Mechanism of increased tubular Na-K-ATPase during streptozotocin-induced diabetes

Since the mechanisms responsible for stimulation of kidney Na-K-ATPase during streptozotocin-induced diabetes are unknown, we studied the possible role(s) of kidney hyperfiltration and hypertrophy and of hyperaldosteronism on Na-K-ATPase induction. For this purpose, we studied the relationship between Na-K-ATPase activity in individual nephron segments and alterations of glomerular filtration rate during the early phase of diabetes. Within 2 days after streptozotocin administration, Na-K-ATPase activity markedly increased in the proximal convoluted tubule, medullary thick ascending limb and cortical and outer medullary collecting tubule, but not in the proximal straight tubule, cortical thick ascending limb and distal convoluted tubule. Streptozotocin administration also markedly enhanced the glomerular filtration rate but only after 4 days following initiation of treatment. Changes in Na-K-ATPase were specific since the activity of adenylate cyclase, another marker of basolateral membranes, was not altered. Finally, when animals were adrenalectomized prior to streptozotocin treatment, Na-K-ATPase stimulation was curtailed in the collecting tubule but not in more proximal segments. These results suggest that diabetes alters Na-K-ATPase activity in specific nephron segments independent of alterations of glomerular filtration rate and of kidney hypertrophy, and that the stimulation of collecting tubule Na-K-ATPase is secondary to hyperaldosteronism.     

Glucagon receptors along the nephron: [125I]glucagon binding in rat tubules

Binding of [¹²⁵I]glucagon was measured in microdissected pieces of tubules from the rat nephron. Specific glucagon binding sites were found only in nephron segments containing a glucagon-sensititive adenylate cyclase activity. At 7.5 nM labelled hormone, higher levels of specific binding (16–27×10^–18 mol mm^–1) were found in the thick ascending limb of the Henle's loop and in the distal convoluted tubule and lower binding levels (2–5×10^–18 mol mm^–) in the collecting tubule whereas specific binding could not be detected in the proximal tubule and in the thin segments of the Henle's loop. In the medullary thick ascending limb, Scatchard analysis of specific [¹²⁵I]glucagon binding indicated an apparent equilibrium dissociation constant of 2.4 nM. The stereospecificity of binding sites in medullary thick ascending limbs and medullary collecting tubules, was assessed by competition experiments using unlabelled glucagon, enteroglucagon and unrelated hormones (vasopressin, calcitonin, parathyroid hormone and insulin); in both segments, glucagon was more active than enteroglucagon in displacing labelled glucagon from its tubular binding sites, whereas all other hormones tested were inactive. These results indicate that tubule binding sites might be the physiological receptors for glucagon involved in adenylate cyclase activation.Abbreviations PCT proximal convoluted tubule - TDL thin descending limb - TAL thin ascending limb - MAL medullary thick ascending limb - CAL cortical ascending limb - DCT distal convoluted tubule - CCT cortical collecting tubule - MCT medullary collecting tubule     

Segmental staining of the murine nephron by monoclonal antibodies directed against the GP-2 subunit of laminin

Monoclonal antibodies to GP-2, the 220,000-dalton subunit of laminin, were used in an immunohistologic study to investigate structural variations in basement membranes. Mouse kidney was used because of the wide range of basement membranes represented. Two rat/mouse monoclonal antibodies, designated LAM-I and LAM-II were compared with rabbit polyclonal anti-GP-2 in a light and electron microscopic study that identified nephron segments by morphology, by topography, and by the use of markers specific for individual segments. LAM-I staining is demonstrable on all tubular and glomerular basement membranes but not on those of blood vessels or smooth muscle, differing in this respect from anti-GP-2. LAM-II staining is confined to the basement membranes of the convoluted portion of proximal tubule segments (S1 and S2), not the straight (S3) portion; to the thin limb and the thick ascending limb of Henle's loop, but not distal convoluted tubules or collecting ducts. The heterogeneity of GP-2 localization may be due to differing conformations of the molecule at these sites.     

Sites of thyroid hormone action on Na−K-ATPase along the rabbit nephron

Although Ismail-Beigi and Edelman demonstrated in 1971 that thyroid hormones control the activity of Na–K-ATPase in the mammalian kidney, the actual site of this regulation inside the organ was not located. We therefore decided to study the relationship between thyroid hormones and Na–K-ATPase activity in individual nephron segments obtained by microdissection of collagenase-treated rabbit kidneys. For this purpose, the changes in the activity and number of catalytic sites of Na–K-ATPase in response to thyroidectomy or triiodothyronine administration were examined. Eight to 12 days after thyroidectomy, Na–K-ATPase activity had dropped by 40 to 80% in the convoluted and straight portions of the proximal tubules, and in the cortical and outer medullary collecting tubules, but not in the thick ascending limbs of Henle's loops or distal convoluted tubules. The apparent number of catalytic sites for Na–K-ATPase, as measured by specific binding of³H-ouabain, decreased in parallel with Na–K-ATPase activity, and therefore this enzyme's specific activity was not altered. Fourty eight hours after injection of thyroidectomized animals with a single dose of either 100 or 500 g/kg triiodothyronine, Na–K-ATPase activity in target segments was restored to the level measured in control animals. These effects of thyroid hormone were specific for Na–K-ATPase, since the activity of adenylate cyclase, another marker of the basolateral membrane, was not altered by thyroidectomy. The results obtained indicate that triiodothyronine controls Na–K-ATPase activity in specific nephron segments, by altering the number of this enzyme's catalytic sites.Abbreviations PCT proximal convoluted tubule - PR pars recta - MAL medullary thick ascending limb of Henle's loop - CAL cortical thick ascending limb - DCT_b initial bright portion of the distal convoluted tubule - DCT_g granular portion of the distal convoluted tubule - CCT cortical collecting tubule - MCT outer medullary collecting tubule - TX thyroidectomized - T₃ triiodothyronine - AVP arginine vasopressin - PTH parathyroid hormone - ISO isoproterenol     

Corticosterone binding sites along the rat nephron

Glucocorticoids influence numerous kidney functions but the precise location of glucocorticoid receptors in the nephron is not known. To identify the renal binding sites of corticosterone, the natural glucocorticoid in the rat, we measured the binding of [3H]corticosterone to discrete nephron segments microdissected from adrenalectomized rats. Highest specific binding capacity at 25 degrees C (expressed as fmol X cm-1 +/- SE) was found in the cortical collecting tubule (9.69 +/- 0.77) followed in decreasing order by the distal convoluted tubule (2.70 +/- 0.49), medullary collecting tubule (2.58 +/- 0.64), proximal convoluted tubule (1.09 +/- 0.10), and pars recta (0.57 +/- 0.08). Binding was lowest in the thick ascending limb of Henle's loop, with comparable values in the medullary (0.27 +/- 0.05) and cortical (0.26 +/- 0.05) portions of this segment. The apparent maximal binding capacity of the cortical collecting tubule for corticosterone exceeded by nearly two orders of magnitude that of aldosterone previously measured by us in this structure, which is in agreement with the observations of other investigators in kidney cytosol. Specific binding of corticosterone can be demonstrated along the entire rat nephron, but binding sites are concentrated in the cortical collecting tubule. This segment appears to be the main target site for corticosterone as it is for aldosterone.     

Primary culture of mammalian nephron epithelia

Nephron segments were dissected from fetal or from early postnatal rabbit kidneys (n=86) and explanted individually into primary tissue culture. Outgrowth of epithelioid cells and proliferation in monolayers from the collecting tubule and the thick ascending limb of Henle's loop occurred regularly if a special substratum and matrix were used. Media were either supplemented with non-proteins (defined media) or with embryo extract and fetal sera (undefined media). Outgrowth and monolayer spreading were recorded by Differential Interference Contrast Microscopy. Monolayer cells resemble those seen with the same recording technique during tubule in vitro of convoluted tubule segments from embryonic Nephron Anlagen and outgrowth of cells from proximal straight tubules, in contrast to the distal nephron segments. The specific functional and morphological properties of cells derived from primary culture of nephron segments are under study.     

Effect of chronic metabolic acidosis on ammonia production fromL-glutamine in microdissected rat nephron segments

To evaluate the role of each nephron segment in renal ammoniagenesis, distribution of renal ammoniagenic activity along the nephron in control and acidotic rats was examined. We used our original aerobic incubation system and ammonia produced from glutamine in 7 defined segments of microdissected nephron was measured using the enzymatic cycling method.When ammonia production in the control was compared in each nephron segment, the highest specific activity of ammoniagenesis per mm tubular length and that per g protein were observed in the proximal straight tubule (PST) and the thick ascending limb of Henle's loop, respectively. Chronic metabolic acidosis increased ammonia production per mm tubular length markedly in the proximal convoluted tubule (PCT) (+171%), moderately in the medullary collecting tubule (+123%) and PST (+77%), and slightly in the distal convoluted tubule (+52%), revealing that the highest activity of ammoniagenesis was located in PCT and PST in acidosis.These data indicate that proximal tubules have major roles in renal ammoniagenesis both in the control and in acidosis. From the early observation of glutaminase I isoenzyme distribution along the nephron, our data suggest that not only phosphate-dependent glutaminase but also phosphate-independent glutaminase may have important roles in renal ammoniagenesis.This study was presented in part at 26th Congress of Japanese Society of Nephrology, Kyoto, 1983     

Na-K-activated ATPase: activity maturation in rabbit nephron segments dissected in vitro.

Single, defined nephron segments were dissected in vitro from fresh slices of neonatal and mature rabbit kidney. Na-K-ATPase was quantified for six different tubule segments with an ultramicromethod. The enzyme distribution pattern in the neonatal nephron was similar to that in the mature nephron. Activity in distal segments was 2-5 times higher than proximal tubule activity referred to tissue dry weight, and 2 times higher referred to tubule length. Neonatal enzyme activity was lower than mature in all segments. Some segments carried only 23% of mature activity. Enzyme activity in the proximal convoluted tubule was constant during maturation when referred to the basal and lateral membrane area measured in the same developmental stages. In vitro activities of these tubules were similar to the enzyme activity measured previously in freeze-dried slices. The Vmax during development of Na-K-ATPase was higher by a factor of 5 in the mature tubule, whereas the Km was identical in the neonatal and mature tubule. The ouabain-insensitive ATPase did not show a maturational activity change.     

Mineralocorticoid effects on Na-K-ATPase in individual nephron segments

A fluorometric micromethod for the assay of Na-K-ATPase was used to determine Na-K-ATPase activity in 11 segments of the rabbit nephron. The Na-K-ATPase activity was found to be highest (greater than 100 pmol.mm1-.min-1) in proximal convoluted tubules (S1), medullary thick ascending limbs, distal convoluted tubules, and connecting tubules. There was a good correlation (r = 0.94) between Na-K-ATPase activity in different segments and net Na transport previously measured by in vitro tubule perfusion. Treatment of rabbits with deoxycorticosterone (DOCA) for 8-11 days produced increases in Na-K-ATPase activity of 100 and 500% in connecting tubules and cortical collecting ducts, respectively, without significant change in other segments. Maintenance on a low sodium diet for 12-18 days was associated with a 200% increase in Na-K-ATPase activity in cortical collecting ducts only. We conclude that the Na-K-ATPase activity is stimulated by mineralocorticoids in the cortical collecting duct in association with the previously observed changes in sodium transport.     

Short-term effects of aldosterone and dexamethasone on Na−K-ATPase along the rabbit nephron

Both glucocorticoids and mineralcorticoids stimulate the renal Na-K-ATPase. However, the exact site of their respective action is not precisely determined and it is still unknown whether these effects are cumulative or not. We studied the effects of dexamethasone and aldosterone on Na-K-ATPase activity in microdissected nephron segments from adrenalectomized rabbits. In proximal convoluted tubule (PCT) the enzyme activity was altered neither by adrenalectomy nor by any steroid replacement. In the medullary thick ascending limb of the loop of Henle (MAL) and the distal convoluted tubule (DCT), Na-K-ATPase activity decreased by 40% after adrenalectomy, and was restored to control level three hours after administration of dexamethasone (100 micrograms/kg) but not by aldosterone (up to 10 micrograms/kg). In the cortical (CCT) and medullary (MCT) collecting tubule the enzyme activity decreased by 75% after adrenalectomy but in contrast with the MAL and the DCT, these two segments were sensitive to both dexamethasone (100 micrograms/kg) and aldosterone (10 micrograms/kg) and recovered their activities within 3 h after the hormone injection. These effects were not additive. Spironolactone (100 micrograms/kg) abolished the action of each of the two hormones on the CCT and MCT. In contrast, spironolactone did not curtail the effect of dexamethasone on MAL and DCT. These results indicate that whereas glucocorticoid action is localized in MAL, DCT, CCT and MCT, the mineralocorticoid effect is restricted to the CCT and MCT exclusively. They also suggest that, in the CCT and MCT, the two types of hormones share the same receptors.     

Effect of water intake on Na−K-ATPase in nephron segments of the desert rodent,Jaculus orientalis

Na–K-ATPase activity was measured in individual pieces of nephron microdissected from collagenase-treated kidneys of jerboas,Jaculus orientalis. Na–K-ATPase activity was high in the distal convoluted tubule, intermediate in the thick ascending limb of the loop of Henle and low in the proximal and collecting tubule. When jerboas were adapted for several weeks to a hydrated diet and excreted a more diluted urine, Na–K-ATPase activity was altered in specific segments of the nephron: 1. In the proximal convoluted tubule, Na–K-ATPase activity decreased, especially in the juxtamedullary nephrons, suggesting that internephron heterogeneity was diminished; 2. In the medullary thick ascending limb, but not in the cortical portion, Na–K-ATPase activity decreased by 30%; 3. Na–K-ATPase was also diminished in the cortical collecting tubules (by 20%) but not in the medullary collecting tubule. Morphometric measurements also indicate that changes in Na–K-ATPase activity observed in the thick ascending limb are correlated to a cell atrophy, whereas in the collecting tubule, they occur independently of any visible morphological alteration. These differences in Na–K-ATPase activity are likely to be secondary to the changes in the plasma concentration of vasopressin previously described during such adaptation and to be involved in the control of water and sodium handling.     

Determination of Na-K-ATPase activity in single segments of the mammalian nephron.

A micromethod for the determination of Na-K-ATPase in discrete segments of nephrons from rabbit, rat, and mouse kidneys is described. To facilitate tubule microdissection, the kidneys were perfused with collagenase after it had been verified that collagenase had no effect on ATPase activity. Individual tubule segments were dissected under stereomicroscopic observation, exposed to a hypotonic environment followed by rapid freezing, and incubated in 1 microliter assay medium. Enzyme activity was determined by direct measurement of labeled inorganic phosphate release by the hydrolysis of [gamma-32P]ATP and was expressed as a function of tubule length. This method is technically simple enough to permit simultaneous measurement of the enzyme in large numbers of tubules and sufficiently sensitive to determine its activity in each region of the nephron. Correlation of Na-K-ATPase activity in single tubules with functional measurements obtained in the corresponding segment of the nephron with the perfused tubule or micropuncture techniques should help define the role of this enzyme in tubular ion transport.     

Changes in tubular basolateral membrane markers after chronic DOCA treatment

Chronic administration of DOCA to rabbits is known to increase the surface area of the basolateral membrane and the Na-K-ATPase activity of the cortical collecting tubule (CCT). We attempted to ascertain 1) whether Na-K-ATPase is the only basolateral membrane marker induced by DOCA, and 2) whether CCT is the only nephron segment affected by this steroid. We measured the activity of Na-K-ATPase and adenylate cyclase (AC) and the protein content of nephron segments microdissected from control and DOCA-treated rabbits. Morphogenic effects of DOCA, assessed by 30-60% increases in protein content, were specifically observed in the distal convoluted tubule, CCT, and medullary collecting tubule. When expressed as a function of tubular length, Na-K-ATPase activity rose from 80 to 200% in all these segments, whereas the increments in AC of 40-70%, observed in response to four different hormones, occurred only in some of them. When expressed as a function of protein content, Na-K-ATPase activity increased but AC activity remained unchanged. This study indicates that the morphogenic action resulting from chronic DOCA administration affects the entire rabbit distal nephron. During this action Na-K-ATPase is the preferentially induced enzyme.     

Effect of low potassium-diet on Na−K-ATPase in rat nephron segments

Na–K-ATPase activity was determined in 10 segments of the rat nephron using a fluorometric microassay method [4]. The enzyme activity showed three peaks (>200 pmol ADP min^–1 mm^–1) along the nephron of normal rats. These peaks were in the S₁ portion of the proximal tubule, the medullary thick ascending limb from the inner stripe and the distal convoluted tubule. Feeding the rats a low potassium diet for 8 weeks produced a significant decrease in Na–K-ATPase activity in the cortical collecting duct, but no significant change in this enzyme in any other segment. The low potassium diet did not produce a significant change in Mg-ATPase in any nephron segments. We conclude that Na–K-ATPase activity along the rat nephron shows a pattern that is qualitatively similar to that seen in the rabbit nephron [4]. However, quantitatively the Na–K-ATPase activity in the rat nephron is greater than in the corresponding segments of the rabbit nephron. The results are consistent with the greater rate of glomerular filtration and Na⁺ reabsorption per rat nephron. Furthermore, our results suggest that the decrease in potassium excretion during potassium deficiency is modulated, at leat in part, by the level of Na–K-ATPase activity in the cortical collecting duct.     

Lack of stimulation of kidney Na−K-ATPase by thyroid hormones in long-term thyroidectomized rabbits

The effects of long-term thyroidectomy and of subsequent triiodothyronine administration on kidney Na–K-ATPase were studied at the level of single nephron segments and were compared to the short-term effects previously reported. After 8–11 weeks, thyroidectomy resulted in a marked decrease in Na–K-ATPase activity in all the segments of the rabbit nephron, the proximal tubule, the thick ascending limb of Henle's loop, the distal convoluted tubule and the collecting tubule. Within this delay, thyroidectomy also decreased the ouabain-insensitive Mg-ATPase activity, the basal and hormone-stimulated adenylate-cyclase activity, and the volume of tubular epithelium in all the segments where these parameters were measured. Administration of 50 g/kg body weight triiodothyronine to 8–11 weeks thyroidectomized rabbits did not restore Na–K-ATPase activity in any nephron segment within 48 h. These observations are different from those reported in animals thyroidectomized only 1 week before study since, within this latter delay, (1) thyroidectomy altered specifically Na–K-ATPase activity, (2) this action was observed on the proximal and collecting tubules exclusively and, (3) triiodothyronine administration corrected Na–K-ATPase alterations after 49 h. Results of the present study indicate that in the long term, thyroidectomy has a wide spectrum of renal effects which involves the whole nephron and most cellular functions. The tubular involution induced by longterm thyroidectomy is probably responsible for the inability of kidney cells to quickly increase their Na–K-ATPase activity in response to hormonal stimulation.     

Effects of acute cisplatin administration on renal ATPase activities and magnesium excretion of rats

Male Wistar rats were killed 1, 2, or 4 days after a single intraperitoneal injection of cisplatin (5 mg/kg). Functional renal indices, enzymatic activities, and morphological variables were studied. One day after the injection, the treated group showed an increase in the magnesium and phosphate fractional urinary excretion (FE) vs the control group (FE Mg = 5.2 +/- SEM 0.5% vs 13.0 +/- 1.7%; P less than 0.01; and FE P = 4.7 +/- 0.7% vs 14.0 +/- 1.9%; P less than 0.01). Two days after cisplatin administration, a decrease in creatinine clearance of treated animals was found, to 0.33 +/- 0.03 vs 0.51 +/- 0.03 ml/min; P less than 0.05. Na-K-ATPase and ouabain-insensitive ATPase activities were studied in the proximal convoluted tubule, the medullary thick ascending limb of the Henle's loop (mTAL), and the distal convoluted tubule. Only in mTAL one day after the cisplatin injection was there a decrease in Na-K-ATPase activity in the treated group vs controls (1103 +/- 145 vs 1734 +/- 189 pmol Pi/mm.h; P less than 0.05). Morphological studies showed a decrease in mTAL diameters on day 1, and an increase in proximal convoluted tuble diameters at day 2 of treated rats vs controls, at 27.8 +/- 0.6 vs 31.4 +/- 0.7 microns; P less than 0.05, and 50.4 +/- 1.2 vs 47.4 +/- 0.2 microns; P less than 0.05 respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

PTH sensitive adenyl cyclase activity in different segments of the rabbit nephron

Summary PTH sensitive adenylate cyclase activity was measured in 9 different segments of the nephron, isolated by microdissection from collagenase-treated rabbit kidney slices.The enzyme of the following segments was stimulated by PTH, 1 U/ml: PCT. (proximal convoluted tubule); PR (pars recta); CAL (cortical portion of the thick ascending limb); DCT (distal convoluted tubule); BCT (first, branched portion of the collecting tubule); the segments which did not respond to PTH were: TDL (thin descending limb); MAL (medullary portion of the thick ascending limb); CCT (cortical portion of the collecting tubule distally adjacent to BCT); MCT (collecting tubule from the outer medulla).PTH sensitive adenylate cyclase per mm tubule in PR was half that measured in PCT.Half maximal stimulation corresponded to 50–100 mU/ml PTH (1–2×10^–8M) in both PCT and PR, and to about 350 mU/ml in CAL. PTH (1 U/ml) stimulation factors ranged from 5 to 60 depending on the structures.It is concluded that in addition to PCT and PR, CAL and BCT might be target structures involved in the physiological actions of PTH on the kidney.This work was supported by a grant from the C.N.R.S. to the LRA n^o 219.     

Distribution of enzymes of adenylate and guanylate nucleotide metabolism in rat nephron

In a previous study of discrete segments of rat nephron, we reported the levels of high-energy adenylate and guanylate phosphates to be highest in the distal straight and convoluted tubules. Those findings stimulated the study of the distribution of seven enzymes involved in the following metabolic pathways of these nucleotides [Formula: see text]. The patterns of distribution of enzymes in each pathway differed greatly. The phosphodiesterases, 1 and 2, were high in glomeruli and distal tubular segments and low in proximal segments. Adenylate kinase, 3, in contrast, was high in glomeruli, proximal segments, thick ascending limb of Henle, and distal convoluted tubules. Guanylate kinase levels, 4, however, were similar in all segments. The pattern of nucleosidediphosphate kinase, 5, was high in proximal convoluted, thick ascending limb, and distal convoluted tubules. The pattern of the degradative enzyme, 5'-nucleotidase, 6, whose levels were highest in proximal segments, was opposite from that of AMP deaminase, 7, highest in the distal nephrons. These dissimilar patterns underscore the extent of nephron heterogeneity.     

Effects of human calcitonin on water and electrolyte movements in rat juxtamedullary nephrons: inhibition of medullary K recycling

The effects of synthetic human calcitonin (HCT) on water and electrolyte deliveries to the thin limbs of Henle's loop of juxtamedullary nephrons were investigated by micropuncture in the rat. To avoid undesirable interference with exogenous calcitonin, experiments were performed in hormone-deprived rats with reduced circulating calcitonin, antidiuretic hormone, parathyroid hormone and glucagon, all four of which stimulate the adenylate-cyclase activity in the thick ascending limb and the distal tubule. Administration of HCT (1.0 mU/min·100 g body wt) to such rats significantly reduced the urinary fractional excretion rate of water, Mg, Ca and K. At the tip of the long-looped nephrons, the fractional delivery of water diminished in the presence of HCT, although the glomerular filtration rate of these nephrons was unaltered. Simultaneously, the loop fluid osmolality rose significantly. HCT, however, did not alter the fraction of total filtered solutes remaining in the thin limbs, nor the NaCl fractional delivery. As previously observed in this laboratory with dDAVP, the reduced fractional delivery of water at the hairpin turn was accompanied by a decrease in Mg and Ca deliveries in rats given HCT, indicating that the handling of these two ions along the descending limb may be linked in part to the water movements in this nephron segment. The fractional deliveries of K at the hairpin turn and in urine were significantly correlated, and both decreased in the presence of HCT. Since, as shown previously, HCT reduces the net addition of K along the superfical distal tubule, it is concluded that calcitonin inhibits the medullary recycling of K between the nephron terminal segments and the loop of Henle of juxtamedullary nephrons.     

Aldosterone binding along the rabbit nephron: an autoradiographic study on isolated tubules

The sites of action of aldosterone (A) along the tubule of rabbit kidney were studied by autoradiographic localization of mineralocorticoid-binding sites on microdissected tubular segments. Kidney pyramids were incubated at 30 degrees C for 1 h in a collagenase solution with [3H]aldosterone at a concentration of 1.5 X 10(-9) M with and without an excess unlabeled A. Tubular segments were then microdissected and transferred onto dry film; fixation and staining were done only after exposure of the film 4 mo later in order to avoid diffusion. Specific nuclear labeling was 19.0 +/- 1.3 silver grains/100 micrometers2 in distal convoluted tubules (n = 28) and 21.0 +/- 1.8 in cortical collecting ducts (n = 18). No difference between these two structures was observed (P greater than 0.1, paired t test, n = 15). No specific binding was found in the proximal tubule (0.5 +/- 0.4, n = 17). In the thick ascending limb of Henle's loop, the labeling was low (3.9 +/- 0.9, n = 16). We conclude that, in the rabbit kidney, nuclear mineralocorticoid-binding sites, presumably receptors, are present in the distal and cortical collecting tubule.