Signaling pathways involved in the rapid biphasic effect of aldosterone on Na/H exchanger in rat proximal tubule cells.

The receptors and signaling pathways for nongenomic effects of aldosterone (Aldo) on the proximal Na/H exchanger are still unknown; therefore, the aim of this study was to investigate the mineralocorticoid receptor (MR) and/or glucocorticoid receptor (GR) participation in rapid Aldo effects on NHE1 (basolateral Na/H exchanger isoform) and cytosolic calcium concentration ([Ca]). In addition, phospholipase C (PLC), protein kinase C (PKC), and mitogen-activated protein kinase kinase (MEK) involvement in signaling pathways of such effects was evaluated, using immortalized proximal tubule cells of rat (IRPTC) as an experimental model. MR and GR expression was investigated using reverse transcription polymerase chain reaction and immunoblotting.

The Role of β-Adrenergic Overstimulation in the Early Stages of Renal Injury.

Background/aims: To assess the possible contribution of the β-adrenergic overstimulation in early stages of renal injury, the present study evaluated, in rats, the effects of the β-adrenoceptor agonist isoproterenol (ISO) on renal function and morphology, as well as the renal mRNA and protein expression of the NADPH oxidase isoform 4 (Nox 4) and subunit p22phox, endoplasmic reticulum (ER) stress, pro-inflammatory, pro-apoptotic and renin-angiotensin system (RAS) components.

Methods: Wistar rats received ISO (0.3 mg.

The effects of angiotensin-(1-7) on the exchanger NHE3 and on [Ca] in the proximal tubules of spontaneously hypertensive rats.

The acute effects of angiotensin-1-7 [ANG-(1-7)] on the reabsorptive bicarbonate flow (J[Formula: see text]) were evaluated using stationary microperfusion in vivo in the proximal tubules of spontaneously hypertensive rats (SHR) and their normotensive controls, Wistar-Kyoto (WKY) rats, using a microelectrode sensitive to H In WKY rats, the control J[Formula: see text] was 2.40 ± 0.10 nmol·cm·s ( = 120); losartan (10 M) or A779 (10 M, a specific Mas antagonist), alone or in combination with losartan, decreased the J[Formula: see text] ANG-(1-7) had biphasic effects on J[Formula: see text]: at 10 M, it inhibited, and at 10, it stimulated the flow.

Dose-dependent effects of angiotensin-(1-7) on the NHE3 exchanger and [Ca(2+)](i) in in vivo proximal tubules.

The acute direct action of angiotensin-(1-7) [ANG-(1-7)] on bicarbonate reabsorption (JHCO(3)(-)) was evaluated by stationary microperfusions on in vivo middle proximal tubules in rats using H ion-sensitive microelectrodes. The control JHCO(3)(-) is 2.82 ± 0.

Action of ANP on the nongenomic dose-dependent biphasic effect of aldosterone on NHE1 in proximal S3 segment.

The rapid (2 min) nongenomic effects of aldosterone (ALDO) and/or spironolactone (MR antagonist), RU 486 (GR antagonist), atrial natriuretic peptide (ANP) and dimethyl-BAPTA (BAPTA) on the intracellular pH recovery rate (pHirr) via NHE1 (basolateral Na⁺/H⁺ exchanger isoform), after the acid load induced by NH₄Cl, and on the cytosolic free calcium concentration ([Ca²⁺](i)) were investigated in the proximal S3 segment isolated from rats, by the probes BCECF-AM and FLUO-4-AM, respectively. The basal pHi was 7.15±0.

Genomic and nongenomic stimulatory effect of aldosterone on H+-ATPase in proximal S3 segments.

The genomic and nongenomic effects of aldosterone on the intracellular pH recovery rate (pHirr) via H(+)-ATPase and on cytosolic free calcium concentration ([Ca(2+)](i)) were investigated in isolated proximal S3 segments of rats during superfusion with an Na(+)-free solution, by using the fluorescent probes BCECF-AM and FLUO-4-AM, respectively. The pHirr, after cellular acidification with a NH(4)Cl pulse, was 0.064 ± 0.

Regulation of Na+/H+ exchanger isoform 1 (NHE1) by calmodulin-binding sites: role of angiotensin II.

We examined the effect of Angiotensin II (Ang II) on the interaction between the Ca(2+)/CaM complex and hNHE1. Considering that calmodulin binds to NHE1 at two sites (A and B), amino acids at both sites were modified and two mutants were constructed: SA(1K3R/4E) and SB(1K3R/4E). Wild type and mutants were transfected into PS120 cells and their activity was examined by H(+) flux (J(H+)).

Direct action of aldosterone on bicarbonate reabsorption in in vivo cortical proximal tubule.

The direct action of aldosterone (10(-12) M) on net bicarbonate reabsorption (J(HCO(3)(-))) was evaluated by stationary microperfusion of an in vivo middle proximal tubule (S2) of rat kidney, using H ion-sensitive microelectrodes. Aldosterone in luminally perfused tubules caused a significant increase in J(HCO(3)(-)) from a mean control value of 2.84 +/- 0.

Genomic and nongenomic dose-dependent biphasic effect of aldosterone on Na+/H+ exchanger in proximal S3 segment: role of cytosolic calcium.

The effects of aldosterone on the intracellular pH recovery rate (pHirr) via Na+/H+ exchanger and on the [Ca2+]i were investigated in isolated rat S3 segment. Aldosterone [10(-12), 10(-10), or 10(-8) M with 1-h, 15- or 2-min preincubation (pi)] caused a dose-dependent increase in the pHirr, but aldosterone (10(-6) M with 1-h, 15- or 2-min pi) decreased it (these effects were prevented by HOE694 but not by S3226). After 1 min of aldosterone pi, there was a transient and dose-dependent increase of the [Ca2+]i and after 6-min pi there was a new increase of [Ca2+]i that persisted after 1 h.

The effect of angiotensin II on intracellular pH is mediated by AT(1) receptor translocation.

The effect of ANG II on intracellular pH (pH(i)) recovery rate and AT(1) receptor translocation was investigated in transfected MDCK cells. The pH(i) recovery rate was evaluated by fluorescence microscopy using the fluorescent probe BCECF-AM. The human angiotensin II receptor isoform 1 (hAT(1)) translocation was analyzed by immunofluorescence and confocal microscope.

Signaling pathways in the biphasic effect of ANG II on Na+/H+ exchanger in T84 cells.

The effect of ANG II on pH(i), [Ca(2+)](i) and cell volume was investigated in T84 cells, a cell line originated from colon epithelium, using the probes BCECF-AM, Fluo 4-AM and acridine orange, respectively. The recovery rate of pH(i) via the Na(+)/H(+) exchanger was examined in the first 2 min following the acidification of pH(i) with a NH(4)Cl pulse. In the control situation, the pH(i) recovery rate was 0.

Effect of D-alpha-tocopherol on tubular nephron acidification by rats with induced diabetes mellitus.

The objective of the present study was to determine if treatment of diabetic rats with D-alpha-tocopherol could prevent the changes in glomerular and tubular function commonly observed in this disease. Sixty male Wistar rats divided into four groups were studied: control (C), control treated with D-alpha-tocopherol (C + T), diabetic (D), and diabetic treated with D-alpha-tocopherol (D + T). Treatment with D-alpha-tocopherol (40 mg/kg every other day, ip) was started three days after diabetes induction with streptozotocin (60 mg/kg, ip).

Action of ANG II and ANP on colon epithelial cells.

The interaction of angiotensin II (ANG II) and atrial natriuretic peptide (ANP) on intracellular pH (pH(i)) and calcium ([Ca2+](i)) was investigated in T84 cells (a permanent cell line derived from human colon epithelium) using the fluorescent stains BCECF/AM and Fluo 4/AM, respectively. pH(i) recovery rate mediated by the Na(+)/H+ exchanger (NHE) was examined following an NH4Cl pulse. Under control conditions pH(i) recovered at 0.

Signaling path of the action of AVP on distal K+ secretion.

Background: Previous studies from our laboratory have shown that luminal perfusion with arginine vasopressin (AVP) stimulates distal tubule secretory potassium flux (JK) via V1 receptors (Am J Physiol 278:F809-F816, 2000). In the present work, we investigate the cell signaling mechanism of this process.

Methods: In vivo stationary microperfusion was performed in rat cortical distal tubules and luminal K+ was measured using double K+ resin/reference microelectrodes.

Arginine vasopressin stimulates H+-ATPase in MDCK cells via V1 (cell Ca2+) and V2 (cAMP) receptors.

The effect of arginine vasopressin (AVP) and/or atrial natriuretic peptide (ANP) on the regulation of intracellular pH (pHi) via H+-ATPase and of cytosolic calcium ([Ca2+]i) was investigated in Madin-Darby canine kidney (MDCK) cells by the fluorescent probes BCECF-AM and fluo-4-AM, respectively. The pHi recovery rate was examined after intracellular acidification following an NH4Cl pulse, in the presence of zero Na+ plus Schering 28080 (a specific inhibitor of H+-K+-ATPase). AVP (10-12-10-6 M) increased the rate of pHi recovery and [Ca2+]i in a dose-dependent manner.

Atrial natriuretic peptide impairs the stimulatory effect of angiotensin II on H+-ATPase.

Background: Angiotensin II (Ang II) action on H+-ATPase is not clearly defined, and may vary with renal tubule segment and hormonal doses being studied. Since an increase of cytosolic calcium ([Ca2+]i) can stimulate acid vesicle movement and exocytotic insertion of proton pumps, and it has been shown that Ang II increases [Ca2+]i while atrial natriuretic peptide (ANP) reduces it, there may be some interaction between Ang II and ANP in the regulation of intracellular pH (pHi) mediated by H+-ATPase.

Methods: The effects of Ang II and/or ANP on the regulation of pHi via H+-ATPase and of [Ca2+]i was investigated in Madin-Darby canine kidney cells (MDCK) by the fluorescent probes BCECF-AM and Fluo-4/AM, respectively.

Distal tubule bicarbonate transport.

The superficial cortical distal tubule, accessible to in vivo micropuncture and microperfusion, has been the site of a considerable number of investigations. An important fraction of renal tubule bicarbonate reabsorption, about 8 to 9% of the filtered load of this ion in the rat, takes place in this segment. The present review addresses several aspects of bicarbonate transport by distal tubule.

Peritubular AVP regulates bicarbonate reabsorption in cortical distal tubule via V(1) and V(2) receptors.

Peritubular arginine vasopressin (AVP) regulates bicarbonate reabsorption in the cortical distal tubule via V(1) and V(2) receptors. The dose-dependent effects of peritubular AVP on net bicarbonate reabsorption (J(HCO)) were evaluated by stationary microperfusion of in vivo early (ED; distal convoluted tubule) and late distal (LD; connecting tubule and initial collecting duct) segments of rat kidney, using double-barreled H(+)-sensitive, ion-exchange resin/reference (1 M KCl) microelectrodes. AVP (10(-11) M) perfused into peritubular capillaries increased J(HCO), compared with basal levels during intact capillary perfusion with blood, in ED and LD segments.

Effect of arginine vasopressin and ANP on intracellular pH and cytosolic free [Ca2+] regulation in MDCK cells.

Background: The effects of arginine vasopressin (AVP) on intracellular pH (pHi) are not clearly defined, and may vary with cell membrane surface and the hormonal doses being studied. Since cytosolic free calcium concentration ([Ca2+]i) has an important effect on cellular H+ extrusion and it was shown that AVP increases [Ca2+]i while atrial natriuretic peptide (ANP) reduces it, there may be some interaction between AVP and ANP during the regulation of pHi.

Methods: The effects of AVP and/or ANP on pHi and [Ca2+]i were investigated in Madin-Darby canine kidney (MDCK) cells by the fluorescent probes BCECF-AM and Fluo 4-AM, respectively.

Interaction of angiotensin II and atrial natriuretic peptide on pH(i) regulation in MDCK cells.

The effect of ANG II and atrial natriuretic peptide (ANP) on intracellular pH (pH(i)) and cytosolic free calcium concentration ([Ca(2+)](i)) was investigated in Madin-Darby canine kidney cells by using the fluorescent probes 2', 7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein-acetoxymethyl ester (AM) and fura 2-AM or fluo 4-AM. pH(i) recovery rate was examined in the first 2 min after the acidification of pH(i) with a NH(4)Cl pulse. In the control situation, the pH(i) recovery rate was 0.

Luminal arginine vasopressin stimulates Na(+)-H+ exchange and H(+)-ATPase in cortical distal tubule via V1 receptor.

Bicarbonate reabsorption was evaluated by stationary microperfusion of in vivo early distal (ED) and late distal (LD) segments of rat kidney. Intratubular pH was recorded by double-barreled H ion-exchange resin/reference (1 M KCl) microelectrodes for the determination of HCO3- reabsorption. In the presence of luminal arginine vasopressin (AVP, 10(-9) M), a significant increase in HCO3- reabsorption was observed both in ED (from 0.

Interactions of ANP and ANG II in tubular nephron acidification.

In order to examine the effects and the interaction of angiotensin II (ANG II, 1 pM) and atrial natriuretic peptide (ANP, 1 microM) on the kinetics of bicarbonate reabsorption in the rat middle proximal tubule, we performed in vivo experiments using a stopped-flow microperfusion technique with the determination of lumen pH by Sb microelectrodes. These studies confirmed that ANG II added to the luminal or peritubular capillary perfusion fluid stimulates proximal bicarbonate reabsorption and showed that ANP alone does not affect this process, but impairs the stimulation caused by ANG II. We also studied the effects and the interaction of these hormones in cortical distal nephron acidification.

Alterations of the renal handling of H+ in diabetic rats.

Renal acid excretion and proximal and distal nephron acidification were evaluated 20 days after induction of diabetes, in rats, by intraperitoneal injection of streptozotocin (45 mg/kg). Titratable acidity in urine was measured by microtitration and ammonium excretion (NH4+) by spectrophotometry. Proximal tubular acidification was evaluated by the kinetics of reabsorption of perfused HCO3-.

Effect of luminal angiotensin II and ANP on early and late cortical distal tubule HCO3- reabsorption.

Bicarbonate reabsorption was evaluated by stationary microperfusion "in vivo" early distal (ED) and late distal (LD) segments of at kidney. Intratubular pH was recorded by double-barreled of H+ exchange resin/reference (1 M KCl) microelectrodes for the determination of HCO3- reabsorption. In the presence of angiotensin II (ANG II) (10(-12) M), a significant increase in HCO3- reabsorption was observed both in ED (from 0.

Measurement of glomerular filtration rate using inulin prepared from Vernonia herbacea, a Brazilian native species.

Vernonia herbacea (Vell.) Rusby (Asteraceae) is a perennial herb native to the cerrado vegetation of tropical areas in Brazil, which accumulates inulin in the underground reserve organs. The aim of this paper was to determine whether the inulin extracted from V.