Cushny(4) has shown, however, that the amount of urea in the kidney. by some vital process, retain those diflusible substances which are of

Transcription

1 THE FUNCTION OF THE TUBULES IN KIDNEY EXCRETION. BY E. B. MAYRS. (From the Department of Pharmacology, Edinburgh.) IT is becoming generally recognised that filtration through the glomeruli and some degree of re-absorption in the tubules are essential features of renal activity. The questions at issue are, in the first place, whether the kidney secretes at all; and secondly, whether the glomerular membrane is a simple ultra-filter in the ordinary physical sense, or can, by some vital process, retain those diflusible substances which are of value to the organism. Evidence of the constant presence of glucose in the glomerular fluid is now fairly strong; and since this is not a waste product but a substance of high physiological importance, the cells of Bowman's membrane have apparently no power of vital selection, but allow all the crystalloids of the plasma to pass through or between them without any change in concentration. It is intended, therefore, to deal in this paper with the question of secretion by the tubule cells. Interesting observations have recently been published by Marshall and Vickers(1) and also by de Haan(2), which seem to show that certain dyes are secreted by the cells of the convoluted tubules. It should be mentioned, however, that de Haan does not invoke secretion to interpret his results, but prefers the unlikely theory that proteins can pass through the glomerular membrane. This hypothesis does not explain certain results obtained by Marshall and Vickers. Neither is it in accord with the fact that gum arabic, if present in the blood in sufficient concentration, can produce almost complete anuria; for gum should pass through a membrane which allows the passage of protein. Accumulation of dye in the cells of the kidney was observed many years ago by Hei denhain(3), who regarded this as a stage in the process of secretion and concluded that urea is concentrated in a similar way. Cushny(4) has shown, however, that the amount of urea in the kidney is not sufficient to admit of the theory of storage, especially wien allowance is made for the urea content of the fluid in the tubules. He has examined the excretion of glucose also, with similar results. The recent evidence of secretion of dyes is more conclusive than that of Heidenhain, and justifies further search for analogies to normal excretion. A

2 KIDNEY EXCRETION. 423 series of experiments has therefore been carried out in order to decide whether, if this power of secretion exists, the kidney can apply it in dealing with ordinary constituents of the plasma. The experiments bear a general resemblance to those of Cushny, but special efforts have been made to -reduce as far as possible the extra-cellular portions of the substances estimated in the kidney. Tle observations were also more quantitative in character, since plasma concentrations were used for comparison with those of the kidney fluid. On the other hand, more operative interference with the kidney was required, and considerable diuresis was essential. During the rapid excretion of urea and sulphate the kidney was quickly freed from blood and its total content of these substances determined, so that any storage which might occur in the renal cells could be observed. The error due to partially formed urine in the tubules was unavoidable, but was reduced to a minimum by diuresis, which prevented much concentration of this fluid. Method. Rabbits were ancesthetised with urethane and eviscerated. A loose ligature was placed round the aorta above the left renal artery, and another was arranged to include the left ureter and both the left renal vessels close to the hilum of the kidney. A cannula was tied into the aorta below the renal arteries and was connected by.rubber tubing to a large syringe. A capillary tube was introduced into the left ureter. A solution containing sodium sulphate and urea was then injected into the jugular vein, and soon after the injection was complete urine was collected from the left kidney for a period of 1 to 4 minutes. Blood was taken from the carotid artery, and at the same time the ligature on the aorta was tightened. Air was at once forced through the kidney from the syringe, and when bubbles appeared in the renal vein the ligature at the hilum was tied. The kidney was now dissected out, and, after removal of the vessels, ureter, and adherent fat, was weighed, transferred to a mortar, and ground to a homogeneous semi-fluid pulp. This pulp, or an aliquot part of it, was washed into an evaporating basin and dried at 900 to 1000 C. It was again weighed and then thoroughly extracted by grinding with five or six changes of warm water, the total volume of which was known. Most of the suspended matter in the extract was removed by the centrifuge, and urea and sulphate were determined in suitable quantities of the fluid. Urea was estimated by hydrolysis with urease followed by vacuum distillation, and sulphate by the gravimetric method, the organic matter in the extract being first oxidised with potassium chlorate and hydrochloric acid. The urea and sulphate of the

3 424 E. B. MA YRS. plasma and urine were determined in the same way, except that before sulphate estimations in the plasma the proteins were destroyed by incineration. The total fluid of the kidney is represented by the weight lost in drying. Part of this fluid was in the renal cells and part in the tubules; traces probablyin the smaller b,iood vessels and in intercellular lymphatics. The urea and sulphate in the mixed fluid were calculated from the results, and compared with the concentrations found in the plasma and urine. The figures obtained in a number of experiments are given in the accompanying table. Vol. of I/0 0/0 0/0 0/0 0/o 0/o Wt. of urine. urea urea in urea Na.2S04 Na 'O40 Na2SO4 rabbit, Injection. c.c. per in ki(dney in in in kidnley in No. kilos c.c. minute plasma fluid urinie plasma fluid urine Urea 5 0% *380 *258 *605 * Na2S ,, *272 *272 *717 *511 *452 1* Urea 10 0/ *475 *412 *984 *752 * Na2S04 10 %// 4 2-0,, *260 * *489 *515 1* Urea 5 % 27-0 *53 *150 *206 * Na2SO4 10 % 6 2-1,, *201 *243 *520 *511 * ,, *177 *288 *365 *583 * ,, *308 *336 * Average *598 * It is clear from these results that during excretion of urea and sulphate, under the experimental conditions described, their concentrations in the total fluid of the kidney do not differ much from their concentrations in the plasma. Where any material differences are observed, the records of exceptionally high and exceptionally low kidney concentrations are about equally divided. If the error of individual experiments is eliminated as far as possible by taking the average results of the series, the concentrations in the kidney and plasma are found to be practically the same. When it is remembered that the urea and sulphate in the partially formed urine of the renal tubules must have undergone some concentration, the small total content of these substances is even clearer evidence that there is no concentration in the renal cells. It may be objected that urea is perhaps shielded from the action of urease by forming a compound with the protein of secreting cells; but no similar explanation will cover the shortage of sulphate, since organic matter is oxidised before the estimation is begun. The process of elimination of urea and sulphate is, therefore, entirely different from that which Marsh all and Vi c k er s describe in the secretion of dye; for these observers conclude that a certain amount of dye

4 KIDNEY EXCRETION. 425 is stored in the renal cells before concentration in the urine occurs. Urea and sulphate, on the contrary, do not appear to be stored in the renal cells, and their concentrations in these cells may even be lower than their concentrations in the plasma. Two objections may be raised to the technique recorded above. In the first place, it may be argued that the interference with the kidney necessary for arranging ligatures on its vessels was so great as to prevent secretion from taking place. In some of Marshall and Vickers' experiments, however, in which part of the spinal cord had been destroyed, the kidney must have been in a much more unfavourable condition for secretory efforts; yet storage of dye in the renal cells was found to occur. Secondly, it may be objected that the concentrations of urea and sulphate in the plasma were already so high that storage in the renal cells at a greater concentration was impossible, owing to the large amount of work against osmotic tension which this would involve. But the concentrations of these substances were increased by some means during the formation of urine, and if the kidney can secrete them, part at least of this increase might have been expected to take place within the cells. This introduces the question of whether true secretion of any constituent of the urine might be possible without intracellular concentration; for the process might consist simply in retention of water while the substance is passing through the free border of the cell. The secretion of hydrochloric acid by the stomach may be an example of some such process, alkali in this case being held back. In view of the toxicity of certain products which the kidney must excrete, the value of an arrangement of this kind is clear; although the advantage of concentration by. absorption of water from the tubules is still more obvious. But in the experiments of Marshall and Vickers (the only evidence of secretion by the mammalian kidney on which any reliance can be placed) concentration of dye in the renal cells undoubtedly occurred; and the inference can be drawn that if the kidney could secrete the normal constituents of the urine, it would do so by a similar method. At present, therefore, concentration in the cells must be regarded as a necessary criterion of renal secretion. A striking characteristic of the excretion of dyes, well shown in the experiments of de Haan, is the extent to which the concentration of free dye is raised in its passage through the kidney. The concentration may be 8000 times as high in the urine as in an ultrafiltrate from the plasma. If secretion ordinarily takes place in the kidney, some, at least,

5 426 E. B. MA YRS. of the constituents of the urine should be concentrated to a comparable degree. But even when the volume of urine is small, no increase of this magnitude is found to occur. Any doubt which exists in the case of creatinine, owing to the negligible amount in the plasma, can be removed by injecting this substance and observing its elimination; and the same method can be applied to ammonia also. In neither instance is the concentration very great. Hence, while it is not possible at present to dispute the secretion of dyes, it has yet to be shown that this process has any resemblance to the ordinary formation of urine. In conclusion it is suggested that the primitive kidney may have had both secreting and absorbing functions, but that in the mammal a method of excretion has been selected which depends on filtration and re-absorption; only a few traces surviving of the earlier secretory power. The secretion of readily adsorbed substances, which exert little osmotic resistance, is possibly a relic of the past; perhaps also the synthesis of hippuric acid and the hydrolysis of urea(5); for none of these processes seem to have much value as functions of the mammalian kidney. In the bird absorption of fluid is, for the most part, effected in the cloaca, and the kidney may have preserved its secretory function. Some evidence of secretion of uric acid has recently been obtained in the fowl. If the view advanced here is a correct estimate of the position of secretory activity in the kidney, the value of phenol sulphone phthalein as a test for renal efficiency must be doubted; since this substance is apparently eliminated by secretion, and gives no evidence of functional capacity in dealing with ordinary waste products. SUMMARY. No intracellular concentration has been found to occur during excretion of urea and sulphate by the kidney. Evidence obtained by other observers that phenol sulphone phthalein is secreted shows that concentration of the dye takes place in the renal cells. It is therefore concluded that urea and sulphate are eliminated by a process other than secretion; and, in view of the relatively small increase in concentration which occurs in excretion of waste products as compared to dyes, this conclusion is extended to the urine as a whole. The conception of renal activity which justifies the employment of dyes in efficiency tests is probably erroneous. Most of the expenses of this research have been covered by a grant from the Moray Fund of Edinburgh University.

6 KIDNEY EXCRETION. 427 REFERENCES. (1) Marshall and Vickers. Bull. Johns Hopkins Hosp. 34. p (2) de Haan. Journ. Physiol. 56. p (3) Heidenhain. Pfluiger's Arch. 9. p (4) Cushny. Journ. Physiol. 51. p (5) Nash and Benedict. Journ. Biol. Chem. 48. p