bluuebunny/crossref_metadata_2025 huggingface dataset:
Localization of the extracellular Ca2+/polyvalent cation-sensing protein in rat kidney
Authors: Riccardi Daniela , Hall Amy E. , Chattopadhyay Naibedya , Xu Jason Z. , Brown Edward M. , Hebert Steven C.
Published: December 2017

We previously identified transcripts encoding a G protein-coupled, extracellular calcium/polyvalent cation-sensing receptor, RaKCaR, in rat kidney (D. Riccardi, J. Park, W.-S. Lee, G. Gamba, E. M. Brown, and S. C. Hebert. Proc. Natl. Acad. Sci. USA 92: 131–135, 1994), which was proposed to provide the mechanism for modulating a variety of renal functions in response to changes in extracellular Ca2+ (E. M. Brown. In: Handbook of Physiology. Bethesda, MD: Am. Physiol. Soc., 1992, sect. 8, vol. 2, chapt. 39, p. 1841–1916; and S. C. Hebert. Kidney Int. 50: 2129–2139, 1996). Here, we examine the cellular and regional distribution of receptor protein by immunofluorescence microscopy using a polyclonal antibody raised against a 22 amino acid region of the NH2 terminus of the receptor. The most intense fluorescence was seen at the basolateral border of cortical thick ascending limb cells. Basolateral staining for the receptor was also detected in medullary thick ascending limbs, in macula densa cells identified by costaining with antibody to brain nitric oxide synthase, NOS-B1, and in distal convoluted tubule cells distinguished by costaining for the apical thiazide-sensitive Na+-Cl−cotransporter. Apical anti-RaKCaR staining was detected at the base of the brush border of proximal tubules with decreasing intensity from S1 to S3 segments. In cortical collecting ducts, anti-RaKCaR staining was detected in some, but not all, type A intercalated cells identified by costaining with anti-H+-ATPase and anti-AE1 Cl−/[Formula: see text]exchanger antibodies. The present study demonstrates that RaKCaR protein is expressed in many different nephron segments and that the polarity of receptor expression varies with cell type along the nephron. These results suggest potential roles for the extracellular Ca2+/polyvalent cation-sensing receptor in responding to both circulating and urinary concentrations of divalent minerals and potentially other polyvalent cations (e.g., aminoglycoside antibiotics) to modulate nephron function.
Dietary salt modulates renal production of transforming growth factor-β in rats
Authors: Ying Wei-Zhong , Sanders Paul W.
Published: December 2017

Transforming growth factors (TGF) are potent multifunctional polypeptides that are involved in renal function and glomerular sclerosis. We postulated that dietary salt modified renal production of TGF-β. An increase in dietary salt produced sustained increases in steady-state levels of mRNA for TGF-β1, -β2, and -β3 in the rat kidney. While serum concentration of TGF-β1 did not change, the 8.0% NaCl diet increased urinary excretion of TGF-β1, indicating enhanced renal production was the source of TGF-β1. Increasing urinary flow rates with diuretics did not further increase synthesis of TGF-β1 in animals receiving the 8.0% NaCl diet. The 8.0% NaCl diet increased production of TGF-β1 in both glomeruli and tubules, although active TGF-β1 was secreted in greater amounts only from glomeruli. Enhanced glomerular production of both inactive and active TGF-β1 induced by the 8.0% NaCl diet was inhibited by tetraethylammonium (TEA) and not glybenclamide. Cardiac production of TGF-β1 also increased on the 8.0% NaCl diet but was not affected by TEA. The results demonstrated that increased dietary salt augmented glomerular TGF-β production by a mechanism that included a TEA-sensitive potassium channel. Dietary salt, by facilitating glomerular expression of TGF-β, may directly promote development of glomerulosclerosis.
Downregulation of nitric oxide synthase in chronic renal insufficiency: role of excess PTH
Authors: Vaziri N. D. , Ni Z. , Wang X. Q. , Oveisi F. , Zhou X. J.
Published: December 2017

The available data on the effect of chronic renal failure (CRF) on nitric oxide (NO) metabolism are limited and contradictory. We studied rats with CRF 6 wk after a five-sixths nephrectomy and compared the results with those in the sham-operated controls, felodipine-treated CRF, and parathyroidectomized (CRF-PTX) animals. CRF was produced by surgical resection of the upper and lower thirds of the left kidney, followed by contralateral nephrectomy. We chose this model, as opposed to that produced by renal artery branch ligation, because the latter causes exuberant hypertension (HTN), which independently affects NO metabolism. The CRF group exhibited a mild HTN coupled with elevated basal platelet cytosolic Ca2+concentration ([Ca2+]i), blunted hypotensive response tol-arginine, decreased hypertensive response to NO synthase (NOS) inhibitor, NG-monomethyl-l-arginine, and normal hypotensive response to NO donor, sodium nitroprusside. This was associated with a significant reduction in urinary excretion of stable NO metabolites (NOX) and depressed NOS activity, as well as endothelial and inducible NO synthase (eNOS and iNOS, respectively) protein contents of thoracic aorta and the remnant kidney in the CRF animals. Calcium channel blockade and PTX lowered blood pressure, increased urinary NOX, and enhanced vascular NOS activity, as well as eNOS and iNOS protein expressions in the tested tissues. Thus CRF animals exhibited significant reductions in vascular NOS activity and eNOS and iNOS expressions. These abnormalities were reversed by calcium channel blockade and PTX, suggesting the possible causal role of CRF-induced dysregulation of [Ca2+]i.
Effect of growth hormone on renal and systemic acid-base homeostasis in humans
Authors: Sicuro Anita , Mahlbacher Katia , Hulter Henry N. , Krapf Reto
Published: December 2017

The effects of recombinant human growth hormone (GH, 0.1 U ⋅ kg body wt−1 ⋅ 12 h−1) on systemic and renal acid-base homeostasis were investigated in six normal subjects with preexisting sustained chronic metabolic acidosis, induced by NH4Cl administration (4.2 mmol ⋅ kg body wt−1 ⋅ day−1). GH administration increased and maintained plasma bicarbonate concentration from 14.1 ± 1.4 to 18.6 ± 1.1 mmol/l ( P < 0.001). The GH-induced increase in plasma bicarbonate concentration was the consequence of a significant increase in net acid excretion that was accounted for largely by an increase in renal [Formula: see text]excretion sufficient in magnitude to override a decrease in urinary titratable acid excretion. During GH administration, urinary pH increased and correlated directly and significantly with urinary[Formula: see text] concentration. Urinary net acid excretion rates were not different during the steady-state periods of acidosis and acidosis with GH administration. Glucocorticoid and mineralocorticoid activities increased significantly in response to acidosis and were suppressed (glucocorticoid) or decreased to control levels (mineralocorticoid) by GH. The partial correction of metabolic acidosis occurred despite GH-induced renal sodium retention (180 mmol; gain in weight of 1.8 ± 0.2 kg, P< 0.005) and decreased glucocorticoid and mineralocorticoid activities. Thus GH (and/or insulin-like growth factor I) increased plasma bicarbonate concentration and partially corrected metabolic acidosis. This effect was generated in large part by and maintained fully by a renal mechanism (i.e., increased renal NH3 production and[Formula: see text]/net acid excretion).
Anomalous decrease in dextran sulfate clearance in the diabetic rat kidney
Authors: Burne Melissa J. , Adal Yalçin , Cohen Neale , Panagiotopoulos Sianna , Jerums George , Comper Wayne D.
Published: December 2017

The anomalous increase in charge selectivity as previously observed with reduced dextran sulfate clearances in diabetic rats (L. D. Michels, M. Davidman, and W. F. Keane. Kidney Int. 21: 699–705, 1982) was confirmed in 4-wk streptozotocin (STZ) diabetic Sprague-Dawley rats using the isolated perfused kidney technique. The apparent charge selectivity in both control and diabetic rats could be abolished by increasing the dextran sulfate concentration to 200 μg/ml in the perfusate. This was demonstrated by a high rate of processing of dextran sulfate (∼1,700 ng ⋅ min−1⋅ kidney−1) by glomeruli in both control and diabetic kidneys and by the fact that charge interaction could not explain the concentration dependence. The amount of urinary desulfation of dextran sulfate was also found to be significantly less in the diabetic kidney as was glomerular sulfatase activity compared with controls. Dextran sulfate glomerular processing is therefore altered in the STZ diabetic rat kidney but could be rationalized in terms of previous models of endothelial cell receptor-mediated uptake of dextran sulfate. The results are consistent with recent work demonstrating that there is little or no electrostatic charge interaction operating on dextran sulfate or other negatively charged molecules at the glomerular capillary wall.
Stimulation of renin secretion by NO donors is related to the cAMP pathway
Authors: Kurtz Armin , Götz Karl-Heinz , Hamann Marlies , Kieninger Martin , Wagner Charlotte
Published: December 2017

This study aimed to characterize the cellular pathways along which nitric oxide (NO) influences the secretion of renin from the kidney. Using the isolated perfused rat kidney model, we found that the NO donor sodium nitroprusside (SNP) (1–30 μmol/l) induced a prompt, concentration-dependent fourfold increase of basal renin secretion. The membrane-permeable cGMP analogs 8-bromo-cGMP and 8-(4-chlorophenylthio)-cGMP (8-pCPT-cGMP; each 5–50 μmol/l) inhibited basal renin secretion and attenuated the stimulation of renin secretion by SNP. Conversely, the renin stimulatory effect of SNP was enhanced in the presence of the G kinase inhibitor Rp-8-CPT-cGMPS (10 μmol/l). The renin stimulatory effect of SNP was amplified in nominally calcium-free perfusate and was abolished in the presence of angiotensin II (1 nmol/l). Renin secretion stimulated by SNP was clearly attenuated by the A kinase inhibitor Rp-8-CPT-cAMPS (25 μmol/l). These findings indicate that the renin stimulatory effect of NO donors in renal juxtaglomerular cells cannot be explained by activation of G kinase and is also less likely to be causally related to the regulation of renin secretion by calcium. Because A kinase activity is required for the stimulation of renin secretion by SNP, it appears as if the renin stimulatory effect is causally related to the cAMP pathway controlling renin secretion.
A role for PKCε and MAP kinase in bradykinin-induced arachidonic acid release in rabbit CCD cells
Authors: Lal Mark A. , Proulx Pierre R. , Hébert Richard L.
Published: December 2017

Arachidonic acid (AA) release is the rate-limiting step in the production of prostaglandins, an important class of autocrine/paracrine factors that modulate collecting duct function. Previous results from this laboratory have established cytosolic phospholipase A2(cPLA2) as the enzyme responsible for bradykinin (BK)-stimulated AA mobilization in rabbit cortical collecting duct (RCCD) cells, and the present study pursues the intracellular signaling mechanisms responsible for its activation. Pretreatment of cells with Ro-31-8220, an inhibitor of protein kinase C (PKC), or PD-98059, an inhibitor of the mitogen-activated protein kinase (MAPK) cascade, resulted in a 50–60% reduction in BK-stimulated AA release. Incubation of RCCD cells with a combination of both Ro-31-8220 and PD-98059 did not achieve a greater inhibition of either BK-stimulated AA release or cPLA2activity, possibly indicating that MAPK activation was dependent upon prior activation of PKC. This was supported by the observation that BK-induced MAPK activation could be reversed by either inhibitor. Additional experiments dealing with immunoblots for PKC isozymes revealed that RCCD cells express PKC species α, γ, ε, and ζ. Following BK stimulation, only PKCε translocated to the particulate fraction. Based on these results, it appears that PKC is activated and involved in the sequential activation of MAPK and cPLA2following BK treatment. The results also suggest that PKCε may be the isozyme implicated in the process.
Renal potassium transport: mechanisms and regulation
Authors: Giebisch Gerhard
Published: December 2017

The regulation of potassium metabolism involves mechanisms for the appropriate distribution between the intra- and extracellular fluid compartments and for the excretion by the kidney. Clearance and single-nephron studies show that renal excretion is determined by regulated potassium secretion and potassium reabsorption, respectively, in principal and intercalated cells of the distal nephron. Measurement of the electrochemical driving forces acting on potassium transport across individual cell membranes and characterization of several ATPases and potassium channels provide insights into the transport and regulation of renal potassium excretion.
Angiotensin (AT1A) receptor-mediated increases in transcellular sodium transport in proximal tubule cells
Authors: Thekkumkara Thomas J. , Cookson Rochelle , Linas Stuart L.
Published: December 2017

Angiotensin II (ANG II), acting through angiotensin type 1A receptors (AT1A), is important in regulating proximal tubule salt and water balance. AT1Aare present on apical (AP) and basolateral (BL) surfaces of proximal tubule epithelial cells (PTEC). The molecular mechanism of AT1Afunction in epithelial tissue is not well understood, because specific binding of ANG II to intact PTEC has not been found and because a number of isoforms of AT receptors are present in vivo. To overcome this problem, we developed a cell line from opossum kidney (OK) proximal tubule cells, which stably express AT1A( Kd= 5.27 nM, Bmax= 6.02 pmol/mg protein). Characterization of nontransfected OK cells revealed no evidence of AT1AmRNA (reverse transcriptase-polymerase chain reaction analysis) or protein (125I-labeled ANG II binding studies) expression. In cells stably expressing AT1A, ANG II binding was saturable, reversible, and regulated by G proteins. Transfected receptors were coupled to increases in intracellular calcium and inhibition of cAMP. To determine the polarity of AT1Aexpression and function in proximal tubules, transfected cells were grown to confluence on membrane inserts under conditions that allowed selective access to AP or BL surfaces. AT1Awere expressed on both AP ( Kd= 8.7 nM, Bmax= 3.33 pmol/mg protein) and BL ( Kd= 10.1 nM, Bmax= 5.50 pmol/mg protein) surfaces. Both AP and BL AT1Areceptors underwent agonist-dependent endocytosis (AP receptor: t1/2= 7.9 min, Ymax= 78.5%; BL receptor: t1/2= 2.1 min, Ymax= 86.3%). In cells transfected with AT1A, ANG II caused time- and concentration-dependent increases in transepithelial22Na transport (2-fold over control at 20 min) by increasing Na/H exchange. In conclusion, we have established a stable proximal tubule cell line that expresses AT1Aon both AP and BL surfaces, undergoes agonist-dependent receptor endocytosis, and is functional, as evidenced by inhibition of cAMP and increases in cytosolic calcium mobilization and transepithelial sodium movement. This cell line should prove useful for understanding the molecular and biochemical regulation of AT1Aexpression and function in PTEC.
Effects of acute AT1receptor blockade by candesartan on arterial pressure and renal function in rats
Authors: Cervenka Ludek , Wang Chi-Tarng , Navar L. Gabriel
Published: December 2017

Experiments were performed on normal anesthetized rats to determine the effects of candesartan, a novel AT1receptor antagonist, on the arterial pressure and renal hemodynamic responses to bolus doses of angiotensin II (ANG II) and on renal hemodynamics and sodium excretion. Control arterial pressure responses to bolus ANG II doses of 10, 50, 100 and 1,000 ng were 26 ± 6, 54 ± 7, 57 ± 7, and 79 ± 7 mmHg; the decreases in cortical renal blood flow (CRBF), measured with laser-Doppler flowmetry, were 47 ± 9, 64 ± 8, 71 ± 6, and 82 ± 6%. The vasoconstrictor responses to ANG II up to 1,000 ng were completely blocked by candesartan doses of 1 and 0.1 mg/kg, whereas treatment with 0.01 mg/kg candesartan attenuated the arterial pressure and CRBF responses. The higher doses of candesartan (1 and 0.1 mg/kg) elicited rapid decreases in arterial pressure, leading to associated decreases in sodium excretion. Renal blood flow (RBF), glomerular filtration rate (GFR), and urine flow also decreased following treatment with candesartan at 1 mg/kg. In contrast, when candesartan was given at 0.01 mg/kg, which did not decrease arterial pressure significantly, there were significant increases in GFR (16 ± 4), RBF (9 ± 2), urine flow (11 ± 2), sodium excretion (35 ± 7), and fractional sodium excretion (39 ± 8%). The inability to overcome blockade, even with very high ANG II doses, indicates that candesartan is a potent noncompetitive blocker of ANG II pressor and renal vasoconstrictor effects. The lower candesartan dose that did not cause significant hypotension elicited substantial increases in RBF, GFR, and sodium excretion, revealing the direct renal vasodilator and natriuretic effects of AT1receptor blockade.
Nitric oxide increases the activity of the apical 70-pS K+channel in TAL of rat kidney
Authors: Lu Ming , Wang Xiaohong , Wang Wenhui
Published: December 2017

We have previously shown that nitric oxide (NO) mediates the stimulatory effect of angiotensin II on the apical 70-pS K+channel in the thick ascending limb (TAL) of Henle’s loop of the rat kidney (12). In the present study, we used the patch-clamp technique to examine the effects of NO on the 70-pS K+channel. Addition of 10 μM S-nitroso- N-acetylpenicillamine (SNAP), a NO donor, increased the channel activity in cell-attached patches. In contrast, application of 100 μM Nω-nitro-l-arginine methyl ester (l-NAME), an inhibitor of nitric oxide synthase (NOS), reduced the channel activity by 75 ± 7%. The effect of l-NAME was the result of inhibiting NOS, since d-NAME, which does not block NOS activity, had no effect on the channel activity. In addition, the effect ofl-NAME was abolished in the presence of 1 mM l-arginine or by addition of 10 μM SNAP, further supporting the role of NO. Finally, the l-NAME-induced inhibition was also reversed by adding 8-bromoguanosine 3′,5′-cyclic monophosphate (8-BrcGMP). That the effect of NO is mediated by the cGMP-dependent pathway is also suggested by experiments in which inhibition of guanylate cyclase abolished the effect of SNAP. Finally, 10 μM SNAP significantly increased cGMP concentration of the medullary TAL from 12.4 fM/μg protein to 38.9 fM/μg protein, as measured with ELISA. We conclude that NO is involved in regulating the activity of the apical 70-pS K+channel in the TAL of the rat kidney.
Monoclonal antibody P-31 recognizes a novel intermediate filament-associated protein (p250) in rat podocytes
Authors: Kurihara Hidetake , Sunagawa Norio , Kobayashi Tatsuo , Kimura Kazuhiro , Takasu Nobuo , Shike Tsutomu
Published: December 2017

The visceral glomerular epithelial cells (GECs) or podocytes of the renal glomerulus constitute a highly specialized epithelium. To study the nature of podocytes, we established mouse monoclonal antibodies against GEC. Clone P-31 reacted exclusively with the cytoplasm of GEC by immunofluorescence. Immunoblot analysis with P-31 showed that a single band of 250 kDa was detectable in a glomerular lysate. The 250-kDa polypeptide (p250) was recovered from Triton X-100-insoluble fractions of isolated glomeruli, suggesting that this molecule is associated with the cytoskeleton. Immunogold staining with P-31 demonstrated that the gold particles were located at the intersections of vimentin-type intermediate filaments of podocytes. In developing kidney, this protein first appeared in immature GECs during the S-shaped body stage. In puromycin aminonucleoside nephrosis, p250 was dramatically increased in glomeruli where enhanced desmin expression was observed in GECs. These results indicate that p250 is a novel intermediate filament-associated protein and plays a role in the organization of the intermediate filament network in both normal and diseased conditions.
P2 purinoceptor localization along rat nephron and evidence suggesting existence of subtypes P2Y1and P2Y2
Authors: Cha Seok Ho , Sekine Takashi , Endou Hitoshi
Published: December 2017

Effects of extracellular ATP on intracellular free calcium concentration ([Ca2+]i) were examined in rat single nephron segments using the fura 2-AM. ATP (10 μM) induced a significant transient increase in [Ca2+]iin the glomerulus, the early proximal convoluted tubule (S1), the cortical collecting tubule (CCT), and the outer medullary collecting tubule (OMCT). The magnitude of the response was the greatest in the OMCT among four segments. ATP induced an increase in the [Ca2+]iin a dose-dependent manner in S1 and OMCT. In the OMCT, ATP caused a biphasic increase in [Ca2+]iconsisting of an initial rapid rise and a sustained phase. Removal of calcium from the medium resulted in an attenuation of the sustained phase of [Ca2+]iand an ∼30% reduction in the height of the initial [Ca2+]ipeak in response to 10 μM ATP. Effects of ATP, its analogs, and its metabolites were tested in the S1 and OMCT. ATP, 2-methylthio-ATP (2-MeS-ATP), ADP, and UTP increased [Ca2+]idose dependently. AMP and adenosine did not affect [Ca2+]iin the S1 and OMCT. The ATP- or 2-MeS-ATP-induced [Ca2+]iincrease was inhibited by the pretreatment of the S1 and OMCT with suramin or reactive blue 2. Neomycin, a phospholipase C inhibitor, attenuated the ATP-induced [Ca2+]iincrease. To investigate the hormonelike action of ATP in OMCT, a heterologous cross desensitization was performed. The pretreatment of OMCT with ATP inhibited increases in vasopressin-, ANG II-, endothelin-1-, or bradykinin-induced [Ca2+]iincrease. These findings suggest that ATP might affect the above peptidyl agonist-activated calcium mobilizations.
Ornithine decarboxylase along the mouse and rat nephron
Authors: Levillain Olivier , Hus-Citharel Annette
Published: December 2017

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 withoutd,l-2-(difluoromethyl)ornithine (DFMO), a selective inactivator of ODC. Tubules from either control or DFMO-treated animals were incubated with 100 μMl-[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.
Physiological disposal of the potential alkali load in diet of the rat: steps to achieve acid-base balance
Authors: Lin Shih-Hua , Cheema-Dhadli Surinder , Chayaraks Sorasak , Chen Ching-Bun , Gowrishankar Manjula , Halperin Mitchell L.
Published: December 2017

The purpose of this study was to provide a better understanding of the physiological role of endogenous net organic acid production in rats consuming their usual diet. Balance studies were performed over 24 h, and urine was collected in the day and night portions of the diurnal cycle. A supplemented low-electrolyte diet (LED) was fed to determine whether urinary organic anions were identical to those in the diet. A titration procedure was developed to determine the p K of titratable groups in the urine of rats studied with and without an acid load. Although normal rats excreted net acid (NAE), the latter was inversely related to the amount of food consumed. The rates of excretion of bicarbonate ([Formula: see text]), citrate, unmeasured organic anions, and [Formula: see text] were higher in the night portion of the diurnal cycle. NAE rose dramatically when alkali intake was decreased by consuming the LED. Dietary and urinary organic anions were not identical because rats fed the LED supplemented with potassium citrate excreted <10% of this alkali load as citrate and <25% as[Formula: see text]. In the 24 h after 3,000 μmol NH4Cl was given intraperitoneally, H+did not appear to be retained, yet NAE rose by only close to 2,000 μeq. The rate of excretion of titratable groups with a p K in the 3 to 5 pH range fell by close to 1,000 μeq; most of these changes occurred in the first 7 h after NH4Cl was given. We conclude that rat chow provides a large net alkali load. There appear to be two types of endogenous acid production, a form associated with a rise in NAE (e.g., sulfuric acid) and dietary alkali-driven endogenous net acid production, which titrates this alkali. Renal excretion of organic anions makes these acids end products of metabolism.
Elevated glucose increases mesangial cell sensitivity to insulin-like growth factor I
Authors: Horney Mark J. , Shirley David W. , Kurtz David T. , Rosenzweig Steven A.
Published: December 2017

To determine the effects of glucose on insulin-like growth factor I (IGF-I)-induced mesangial cell (MC) proliferation, we have examined the relationships between IGF binding protein 2 (IGFBP-2) secretion and proliferation in murine MCs (MMCs). MMCs incubated in high glucose (HG, 25 mM) exhibited a 25–30% reduction in IGFBP-2 secretion compared with cells in normal glucose (NG, 5.6 mM). This loss was not due to cell surface binding; it correlated with a 3.1-fold decrease in IGFBP-2 mRNA. IGFBP-2 secretion was stimulated by IGF-I in NG but was unaltered in HG. Insulin treatment yielded similar results at 10-fold higher doses, indicating that this response is IGF-I receptor dependent. MMCs in HG displayed increased IGF-I-stimulated insulin receptor substrate-1/2 phosphorylation and activator protein-1 transcriptional activity compared with NG controls. Accordingly, although IGF-I was not proliferative in NG, it increased [3H]thymidine incorporation and cell number in HG to an extent proportional to the decrease in IGFBP-2. Thus hyperglycemia, as seen in diabetes, may increase MC IGF-I sensitivity by reducing IGFBP-2 expression, in turn increasing its proliferative and secretory responses and contributing to the development of diabetic glomerulosclerosis.
Characterization of the Hsp110/SSE gene family response to hyperosmolality and other stresses
Authors: Santos Bento C. , Chevaile Alejandro , Kojima Ryoji , Gullans Steven R.
Published: December 2017

Hsp110, Osp94, and Hsp70RY are members of the recently described Hsp110/SSE subfamily of (heat and osmotic) stress proteins whose members are structurally related to the Hsp70/BiP gene superfamily. To date, little is known about the response of this gene family to stresses in vitro or in vivo. In this study, an analysis of mRNA expression showed that Hsp110 and Osp94, like Hsp70, are induced in renal murine inner medullary collecting duct (mIMCD3) epithelial cells by heat shock, hyperosmotic NaCl, and cadmium, whereas low pH had a suppressive effect on Osp94. H2O2decreased expression of Osp94 while inducing levels of Hsp110 and Hsp70 message. Tunicamycin, hypertonic urea, and tumor necrosis factor-α had no effects. Hsp70RY was responsive exclusively to cadmium chloride. Moreover, enhanced expression of Hsp110 and Osp94 was subsequent to induction of Hsp70 and was suppressed by inhibition of protein synthesis by cycloheximide. RT-PCR analysis showed Hsp110, Osp94, and Hsp70RY are ubiquitously expressed in mouse tissues. In murine kidney, there was a corticomedullary gradient of expression of Hsp110, Osp94, Hsp70RY, and Hsp70 but not Hsc70 or BiP. Furthermore, dehydration increased inner medullary expression of Hsp110 and Osp94. An analysis of stress tolerance in mIMCD3 cells showed that heat shock and hyperosmotic NaCl stress are cross-tolerant stresses, suggesting hyperosmolality is a physiological correlate of heat shock in mammalian kidney. Thus Hsp110 and Osp94 behave as heat shock proteins, although they are regulated differently than Hsp70.
Nephrogenesis and angiotensin II receptor subtypes gene expression in the fetal lamb
Authors: Gimonet Valérie , Bussieres Laurence , Medjebeur Anissa A. , Gasser Bernard , Lelongt Brigitte , Laborde Kathleen
Published: December 2017

To investigate the role of angiotensin II (ANG II) in nephrogenesis, a developmental study of renal AT1and AT2receptor mRNA expression was performed in parallel with the quantitative and qualitative analysis of metanephros development in fetal lamb from 60 to 140 days of gestation. Both ANG II receptor subtypes were expressed early during nephrogenesis but displayed specific spatial and temporal distribution during gestation. High-AT2mRNA expression took place in the outermost nephrogenic area and in the undifferentiated mesenchymal cells surrounding the ampulla; level of AT2expression in this localization followed closely glomeruli proliferation rate and disappeared after nephrogenesis completion (>120 days). AT2mRNA was also detected in the differentiated epithelial cells of macula densa of maturing glomeruli. Although most of AT1mRNA labeling was found in the mesangial cells of maturing glomeruli, where it persisted after nephrogenesis completion, additional labeling was found in undifferentiated cells, in cells invading the inferior cleft of S-shaped bodies (80 days), and in medullar cells between tubules (120 days). Our results suggest that each receptor subtype has a specific role in renal morphogenesis, i.e., AT2in mesenchymal proliferation or apoptosis and AT1in vascular smooth muscle cells differentiation.
Neuropeptide Y shifts equilibrium between α- and β-adrenergic tonus in proximal tubule cells
Authors: Holtbäck Ulla , Ohtomo Yoshiyuki , Förberg Petter , Sahlgren Bo , Aperia Anita
Published: December 2017

Renal sympathetic nerves play a central role in the regulation of tubular Na+reabsorption. Norepinephrine (NE) and neuropeptide Y (NPY) are colocalized in renal sympathetic nerve endings. The purpose of this study is to examine the integrated effects of these neurotransmitters on the regulation of Na+-K+-ATPase, the enzyme responsible for active Na+reabsorption in renal tubular cells. Studies were performed on proximal tubular segments, which express adrenergic α- and β-receptors, as well as NPY-Y2receptors. It was found that α- and β-adrenergic agonists had opposing effects on Na+-K+-ATPase activity. β-Adrenergic agonists induced a dose-dependent inhibition of the Na+-K+-ATPase activity, whereas α-adrenergic agonists stimulated the enzyme. NPY abolished β-agonist-induced deactivation of Na+-K+-ATPase and enhanced α-agonist-induced activation of Na+-K+-ATPase. The β-adrenergic agonist appeared to inhibit Na+-K+-ATPase activity via a cAMP pathway. NPY antagonized β-agonist-induced accumulation of cAMP. In our preparation, NE alone had no net effect but stimulated the Na+-K+-ATPase activity in the presence of β-adrenergic antagonists, as well as in the presence of NPY. The results indicate that, in renal tissue, NPY determines the net effect of its colocalized transmitter, NE, by its ability to attenuate the β- and enhance the α-adrenergic effect.
Dynamic autoregulation in the in vitro perfused hydronephrotic rat kidney
Authors: Cupples William A. , Loutzenhiser Rodger D.
Published: December 2017

Renal autoregulation is mediated by tubuloglomerular feedback, operating at 0.03–0.05 Hz, and a faster system, operating at 0.1–0.2 Hz, that has been attributed by exclusion to myogenic vasoconstriction. In this study, we examined dynamic autoregulation in the hydronephrotic rat kidney, which lacks tubuloglomerular feedback but exhibits pressure-induced afferent arteriolar vasoconstriction. Kidneys were harvested under anesthesia from Sprague-Dawley rats and perfused in vitro using defined, colloid-free medium. Renal perfusate flow was assessed during forced pressure fluctuations at mean pressures of 60–140 mmHg. Transfer function analysis revealed passive behavior at 60 mmHg and active, pressure-dependent responses at higher pressures. In all cases, coherence was high (0.89 ± 0.03 between 0.01 and 0.9 Hz). There was a resonance peak in admittance gain at ≈0.3 Hz and an associated broad peak in phase angle. Below this frequency, gain declined progressively. The minimum gain achieved at 0.01–0.05 Hz was pressure sensitive, being 1.08 ± 0.02 at 60 mmHg and 0.71 ± 0.04 at 140 mmHg. These findings are consistent with in vivo results and with model-based predictions of the dynamics of myogenic autoregulation, supporting the postulate that the rapid component of autoregulation reflects operation of a myogenic mechanism.
Dietary salt enhances glomerular endothelial nitric oxide synthase through TGF-β1
Authors: Ying Wei-Zhong , Sanders Paul W.
Published: December 2017

Dietary salt controls production of nitric oxide (NO), a potent paracrine relaxation factor involved in glomerular filtration and salt excretion. We hypothesized that glomerular NO production was enhanced through endothelial nitric oxide synthase (NOS3). Rats in metabolic cages were studied after 4 days on 0.3% (Lo-salt) or 8.0% (Hi-salt) NaCl diet. Steady-state mRNA and protein levels of NOS3 and calcium-dependent NO production of isolated glomeruli from Hi-salt animals were greater than those values observed in glomeruli from Lo-salt rats. Because dietary salt enhanced glomerular production of transforming growth factor-β1 (TGF-β1) [W.-Z. Ying and P. W. Sanders. Am. J. Physiol. 274 ( Renal Physiol. 43): F635–F641, 1998], studies were then conducted to examine the interaction between NOS3 and TGF-β1. Glomerular steady-state levels of mRNA of NOS3 and TGF-β1 directly correlated ( r2= 0.946; P < 0.0001). A neutralizing antibody to TGF-β reduced NOS3 protein and NO production in cultured glomeruli from Hi-salt animals to levels seen in the Lo-salt glomeruli. Thus dietary salt increased glomerular expression of TGF-β1, which in turn augmented NO production through NOS3.
Effect of norepinephrine on intracellular pH in kidney proximal tubule: role of Na+-( HCO 3 − ) n cotransport
Authors: Abdulnour-Nakhoul Solange , Khuri Raja N. , Nakhoul Nazih L.
Published: December 2017

We examined the effect of norepinephrine (NE) on intracellular pH (pHi) and activity of Na+([Formula: see text]) in the isolated perfused kidney proximal tubule of Ambystoma, using single-barreled voltage and ion-selective microelectrodes. In control[Formula: see text] Ringer, addition of 10−6 M NE to the bath reversibly depolarized the basolateral membrane potential ( V 1), the luminal membrane potential ( V 2), and the transepithelial potential difference ( V 3) and increased pHi by 0.14 ± 0.02. These effects were mimicked by isoproterenol but were abolished after pretreatment with SITS or in the absence of CO2/[Formula: see text]. Removal of bath Na+ depolarized V 1 and V 2, hyperpolarized V 3, and decreased pHi. These effects are largely mediated by the electrogenic Na+-([Formula: see text]) n cotransporter. In the presence of NE, the effects of Na+ removal on membrane potential differences and the rate of change of pHi were significantly smaller. Reducing bath [Formula: see text] concentration from 10 to 2 mM at constant CO2 (pH 6.8) depolarized V 1 and V 2, decreased pHi, and lowered[Formula: see text]. These changes are also due to Na+-([Formula: see text]) n . In the presence of NE, reducing bath [[Formula: see text]] caused a smaller depolarizations of V 1 and V 2, and the rate of pHi decrease was significantly reduced. Our results indicate: 1) NE causes an increase in pHi; 2) the NE-induced alkalinization is mediated by a SITS-sensitive and[Formula: see text]-dependent transporter on the basolateral membrane; and 3) in the presence of NE, the reduced effects caused by basolateral[Formula: see text] changes or Na+ removal are indicative of an inhibitory effect of NE on Na+-([Formula: see text]) n cotransport.
Effect of dietary salt on neuronal nitric oxide synthase in the inner medullary collecting duct
Authors: Roczniak Agnes , Zimpelmann Joseph , Burns Kevin D.
Published: December 2017

Nitric oxide (NO) derived from neuronal NO synthase (nNOS) in the kidney inner medulla has been implicated in the regulation of arterial blood pressure. The purpose of the present study was to determine the effect of high dietary NaCl on the expression of nNOS in the rat inner medullary collecting duct (IMCD). After 3 days or 3 wk of high (4.0%)-NaCl diet in rats, urinary[Formula: see text]/[Formula: see text]excretion significantly increased. In freshly microdissected IMCD, nNOS was readily detected by immunofluorescence with polyclonal antibody, an effect that was completely blocked by neutralization of antibody with immunizing antigen. In rats fed a 4.0% NaCl diet for 3 days, IMCD nNOS mRNA, detected by RT-PCR, did not change from control values (0.3% NaCl, 19.84 ± 1.57 × 103, vs. 4.0% NaCl, 20.44 ± 3.14 × 103cpm; P = not significant, n = 3). By Western blotting however, nNOS protein expression significantly increased (0.3% NaCl, 0.51 ± 0.12, vs. 4.0% NaCl, 0.92 ± 0.14 arbitrary units; P < 0.05, n = 5). After 3 wk of 4.0% dietary NaCl, expression of nNOS mRNA and protein in IMCD did not differ significantly from control values. In contrast to these data, renal cortical expression of nNOS mRNA and protein was significantly decreased after 4.0% NaCl diet for 3 days. High dietary NaCl had no significant effect on expression of mRNA for inducible NO synthase (iNOS) in IMCD after either 3 days or 3 wk. In summary, our data indicate that nNOS mRNA and protein are expressed in IMCD and that high dietary NaCl differentially regulates nNOS expression in IMCD and cortex. The early increase in nNOS protein in IMCD may contribute to enhanced local production of NO and thereby represent an adaptive response to salt intake.
Renal handling of circulating nitrates in anesthetized dogs
Authors: Godfrey Murrell , Majid Dewan S. A.
Published: December 2017

Nitric oxide (NO) is rapidly oxidized to nitrite ([Formula: see text]) and then to nitrate ([Formula: see text]) in biological tissues. Although urinary excretion rates of[Formula: see text] are often used as an index of NO production in the body, very little is known regarding the kidney’s ability to excrete circulating [Formula: see text]. We have evaluated the renal responses to systemic administration of sodium nitrate (NaNO3) in eight anesthetized dogs treated with the NO synthase inhibitor, nitro-l-arginine (NLA; 50 μg ⋅ kg−1⋅ min−1), intrarenally to minimize renal production of NO. Urinary and plasma concentrations of[Formula: see text]/[Formula: see text](NOX) were determined by the Greiss reaction after enzymatic reduction of [Formula: see text] to[Formula: see text]. NLA treatment alone resulted in reductions in urinary NOX excretion rates (UNOXV, 1.13 ± 0.2 to 0.53 ± 0.1 nmol ⋅ min−1⋅ g−1) and an increase in fractional reabsorption of NOX (FRNOX, 93.8 ± 0.6 to 97 ± 0.6%) without changes in arterial plasma concentrations (ANOX, 18.7 ± 1.4 to 21.2 ± 3.7 μM). Administration of NaNO3(10, 20, 30, and 40 μg ⋅ kg−1⋅ min−1) resulted in dose-dependent increases in ANOX(34.5 ± 8.0, 46.4 ± 7.3, 60.7 ± 6.3, and 78.1 ± 6.3 μM), UNOXV (1.8 ± 0.7, 4.2 ± 1.8, 7.0 ± 2.0, and 11.4 ± 3.3 nmol ⋅ min−1⋅ g−1), and decreases in FRNOX(93.8 ± 2.3, 90.3 ± 3.5, 88.6 ± 3.2, and 84.6 ± 3.5%). Absolute net tubular reabsorption of [Formula: see text] showed a linear relationship with filtered loads, with no evidence of a transport maximum. These data show that, in the absence of additions from intrarenal sources, urinary excretion rates of nitrate increases progressively in response to increases in its circulating levels without exhibiting a transport maximum but with progressive decreases in fractional reabsorption.
Low-NaCl diet increases H-K-ATPase in intercalated cells from rat cortical collecting duct
Authors: Silver Randi B. , Choe Han , Frindt Gustavo
Published: December 2017

Extracellular K+-dependent H+extrusion after an acute acid load, an index of H/K exchange, was monitored in intercalated cells (ICs) from rat cortical collecting tubule (CCT) using ratiometric fluorescence imaging of the intracellular pH (pHi) indicator, 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). The hypothesis tested was that 12- to 14-day NaCl deprivation increases H-K-ATPase in rat ICs. The rate of H/K exchange in the low-NaCl ICs was double that of controls. In control ICs, H/K exchange was inhibited by Sch-28080 (10 μM). In the low-NaCl ICs, it was partially blocked by Sch-28080 or ouabain (1 mM). Simultaneous addition of both inhibitors abolished K-dependent pHirecovery. The induced H/K exchange observed with NaCl restriction was not due to elevated plasma aldosterone as evidenced by experiments on ICs from rats implanted with osmotic minipumps administering aldosterone such that plasma levels were comparable to those of NaCl-deficient rats. The results suggest that NaCl deficiency induces two isoforms of H-K-ATPase in ICs and that this effect is not mediated by elevated plasma aldosterone.
Natriuretic and kaliuretic activities of guanylin and uroguanylin in the isolated perfused rat kidney
Authors: Fonteles Manasses C. , Greenberg Richard N. , Monteiro Helena S. A. , Currie Mark G. , Forte Leonard R.
Published: December 2017

Guanylin and uroguanylin are novel peptides that activate membrane guanylate cyclases found in the kidney and intestine. We compared the effects of these peptides in the isolated perfused rat kidney. Both peptides are natriuretic and kaliuretic in this preparation. Uroguanylin (0.19–1.9 μM) increased glomerular filtration rate from 0.77 ± 0.07 to 1.34 ± 0.3 ml ⋅ g−1⋅ min−1at the highest concentration. A maximal increase in Na+excretion was achieved at 0.66 μM uroguanylin, with a reduction in fractional Na+reabsorption from 78.7 ± 1.7 to 58.8 ± 4.4%. The highest dose of uroguanylin increased kaliuresis by 50%. Osmolar clearance doubled at the highest concentration of uroguanylin tested ( P< 0.05). Guanylin also elicited a natriuresis and kaliuresis but appeared to be less potent than uroguanylin. The highest concentration of guanylin (1.3 μM) decreased fractional Na+reabsorption from 73.9 ± 2.4 to 64.5 ± 4.0%, but lower doses were ineffective. Guanylin stimulated urine K+excretion at the lowest concentration tested (0.33 μM) without any effect on Na+excretion. These peptides may influence salt and water homeostasis by biological effects in the kidney that are mediated by the intracellular second messenger, cGMP.
Thromboxane A2modulates the fibrinolytic system in glomerular mesangial cells
Authors: Coffman Thomas M. , Spurney Robert F. , Mannon Roslyn B. , Levenson Richard
Published: December 2017

We examined the effects of thromboxane A2(TxA2) on the activities of the plasminogen-plasmin system in glomerular mesangial cells. When mesangial cells are exposed to the TxA2agonist U-46619, a substantial increase in production of plasminogen activator inhibitor-1 (PAI-1) protein is observed that is significantly greater than that induced by 10% serum alone. This increase in PAI-1 protein production is accompanied by an increase in steady-state levels of PAI-1 mRNA. This stimulation is specifically mediated by TxA2(thromboxane prostanoid, TP) receptors, since U-46619 also stimulates PAI-1 expression in cells that are transfected with TP receptors, and this stimulation of PAI-1 production is completely blocked by the TxA2receptor antagonist, SQ-29,548. Despite the increase in PAI-1 production, there was net stimulation of plasmin activity in the medium of mesangial cells that had been exposed to U-46619. Furthermore, U-46619 also caused an increase in tissue plasminogen activator (tPA) mRNA levels. Thus TxA2stimulates the production of PAI-1 and plasminogen activators by mesangial cells through a receptor-dependent mechanism. In inflammatory renal diseases, the balance of these effects may modulate glomerular thrombosis and renal fibrosis.
Acute effects of vasopressin V2-receptor antagonist on kidney AQP2 expression and subcellular distribution
Authors: Christensen Birgitte Mønster , Marples David , Jensen Uffe Birk , Frøkiaer Jørgen , Sheikh-Hamad David , Knepper Mark , Nielsen Søren
Published: December 2017

The acute effect of treatment with the vasopressin V2-receptor antagonist OPC-31260 (OPC) on aquaporin-2 (AQP2) distribution and expression in rat kidney was examined. Immunofluorescence and semi-quantitative immunoelectron microscopy revealed that 15 and 30 min of OPC treatment resulted in significant reduction in apical plasma membrane labeling of AQP2, with a concomitant increase in labeling of vesicles and multivesicular bodies. In parallel, OPC treatment induced a large increase in urine output [0.6 ± 0.2 vs. 8.3 ± 1.0 ml/h ( n = 4)]. Northern blotting using a32P-labeled AQP2 cDNA probe and a digoxigenin-labeled AQP2 RNA probe revealed a band of ∼1.6 kb corresponding to the predicted size of AQP2 mRNA. In control experiments, thirsting increased, whereas water loading decreased AQP2 mRNA levels. Treatment of rats with OPC caused a significant reduction in AQP2 mRNA within 30 min (52 ± 21%, n = 8, P < 0.025) and 60 min (56 ± 7%, n = 4, P < 0.001) of treatment compared with intravenous saline-injected controls. Thus a very rapid reduction in AQP2 mRNA was observed in response to vasopressin-receptor antagonist treatment. The reduction in AQP2 mRNA persisted after 24 h (40 ± 17%, n = 5, P < 0.05) of OPC treatment. There was a parallel increase in diuresis and reduction in urine osmolality. In conclusion, V2-receptor blockade produced a rapid internalization of AQP2 parallel with a rapid increase in urine output. Furthermore, OPC treatment caused a rapid and significant reduction in AQP2 mRNA expression, demonstrating that for rapid regulation of AQP2 expression, modulation of AQP2 mRNA levels is regulated via vasopressin-receptor signaling pathways.
Roles of Na-K-2Cl and Na-Cl cotransporters and ROMK potassium channels in urinary concentrating mechanism
Authors: Hebert Steven C.
Published: December 2017

The cloning of the apical transport proteins involved in NaCl absorption in the thick ascending limb of Henle and the distal convoluted tubule has ushered in a new era that promises to provide molecular details of the urinary concentrating and diluting processes. We now have at hand tools to examine the regulation of these proteins that can affect the concentrating and diluting capacity.
Cellular mechanisms of aquaporin trafficking
Authors: Brown Dennis , Katsura Toshiya , Gustafson Corinne E.
Published: December 2017

Aquaporins (AQPs) are a family of functionally important water channel proteins that are of special cell biological interest because of their diverse intracellular targeting and trafficking properties. AQPs have been found in many different cells and tissues. This short review summarizes recent work that addresses the regulation of AQP2 trafficking in response to vasopressin.