Glomerular filtration in single nephrons

Transcription

1 Kidney International, Vol. 1 (1972), p EDITORIAL REVIEW Glomerular filtration in single nephrons In recent years as many investigators have turned their attention to the study of tubular transport processes in individual nephrons, it has become increasingly important to measure and to understand the factors that determine the filtration rate of single nephrons (SNGFR). More frequent measurements of SNGFR have yielded a great deal of new information concerning the function of individual glomeruli. An inevitable consequence of more information, however, seems to be controversy. Controversial matters that have arisen in recent studies of SNGFR include questions about the most appropriate technique of making the measurements, as well as disagreement about conclusions that have been drawn from the results Of these measurements. Knowing the rate of glomerular filtration in single nephrons is crucial to the determination of absolute rates of reabsorption or secretion by the renal tubules. The excretion rate of sodium, for example, has long been known to be dependent on the filtered load of sodium. Assessment of factors involved in the control of sodium reabsorption depends on knowing the rate at which sodium enters and leaves a tubule segment. Measurement of SNGFR is also important to investigation of the regulation of the filtration process itself. It is likely that future clarification of the still unknown mechanisms controlling filtration rate will come from studies where SNGFR is measured as the functional environment of that nephron unit is controlled. Factors that affect SNGFR can presently be grouped into those determined by the functional state of the experimental animal and those that result from technical difficulties and artifacts. It appears that SNGFR is influenced by age [1, 2], body weight [1, 2], the number of kidneys present [3], hydration [4], extracellular fluid volume [5], and the genetic makeup [6, 7]. The nephron site at which micropuncture collections are made proximal or distal, superficial or deep also has to be considered. 1972, by the International Society of Nephrology. Technical d(fficulties and art?facls. Several investigators using micropuncture methods have examined the degree to which variations in sampling technique can affect measurements of SNGFR. Some of the results of these efforts have been published and other more recent findings were discussed in November, 1971, at a Micropuncture Workshop in New Haven.1 Potential sources of artifactual results that have been considered include variations of intratubular pressure at the collection site, collection from more than one tubule, the retrograde flow of tubule fluid during sampling, and the effect of repeated sampling at the same site. Tables 1 and 2 show the reported values for SNGFR that have been derived from micropuncture measurements in superficial proximal tubules of adult, hydropenic rats and dogs. Most values for rats range between 25 and 45 ni/mm (Table I). The level of SNGFR in the dog is approximately twice as high (Table 2). The rather wide range of reported values has not been narrowed very much after more than five years' experience with the method. Age, weight, hydration, collection site and collection technique are similar in these studies. Differences between strains of inbred rats may well provide a source of some of the variation in Table I. The limited attention that has been given to the question of genetic differences has shown that there may be inherited variations in filtration rate [6, 7]. Although deliberate attempts to vary SNGFR by changing the sampling method have often shown only small effects, it remains likely that some of the variation in Table 1 is due to differences in technique. Schnermann, Horster and Levine found that SNGFR could be increased approximately 25% by reducing intratubular pressure during sampling by "aspirating" [13]. Brenner, Daugharty, IJeki and Troy [28] found that "excessive suction" may have increased SNGFR by about lo%, but concluded that normal collection The presentations at the Workshop on Renal Micropuncture Techniques will be published in a future issue of the Yale Journal of Biology and Medicine. 201

2 202 Wright / Giebisch Table 1. Filtration rate of superficial nephrons in rats U SNGFR ni/mm Reference SNGFR ni/mm Reference 32.7b C a Results of free flow micropuncture collections from surface proximal tubules of adult rats under non-diuretic conditions. Most values are means reported by authors or weighted averages of reported group means. a Uninephrectomized rats were used. One-quarter of a mean value per kg body wt. Table 2. Filtration rate of superficial nephrons in dogs a SNGFR ni/mhz Reference SNGFR ni/mm Reference a Measurements in dog experiments were made under conditions similar to those in Table 1. techniques using "controlled suction" at late proximal sites did not substantially alter early proximal tubule pressure or SNGFR. Schnermann [31, 47] has re-examined the effect of aspiration on measurements of SNGFR, and he has found that SNGFR can be increased about l4% by aspirating. He agrees with Brenner that this is not an important source of error. Davidman, Lalone, Alexander and Levinsky [32] and Andreucci, Herrera-Acostä, Rector and Seldin [35] and Andreucci and Rector [48] have found no correlation between reduction of intratubular pressure of six to seven cm H20 and measurements of SNGFR. One explanation for the lack of effect is that offered by Brenner et al [28]. By measuring hydrostatic pressure at two sites in the same proximal tubule he observed that reduced pressure in the late proximal tubule is not transmitted to early proximal segments because of the tendency of the intervening segment to collapse. It appears, therefore, that a reduction of late proximal pressure will not falsely elevate the SNGFR. However, changes of intraluminal pressure can probably affect filtration in two other situations: 1) aspiration during collections from early proximal segments would be expected to increase SNGFR, and 2) elevations in late proximal pressure caused by blocking tubule fluid flow would be expected to be transmitted back to the glomerulus and thereby decrease SNGFR. Schnermann, Levine and Horster showed that obstruction of a single nephron cciuld lower its SNGFR by 39% [12] and have concluded [31] that elevations in collection line pressure can be responsible for the low estimates of SNGFR that were reported by Gertz, Braun-Schubert and Brandis [14] and Gertz et al [49]. ideally, SNGFR should be measured at the luminal pressure existing prior to puncture. In practice, accomplishing this goal seems to require an initial measurement of the hydrostatic pressure of the tubule under study, and then a collection of fluid at that pressure. This approach would require the use of a second micropipet in place during the collection. Discussion at the Micropuncture Workshop indicated that most investigators at the present time think it reasonable to measure SNGFR simply by collecting fluid from the late proximal tubule if care is taken by gentle aspiration, to avoid raising intratubular pressure. The problem of proximal versus distal collection sites will be discussed later. 'The possibility that falsely high values for SNGFR can be due to collections from more than one tubule has recently been discussed by Andreucci et al [35, 48]. They suggest that inadvertent puncture of a second nephron below the kidney surface may create a persisting leak into the nephron of interest. The fluid flow rate would then be spuriously elevated. Visual control of the pipet tip is essential to be sure that underlying tubule segments have not been punctured. Suspicion that other segments were punctured should be grounds for discarding the sample before analysis. Also, subsequent microdissection of a latex cast of the tubule will aid in determining whether adjacent segments were punctured. An apparently less troublesome matter of technique is the threat that one may collect fluid from two directions at once. Brenner and co-workers [5, 15] concluded that falsely high values for SNGFR could

3 Single nephron GFR 203 be due to a retrograde leak of distal fluid around an oil block too short to seal the collection site from distal nephron segments. They found high values for SNGFR more frequently with short blocks of polymer oil (Kel-F oil) than with longer ones or castor oil blocks. They assumed that less reflux occurred when long castor oil blocks were used [15]. More recently direct tests of the possibility of retrograde flow have been carried out. Schnermann et al [31] found that, with mineral oil, even blocks as short as 50 itwere adequate to prevent reflux. Additional reassurance is given by Andreucci and Rector [35, 48] who also tested the possibility of retrograde flow by direct experiments. Using mineral oil blocks of two to four tubule diameters and creating pressure gradients favoring retrograde flow, they found no back leakage of tubule fluid. These studies thus provide evidence that retrograde flow is not an important source of error in studies where mineral oil or castor oil is used to block the tubule. No direct experiments, however, have been done that show whether less viscous oils such as Kel-F can be used safely. The possibility of an additional artifact in SNGFR measurement, though not one that contributes to the variability in Tables 1 and 2, was pointed out by Mandin et al [50]. They found in dogs that recollection from a previously punctured tubule was associated with a spuriously elevated SNGFR especially after volume expansion with saline. Presumably, surface fluid enters the collecting pipet in addition to tubule fluid during repuncture because the hole in the tubule wall is larger and "leaky". Several investigators have examined this possible source of error in rats and agree that it is not important [2, 6, 28, 32]. The matter remains controversial, however, in dog experiments. Auld, Alexander and Levinsky [40], as did Mandin et a! [50], found higher values in repunctured tubules than in fresh tubules. Schneider et al. [43], however, found SNGFR to be the same in repunctured and fresh tubules after saline loading. Respiratory and pulsatile movements are more troublesome in dog experiments and they may make it more difficult to obtain an adequate seal around the pipet. It has been suggested [35, 51] that repuncture at a slightly more proximal tubule site may minimize this source of error. Eliminating pulsatile movements as Seely and Boulpaep have done by autoperfusion [52] is another approach that may be useful in some experiments with dog kidneys. Further investigation should clarify whether previous impalement of a tubule raises the SNGFR of its glomerulus. Heterogeneity of nephron populations. New information about the function of individual nephrons has come from several different experimental approaches. It is now clear that, in addition to morphological differences among nephrons, there is functional heterogeneity as well: filtration rates differ at different levels in the renal cortex; the response of SNGFR to a variety of physiological changes may be different for different nephron populations; and different control mechanisms may mediate these changes. The suspicion that nephron heterogeneity might play a significant role in renal function [53, 54] has been investigated extensively in recent years. A microdissection technique described by Hanssen [55] provided evidence that nephrons deep in the cortex (juxtamedullary) had higher filtration rates than superficial nephrons. De Rouffignac and co-workers [56 58], using modifications of this technique have obtained values for juxtamedullary SNGFR ranging from 38.5 to 49 nl/min and values for superficial SNGFR ranging from 27.9 to 32.7 ni/mm. The lower values are the more recent and are considered to be better estimates. Using de Rouffignac's modification of the Hanssen method, Davis and Schnermann [59] found even lower values for juxtamedullary and superficial filtration rates. They found that the SNGFR was 30.1 nl/min for deep nephrons, and 24.5 nl/min for those on the surface. Concurrent with this extensive application of the Hanssen ferrocyanide technique, other workers have studied nephron heterogeneity using micropuncture techniques. In 1968 Horster and Thurau [60] reported SNGFR determined from micropuncture of both surface proximal tubules and from loops of Henle punctured near the tip of the renal papilla in young rats. They found that the juxtamedullary SNGFR averaged 58.2 nl/min/g kidney wt, more than twice the superficial SNGFR of 23.5 nl/min/g kidney wt. Jamison and co-workers [2, 61, 62] and Stumpe, Lowitz and Ochwadt [16] have made similar observations. They have reported mean values for juxtamedullary SNGFR's ranging from 50.9 to 64.9 nl/min/g kidney wt and superficial SNGFR's ranging from 21.3 to Investigators using microdissection and micropuncture methods agree, therefore, that superficial and deep SNGFR's differ. Results from the two approaches are not identical, however, and it should be noted that the ratio of superficial to juxtamedullary SNGFR in micropuncture experiments is less than 0.5, whereas the ratio is approximately 0.7 when the Hanssen technique is used. Also, whereas the superficial

4 204 Wright / Giebisch SNGFR determined by micropuncture is usually greater than 30 ni/mm, that measured with the microdissection technique is usually less than 30 nl/min. Some of this difference may be related to differences in kidney size, but it is also of interest that, although the micropuncture determination are higher, de Rouffignac and Bonvalet believe that the ferrocyanide technique may overestimate the true value for SNGFR by about 11% [63]. Does distribution of SNGFR vary? Given that there are functionally distinct nephron populations, the behavior of these different nephron types is of interest. Are changes of whole kidney GFR, for example, the result of roughly parallel increases or decreases in superficial and deep SNGFR, or is the initially unequal functional load redistributed among different nephron types when conditions are altered? Goodyer and Jaeger [53] suggested that such changes in distribution could account for changes in sodium excretion when total GFR was not changed. The effects of changes in sodium intake, volume expansion, hemorrhage, aortic constriction, and administration of ADH on the distribution of superficial and juxtamedullary blood flow and filtration rate have been examined. To limit this discussion we will consider only those studies in which filtration rate was measured. In general the more recent studies seem to find less evidence for redistribution than was seen earlier, but this is not strictly true and disagreement still persists. A recent stimulus to much of the study of possible redistribution of flow and filtration rate was the finding of Horster and Thurau [60] that a chronic high sodium intake resulted in a 50 % increase of superficial SNGFR and a nearly four-fold decrease in juxtamedullary SNGFR. There has been no subsequent report of a similar study of the effect of chronic high salt intake. However, the changes in SNGFR caused by an acute infusion of NaC1 solution have been investigated in several laboratories and disagreements persist as to whether a redistribution of GFR occurs. Several investigators have found in rats that a larger fraction of total GFR is diverted to superficial nephrons following extracellular volume expansion. These observations have been made either by comparing superficial SNGFR determined by micropuncture methods with whole kidney inulin clearance [17, 64, 65], by comparing micropuncture measurements of both superficial and deep SNGFR [2], and by comparing ferrocyanide microdissection measurements of superficial and juxtamedullary SNGFR [58, 63, 661. In contrast, in recent micropuncture and microdissection experiments Brenner et al [28] and Carriere, Boulet, and Brunett [67] have found no indication of redistribution following saline loading in rats. In dogs, the weight of evidence seems to be against a significant redistribution of filtration. Auld, Alexander and Levinsky found a disproportionate increase in superficial SNGFR only in repunctured nephrons [41], not in freshly punctured tubules [40]. Mandin et al [50] and Schneider et al [43] have also reported that when GFR increases in dogs after saline loading, superficial SNGFR increases to about the same extent. Finally, acute volume loading experiments with rats have given no indication that SNGFR of juxtamedullary nephrons is reduced as a part of the natriuretic response [2, 58]. At this time, therefore, a role for redistribution of SNGFR within the kidney during natriuretic states is not established. Natriuresis can occur without redistribution; however, the disproportionate increases in superficial SNGFR seen in some rat experiments seem to be real changes. One might speculate that genetic differences may be responsible for some of the differences seen in different rat experiments. Disagreement also continues over whether redistribution of SNGFR participates in the renal conservation of sodium, for example, when extracellular volume or arterial blood pressure is reduced. Brenner and Berliner [17] reduced extracellular volume by intraperitoneal instillation of polyethylene glycol. In their study a 15 % decrease in kidney GFR was associated with a 21 % decrease in superficial SNGFR suggesting at most a modest redistribution of filtration to deeper nephrons. Other studies of the effect of extracellular fluid volume reduction have employed hemorrhage and have yielded conflicting results. Gertz et al [49] found a 64 % reduction in superficial SNGFR associated with a 37 % decline in kidney GFR. Coehlo et al [66], however, found no evidence of redistribution in a study using the Hanssen technique; the ratio of superficial/juxtamedullary SNGFR was not reduced after hemorrhage. There has also been disagreement about the effect of lowering blood pressure on the distribution of SNGFR. Using phentolamine to lower blood pressure in dogs, Liebau, Levine, and Thurau [36] found that kidney GFR was reduced 56 % while the superficial SNGFR was reduced 76 %. Also Brenner et al [5] observed a disproportionate decrease in superficial SNGFR in rats when perfusion pressure was lowered by aortic constriction. Other studies of the effect of aortic constriction, however, have failed to demonstrate redistribution of SNGFR toward the

5 Single nephron GFR 205 inner cortex. Landwehr et al [11], Arrizurieta et al [19], and Rodicio et al [20] all reported slightly larger reductions of kidney GFR than of superficial SNGFR. In a direct comparison of superficial and juxtamedullary SNGFR's using the ferrocyanide microdissection method, Bonvalet, Bencsáth and de Rouffignac [68] found no evidence of redistribution. The weight of evidence seems to indicate that when kidney GFR is reduced a proportionate reduction in SNGFR occurs throughout the cortex. Finally, the available answers to a possibly related question, i.e., the effect of ADH on the distribution of filtration, also conflict. Davis and Schnermann [59], using a ferrocyanide method found that ADH increased the SNGFR of juxtamedullary nephrons and suggested that this effect could assist in the concentrating process by increasing the sodium load to the loops of Henle of deep nephrons. Jamison et al [62, 69], however, using micropuncture methods found no change in either superficial or juxtamedullary SNGFR after injecting ADH. Effective filtration pressure. New information has also emerged regarding the forces responsible for filtrate formation and the mechanisms that may be involved in regulating filtration. For the past six years the hydrostatic and oncotic pressures across the glomerular capillary wall have been estimated only indirectly and with an uncomfortable range of variation. Recently, however, two groups have managed to measure glomerular capillary pressure (PG) by direct puncture of surface glomeruli in rats. Brenner, Troy and Daugharty [30] and Blantz, Israelit, Rector and Seldin [70], have reported values for G of 44 and 48 mm Hg. These direct measurements, and the values for PG calculated by these authors from stopflow measurements [30, 33], confirm previous stopflow measurements of some investigators [71 74] but they are substantially lower than the calculated values of about 80 mm Hg obtained by others [14, 75 77]. In these earlier considerations of glomerular filtration pressures it was customary to assume a value for the mean oncotic pressure opposing filtration (xg) of 25 mm Hg [75]. That the assumption was justified seems clear since Brenner et al [30] have calculated a value of ltg kg(7a+te)!2] of 26 mm Hg from measurements of protein concentration in arterial and efferent capillary blood. A similar value for lrg was calculated by Andreucci et al [33] from measurements of arterial protein concentration and kidney filtration fraction, If a G of 45 to 50 mm Hg can be taken as established fact, the important new result that follows is that the effective filtration pressure is approximately 10 mm Hg [30, 33] instead of 30 to 40 mm Hg as calculated by Gertz et a! [14]. It follows from this that glomerular capillary permeability must be higher than previously calculated [30], and variations in tubule pressure [33, 51] and arteriolar resistance will have proportionately larger effects on SNGFR than previously assumed. Regulation of SNGFR. Although changes in arterial blood pressure can change filtration rate, GFR can also vary while renal perfusion pressure remains unchanged. Variations of GFR independent of arterial pressure have been observed in a variety of circumstances such as extracellular fluid volume expansion [28, 33], after administration of diuretics [15, 381, or following protein feeding [78] and suggest that intrarenal regulatory mechanisms accomplish this control. The mechanisms controlling GFR have defied many investigative efforts and they remain largely unexplained. One type of control mechanism might reside within the vasculature of the glomerular pole itself. Although there is little evidence to support such a speculation, changes in extracellular volume might be sensed locally as changes in interstitial pressure which effect myogenic changes in vascular resistance. Speculation about another type of control mechanism has arisen from the long recognized close anatomical association of the macula densa region of the distal tubule with the vascular structures of the glomerulus of the same nephron [79, 80]. This anatomical arrangement provides an avenue along which information from the distal nephron might be transmitted to the glomerular vessels. The feedback hypothesis for control of GFR would thus predict that variations in flow rate through the ioop of Henle might cause changes in the filtration rate of the same nephron. Evidence that some function of loop flow rate does affect SNGFR was obtained by Schnermann et al [81] although in a similar study Morgan [82] observed no changes in SNGFR that were suggestive of feedback control. One difference in the methods employed in these two studies may be of importance in the light of newer information that sampling technique can affect SNGFR when collections are made in early proximal tubules. Schnermann et al [81] monitored luminal pressure during their collections whereas Morgan's experiments [82] were not designed in this way. The feedback control hypothesis would also predict that sampling tubule fluid at late proximal or distal sites should yield different values for SNGFR. Blocking the

6 206 Wright / Giebisch loop of Henle during proximal collections might activate a mechanism that would act to increase filtration rate. This postulated effect of loop blockage implies that most micropuncture measurements of SNGFR are probably higher than the "true" value for normal conditions. Landwehr et al [11], and Jamison and Lacy [21 found with random collections that distally measured SNGFR tended to be lower than proximally measured values but the mean values were not different statistically. The only study in which distal and proximal samples were collected from the same nephron has been reported by Schnermann et al [31]. Distally measured SNGFR was 25.2 ni/mm, a value that was significantly less than that of 34.5 ni/mm obtained from proximal collections.2 A third prediction of the feedback hypothesis might be that variations in loop flow should change PG. Hierholzer has reported experiments [83] in which increased flow through Henle's loop was associated with decreased proximal stop-flow pressure in 16 of 25 tubules studied. Reductions in loop flow rate did not change stop flow pressure. In agreement with this last observation, Blantz et al [70] observed that blocking the proximal tubule with oil changed neither the stop flow pressure in that tubule nor the directly measured P0. Further investigation will be needed to determine whether loop flow affects PG and whether it does so only in one direction or only in some nephrons. It may also be that filtration can be varied without changes in P. Daugharty et al [84] found that, even though superficial SNGFR was doubled after volume expansion, there was no change in the directly measured P0. They speculated that the change in GFR might have been due to an increase in the surface area for filtration pressure. Earlier Gertz et al [49] had found disproportionate changes in SNGFR and calculated effective filtration. They concluded that glomerular capillary permeability had changed acutely when GFR was increased by saline injection or decreased by hemorrhage. Future investigations of the regulation of filtration rate will have to consider the effective filtration pressure, the point along the glomerular capillary at which filtration equilibrium is reached [30, 33, 84], the number of functioning capillary loops [84], the glomerular plasma 2 Others do not agree that distally measured SNGFR is lower because of a feedback control mechanism. Bartoli and Earley have concluded in abstract form that the lower values for distal measurements are due to systematic error (Am. Soc. Nephrol. 5: 6, 1971; Clin. Res. 20: 167, 1972). flow rate [51] and the permeability of the capillary wall [49]. Methods currently available for measuring SNGFR will continue to be useful in studies of single nephron function. Investigators must be aware that filtration rate may vary with age, kidney size, number of kidneys present, hydration, extracellular fluid volume, and genetic make-up. Also, the measurement of SNGFR may not accurately reflect the true rate of filtration if effective filtration pressure is disturbed by changes in intratubular pressure or by interference with an intrarenal control mechanism. Some authors suspect that collecting from the proximal tubule may interfere with the regulation of SNGFR. Future work may confirm that distal collections give the best estimate of the true filtration rate. At this time, however, for experimental measurement of the filtered load presented to a single nephron, we think that an acceptable technical approach for quantitative collection of tubule fluid is as follows: collect from a segment at the end of a proximal tubule, block distal to the collection site with an oil at least as viscous as mineral oil, make the oil column several diameters in length, use gentle aspiration if necessary to maintain a collection rate and prevent increases in tubule pressure, collect over a period at least 20 times longer than the initial period of blocked flow, avoid the possibility of aspirating surface fluid along with tubule fluid when collecting from previously punctured segments, and be sure that no adjacent tubule segment has been punctured. The recent work of a large number of investigators has clarified the importance of these precautions. The first reported study of mammalian kidney tubules [8] used most of them. Fred S. Wright and Gerhard Giebisch Department of Physiology Yale University School of Medicine New Haven, Connecticut 06510, U.S.A. References 1. Horster, M., and Valtin, H.: Postnatal development of renal function: Micropuncture and clearance studies in the dog. J. Clin. Invest. 50: , Jamison, R. L., and Lacy, F. B.: Effect of saline infusion on superficial and juxtamedullary nephrons in the rat. Am. J. Physiol. 221: , Hayslett, J. P., Kashgarian, M., and Epstein, F. H.: Functional correlates of compensatory renal hypertrophy. J. Clin. Invest. 47: , 1968.

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8 208 Wright / Giebisch 30. Brenner, B. M., Troy, J. L., and Daugharty, T. M.: The dynamics of glomerular ultrafiltration in the rat. J. Clin. Invest. 50: , Schnermann, J., Davis, J.M., Wunderlich, P., Levine, D.Z., and Horster, M.: Technical problems in the micropuncture determination of nephron filtration rate and their functional implications. Pflugers Arch. ges. Physiol. 329: , Davidman, M., Lalone, R.C., Alexander, E.A., and Levinsky, N. G.: Some micropuncture techniques in the rat. Am. J. Physiol. 221: , Andreucci, V.E., Herrera-Acosta, J., Rector, F.C., Jr., and Seldin, D. W.: Effective glomerular filtration pressure and single nephron filtration rate during hydropenia elevated ureteral pressure and acute volume expansion with isotonic saline. J. Clin. Invest. 50: , Bartoli, E., and Earley, L. F.: Effect of tubular occlusion on nephron filtration rate as measured by collection from proximal and distal tubules. Am. Soc. Nephrol. 5: 6, 1971 (abstract). 35. Andreucci, ye., Herrera-Acosta, J., Rector, F.C., Jr., and Seldin, D.W.: Measurement of single nephron glomerular filtration rate by micropuncture: Analysis of error. Am. J. Physiol. 221: 155i 1559, Liebau, G., Levine, D.Z., and Thurau, K.: Micropuncture studies on the dog kidney. I. The response of the proximal tubule to changes in systemic blood pressure within and below the autoregulatory range. Pflugers Arch. ges. Physiol. 304: 57 68, Wright, F.S., Knox, F.G., Howards, S.S., and Berliner, R.W.: Reduced sodium reabsorption by the proximal tubule of Doca-escaped dogs. Am. J. Physiol. 216: , Knox, F.G., Wright, F.S., Howards, S.S., and Berliner, R.W.: Effect of furosemide on sodium reabsorption by proximal tubule of the dog. Am J. Physiol. 217: , Wright, F.S., Howards, S.S., Knox, F.G., and Berliner, R.W.: Measurement of sodium reabsorption by proximal tubule of the dog. Am. J. Physiol. 2/7: , Auld, R. B., Alexander, E., and Levinsky, N. G.: Nephron filtration and proximal tubular function in normal and chronic TVC dogs during saline infusion. Clin. Res. /8: 745, 1970 (abstract). 41. Auld, RB., Alexander, E.A., and Levinsky, N.G.: Nephron filtration and proximal reabsorption during saline infusion, arterial clamping and hemorrhage in the dog. J. Clin. Invest. 49: 5, 1970 (abstract). 42. Knox, F. G., Schneider, E.G., Dresser, T. P., and Lynch, R. E.: Natriuretic effect of increased proximal delivery in dogs with salt retention. Am. J. Physiol. 2/9: , Schneider, E.G., Lynch, R. E., Dresser, T. P., and Knox, F.G.: Effect of saline infusion on superficial nephron filtration rate in the dog. Physiologist. 13: 303, 1970 (abstract). 44. Stein, J. M,, Reineck, J. M., Osgood, R. W., and Ferris, T.F.: Effect of acetylcholine on proximal tubular sodium reabsorption in the dog. Am. J. Physiol. 220: , Schneider, E.G., Dresser, T. P., Lynch, R. E., and Knox, F. G.: Sodium reabsorption by proximal tubule of dogs with experimental heart failure. Am. J. Physiol. 220: , Auld, R.B., Alexander, E.A., and Levinsky, N.G.: Proximal tubular function in dogs with thoracic caval constriction. J. Gun. Invest. 50: , Schnermann, J.: Methodological aspects of filtrate determination by the micropuncture technique. Yale J. Biol. Med. (In press). 48. Andreucci, V. E., and Rector, F. C., Jr.: Some artifacts in measuring single nephron glomerular filtration rate. Yale J. Biol. Med. (In press). 49. Gertz, K. H., Brandis, M., Braun-Schubert, G., and Boylan, J. W.: The effect of saline infusion and hemorrhage on glomerular filtration pressure and single nephron filtration rate. Pflflgers Arch. ges. Physiol. 310: , Mandin, H., Israelit, A. H., Rector, F. C., Jr., and Seldin, D.W.: Effect of saline infusions on intrarenal distribution of glomerular filtrate and proximal reabsorption in the dog. J. Clin. Invest. 50: , Rector, F.C., Jr., Andreucci, V.E., Herrera-Acosta, J., and Seldin, D.W.: Potential sources of error in measuring single nephron glomerular filtration rate. Yale J. Biol. Med. (In press). 52. Seely, J. F., and Boulpaep, E. L.: Renal function studies on the isobaric autoperfused dog kidney. Am. J. Physiol. 22/: , Goodyer, A.V.N., and Jaeger, C.A.: Renal response to non-shocking hemorrhage. Am. J. Physiol. /80: 69 74, Bradley, S.E., and Wheeler, HO.: On the diversities of structure, perfusion and function of the nephron population. Am. J. Med. 24: 692, Hanssen, 0. E.: The relationship between glomerular filtration and length of the proximal convoluted tubules in mice. Acta Pathol. Microbiol. Scand. 53: , Baines, A. D., and Rouffignac, C. de: Functional heterogeneity of nephrons. II. Filtration rates, intralurninal flow velocities and fractional water reabsorption. PflUgers Arch. ges. Physiol. 308: , Rouffignac, C. de, Deiss S., and Bonvalet, J. P.: Determination du taux individuel de filtration glomérulaire

9 Single nephron GFR 209 des néphrons accessibles et inaccessibles a Ia microponction. Pflügers Arch. ges. Physiol. 315: , Rouffignac, C. de, and Bonvalet, J.P.: Etude chez le rat des variations du debit individuel de filtration glomérulaire des néphrons superficiels et profonds en fonction de l'apport sodé. Pflugers Arch. ges. Physiol. 317: , Davis, J.M., and Schnermann, J.: Juxtamedullary filtration rate and urinary concentrating ability. mt. Cong. of Physiol. 9: 132, 1971 (abstract). 60. Horster, M., and Thurau, K.: Micropuncture studies on the filtration rate of single superficial and juxtamedullary glomeruli in the rat kidney. Pflugers Arch. ges. Physiol. 301: , Jamison, R.L.: Micropuncture study of superficial and juxtamedullary nephrons in the rat. Am. J. Physiol. 218: 46 55, Jamison, R.L.: Evidence for functional intrarenal heterogeneity obtained by the micropuncture technique. Yale J. Biol. Med. (In press). 63. Rouffignac, C. de, and Bonvalet, J. P.: Use of sodium ferrocyanide as glomerular indicator to study the functional heterogeneity of nephrons. Yale J. Biol. Med. (In press). 64. Stein, J.H., Barton, L.J., Mandin, H., Lackner, L.M., Rector, F.C., Jr., and Seldin, D.W.: Effect of extracellular volume expansion on proximal tubular sodium reabsorption and distribution of renal blood flow and glomerular filtrate in the dog. Clin. Res. /7: 449, 1969 (abstract). 65. Herrera-Acosta,J., Rector, F.C., Jr., and Seldin, D.W.: The influence of extracellular volume on GFR and proximal reabsorption in the rat. Am. Soc. Nephrol. 3: 27, 1969 (abstract). 66. Coehlo, J. B., Stella, S. R., Kuang-Chung Hu Chien, and Bradley, S. E.: Glomerular filtration rate in superficial and juxtamedullary nephrons during salt loading and hemorrhagic hypotension. Clin. Res. /8: 496, 1970 (abstract). 67. Carriere, S., Boulet, P., and Brunett, M.G.: Isotonic saline loading and intrarenal distribution of single nephrori glomerular filtration rate in dogs. Am. Soc. Nephrol. 5: 13, 1971 (abstract). 68. Bonvalet, J. P., Bencsáth, P., and Rouffignac, C. de: Glomerular filtration rate of superficial and deep nephrons during aortic constriction. Am..1. Physiol. (In press). 69. Jamison, R., Buerkert, J., and Lacy, F.: Micropuncture of Henle's loop in rats with hereditary diabetes insipidus. Am. Soc. Nephrol. 5: 35, 1971 (abstract). 70. Blantz, R.C., Israelit, A.M., Rector, F.C., Jr., and Seldin, D.W.: Influence of distal sodium delivery on glomerular hydrostatic pressure in the rat. Am. Soc. Nephrol. 5: Il, 1971 (abstract). 71. Selkurt, E.E., Deetjen, P., and Brechtelsbauer, H.: Tubular pressure gradients and filtration dynamics during urinary stop flow in the rat. Pflugers Arch. ges. Physiol. 286: 19 35, Henry, L.N., Lane, C.E., and Kashgarian, M.: Micropuncture studies of the pathophysiology of acute renal failure in the rat. Lab. Invest. 19: , Jaenike, J. R.: Micropuncture study of methhemoglobin-induced acute renal failure in the rat. J. Lab. Clin. Med. 73: , Haylett, J. P., Domoto, D. T., Kashgarian, M., and Epstein, F.: Role of physical factors in the natriuresis induced by acetylcholine. Am. J. Physiol. 2/8: , Gertz, K.H., Mangos, J.A., Braun, G., and Pagel, H. D.: Pressure in the glomerular capillaries of the rat kidney and its relation to arterial blood pressure. Pflugers Arch. ges. Physiol. 288: , Koch, K. M., Dume, T., Krause, H. M., and Ochwadt, B.: lntratubulärer Druck, glomerularer Capillardruck und Glomerulumfiltrat während Mannit-Diurese. Pflügers Arch. ges. Physiol. 295: 72 79, Krause,H.M., Dumme,T., Koch, K.M., and Ochwadt, B.: lntratubularer Druck glomerularer Capillardruck and Glomerulum filtrat nach Furosemid und Hydrochiorothiazid. Pflugers Arch. ges. Physiol. 295: 80 89, Lindheimer, M. D., Lalone, R. C., and Levinsky, N. G.: Evidence that an acute increase in glomerular filtration has little effect on sodium excretion in the dog unless extracellular volume is expanded. J. Clin. Invest. 46: , Guyton, A.C., Langston, J.B., and Navar, G.: Theory for renal autoregulation by feedback of the juxtaglomerular apparatus. Circ. Res. 15, Suppl. I, , Thurau, K.: Renal hemodynamics. Am. J. Med. 36: , Schnermann, J., Wright, F.S., Davis, J. M., Stackelberg, W. v., and Grill, G.: Regulation of superficial nephron filtration rate by tubuloglomerular feedback. Pflugers Arch. ges. Physiol. 3/8: 147 l75, I Morgan, T.: A microperfusion study of influence of macula densa on glomerular filtration rate. Am. J. Physiol. 220: , Hierholzer, K., Butz, M., Müller-Suur, R., and Lichtenstein, I.: Pressure measurements in proximal surface tubules of the rat: single nephron filtration rate and tubuloglornerular feedback. Yale J. Biol. Med. (In press). 84. Daugharty, T., Troy, J., and Brenner, B. M.: Glomerular dynamics and the concept of filtration pressure equilibrium. Am. Soc. Nephrol. 5: 17, 1971 (abstract).