[Contribution of apelin to water balance, blood glucose control, and cardiovascular functions]. / Participation de l'apéline à la régulation de l'équilibre hydrique, de l'homéostasie glucidique et des fonctions cardiovasculaires.

The discovery of apelin, an endogenous ligand of the orphan APJ receptor, constitutes an important advance in both fundamental research and clinical medicine. Experimental data have shown that apelin has a diuretic effect via its central and renal actions: by inhibiting the phasic activity of vasopressinergic neurons and systemic secretion of vasopressin and its direct effect on the renal microcirculation and probably tubular function. Besides its diuretic action, when injected into the blood stream, apelin decreases blood pressure and increases the contractile force of the myocardium while decreasing pre- and post-load, actions opposing those of vasopressin and angiotensin II. Taken together, these data show that this new circulating vasoactive (neuro)peptide could play a crucial role in maintaining water and electrolyte balance and cardiovascular functions. Finally, a systemic injection of apelin in insulin-resistant mice decreases glycemia and enhances glucose uptake in skeletal muscle and adipose tissue, contributing to homeostatic control of blood glucose. Clinically, the development of non-peptide analogs of the apelin receptor could provide new therapeutic tools potentially useful for the treatment of heart failure, states of water and/or electrolyte retention, and type 2 diabetes mellitus.

P2Y receptors present in the native and isolated rat glomerulus.

Extracellular ATP can mobilize intracellular calcium in rat glomeruli by interacting with P2Y receptors. However, the identity of the receptor subtypes involved is not known. In the present study, we have used RT-PCR to identify mRNAs for specific P2Y receptor subtypes expressed in the rat glomerulus: mRNA for P2Y1, P2Y2, P2Y4 and P2Y6 receptors was detected. Functional expression of P2Y1 and P2Y2/P2Y4, but not P2Y6, receptors in intact glomeruli was confirmed by measuring the relative stimulation of the inositol phosphate pathway induced by selective agonists of a particular receptor subtype. Finally, we have used available polyclonal antibodies to confirm the expression of P2Y1 and P2Y2 in the glomerulus, in mesangial cells and glomerular epithelial cells (podocytes), respectively; but we could not demonstrate P2Y4 or P2Y6 receptor expression by this means. In a separate series of experiments, we have examined the possibility that intra-renal sympathetic nerve terminals are a source of extracellular ATP and that this would be supported, though not excluded, by supersensitivity to ATP following denervation. Nucleotide-induced stimulation of the inositol phosphate pathway was measured in both control rats and rats that had been sympathectomized by intraperitoneal injection of 6-hydroxydopamine. The response to norepinephrine was measured as a positive control. In the sympathectomized rats, the effect of norepinephrine was significantly enhanced, whereas ATP-induced inositol phosphate production was unaffected, being similar in both groups of animals.

Desensitization of type 1 angiotensin II receptor subtypes in the rat kidney.

Differences involving serine residues in the sequence of the carboxyl-terminal tail of type 1 angiotensin II (Ang II) receptor subtypes AT(1A) and AT(1B) suggest differences in desensitization ability. We examined the Ang II-induced homologous desensitization patterns of both receptor subtypes in freshly isolated renal structures: glomerulus (Glom), afferent arteriole, and cortical thick ascending limb (CTAL), whose content in each subtype mRNA is different, by measuring variations in intracellular calcium concentration. A preexposure to a maximal dose of Ang II, followed by a second application of the same concentration, induced: 1) a complete desensitization in Glom, where AT(1A) and AT(1B) mRNAs were expressed in similar proportions, and 2) no or partial desensitization in afferent arteriole and CTAL, where AT(1A) mRNA was predominant. In the absence of nephron structure containing only AT(1B) mRNA, we studied rat anterior pituitary cells that exhibit high content in this subtype and observed that desensitization was not complete. In Glom, CTAL, and pituitary cells, desensitization proceeded in a dose-dependent manner. In Glom and CTAL, desensitization occurred via a PKC-independent mechanism. These results suggest that desensitization does not depend on the nature of Ang II receptor subtype but either on the proportion of each subtype in a given cell and/or on cell specific type. This could allow adaptive biological responses to Ang II appropriate to the specific function of a given cell type.

Aminopeptidase A activity and angiotensin III effects on [Ca2+]i along the rat nephron.

BACKGROUND: This study examined the specific effects of angiotensin III (Ang III) along the nephron. METHODS: We examined the distribution of aminopeptidase A (APA) activity by using a specific APA inhibitor and by immunostaining with an antirat kidney APA antibody, the Ang III-induced variations of [Ca2+]i by using fura-2 and the characterization of the receptor subtype involved in the response to Ang III in cortical thick ascending limb (CTAL). RESULTS: APA activity was found all along the nephron but was higher in the cortex than in the medulla. This was confirmed by immunostaining. Increases in [Ca2+]i elicited by 10(-7) mol/liter Ang III were observed all along the nephron. The characterization of the receptor subtype involved in the [Ca2+]i response to Ang III in CTAL indicated that EC50 values for Ang III and Ang II were similar (13.5 and 10.3 nmol/liter, respectively), and Ang III-induced responses were totally abolished by AT1 receptor but not by AT2 receptor antagonists. There was a cross-desensitization of [Ca2+]i responses to 10(-7) mol/liter Ang III and Ang II, and the [Ca2+]i responses to 10(-7) mol/liter Ang II and Ang III were not additive. CONCLUSION: These results show that in CTAL, the [Ca2+]i responses to Ang II and Ang III occur through the same AT1a receptor because this subtype is predominant in this segment. Taken together, these data suggest that APA could be a key enzyme to generate Ang III from Ang II in the kidney.

Identification of two arginase isoenzyme activities along the nephron of Meriones shawi.

Conflicting theories on the existence of several renal arginase isoenzymes remain in debate. Because the activity of arginase is high in two embryologically different nephron segments of the Meriones shawi kidney, namely the cortical (CPST) and medullary (OSPST) proximal straight tubule and the outer medullary collecting duct (OMCD), we postulate that these nephron segments may contain different isoforms. Isolated nephron segments were dissected from collagenase-treated kidneys. Tubules were permeabilized with Triton X-100 (0.25%) and incubated with increasing Arg concentrations to characterize the arginase activity. The results were as follows: (1) in OMCD, one arginase isoform (E1), characterized by a high Arg affinity (1.160 mM), was present; (2) in CPST, two arginase isoforms were discovered - one, E1, had a similar Km (1.407 mM) to that found in OMCD whereas the other (E2) had a low affinity for Arg (Km =18.8 mM); and (3) in OSPST, two isoenzymes were present - E1 which had a Km of 1.478 mM and the second isoform that we named E2 which had a Km of 9.07 mM. In addition, arginase located in CPST and OMCD was strongly inhibited by Orn and Lys. The Ki value for Lys varied between 1.635 and 2.288 mM. Therefore, this work demonstrates that two arginase isoforms are present in the kidney of Meriones shawi. Isoform E1 is present in the proximal tubule and the collecting duct whereas isoform E2 is restricted to the proximal tubule.

Ornithine decarboxylase along the mouse and rat nephron.

Renal arginase activity is a potent source of ornithine (Orn) for polyamine synthesis. Ornithine decarboxylase (ODC) was localized along the mouse and rat nephron by incubating viable nephron segments isolated by microdissection from collagenase-treated kidneys with or without D,L-2-(difluoromethyl)ornithine (DFMO), a selective inactivator of ODC. Tubules from either control or DFMO-treated animals were incubated with 100 ¿M L-[1-14C]Orn. In control mice, Orn decarboxylation occurred mainly in the proximal convoluted tubule (PCT). In DFMO-treated mice, Orn decarboxylation was dramatically reduced in PCT and in proximal straight tubules (PST). In rats, Orn decarboxylation also occurred predominantly in the proximal tubule. Addition of 10 mM DFMO to isolated tubules dramatically decreased Orn decarboxylation in PCT and in PST. Thereafter, ODC activity was demonstrated in permeabilized tubules. In Triton X-100-treated tubules from control mice, ODC was exclusively found in proximal tubules (PCT > PST). This ODC activity was strongly inhibited in DFMO-treated mice. In conclusion, the highest ODC activity was found in rat and mouse PCT, a segment devoid of arginase. We hypothesize that the filtered Orn, which is reabsorbed along the PCT,is the main source of Orn for ODC.

Expression of type 1 angiotensin II receptor subtypes and angiotensin II-induced calcium mobilization along the rat nephron.

The localization of two type 1 angiotensin II receptor subtype mRNA, AT1A and AT1B, was determined by reverse transcription-PCR on microdissected glomeruli and nephron segments. The coupling sensitivity of these two receptor subtypes was evaluated by measuring variations in intracellular calcium ([Ca2+]i) elicited by angiotensin II (Ang II) in structures expressing either AT1A or AT1B mRNA, using Fura-2 fluorescence. The highest expression of AT1 mRNA was found in glomerulus, proximal tubule, and thick ascending limb. In glomerulus, AT1A and AT1B mRNA were similarly expressed, whereas in all nephron segments AT1A mRNA expression was dominant (approximately 84%). The increase in [Ca2+]i elicited by 10(-7) mol/L Ang II was highest in proximal segments (delta [Ca2+]i is approximately equivalent to 300 to 400 nmol/L) and thick ascending limb (delta [Ca2+]i is approximately equivalent to 200 nmol/L). In glomerulus and collecting duct, the response was lower (delta < 100 nmol/L). The median effective concentrations for Ang II were of the same order of magnitude in glomerulus (12.2 nmol/L), in which both AT1A and AT1B are expressed, and in cortical thick ascending limb (10.3 nmol/ L), in which AT1A is almost exclusively expressed. The Ang II-induced calcium responses were totally abolished by the AT1 receptor antagonist losartan (1 mumol/L) but not by the AT2 antagonist PD 123319 (1 mumol/L). In the absence of external Ca2+, the peak phase of the response induced by 10(-7) mol/L Ang II was reduced and shortened, suggesting that a part of the [Ca2+]i increase originated from the mobilization of the intracellular Ca2+ pool. In conclusion, these results demonstrate that in the rat kidney: (1) AT1A is the predominant AT1 receptor subtype expressed in the nephron segments, (2) glomerulus is the only structure with a relatively high AT1B mRNA content, and (3) AT1A and AT1B receptor subtypes do not differ in their efficiency for the activation of calcium second-messenger system.

Biochemical heterogeneity of arginine metabolism along kidney proximal tubules.

By using an in vitro single tubule micromethod of high specificity, in four different species of mammals it has been observed that (a) arginine synthesis from citrulline (arginine synthase activity, E.C. 6.3.4.5 and E.C. 4.3.2.1) is restricted to the early portions of proximal convoluted tubules, whereas (b) urea production from arginine (arginase activity, E.C. 3.5.3.1.) is present mainly in the cortical (CPST) and even more in the outer medullary (OSPST) portions of straight proximal tubules. The data suggest that (a) in early PCT cells, the citrulline reabsorbed from glomerular filtrate is converted into arginine, which in turn crosses peritubular cell membranes together with reabsorbed arginine, and (b) the urea formed in CPST and OSPST cells might passively diffuse into the luminal fluid entering Henle's loops. Such urea secretion might contribute to sustain the process of urea recycling in kidney medulla and thereby participate in the mechanism of urine concentration.

Arginine metabolism in cat kidney.

1. Arginine is essential for growth in the kitten and, because of the resulting hyperammonaemia, in the adult cat an arginine-free diet is life threatening. 2. The kidney is the main site of arginine synthesis. 3. This study was performed to determine whether the cat kidney synthesizes arginine and to establish which factors, such as low citrullinaemia, defects of argininosuccinate synthase and lyase activities or high renal arginase activity, might limit renal arginine production. 4. Identified nephron segments were isolated by microdissection from collagenase-treated cat kidney. 5. Arginine metabolism was studied by incubating the nephron segments with either physiological concentrations of L-[ureido-14C]citrulline (anabolism) or L-[guanido-14C]-arginine (catabolism). Arginine and urea were measured by a micro-enzymatic method. Amino acids were measured by HPLC. 6. In cat blood, the citrulline, but not the arginine, concentration was very low by comparison with other species. 7. Arginine synthesis occurred almost entirely in the proximal tubule, the highest rate occurring in the proximal convoluted tubule and the lowest in the medullary straight proximal tubule. 8. Arginase activity was restricted to the proximal tubule. Urea production increased from the convoluted towards the medullary straight tubule. 9. The limited capacity of the cat kidney to produce arginine in vivo may result from the low blood concentration of citrulline and from the high arginase activity in the various proximal cells with the ability to synthesize arginine.

Sites of arginine synthesis and urea production along the nephron of a desert rodent species, Meriones shawi.

The distribution of arginine synthase and arginase activities along the successive nephron segments of Meriones kidney was measured in vitro with single tubule enzymatic microtechniques making use of either L-[ureido-14C] Citrulline (0.108 mM) or L-[guanidino-14C]arginine (0.2 mM) as the respective substrates. Arginase activity (fmol urea formed per min per mm of tubule) was very low (5-25 fmol.min-1.mm-1) in most nephron segments including the early portions of proximal convoluted tubules (early PCT). It increased progressively after 3 mm of the PCT to reach a value of 200 fmol.min-1.mm-1 in the cortical portion of the straight proximal tubule (CPST), with a further increase, along the pars recta, of up to 250 fmol.min-1.mm-1 in the outer medullary portion (OSPST). In addition, arginase activity in OSPST and the adjacent descending thin limb (DTL) was higher in juxtamedullary nephrons (JN) than in the corresponding portions of superficial nephrons (SN). Arginine synthase activity (fmol arginine formed per mm of tubule per min) was present in proximal tubules exclusively, with a gradient decreasing along the PCT (about 600 fmol.min-1.mm-1 in the 1st mm, 65 fmol.min-1.mm-1 in CPST and 30 fmol.min-1.mm-1 in OSPST). It has been checked that CPST and OSPST (where the two enzyme systems are present) are able to convert citrulline directly into urea with a yield of 65%.(ABSTRACT TRUNCATED AT 250 WORDS)

Urea production by kidney collecting ducts in vitro: effect of amino acid addition.

Urea production by cortical (CCD) and medullary (OMCD) collecting ducts of the rat kidney was measured in vitro by incubating single microdissected pieces of tubule in the presence of L-[guanido-14C]arginine (0.2 mM). The [14C]urea released from the cells was hydrolysed in presence of urease added to the incubation medium and the 14CO2 formed was trapped in KOH and counted. The effect of various amino acids (AA) on urea production was investigated by adding unlabelled AA (either in combination or singly) at concentrations close to those present in blood plasma. A mixture of 17 AA decreased urea production from [14C]arginine by 46% in CCD and by 58% in OMCD. When lysine and proline were omitted from the mixture, the inhibition was less marked (19% in CCD and 43% in OMCD, respectively). When AA were tested singly, lysine induced the larger inhibition (40% in CCD and 45% in OMCD), than ornithine and glutamine (about 15% each, in CCD and OMCD), whereas proline inhibition (7% in CCD, 10% in OMCD) was not statistically significant. Branched-chain amino acids (BCAA) in combination (leucine, isoleucine and valine) also markedly reduced urea production by CCD and OMCD. Their effect was dose dependent. Solubilization of CCD and OMCD cell membranes with Triton X-100 resulted in a twofold increase in urea production by control samples; the relative inhibition (per cent) induced by BCAA was enhanced, whereas that induced by lysine was decreased. The data suggest that, in living tubules, the inhibition obtained with lysine resulted, for a large part, from competition between lysine and arginine for cell uptake via a common membrane carrier, whereas the inhibition induced by BCAA corresponded to an effect on arginase activity itself.

Arginine synthesis in mouse and rabbit nephron: localization and functional significance.

In the rat kidney, arginine (Arg) synthesis is restricted to the proximal tubule with a decreasing intensity from its convoluted (PCT) to its straight part (PST). The present study was designed to investigate the pattern of Arg synthesis along the nephron in other mammals, the mouse and rabbit. Microdissected representative nephron segments were incubated with 0.1 mM L-[ureido-14C]citrulline in a sealed chamber. Addition of arginase and urease to the incubation medium led to the hydrolysis of Arg into ornithine, NH3, and 14CO2. The latter was trapped in KOH and counted (results are in fmol Arg.min-1.mm tubular length-1). As in the rat, the main site of Arg synthesis in both species was found to be the PCT (mouse, 191; and rabbit, 57). A lower production was observed in rabbit and mouse PST and in rabbit distal segments. Along the PCT (from 1st to 4th mm after the glomerulus), a steep decrease is observed in mouse (595 and 37, respectively) but not in rabbit (57 and 23). The fate of the newly synthesized Arg probably depends on its site of production. Intracellular arginase activity is known to be present in the cortical (C) and medullary (OS) PST, in both mouse and rabbit. In rabbit only, arginase activity is also found in the PCT. We observed that a large part of Arg was further hydrolyzed into urea and ornithine in CPST and OSPST of mouse (66 and 80%, respectively) and rabbit (40 and 70%) but not in rabbit PCT (8%). Thus Arg produced by PCT in both species is probably released in the cortical blood, whereas Arg produced in PST may serve locally to produce urea and ornithine, and the latter could be used for polyamine synthesis.

Localization of urea and ornithine production along mouse and rabbit nephrons: functional significance.

Hydrolysis of arginine into urea and ornithine (Orn) was observed to take place in several segments of the rat nephron including cortical and medullary pars recta of the proximal tubule (PST) and collecting duct (CD). This work was now extended to the adult mouse and rabbit. Representative nephron segments, obtained by microdissection of collagenase-treated kidneys, were incubated with L-[guanido-14C]arginine (216 microM). Addition of urease produced 14CO2 + 2 NH3 from the newly formed urea released in the incubate. 14CO2 was trapped in KOH and counted. In both species, as well as in the rat, the PST was the site of the highest urea + Orn production, with an intensity increasing from cortex to medulla. For other nephron segments, the pattern was not similar in all species. Significant production of urea + Orn was observed in the proximal convoluted tubule and the medullary thick ascending limb in the rabbit, but not in the CD of either the rabbit or the mouse. The functional significance of this urea + Orn production remains unclear. The total amount of urea generated intrarenally by this reaction does not seem sufficient to play a significant role in the urinary concentrating mechanism. It may be assumed that Orn could be further metabolized to polyamines and play a role in maintaining cell integrity and function in the PST, especially in its medullary part, exposed to hypertonicity and poor oxygen supply.

Localization of arginine synthesis along rat nephron.

Arginine production was measured in isolated rat nephron segments. Segments were incubated with 0.3 mM aspartate and 0.1 mM L-[ureido-14C]-citrulline in a sealed chamber. Arginase and urease were added to the medium to hydrolyze arginine and to release 14CO2, which was trapped in KOH and counted. Arginine synthesis was found only in the proximal tubule, with decreasing intensity from proximal convoluted (PCT) to proximal straight tubule (PST). Results were as follows (in fmol.min-1.mm tubule length-1): PCT, 122 +/- 15; cortical PST, 71 +/- 6; outer medullary PST, 41 +/- 4; all other segments, less than 6. Arginine synthesis changed almost proportionally with precursor concentration of less than or equal to 0.4 mM. We had shown previously that PST but not PCT was able to hydrolyze arginine into urea and ornithine. In this study arginine was further hydrolyzed in cortical (40%) and medullary (64%) PST but not in PCT. These observations suggest that the arginine formed in PCT contributes to the maintenance of the whole body arginine pool, whereas most of the arginine formed in PST might contribute, by its conversion to urea, to the process of urine concentration.

Production of urea from arginine in pars recta and collecting duct of the rat kidney.

Urea production from arginine was studied in vitro in the kidney of normal rats in tubule suspensions of the four different renal zones (cortex, outer and inner stripe of outer medulla, and inner medulla), and in individual microdissected nephron segments. Tissue was incubated with L-[guanido-14C]-arginine to measure cellular arginase activity. Addition of urease to the incubate freed 14CO2 from the 14C-urea formed by arginase and released from the cells. CO2 was trapped in KOH and counted. These experiments revealed that significant amounts of urea are produced in the outer stripe and in the inner medulla. This intrarenal urea generation takes place mainly in the proximal straight tubule and in the collecting duct, with increasing activity in these two structures from superficial to deep regions of the kidney. Urea is known to play a critical role in the urinary concentrating process. The fact that some urea can be produced in the mammalian kidney, and that the two structures showing this capacity are straight portions of the renal tubular system descending along the corticopapillary axis suggest that this urea production might play a role in the formation and/or maintenance of the medullary urea concentration gradient.

In vitro stimulation of Na-K-ATPase in rat thick ascending limb by dexamethasone.

Dexamethasone has been reported to stimulate Na-K-ATPase activity in the medullary thick ascending limb of adrenalectomized animals within a few hours. The present study was aimed at characterizing the mechanism of this action by investigating the stimulatory effect of the hormone in vitro. Dexamethasone (10(-8) M) added in vitro to segments of the medullary thick ascending limb of Henle's loop, which were microdissected from adrenalectomized rats, restored in a dose-dependent manner the depressed Na-K-ATPase activity within one h of incubation. This stimulation of Na-K-ATPase was inhibited by cycloheximide and actinomycin D. Dexamethasone also stimulated the component of oxidative metabolism coupled to sodium transport. These results, which confirm previous in vivo observations, demonstrate that dexamethasone-induced stimulation of Na-K-ATPase is a direct tubular action of the hormone mediated by protein synthesis. They suggest that this short-term effect of dexamethasone corresponds to the stimulation of sodium reabsorption by the dilution segment.

Coupling of metabolic CO2 production to ion transport in isolated rat thick ascending limbs and collecting tubules.

Metabolic CO2 production from appropriate [U-14C]-labelled substrates (either L-lactate or D-glucose) was measured in single pieces of tubule as previously described (Le Bouffant et al. 1984). Changing the incubate osmotic pressure by mannitol addition resulted in an increase in oxidative metabolism which was more marked in outer-medullary segment MAL and MCT) than in cortical segments (CAL and CCT). Availability of metabolic substrate was not rate limiting under these conditions because FCCP addition (1 mumol X l-1) produced a marked rise in CO2 production in these structures. Ouabain (1 mmol X l-1) decreased by more than 50% the CO2 production by CAL, MAL, CCT and MCT samples, indicating that the larger part of oxidative metabolism was coupled to active Na transport. Furosemide addition (10(-5) mol X l-1) to CAL and MAL samples, or amiloride addition (10(-4) mol X l-1) to CCT and MCT samples reduced the rate of CO2 production to an extent almost similar to that obtained with ouabain, an observation suggesting that apical entry of Na+ was present in these non-perfused tubules. Finally, the effects of changing the concentration of either K+ or Cl- was tested in CAL samples. K+ suppression greatly depressed the rate of CO2 production. Replacement of chloride with sulfate also decreased this rate to an extent similar to that observed with furosemide.(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolic CO2 production by isolated single pieces of rat distal nephron segments.

A method is described which allowed in-vitro measurements of metabolic CO2 production from [U-14C]-substrates by single pieces of kidney tubules. The tubules were isolated by microdissection from collagenase treated rat kidneys. Single pieces of various distal nephrons portions were incubated in 1 microliter of bicarbonate free minimum essential medium containing the required [U-14C]-substrate (about 0.2 mu Ci per sample), and the 14CO2 produced was continuously trapped into a 2-microliter KOH droplet. The KOH droplets were replaced every 30 min. Metabolic CO2 production from the labelled substrate used was calculated as picomoles CO2 per mm of tubular length per minute, by dividing the KOH radioactivity by the specific radioactivity per carbon of the substrate present in the incubate [( U-14C] plus cold substrate concentrations). Under these conditions, it was established that single pieces of tubule could sustain almost constant CO2 production for at least 2 h at 31 degrees C. Experiments testing four different conditions with five to six replicate samples per condition were performed in order to compare oxidative metabolism in medullary (MAL) and cortical (CAL) thick ascending limbs, medullary (MCT) and cortical (CCT) collecting tubules and, in a few instances, proximal convoluted tubules (PCT) and early distal convoluted tubules (DCT).(ABSTRACT TRUNCATED AT 250 WORDS)

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.

Multiple hormonal control of adenylate cyclase in distal segments of the rat kidney.

Using the single tubule adenylate cyclase microassay, we investigated in vitro in three different segments of the rat nephron whether the effects of various hormones are additive when these hormones are tested in combination. In the cortical portion of the thick ascending limb (CAL), no additivity of the effects of glucagon, calcitonin, and PTH was observed. In the medullary portion of the thick ascending limb (MAL), the effects of vasopressin and glucagon were only partly additive, and the effects of vasopressin and calcitonin were fully additive. In the cortical collecting tubule (CCT), the effects of calcitonin and vasopressin were nonadditive in the kidneys in which vasopressin alone induced a high cyclase stimulation, whereas they were fully additive when vasopressin induced a low cyclase stimulation. The data suggest that in each segment, the hormones tested stimulated the same cells: no additivity was observed when cyclase Vmax acted as the limiting factor of the response; partial or full additivity was observed when the number of hormone receptors acted as the limiting factor of the response. As a consequence, calcitonin, glucagon, and PTH should induce the same effects in CAL; vasopressin, glucagon, and calcitonin, the same effects in MAL; and vasopressin and calcitonin, the same effects in CCT.