RICHARD H. KESSLER,2 KLAUS HIERHOLZER3 AND RUTH S. GURD4 Department of Physiolugy, Cornell University Medical College, New Turk City KESSLER, RICHARD H., KLAUS HIERHOLZER AND RUTH S. GURD. Localisation of wale trans~ort in the nephron of the mongrel and Dalmatian dog kidney. Am. J. Physiol. Ig7(3) : 601-603. I g5g.-Localization of urate transport within the nephrons of mongrel and Dalmatian dogs was studied by stop-flow analysis. In mongrel dogs urate concentrations and clearance ratios were lowest in the segment in which PAH was secreted. Urate clearance ratios of 0.7 in free-flow samples were reduced to about 0.3 in stop-flow samples from the proximal segment. In the distal segment urate clearance ratios did not differ significantly from ratios obtained in free-flow. Probenecid, in ,doses sufficient to block PAH secretion, inhibited urate re- absorption thereby increasing urate clearance. In contrast to these findings with mongrel dogs, the Dalmatians exhibited weak but definite urate secretion within the proximal segment. The action of probenecid in this strain of dogs was .to stop all proximal secretory activity for urate thereby reducing urate clearance. It was suggested that mongrel and Dalmatian dogs transport urate by systems that are identical except for direc- tion of urate movement. T HE RENAL CLEARANCE of urate is considerably less than the rate of glomerular filtration. As urate is com- pletely filterable, its low clearance indicates reabsorp- tion by tubule cells. Both reabsorption in dogs and a reabsorptive Tm in human beings have been demon- strated. In contrast to glucose, urate reabsorption is in- complete, and even at low plasma concentrations some urate appears in the urine. In this respect its tubular transport is more akin to the reabsorptive mechanisms of amino acids, sulfate and phosphate. Several agents can increase urate excretion by diminishing its reabsorp- tion. Among the most potent of these is probenecid (Benemid) which, when used in maximally effective dose, can increase urate excretion only 10-15 %. Even Received for publication April 3, I 959. l This study was aided by research grants from the National Heart Institute, National Institutes of Health, and the Life Insurance Medical Research Fund. 2 Senior Fellow of the Nathan Hofheimer Foundation. 3 New York Community Trust Fellow. 4 Life Insurance Medical Research Fund Fellow. 601 with this uricosuric drug the urate clearance is ap- preciably less than glomerular filtration rate. An excep- tion to these generalizations about the renal transport of urate among the mammalia exists in the Dalmatian coach hound. The Dalmatian lacks the ability to re- absorb urate and exhibits urate clearances that approxi- mate glomerular filtration rate. This study reports on the localization of urate transport in the mongrel and Dal- matian dogs. Using the stop-flow method, we have observed urate reabsorption within the proximal seg- ment of the mongrel dog kidney. The action of pro- benecid in impeding urate reabsorption was likewise placed within the proximal segment. The Dalmatians not only failed to reabsorb 1 trate but exhibited modest urate secretion within the proximal segment. This se- cretory activity was abolished by probenecid. METHODS This report is based on 13 experiments performed on seven dogs including two Dalmatians. Details of six ex- periments will be presented. The technique employed was the same as that described by Malvin et al. (I) as modified in our laboratory (2). Animals were infused intravenously with a creatinine and PAH solution and with a solution of 20 % mannitol in saline. Uric acid in concentrations of 0.5 or 1 .o gm/l. and either 0.7 % sodium hydroxide or lithium carbonate were added to each infusion. Resulting plasma urate levels ranged from 3.0 to 14.0 mg % with filtered urate loads of from 0.8 to 4.0 mg/min/kidney. Using a retroperitoneal approach, the right ureter was catheterized with a polyethylene tube. Urinary stasis was produced by clamping the catheter for a 4-min- ute period. The volume of fluid trapped within the kid- ney varied from 20 to 25 ml. On release of the clamp this trapped fluid was rapidly ejected from the kidney and collected in serial samples of about I ml each over the ensuing 3 minutes. A single experiment consisted of this procedure of stasis and serial collection preceded and followed by conventional clearance samples. In all but 602 RICHARD H. KESSLER, KLAUS HIERHOLZER AND RUTH S. GURD #ia A A A .o. a l * AA. a . . &AA Fig. 2 J, 0.20 pNo 0.15 J/PC, 0.10 0#05 1.0 cl9 b 08l G LOM. II ---- PROXIMAL ---- - - - ---- DISTAL ---- --- Fig, I ---w- PROXIMAL -PC----I ---------- DISTAL ----- ----- I A *A .a. **A 0. l . . . AA. mm a A.4 / 0.25 l A A , A p,, O.2 l l l : 4 = : f A . A OJO OJO l 0.05 0.05 u/ u/ 0.7 A 0.7 A (3.6 A * pUR. pUR. 0.6 A * AA AA AA AA AA AA 0.4 0.4 AA AA A A A A A *A 0. **A .A *A AA l l A A l m am l A l J/ 07 per l 0.6 05 0.4 013 l A A . 4 l & A AA l A 0 l *.A . URATE = . . URATE + BENEMID l . CONTROL URATE 0. l a .* 5 . l *. l m .* l l l . . &**A*****A.A.AA&&AAAAAA pPAH 5 - IO l : t * /PC, *: A 5 rat 444 4 / l l l l l per 3 . . . . l : i t l . l * : A A A *A l l 24 * AA l A 04 2 4.4 A x A4 u6u5u4 110 90 70 50 30 IO USUZI a* . l a l . l * mm l A..*. 4 A * * l & l l l mAA*A&mA t :** A4 A I I U6 U5 UL II0 90 70 50 30 IO % Volume within Ktdney u3uzII % Volume When Ktdney FIG. z. Effect of Benemid (probenecid) on urate clearance ratios, This interference with urate reabsorption may be confined to proximal segment. FIG. I. Localization of urate reabsorption to proximal segment. Region of highest PAH clearance ratios mark proximal segment; lowest Na clearance ratios mark distal segment. one instance two consecutive studies were conducted with each animal. The concentration of urate in protein-free plasma filtrates and in diluted urines was measured by the arsenophosphotungstic acid method (3). Other chemical methods and mode of presentation of data have been published (2). Fig. 3 1.6 / UR. k4 u/p 1.2 Cr IL m / I5 PA, Ic / PGr f L /P Cr : .- L GLOW --p-m----- - PROXIMAL c_-_.-I---- -----__ p-w- DISTAL ----c-- -w-- l l m em. .A A.4 A A * ** A * **AA&A** Control = l RESULTS Figure I illustrates the results of two consecutive ex- periments performed on a single animal. The control study is plotted with circles and the study during urate infusion plotted with triangles. Data include U/PCr and clearance ratios for PAH, urate and sodium. At the lateral margins of the figure are the values for the con- ventional clearance periods. Stop-flow results are pre- sented in the center as percentage volume contained within the clamped kidney. Initial or distal segment samples appear to the right while later or proximal seg- ment samples are shown to the left. At the top of the figure a schematic nephron illustrates functional loci based for the distal segment on lowest sodium ratios, for the proximal segment on maximum PAH ratios and for Benemid=r II0 90 70 50 30 IO % Volume wlthm Kidney FIG. 3. Secretion of urate and its suppression by probenccid in proximal segment of the Dalmatian. LOCALIZATION OF URATE TRANSPORT IN NEPHRON IN DOG KIDNEY 603 c the glomerulus on the appearance of inulin injected just prior to collection of the serial samples. During free flow, urate clearance ratios measured 0.6-0.7. Almost identical values were obtained in the distal, stop-flow samples. Reabsorption of urate with ratios approaching 0.3 appeared in those same samples in which PAH ratios were highest. Urate was reabsorbed within the proximal segment. Note also that the alkaline infusion of urate delivered at 7 mg/min, did not alter creatinine, PAH, or sodium ratios. Figure 2 confirms these observations and demonstrates the actions of probenecid. In this example both experi- ments were performed during urate infusion before (circles) and after (triangles) probenecid. The altera- tions effected by this drug included inhibition of PAH secretion and continued but diminished reabsorption of urate. Clearance ratios for urate were higher in distal as well as proximal segment samples. Blockade of urate reabsorption at the proximal site would present to both the distal segment and ureter more urate than that delivered prior to probenecid. One need not invoke a distal segment effect of probenecid to interpret the data. Figure 3 illustrates consecutive experiments per- formed on a Dalmatian before and after a probenecid infusion. In the control study, shown by circles, urate clearance ratios exceeded I .o in all specimens. These ratios attained a value of 1.6 within the proximal seg- ment. The data from this and a companion experiment indicated modest secretion of urate in this strain of dog. Following probenecid, urate secretion was abolished; note that all ratios were less than I .o. DISCUSSION Urate reabsorption is incomplete even at low plasma concentrations. Unlike glucose, urate appears in the urine at filtered loads well below Tm. In this respect the mecha- nism for urate reabsorption is similar to the tubular transport of amino acids, sulfate and phosphate. There are species differences in the capacity to reabsorb urate; the transport maximum in the dog exceeds that of human REFERENCES I. MALVIN, R. L., L. P. SULLIVAN AND W. S. WILDE. Physidogist 1: 58, qj7* 2. PITTS, R. F., R. S. GURD, R. H. KESSLER AND K. HIERHOLZER. Am. J. P&siol. 194: x25, 1958. 3. BUCHANAN, 0. H., W. D. BLOCK AND A. A. CHRISTMAN. 3. Bid. Chem. 157: 181, 1957. beings. Berliner and co-workers have been able to demon- strate a Tm in man of I 5 mg/min/~ .73 m3 (4). In con- trast to this, Friedman and Byers have reported urate clearance ratios in dogs of less than 0.6 at a urate load of 20 mg/min. (5). The relative insolubility of urate limits the quantity that can be administered. As a result, plasma levels of urate sufficient to reach Tm in the dog have not been achieved. Our data placed the site of urate reabsorption within the proximal segment of the mongrel dog kidney. There was no evidence for urate secretion. As might be pre- dicted, the moderate inhibition of urate reabsorption by probenecid occurred at this same site. Probenecid blocked proximal secretion of PAH as it inhibited proxi- mal urate reabsorption, However, there was less inhibi- tion of the secretory system for urate than for PAH. This is apparent in these data as well as in previous clear- ance studies. The Dalmatian is a curiosity among the mammalia. This strain of dog excretes urate in quantities that suggest the absence of a tubular reabsorptive system for urate. Noted first by Benedict in I 915 (6), the high urate excre- tion by the Dalmatian has been more precisely studied by Friedman and Byers. These authors reported CUrate/ Ccr ratios of 0.89 to I. IO over a wide range of plasma levels of urate (5). Weak secretion of urate in the Dal- matian has been observed (personal communication from A. B. Gutman). By the stop-flow technique exag- geration of tubular transport has allowed localization of this urate secretory mechanism to the proximal segment. Probenecid exhibited a more pronounced action on urate clearance ratios in the Dalmatian than in the mongrel dog. Urate transport is in opposite direction in these two strains, yet probenecid apparently interferes with urate transport in both instances. Perhaps all mammals pos- sess a common transport system for urate which in the Dalmatian is directed from tubule cell to lumen. This distinguishes the Dalmatian from other mammalia in which the identical transport system directs urate from lumen to tubule cell. 4. BERLINER, R. W.,J- G. HILTON, T. F. Yij AND T.J. KENNEDY, JR. J. Clin. Invest. 29: 396, 1950. 5. FRIEDMAN, M. AND S. 0. BYERS. 3. Bid. Chem. 175: 727, 1948. 6. BENEDICT, S. R. Harvey Lect. I o : 346, I g 15-16.
Biochemical Factors Concerned in the Functional Activity of the Nervous System: First International Meeting of the International Society for Neurochemistry, Strasbourg, 1967