College of Medicine, Newcastle-upon-Tyne.)

Transcription

1 GLUCOSE ABSORPTION IN THE RENAL TUBULES OF THE FROG. BY G. A. CLARK. (From the Physiological Laboratory of the University of Durham College of Medicine, Newcastle-upon-Tyne.) OPINION is divided on the question of how the kidney prevents the passage into the urine of the glucose normally present in the blood. The modern theory assumes that the blood-sugar is present in the glomerular filtrate and is reabsorbed by the tubule epithelium, while according to Heidenhain the glomerular membrane is impermeable to glucose. Evidence in support of the latter view has been produced by Hamburger(l) who perfused the frog's kidney by way of the renal arteries with a modified Ringer's fluid containing glucose and found that a sugarfree urine was formed when the perfusing fluid contained *08 p.c. (and occasionally *12 p.c.) glucose. Because the glomeruli in the frog are supplied by the renal artery only, it was held that this retention of glucose was due to some function of the glomerular membrane; it was not fully appreciated that, under the conditions of experiment, the tubules were capable of their normal activity. In the frog the efferent vessel from each glomerulus opens into the venous spaces surrounding the tubules, so that the latter must be bathed byanyfluidflowing through the renal artery. Hamburger also determined that when the perfusion contained *25 p.c. glucose or more, the kidney allowed the whole of this glucose to escape into the urine. This was attributed to a "sickening" effect on the glomerular membrane. The experiments described below were undertaken to ascertain if the renal tubule played any part in preventing the blood-sugar from passing into the urine. The method of perfusion employed was that described by Bainbridge, Collins and Menzies(2). Winter frogs which had been kept in a semi-dark tank were used, and whose blood-sugar content was found to vary between *020 p.c. and *027 p.c.; these figures correspond with those determined by Bang(3), whose micro-chemical method for the estimation of sugar was used in these experiments. The perfusing fluid, in which varying amounts of sugar were dissolved, had the following composition: NaCl 5 p.c., KCl *01 p.c., CaCl2 *02 p.c., NaHCO3 *285 p.c.

2 202 G. A. CLARK. The alkalinity of this fluid corresponds to that of normal frog's serum (1) and the calcium content (which varies with the season of the year in the frog) is approximately the same(4). The arterial perfusion pressure was 23 to 25 cms. of water and the venous 10 to 12 cms. As L. Hill(5) has pointed out that the normal pressure found in the frog's glomerular tuft is only 6-7 to 13-5 cms. of water, and that in the tubule capillaries only 2-7 to 4 cms. a number of experiments were carried out at these reduced pressures, but it was found that the results obtained were in no way affected, except that the amount of urine was considerably less. Experiments,were first performed to determine the " renal threshold" for glucose: in these, both aorta and renal-portal vein were perfused with fluid having the same concentration of sugar. It was found that when the perfusing fluid contained *052 p.c. glucose or less, no sugar could be detected in the urine; in two cases slightly higher concentrations were retained (.056 p.c. and *058 p.c.). The whole of the blood-sugar in the frog is said to be contained in the plasma, none being found in the corpuscles(l; the concentration in the plasma of the winter frog would therefore be about -045 p.c. Thus a "threshold" of *052 p.c. would amply provide against escape of glucose into the urine under normal conditions. The value of the "threshold" varies, however, with the calcium content of the perfusing fluid, as Hamburger points out(1); if no calcium be present, the kidney becomes completely permeable to sugar, whereas increased amounts above *02 p.c. appear to have little or no influence. It is of importance to recognise this fact, for, if the perfusing fluid be not freshly prepared, the calcium is slowly precipitated as carbonate owing to the decomposition of the NaHCO3; in one case the whole of the calcium was precipitated in 48 hours. When the perfusing fluid had a glucose content above the " threshold" but less than *23 p.c., the percentage of sugar in the urine was always less than that in the perfusing fluid by 05 p.c. to *06 p.c. With concentrations above *23 p.c. the kidney allows the whole of the glucose to escape in the urine. In order to ascertain the effect of this increased concentration of glucose on the tubules alone, advantage was taken of the fact, described in a recent paper(6), that if the renal-portal vein be perfused with Ringer's fluid containing as much as *5 p.c. glucose, while the renal arteries are simultaneously perfused with Ringer's fluid at twice the venous pressure, no sugar can be detected in the urine. This fact was verified for the present paper. Under these conditions, the venous perfusion does not reach the glomerulus and no sugar passes into the urine through the tubule epithelium. It was found by experiment

3 KIDNEY SECRETION. 203 that with a venous perfusion containing *4 p.c. or *5 p.c. glucose, while the arterial contained less than '1 p.c., the rates of flow of the two were about equal, and in many cases the arterial rate was greater than the venous; the increased sugar-content appeared to retard the flow. Thus, the concentration of glucose in the fluid in the tubule capillaries would depend on the relative amounts of the two perfusions mingling there; this concentration can be determined by estimating the glucose in the mixed perfusate flowing from the vena cava. The renal arteries were perfused with a fluid containing less than *1 p.c. glucose, while the venous perfusion contained *4 p.c. or *5 p.c. From Table I it will be seen that when the mixed perfusate contained *21 p.c. glucose or more, the urine had the same percentage of sugar as the arterial perfusing fluid, that is, the kidney was completely permeable to the sugar in the arterial perfusion. That complete permeability was obtained when the mixed perfusate contained *21 p.c. glucose, while in the previous experiments *23 p.c. was necessary, can be attributed to the fact that the presence of the testes and fat-bodies increased the proportion of arterial perfusion in the mixed perfusate. TABLE I. Percentage of glucose in &- 5~A- Urine Urine per kidney Arterial Mixed per hour Exp. perfusion perfusate Right Left c.c. 1 *021 * * * * '030 '028 *6 4 '065 *205 '055 * *072 *21 '073 * *22 *011 *010 *4 7 *026 *22 '025 '027 '2 8 '070 *22 '068 ' '080 *22 *081 - *1 10 *072 '24 '072 '073 *15 11 *032 *26 '000 '032 *27 12 '036 '35 '038 '035 '15 That the effect of the increased sugar percentage is not on the glomerular membrane is obvious, because the renal-portal perfusion does not reach the glomerulus. Moreover, the sugar in the urine cannot be derived from that in the venous perfusing fluid because it has been shown that no sugar passes through the tubule epithelium (6); the fact that in each case the glucose content of the urine is identical, within the range of experimental error, with that of the arterial perfusing fluid is also evidence against this explanation. Thus, the only possible conclusion is that normally glucose is filtered through the glomerular membrane

4 204 G. A. CLARK. and reabsorbed by the tubule epithelium, but that this reabsorption; cannot proceed under the conditions described. By substituting a venous perfusing fluid having the same concentration of glucose as the arterial, it was found that the kidney gradually regained the power of retaining glucose. Unfortunately it was not possible to maintain the secretion of urine long enough for the return of the normal "threshold," but usually after one hour the kidney could retain about -03 p.c. to -04 p.c. In Exp. 11 (Table I) it is seen that the urine from the right kidney contained no glucose, while that from the left showed the same concentration as the arterial perfusion. This was due to an accident by which the right renal-portal vein was included in the ligature holding the ureteral cannula in position; the right kidney therefore was receiving no venous perfusion and, as would be expected, was capable of its normal activity. Throughout the experiments described, the rate of formation of urine bore no relation to the sugar-content of the perfusing fluid. In the mammal, diuresis in glycosuria is due to the osmotic resistance offered to the absorption of water in the tubule by the glucose content of the glomerular filtrate (7); in the frog, however, no absorption of water occurs in the tubules, as can be seen from Table I, where the urine contains exactly the same percentage of sugar as the arterial perfusion. The absorption of water appears to be actually resisted, for the physical conditions invite the passage of water from the glomerular filtrate to the more concentrated fluid in the tubule capillaries. In only one experiment (not quoted) was the sugar content of the urine higher, but in this case the arterial perfusion pressure was less than twice the venous pressure, thus admitting the possibility of the venous perfusing fluid reaching the glomeruli. The tubule epithelium must therefore be regarded as an irreciprocal membrane, so far as glucose ig concerned, of the same type as that formed by the epithelium of the small intestine. Additional evidence of the function of the renal tubule to absorb glucose was furnished by experiments in which the tubule epithelium was poisoned by mercuric chloride or sodium arsenite. A *01 p.c. solution of the former was used and the latter was prepared by dissolving *5 gm. arsenious oxide in 100 c.c. of Na2CO3 solution. Both aorta and renalportal vein were perfused with fluid containing less than -1 p.c. glucose. When the kidneys had been washed free from blood about 5 c.c. of the toxic solution were passed through the venous cannula. At the end of the experiment the renal-portal vein was perfused with dilute (NH4)2S when the mercuric salt had been used and the kidneys examined in

5 KIDNEY SECRETION. serial sections; when sodium arsenite was employed, *5 p.c. CuSO4 was passed through the venous cannula; with the arsenic this formed the insoluble "Scheele's green," which could be distinguished microscopically. Whenever evidence of the presence of the poison was found in the glomeruli, the experiment was discarded, but the only difference observed between those cases where the tubules alone were affected and those where the glomeruli showed a deposit was that in the latter the rate of formation of urine was slower, even when the perfusion flow was normal, although this last was usually reduced. In every case the urine contained the same percentage of glucose as the perfusing fluid. SUMMARY AND CONCLUSIONS. (1) The glomerular membrane is completely permeable to glucose, even when the latter is present in the blood below the " threshold value." (2) The epithelium of the renal tubules has the power to absorb glucose from the glomerular filtrate up to the normal "threshold value." This absorption is possible until the capillaries surrounding the tubules contain glucose at a concentration of nine or ten times that normally present in the blood. (3) In the absence of calcium the renal tubule is no longer capable of its normal activity. (4) No concentration of the glomerular filtrate by absorption of water occurs in the frog. REFERENCES. (1) Hamburger. Brit. Med. Journ. 1. p (2) Bainbridge, Collins and Menzies. Proc. Roy. Soc. B. 86. p (3) Bang. Der Blutzucker (Wiesbaden), p (4) De Waard. Biochem. Ztsch (5) L. Hill. Brit. Journ. Exp. Path. 2. p (6) Atkinson, Clark and Menzies. Journ. Physiol. 55. p (7) Cushny. The Secretion of Urine, p PH. LVI. 14