (minutes) Fig. 2. The influence of adrenaline on the urine flow and heart frequency in a heart-lungkidney preparation. The first dose of adrenaline (A1), equivalent in concentration to about 1 in 20 million, produced only moderate diuresis but almost maximal acceleration of the heart-rate. The second dose (A2), equivalent to an additional concentration of 1 in 40 million, augmented the diuretic response without producing an appreciable further increase in the frequency of the heart beat. Subsequent increase of the ureter pressure to 20 mm. Hg did not suffice to reduce the urine flow to the value obtaining before the administration of adrenaline, until time had elapsed during which most of the drug had been decomposed.

5 VARIATION OF GLOMERULAR PRESSURE. when an increase of ureter pressure of 20 mm. Hg applied during the influence of adrenaline was insufficient to reduce the urine flow to its initial value. The concentration of adrenaline in the blood at any given moment is difficult to estimate in these experiments, partly owing to the rapidity of its destruction, and partly on account of the uncertainty as to the exact volume of liquid in circulation. It was thought desirable, therefore, to employ the frequency of the heart beat as a physiological indicator of the activity of the drug in the blood stream. Fig. 2 represents an experiment of this kind, in which a first dose of adrenaline produced a moderate diuresis concurrently with a large increase of the heart frequency, while a second dose applied so soon after as to augment the concentration in the blood, produced further diuresis associated with little further effect on the heart rate. The diuretic concentrations of adrenaline, though smaller than the anti-diuretic ones, are therefore about the same or rather larger than those which just produce a maximum increase of the heart frequency; they are not as small as the "minute concentrations of adrenaline" which have been described in connection with the vaso-dilator action of the drug. If a dose of adrenaline, yielding a concentration of about 1:2 x 107, be added to the circulating blood, the diuretic and vaso-constrictor effects appear almost immediately, and gradually lessen until they vanish in about 20 minutes. A similar dose of the drug applied subsequently again elicits these effects. THE DIFFERENCE IN THE RESPONSE TO ADRENALINE BETWEEN KIDNEYS WITH VENOUS AND URETERAL OBSTRUCTION. The physical analysis of the processes in the kidney accompanying the diuretic response to adrenaline depends on the observation that the diuretic effect is less in a kidney when its vein is partly obstructed than in one in which the ureter is obstructed to a degree which has the same initial effect in reducing the urine flow. This effect can be clearly demonstrated on the heart-lung-doublekidney preparation described by Verney [1929] and is shown in Table I, which represents one of a series of experiments of this kind. The experimental technique is exactly the same as that detailed in previous papers [Winton, 1931]. In outline, the procedure is as follows. (1) The two kidneys of one dog are connected in parallel to a heart-lung circulation made from another dog so that both are perfused with defibrinated blood of the 155

6 156 F. R. WINTON. same composition and at the same pressure. (2) Arrangements are made to be able to raise the pressure in the vein of one kidney and in the ureter of the other. (3) Before these pressures are raised, both the urine flows TABLE I. Heart-lung-double-kidney experiment, in which the vein of one and the ureter of the other kidney were submitted to partial obstruction. The effect of adrenaline in producing the greater diuretic action in the kidney with the obstructed ureter is illustrated. Urine flow c.c./10 min. Venous Ureter -_ Adrenaline Stage of pressure pressure Vein Ureter added experiment* mm. Hg mm. Hg obstructed obstructed * See text. and blood flows of the two kidneys must be approximately equal, otherwise the preparation is unsuitable. (4) The venous pressure in one, and the ureter pressure in the other kidney are now raised to such values that they produce well-marked and equal reductions of the urine flow in the two organs. (5) The dose of adrenaline is added to the blood in the venous reservoir of the heart-lung circulation, and means taken to hasten its mixing with the blood. (6) The diuretic responses of the two kidneys are measured, and as shown in Table I are different. (7) To enable the change in glomerular pressure to be measured, the venous and ureter pressures are then readjusted until the two urine flows become equal in the presence of adrenaline. (8) The two pressures are reduced to atmospheric pressure, and time is allowed to pass until the adrenaline has been destroyed (indicated by slowing of the heart rate). If the two urine flows are still approximately equal, it is assumed that the kidneys would have responded alike to the same stimuli throughout the experimental period [Canny, Verney, and Winton, 1930], and the experiment is regarded as "successful." Stages 3 to 8 are illustrated in Table I. Now it has been shown [Winton, 1931 a and b] that the effects of venous and ureter obstruction on the tubular activity are negligible; any influence which adrenaline might exert on the tubule cells would therefore be common to both kidneys. Consequently the difference in their diuretic responses to adrenaline must be attributed to a difference in the change of glomerular pressure-the pressure in the kidney with ureteral obstruction being either increased more or reduced less than that with venous obstruction, by the action of the drug. It can be shown

7 VARIATION OF GLOMERULAR PRESSURE. that the diuretic action of adrenaline is in fact due to an increase of glomerular pressure, and not entirely to a change in tubular activity in the following way. Let p be the arterial pressure, which is the same for both kidneys throughout the experiment, and let the glomerular pressure be defined in terms of the difference of pressure between the renal artery and the glomerulus. Suppose ap be this fall of pressure before, and fip the corresponding fall of pressure after the action of adrenaline. The change in glomerular pressure will be p (a -,), 3, positive value of which will indicate an increase of glomerular pressure. These relations will be substantially unaffected by a change in ureter pressure owing to the small proportion of the liquid flowing through the kidney which emerges from the ureter. If, on the other hand, the venous pressure be raised to a value v, the pressure in the glomerulus will rise to a value a (p - v) below that in the renal artery-an assumption justified by considerations put forward in a previous paper [Winton, 1931 c]. In another paper [Winton, 1931 b] it was shown that the venous pressure is equal to the sum of the increase of glomerular pressure which it produces, and the ureter pressure (u.1) which, in the absence of venous obstruction, produces the same reduction of urine flow, i.e. v = u + av.... (1) If the diuretic action of the adrenaline necessitates a higher ureter pressure (u2) to restore equality in the urine flow of the two kidneysthe venous pressure in the other kidney remaining unchanged as in Table I-the corresponding relation between the pressures in the presence of the drug becomes v = U2 +v.... (2) From (1) and (2) it follows that 2 -ul = v (a -)....(3) 157 Experiment shows that u2 exceeds u1, and since vis a positive pressure, a must exceed /P. Hence the fall of pressure (ap) between the renal artery and the glomerulus is greater before the action of adrenaline than its value (ap) after its action. That is, adrenaline raises the glomerular pressure. GLOMERULAR PRESSURE CHANGES. The reasons for and reservations attaching to the measurement of the glomerular pressure, in terms of the product of the arterial pressure and the ratio of the ureter pressure to its equivalent venous pressure,

8 158 F. R. WINTON. were considered in a previous paper [1931 c]. In the present series of experiments the method was applied to exploring the changes in glomerular pressure consequent upon the action of different concentrations of adrenaline. A single dose of adrenaline induces a rise or fall in the glomerular pressure, according as the dose is small or large; this change follows a time course approximately parallel to the corresponding diuretic or antidiuretic responses. Examples of such variations are given in Table II TABLE II. A series of experiments on heart-lung-double-kidney preparations, arranged in order of increasing concentrations of adrenaline applied to the kidneys. The values of the glomerular pressure before and after the action of adrenaline show that this pressure may be increased from a normal of 60 p.c. of the arterial pressure to about 90 p.c., or it may be reduced to about 30 p.c. without changing the arterial pressure. Glomerular pressure Urine flow c.c./10 min. Arterial mm. Hg mm. Hg Exp. pressure Adrenaline,, no. mm. Hg (concentration) Before After Before After I 115 1:2 x II 115 1: III 116 1:1-5 x *9 IV :1-2x V 130 1: 1 Ox VI 116 1:0:8x VII :0-8x VIII : 07 x which indicates the wide range through which the internal control of glomerular pressure may operate. Fig. 3 represents an experiment in which the diuretic response was determined as the increase of ureter pressure needed to keep the urine flow constant. The measurement of the diuresis in terms of pressure rather than volume changes may be compared with the recording of a muscular contraction isometrically rather than isotonically. It simplifies the analysis of the observed relations; for, in the absence of a change in tubular activity, the rate of flow and composition of the liquid passing down the tubules remain unchanged [1931 a], and consequently the pressure fall between the glomerular capillaries and the ureter may be taken as constant. The changes in glomerular pressure should therefore be reflected accurately in the changes of ureter pressure, so long as the stimulus produces no direct change in the tubular activity. The glomerular pressure recorded in Fig. 3 has been calculated from the ratio of the equivalent ureter and venous pressures. It has been shown [1931 c] that in circumstances in which the glomerular pressure is constant, the ratio is independent of changes in the absolute value of the ureter pressure; the glomerular pressure and ureter pressure curves,

9 VARIATION OF GLOMERULAR PRESSURE. 159 therefore, represent two different ways of estimating the same changes, so long as the tubular activity may be taken as unchanged. If an agent were to produce a diuretic action mainly by an influence on the tubular 100 Em co 34 PU 90 I sos. '4 B0X 6 0 " Time (minutes) Fig. 3. Heart-lung-double-kidney experiment, in which the venous pressure of one, and the ureter pressure of the other kidney were raised to such values that the urine flow of each organ was maintained at 3-6 c.c. per 10 minutes throughout the experiment. The arterial pressure was also constant, at 116 mm. Hg. The glomerular pressure was calculated as the product of the arterial pressure and the ratio of the ureter to the venous pressure. The ureter pressure curve was obtained by adding 30 mm. to the actual values of the ureter pressure for convenience of graphic representation, and so that the hydrostatic pressure fall from glomerular capillaries to ureter could be taken as the difference between the glomerular pressure and ureter pressure curves (30 mm. Hg being the assumed value of the osmotic pressure of the plasma proteins). The vertical lines (A) represent the times at which four successive doses of adrenaline were added to the circulating blood, each dose being equivalent to a concentration of about 1 in 20 million. cells, such as retarding the re-absorption of water in the tubules, the glomerular pressure calculated in this way would still be valid, but the increase of ureter pressure needed to keep the urine flow constant would be disproportionately great and would vary independently of the glomerular pressure.

10 160 F. R. WINTON. The parallelism of the glomerular and ureter pressure curves in Fig. 3 may thus be taken as evidence that the diuretic response to adrenaline is due to an increase of glomerular pressure, and not to an action on the tubule cells. In so far as the curves deviate from the parallel, they do so in the opposite sense from that which would be induced by a diuretic acting on the tubules. This deviation may be due to a progressive increase of diffusion of urine out of the tubules with increasing duration of survival after isolation of the kidney. An analogous process in the frog's kidney was described by Richards [1929], and postulated for the human kidney by Rehberg [1926]. The glomerular pressure and ureter pressure curves thus afford mutual confirmation of the effects of adrenaline described on the glomerular pressure. As illustrated in Fig. 3 the pressure is raised by the first dose of adrenaline, but when successive doses of the drug were added to the blood at such short intervals that the concentration was progressively increased, the glomerular pressure fell and reached a value below its initial value. It was not possible in this experiment to increase the concentration further without reducing the urine flow below its standard value, 3-6 c.c. per 10 minutes. The ends of the curves represent the beginning of the recovery due to destruction of the drug in the blood stream. Subsequent removal of the venous and ureteral obstructions showed that the two kidneys had retained their parallel properties under the same conditions. The response of the glomerular pressure to different concentrations of adrenaline, as shown in Table II, indicates a remarkable regularity in the sensitivity of different kidneys to the drug. This is in striking contrast with the wide variation of sensitivity to pituitary extracts and other substances which is encountered when they act on kidneys isolated under similar conditions. THE MECHANISM OF THE INTERNAL CONTROL OF GLOMERULAR PRESSURE. Since the arterial pressure is kept constant throughout an experiment such as that represented by Fig. 3, the rise of glomerular pressure induced by adrenaline implies diminution of the fall of pressure along the vasa afferentia, which must be due to a reduced velocity of blood flowing along them or to vaso-dilatation. The pressure fall is in fact reduced by 60 p.c., whereas the blood flow is reduced only by 18 p.c., a change which, in the absence of a variation in shape of the vessels, could not bring about so large a reduction in the fall of pressure. The disproportion

11 VARIATION OF GLOMERULAR PRESSURE. 161 between the pressure and blood flow changes in the vasa afferentia is even more marked in other experiments of the series, for the reduction in blood flow more usually amounts only to p.c., while the glomerular pressure changes may be even larger than those in Fig. 3, as shown in Table II. Dilatation of the vasa afferentia in response to diuretic doses of adrenaline must therefore be inferred. Along the vasa efferentia and subsequent vessels, on the other hand, an increased fall of pressure is associated with a reduction in the blood flowing through them, which is an unequivocal indication of vaso-constriction. The larger (anti-diuretic) doses of adrenaline produce a relatively great diminution in the blood flow, and a smaller reduction in the glomerular pressure. Again, taking the values from Fig. 3, the blood flow is reduced to 68 p.c. of its original value and the glomerular pressure to 86 p.c. The pressure fall along the vasa afferentia is increased, and the blood flow through them reduced, indicating vaso-constriction. The pressure fall along the post-glomerular vessels is now reduced, but the diminution of blood flow through them is so disproportionately great that a vaso-constrictor change must again be inferred. Large concentrations of adrenaline, e.g. 1:106, induce intense vaso-constriction in the kidney, the blood flow in one instance being reduced from 140 c.c. per min. to 8-2 c.c. per min. SUMMARY. 1. The reactions of the isolated mammalian kidney, at constant arterial pressure, to adrenaline depend on the concentration. Low concentrations (about 1 in 2 x 107) induce diuresis, and slowing of the blood flow; higher concentrations reduce or abolish the urine flow, and diminish the blood flow further. 2. If in a pair of similar kidneys, perfused with blood from the same circulation, the venous pressure of one is raised, and the ureter pressure of the other is adjusted so that the urine flows are equally reduced in the two organs, the diuretic action of adrenaline is greater on the kidney with ureteral obstruction. Reasons are given for inferring that adrenaline increases the glomerular pressure, and has no detectable action on the tubular activity. 3. At constant arterial pressure, the glomerular pressure is increased by low concentrations, and reduced by high concentrations of adrenaline. The glomerular pressure can be varied in this way over a wide range; experiments are recorded showing increases up to about 90 p.c. and reductions to about 30 p.c. of the arterial pressure. (The normal value under the experimental conditions concerned is about 60 p.c.)

12 162 F. R. WINTON. 4. The quantitative relations between the concurrent changes in the glomerular pressure and the blood flow.through the kidney show that the increase of glomerular pressure is due to dilatation of the vasa afferentia and constriction of the vasa efferentia. Reduction of the glomerular pressure due to the large concentrations of adrenaline is accompanied by constriction of both groups of vessels. I am indebted to the Government Grants Committee of the Royal Society for defraying much of the expense involved in these experiments. REFERENCES. Canny, A. J., Verney, E. B. and Winton, F. R. (1930). J. Phy8iol. 68, 333. Ludwig, C. (1856). Lehrbuch der Phy8iol. 2, 257. Rehberg, P. B. (1926). Biochem. J. 20, 447. Richards, A. N. and Plant, 0. H. (1922). Amer. J. Physiol. 59, 144, 184. Richards, A. N. (1929). Method8 and result8 of direct inve8tigation8 of the function of: the kidney. Baltimore. Starling, E. H. (1912). Principles of human phy8iology. London, 1st ed., p Starling, E. H. and Verney, E. B. (1925). Proc. Roy. Soc. B, 97, 320. Verney, E. B. (1929). Lancet, 216, 645. Verney, E. B. and Winton, F. R. (1930). J. Physiol. 69, 153. Winton, F. R. (1931 a). J. Phy8iol. 71, 382. Winton, F. R. (1931 b). J. Phy8iol. 72, 49. Winton, F. R. (1931 c). J. Phy8iol. 72, 361.