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ABSTRACT: Previous experiments have shown that epithelial cells in the renal medulla produce endothelin-1 (ET-1) and possess ETB receptors. It has been suggested that medullary ET-1 may affect water and sodium absorption along the collecting ducts in an autocrine fashion. To study possible mechanisms responsible for the regulation of medullary ET-1 production, experiments were performed in M-1 cells and mIMCD-K2 cells, cell lines derived from cortical and inner medullary collecting ducts of SV40 transgenic mice, grown to confluence on collagen-coated filter inserts. Both cell lines were found to express ET-1 mRNA and to secrete ET almost exclusively into the basolateral medium as long as the transepithelial resistance was high. Inhibition of transcription with actinomycin D was followed by a decline in both ET mRNA [halftime (t1/2) = 30 min] and ET secretion (t1/2 = approximately 90 min). The addition of arginine vasopressin (AVP, 10(-8) M; 2- or 4-h exposure) or incubation of M-1 cells in hypertonic media (+50 mM NaCl, 4- or 6-h exposure) did not significantly alter ET secretion or ET-1 mRNA expression. In contrast, simultaneously increasing AVP(10(-8) M in the basolateral medium) and tonicity (+50 mM NaCl) for 4 h increased ET secretion (from 28.9 +/- 3.9 to 41.8 +/- 3.8 pg.h-1.mg protein-1; P = 0.029, n = 10) and ET-1 mRNA (control = 2,138 cpm/microliter, log of 3.33 +/- 0.048, n = 4; AVP + NaCl = 3,548.1 cpm/microliter, log of 3.55 +/- 0.09; P = 0.045, n = 5). Exposure of M-1 cells to hypertonic media (+50 mM NaCl or 100 mM mannitol) for 24 h was associated with a marked reduction of ET secretion (-83.9% with NaCl and -78.4% with mannitol; P < 0.0001). This reduction was attenuated, but not prevented, by the presence of AVP in the basolateral medium (-40%). ET-1 mRNA, in contrast, did not change with 24-h exposure to hypertonic media and increased when AVP was present. Results are compatible with the concept that generation of ET by collecting duct cells may contribute in a complex and time-dependent fashion to the paracrine control of collecting duct cell function.
The American journal of physiology 08/1996; 271(2 Pt 2):F330-9.
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ABSTRACT: The newborn has a limited ability to regulate H+/HCO3- homeostasis, due in part to immaturity of the intercalated cells in the distal nephron. We traced the postnatal differentiation of the intercalated cells of the rabbit cortical collecting duct (CCD) and outer medullary collecting duct (OMCD) using MAb to the 31-kD subunit of the vacuolar H(+)-ATPase, membrane portion of erythrocyte band 3, and apical surface of B-intercalated cells (peanut agglutinin [PNA], MAb B63). In the most superficial CCD of the newborn there was no binding to these probes, although deeper in the cortex there was faint apical staining with PNA and MAb B63 and a few patterns of H(+)-ATPase and band 3 labeling of neonatal intercalated cells. The OMCD showed mostly apical H(+)-ATPase and both cytoplasmic and basolateral band 3 labeling but B-intercalated cell markers were not seen. By 3 wk of age the staining of the CCD and OMCD was more polarized, resembling those in the adult. Band 3 positive cells (as a percentage of total cells) in the CCD increased from 13 to 17% during maturation, and in the OMCD they increased from 22 to 37%. Some basolateral band 3 and apical H(+)-ATPase staining was also seen in the inner medullary collecting duct of 3-wk-old rabbits to a greater extent than in newborn or adult rabbits. Labeling of intercalated cells in the CCD and OMCD was weakest and least numerous in the newborn, greater in the 3 wk old, and greatest in the adult. Most maturing cortical intercalated cells bound both PNA and H(+)-ATPase MAb, comparable to what has been observed in the adult CCD. PNA-negative cells showing apical H(+)-ATPase labeling, consistent with the classic A-intercalated cell phenotype, comprised only 5% of identified intercalated cells in the newborn CCD compared with 12% in older animals. In or near the developing renal vesicles and ampullary structures were occasional cytoplasmically staining PNA- and B63-positive cells. Whether these cells are precursors of specific renal tubular cells cannot yet be established. Staining for principal cells (ST.9) was less intense in the neonatal cortex than in more mature cortex, but the deep cortex and outer medulla were heavily labeled at all ages. These data indicate that immature intercalated cells, in the CCD and OMCD, may undergo significant postnatal proliferation and differentiation, acquiring mature phenotypes during the first month of life. The A-intercalated cell appears more differentiated than the B cell during the 1st wk of life, suggesting that A-intercalated cells contribute more than B cells to the maintenance of acid-base homeostasis in the newborn.
Pediatric Research 02/1996; 39(1):1-12. · 2.70 Impact Factor
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ABSTRACT: The newborn is limited in its ability to respond to acid-base perturbations. To investigate the development of renal H+/HCO3- transport mechanisms, we probed acid-base-related epitopes in the mesonephric and developing metanephric kidneys of rabbits. Using immunofluorescence with monoclonal antibodies to the vacuolar H+ATPase, band 3-like Cl-/HCO3- exchanger, and apical surface of fully differentiated beta-intercalated cells, and peanut lectin cytochemistry (another marker of beta-intercalated cells), we found that these epitopes were poorly expressed in the nephrogenic zone of the newborn kidney cortex. Deeper in the cortex, collecting ducts showed weak apical staining with beta-intercalated cell antibodies and two patterns of staining with the H+ATPase and band 3 antibodies: polar and circumferential or diffuse. Some cells showed apical staining with H+ATPase while others showed diffuse staining, similar to that observed in the mature cortical collecting duct. Band 3 labeling was basolateral, as observed in the adult, and diffuse, which was rarely seen in mature kidney sections. Newborn outer medullary collecting ducts showed apical labeling with H+ATPase and basolateral staining with band 3 antibodies, similar to the mature outer medulla. Surprisingly, the mesonephric collecting tubule showed cells with apical H+ATPase staining or basolateral band 3 labeling and, less frequently, cells with positive staining for beta-intercalated cells. The relative maturity of the mesonephric collecting tubule and similarity to what is observed in mature metanephric collecting ducts indicates that intercalated cells may be present and functioning in both organs. Thus, the lineage of intercalated cells may be more intricate than previously believed.
Pediatric Nephrology 01/1994; 7(6):792-7. · 2.52 Impact Factor
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ABSTRACT: Although G proteins have been shown to regulate cation channels, regulation of Cl- channels by G proteins has not been demonstrated directly. Accordingly, the objective of this study was to examine whether a G protein regulates Cl- channels in the apical membrane of rabbit kidney CCD cells grown in culture. Previous studies showed that this channel is activated by adenosine and protein kinase C and has a single channel conductance of 305 picosiemens. The PCl-:PNa+ is 9:1 and the PCl-:PHCO3- is 2:1 (Schwiebert, E.M., Light, D.B., Dietl, P., Fejes-Toth, G., Naray-Fejes-Toth, A., and Stanton, B. (1990) Kidney Int. 37,216). In the present study, Cl- channels in the apical membrane of CCD cells were studied by the patch clamp technique. GTP and guanosine 5'-O(3-thiophosphate) (GTP gamma S), a nonhydrolyzable analog of GTP, increased the single channel open probability (Po). In contrast, guanosine 5'-O-(2-thiophosphate), a nonhydrolyzable analog of GDP, and pertussis toxin (PTX) decreased the Po. GTP gamma S, but not GTP, reversed PTX inhibition of the channel. The alpha i-3-subunit of Gi increased the Po in both untreated and PTX-treated membrane patches. Because GTP gamma S activated the Cl- channel in the presence of H8, a protein kinase inhibitor, we conclude that the G protein does not activate the channel by stimulating a protein kinase. Thus, a PTX-sensitive G protein activates a Cl- channel in the apical membrane of renal CCD cells.
Journal of Biological Chemistry 06/1990; 265(14):7725-8. · 4.77 Impact Factor
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ABSTRACT: Ion channels in the apical membrane of cortical collecting duct (CCD) cells in culture were studied by the patch-clamp technique. CCD cells from rabbit kidney were isolated by solid-phase immunoadsorption with a monoclonal antibody. The majority of CCD cells (93%) had phenotypic characteristics similar to intercalated cells (ICC). Although Cl- channels were present in the apical membrane of the ICC cells, they were rarely active in cell-attached patches; however, channels were activated after patch excision. In inside-out patches, the channels exhibited rapid flickering, substrates, and large unitary currents. The single-channel conductance was 303 pS, the Cl(-)-to-Na+ permeability ratio was 10:1 and the Cl(-)-to-HCO3- permeability ratio was 1.5:1. The channel was inhibited by the Cl- channel blockers 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, diphenylamine carboxylic acid, and 5-nitro-2-(3-phenylpropylamino)-benzoic acid. Although a reduction in the cytoplasmic Ca2+ concentration inhibited channel activity in both inside-out patches and cell-attached patches, alterations of Ca2+ within the physiological range did not change the channel open probability. Finally, changing the cytoplasmic pH (6.5 to 8.0) did not alter the open probability. Thus a large conductance anion channel is present in the apical membrane of CCD cells in culture. This channel may be involved in cell volume regulation or in Cl- and HCO3- secretion.
The American journal of physiology 03/1990; 258(2 Pt 2):F273-80.
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ABSTRACT: The effect of in vivo administration of dexamethasone (1 mg/kg/d for 1 wk) was investigated on the ratio of urinary PGE2/PGF2 alpha excretion and on in vitro cortical prostaglandin-9-ketoreductase (PG-9-KR) activity in rabbits. Dexamethasone treatment decreased PGF2 alpha excretion significantly from 32.87 +/- 8.09 to 19.28 +/- 6.14 ng/mg creatinine, p less than 0.05) without causing a significant change in PGE2 excretion (9.38 +/- 2.48 and 13.53 +/- 5.03 ng/mg creatinine before and after dexamethasone, respectively). The ratio of urinary PGE2/PGF2 alpha increased markedly following glucocorticoid administration (from 0.34 +/- 0.08 to 1.03 +/- 0.32, p less than 0.05). The activity of cytosolic PG-9-KR activity, however, was not significantly different in control and dexamethasone-treated rabbits. It is concluded that the glucocorticoid-induced increase in urinary PGE2/PGF2 alpha ratio is probably unrelated to changes in PG-9-KR activity.
Prostaglandins Leukotrienes and Medicine 09/1985; 19(2):133-7.
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ABSTRACT: To investigate the possible role of vasopressin (VP) in the maintenance of DOC-salt hypertension the effect of two VP pressor antagonists on mean arterial pressure and the pressor responsiveness to exogenous VP were studied in conscious, freely moving rats with malignant DOC-salt hypertension. Intravenous injections of either 20 micrograms/kg of dP Tyr(Me)AVP or 10 micrograms/kg of d(CH2)5Tyr(Me)AVP had no significant effect on mean arterial pressure and heart rate, although both antagonists almost completely abolished the pressor response to VP. Furthermore, the animals with DOC-salt hypertension exhibited decreased pressor responsiveness to exogenous VP. The present findings strongly suggest that VP is not essential as a pressor hormone for maintaining blood pressure in malignant DOC-salt hypertension.
Clinical and experimental hypertension. Part A, Theory and practice 02/1985; 7(10):1457-70.
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ABSTRACT: Although it is well established that chronic treatment with antidiuretic hormone increases renal prostaglandin (PG) excretion, the effects of short-term infusions are controversial. Therefore, in the present study the effect of acute administration of arginine vasopressin on urinary PG excretion was investigated in conscious Brattleboro rats and in water-diuresing Long-Evans rats. Water balance was kept constant during arginine vasopressin infusion. Arginine vasopressin caused a significant, dose-related and reversible increase in urinary PG excretion within 20 min in both models. Similar results were obtained during the infusion of 1-deamino-8-D-arginine vasopressin in the Brattleboro rat. Normalization of the hydropenia of Brattleboro rats by infusion of large amounts of hypotonic fluid failed to elevate urinary PG excretion. These results give no support to the hypothesis that the acute enhancement of urinary PG excretion by vasopressin is mediated through either vasoconstriction or volume retention or induction of cyclooxygenase, but rather they indicate that antidiuretic hormone can increase renal PG synthesis through a more direct mechanism in vivo.
Journal of Pharmacology and Experimental Therapeutics 11/1983; 227(1):215-9. · 3.83 Impact Factor
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ABSTRACT: The renal response to arginine vasopressin was investigated with and without the simultaneous administration of the kallikrein inhibitor, aprotinin, in conscious Brattleboro homozygous rats with hereditary diabetes insipidus. Arginine vasopressin caused a marked antidiuretic response (urinary osmolality increased from 118 to 739 mosmol/l) which was accompanied by a significant increase in urinary prostaglandin excretion (prostaglandin E2 and F2 alpha excretion increased by 182 and 441%, respectively). Kallikrein excretion remained unchanged after arginine vasopressin infusion. The infusion of aprotinin diminished urinary kallikrein activity to undetectable levels, decreased potassium excretion significantly and caused a slight fall in urinary prostaglandin excretion. However, aprotinin failed to modify the arginine vasopressin-induced enhancement in prostaglandin excretion (prostaglandin E2 and F2 alpha excretion increased by 168 and 442%, respectively), and the antidiuretic response was also similar to that observed under control conditions. These results indicate that the kallikrein-kinin system is not involved in the renal response to vasopressin in the Brattleboro rat.
The Journal of Physiology 07/1983; 339:585-90. · 4.72 Impact Factor
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Klinische Wochenschrift 10/1982; 60(18):1155-64.
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ABSTRACT: This brief review summarizes the important modulatory effects of endogenous renal prostaglandins (PGs) on renal function. A brief survey of current knowledge of renal PG synthesis reveals the shortcomings of biochemical in vitro studies and points out the necessity to improve methods of in vivo assessment. The problem of measuring circulating levels of PGs and the question of the importance of these PGs for renal function are discussed. Renal blood flow is shown to depend on renal PG's in situations of stress but not under basal conditions. However, in this respect the definition of stress has to include even the mild condition of sodium deprivation. An increasing number of disease states is emerging where renal blood flow is shown to depend on cyclooxygenase activity. Angiotensin II infused or synthesized endogenously seems to be the most likely stress mediator causing enhanced renal PG synthesis which opposes its vasoconstrictor activity. The role of PGs in sodium excretion is not well defined and controversial. In man, most available evidence supports a natriuretic role of renal PGs. However, this role can only be demonstrated under well defined conditions. Inhibitors of PG biosynthesis will cause sodium retention. However, following administration of these drugs a new steady state is reached rapidly characterized by suppressed plasma renin activity and aldosterone. Renal PGs alternate vasopressin effect on urine concentration. The mechanism for this is poorly understood and the role of cAMP in this interaction controversial. Recent evidence showing stimulation of renal PG synthesis by vasopressin independent of its pressor effect is presented. While the stimulatory role of kininogen and kinins on renal PG synthesis has been shown the functional consequences have not been sufficiently defined. Moreover, assessment of the activity of the renal kallikrein-kinin system is at present not possible and no physiological role in renal function has as yet been delineated. Regulation of renal renin release involves PGs. Evidence is presented in support of a role of prostacyclin in baroreceptor and macula densa mediated renin release.Die folgende bersicht beleuchtet die fr die Nierenfunktion wichtigen Aspekte der in der Niere synthetisierten Prostaglandine (PG). Die Analyse des derzeitigen Wissenstandes ber die renale PG-Synthese zeigt die Grenzen der biochemischen in vitro Methoden und die Notwendigkeit der Verbesserung der in vivo Analyse auf. Die Problematik der Messung zirkulierender-PG und die Frage ihrer Bedeutung fr die Nierenfunktion wird diskutiert. Der renale Blutflu hngt unter Stre-Bedingungen von den renalen PG ab, nicht jedoch unter Basalbedingungen. In diesem Zusammenhang beinhaltet die Definition des Stre allerdings auch bereits Natriumarme Ernhrung. Bei einer zunehmenden Anzahl von Erkrankungen wird entdeckt, da PG fr die Aufrechterhaltung der Nierendurchblutung von Bedeutung sind. Angiotensin II ist wahrscheinlich der Stre-Faktor, welcher fr die erhhte PG-Synthese verantwortlich ist, welche die Vasokonstriktion partiell antagonisiert. — Die Rolle der PG bei der Natriumexkretion ist nicht exakt definiert und es liegen widersprchliche Ergebnisse vor. Beim Menschen ist eine natriuretische Wirkung endogener PG wahrscheinlich, kann jedoch nur unter genau definierten Bedingungen nachgewiesen werden. Blocker der PG-Synthese bewirken meist eine Natriumretention. Jedoch kommt es nach Gabe dieser Pharmaka rasch zu einem neuen Fliegleichgewicht fr Natrium mit charakteristisch supprimierten Werten fr die Plasmareninaktivitt und Plasma-Aldosteron.Die renalen PG reduzieren die hydroosmotische Wirkung von Vasopressin (VP). Der Mechanismus dieser Wirkung ist nicht bekannt und die Rolle des cAMP ungewi. Neuere Befunde zeigen eine starke Stimulation der renalen PG-Synthese durch VP unabhngig von seiner pressorischen Wirkung.Stimulation der renalen PG-Synthese durch Kininogen und Kinine ist bekannt, jedoch die funktionelle Bedeutung ist unklar, insbesondere da dem renalen Kallikrein-Kinin-System noch keine physiologische Funktion zugeordnet werden kann. Renale Reninabgabe erfolgt teilweise ber PG-abhngige Mechanismen. Die Bedeutung von Prostazyklin fr die durch den Barorezeptor und die Macula densa vermittelte Reninabgabe wird dargestellt.
Journal of Molecular Medicine 08/1982; 60(18):1155-1164. · 4.67 Impact Factor
