Abstract P269: Sodium Transporter Profile in Mice Lacking AT1A Receptors in the Renal Proximal Tubule

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Alicia A McDonough ◽  
Audrey K Izuhara ◽  
Zhidan Xiang ◽  
Donna L Ralph ◽  
Rishav Adhikari ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Renal Cortex ◽  
Immunoblot Analysis ◽  
Renal Proximal Tubule ◽  
Volume Flow ◽  
Thick Ascending Limb ◽  
Wild Type ◽  
Fluid Reabsorption ◽  
Lower Baseline ◽  
Sodium Transporter

We have reported that mice lacking AT1A receptors (KO) in renal proximal tubule (PT) have 10 mmHg lower baseline BP and less PT fluid reabsorption than wild type (WT). We tested the hypothesis that the lower BP is associated with less abundant renal Na transporters or regulators. Homogenates of renal cortex and medulla (n=6/group) were prepared and 1 and 1/2 protein amounts of each subjected to immunoblot analysis with specific antibodies and quantitated. Results for cortex and medulla, displayed as mean +/- SEM, normalized to mean abundance of WT=1 (*p <0.05), are summarized in figures. In KO vs. WT abundance of PT NHE3, the associated motor myosin VI, the paracellular NaCl transporter claudin 2, and the Na-HCO3 transporter NBCe1 are lower in KO; in the thick ascending limb (TAL) NKCC2 and its associated kinase SPAK are less abundant, and there is a tendency for lower DCT NCC and CCD ENaC in KO. The results support our hypothesis and suggest that KO of PT AT1R reduces transport routes not only in the PT but beyond the PT, in spite of increased volume flow from PT and lower BP.

A γGT-AT1A receptor transgene protects renal cortical structure in AT1 receptor-deficient mice

2004 ◽  
Vol 18 (3) ◽  
pp. 290-298 ◽  
Author(s):  
Thu H. Le ◽  
Michael I. Oliverio ◽  
Hyung-Suk Kim ◽  
Harmony Salzler ◽  
Rajesh C. Dash ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Transgenic Mice ◽  
Proximal Tubule ◽  
Physiological Role ◽  
Renal Proximal Tubule ◽  
Hypoxanthine Phosphoribosyl Transferase ◽  
Wild Type ◽  
Specific Expression ◽  
Low Levels ◽  
Deficient Mice

To understand the physiological role of angiotensin type 1 (AT1) receptors in the proximal tubule of the kidney, we generated a transgenic mouse line in which the major murine AT1 receptor isoform, AT1A, was expressed under the control of the P1 portion of the γ-glutamyl transpeptidase (γGT) promoter. In transgenic mice, this promoter has been shown to confer cell-specific expression in epithelial cells of the renal proximal tubule. To avoid random integration of multiple copies of the transgene, we used gene targeting to produce mice with a single-copy transgene insertion at the hypoxanthine phosphoribosyl transferase ( Hprt) locus on the X chromosome. The physiological effects of the γGT-AT1A transgene were examined on a wild-type background and in mice with targeted disruption of one or both of the murine AT1 receptor genes ( Agtr1a and Agtr1b). On all three backgrounds, γGT-AT1A transgenic mice were healthy and viable. On the wild-type background, the presence of the transgene did not affect development, blood pressure, or kidney structure. Despite relatively low levels of expression in the proximal tubule, the transgene blunted the increase in renin expression typically seen in AT1-deficient mice and partially rescued the kidney phenotype associated with Agtr1a−/− Agtr1b−/− mice, significantly reducing cortical cyst formation by more than threefold. However, these low levels of cell-specific expression of AT1 receptors in the renal proximal tubule did not increase the low blood pressures or abolish sodium sensitivity, which are characteristic of AT1 receptor-deficient mice. Although our studies do not clearly identify a role for AT1 receptors in the proximal tubules of the kidney in blood pressure homeostasis, they support a major role for these receptors in modulating renin expression and in maintaining structural integrity of the renal cortex.

Micropuncture analysis of single-nephron function in NHE3-deficient mice

1999 ◽  
Vol 277 (3) ◽  
pp. F447-F453 ◽  
Author(s):  
John N. Lorenz ◽  
Patrick J. Schultheis ◽  
Timothy Traynor ◽  
Gary E. Shull ◽  
Jürgen Schnermann
Keyword(s):  
Proximal Tubule ◽  
Filtration Rate ◽  
Distal Tubule ◽  
Transport Processes ◽  
Tubuloglomerular Feedback ◽  
Thick Ascending Limb ◽  
Fluid Reabsorption ◽  
Fluid Delivery ◽  
Single Nephron ◽  
Flow Pressure

The Na/H exchanger isoform 3 (NHE3) is expressed in the proximal tubule and thick ascending limb and contributes to the reabsorption of fluid and electrolytes in these segments. The contribution of NHE3 to fluid reabsorption was assessed by micropuncture in homozygous ( Nhe3 −/−) and heterozygous ( Nhe3 +/−) knockout mice, and in their wild-type (WT, Nhe3 +/+) littermates. Arterial pressure was lower in the Nhe3 −/−mice (89 ± 6 mmHg) compared with Nhe3 +/+ (118 ± 4) and Nhe3 +/−(108 ± 5). Collections from proximal and distal tubules demonstrated that proximal fluid reabsorption was blunted in both Nhe3 +/− and Nhe3 −/−mice (WT, 4.2 ± 0.3; Nhe3 +/−, 3.4 ± 0.2; and Nhe3 −/−, 2.6 ± 0.3 nl/min; P < 0.05). However, distal delivery of fluid was not different among the three groups of mice (WT, 3.3 ± 0.4 nl/min; Nhe3 +/−, 3.3 ± 0.2 nl/min; and Nhe3 −/−, 3.0 ± 0.4 nl/min; P < 0.05). In Nhe3 −/−mice, this compensation was largely attributable to decreased single-nephron glomerular filtration rate (SNGFR): 10.7 ± 0.9 nl/min in the Nhe3 +/+ vs. 6.6 ± 0.8 nl/min in the Nhe3 −/−, measured distally. Proximal-distal SNGFR differences in Nhe3 −/−mice indicated that much of the decrease in SNGFR was due to activation of tubuloglomerular feedback (TGF), and measurements of stop-flow pressure confirmed that TGF is intact in Nhe3 −/−animals. In contrast to Nhe3 −/−mice, normalization of early distal flow rate in Nhe3 +/−mice was not related to decreased SNGFR (9.9 ± 0.7 nl/min), but rather, to increased fluid reabsorption in the loop segment ( Nhe3 +/+, 2.6 ± 0.2; Nhe3 +/−, 3.6 ± 0.5 nl/min). We conclude that NHE3 is a major Na/H exchanger isoform mediating Na+ and fluid reabsorption in the proximal tubule. In animals with NHE3 deficiency, normalization of fluid delivery to the distal tubule is achieved through alterations in filtration rate and/or downstream transport processes.

Abstract 79: Caveolin-1 Knockout Mice Have Salt-Sensitive Hypertension and Dopamine-1 Receptor Defects in Renal Cortex and Isolated Renal Proximal Tubule Cells

Hypertension ◽  
2013 ◽  
Vol 62 (suppl_1) ◽  
Author(s):  
John J Gildea ◽  
Nancy L Howell ◽  
Robert E Van Sciver ◽  
Brandon A Kemp ◽  
Robert M Carey ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Proximal Tubule ◽  
Kinase Activity ◽  
Renal Cortex ◽  
Renal Proximal Tubule ◽  
Normal Blood Pressure ◽  
Proximal Tubule Cells ◽  
Caveolin 1 ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells

Dopamine-1 receptors (D1R) are necessary for kidney proximal tubule-dependent natriuresis and maintenance of normal blood pressure, especially under high salt conditions. G-protein coupled receptor kinase 4 (GRK4) is a negative regulator of D1R function and single nucleotide polymorphisms in GRK4 have been associated with both hypertension and salt sensitivity in humans. Caveolin-1 (CAV1) directly binds to GRK4 and decreases kinase activity. We hypothesized that CAV1 knockout mice (CAV1KO) would have increased GRK4 kinase activity due to lack of physical interaction and inhibition of GRK4; thus overactive GRK4 would inactivate the D1R. Mean arterial blood pressure (MAP, in mmHg ±SEM) measured over 5 days was not significantly different for Wild-type mice (WT, 128.9±4.2 mmHg, n=4) vs CAV1KO (129.5±3.5 mmHg, n=4) on normal chow (0.3% sodium). However, on a 4% high sodium diet, the MAP of CAV1KO mice increased in just 2 days by 20.1±4.2 mmHg (p<0.05 vs either Day 0 CAV1KO or Day 2 WT, n=4). The CAV1KO MAP increased by 25.9±6.6 mmHg by day 7 (p<0.05 vs either Day 0 CAV1KO or Day 7 WT, n=4). Hyperphosphorylation and inactivation of the D1R in renal cortex was examined by looking at phospho-serine D1R by immuno-precipitation and Western dot blotting. A 92.5% ± 18.8 SEM increase in phospho-D1R was found in the CAV1KO renal cortex (n=4, p<0.01 vs WT; 14,574/7570 RFU). Cortical slices were made and incubated for 30 minutes with fenoldopam (FEN, 10 μM) with or without LE300 (D1R-like antagonist, 10 μM) or vehicle (VEH). Cyclic AMP was measured by TR-FRET (Lance, Perkin Elmer). FEN significantly increased cAMP 5.6 fold ± 1.2 SEM (n=4, p<0.01 vs VEH; 7.84/1.4 pmole/mg protein) in WT but not in CAV1KO slices, and this effect was completely blocked by LE300. Primary mouse CAV1KO and WT renal proximal tubule cell lines were established and monensin (sodium ionophore, 5 μM, 30 minutes)-induced plasma membrane D1R recruitment increased as measured by confocal microscopy in WT (30.4% ± 7.4 SEM, n=11, p<0.01 vs VEH; 8990/6894 RFU) but not in CAV1KO proximal tubule cells. In summary, CAV1 is necessary in high salt conditions for maintaining normal blood pressure in mice and for preserving normal D1R function in kidney cortex and in mouse renal proximal tubule cells.

scholarly journals Transgenic Overexpression of the Extra-Large Gsα Variant XLαs Enhances Gsα-Mediated Responses in the Mouse Renal Proximal Tubule in Vivo

Endocrinology ◽  
2011 ◽  
Vol 152 (4) ◽  
pp. 1222-1233 ◽  
Author(s):  
Zun Liu ◽  
Hiroko Segawa ◽  
Cumhur Aydin ◽  
Monica Reyes ◽  
Reinhold G. Erben ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Immunohistochemical Analysis ◽  
Renal Proximal Tubule ◽  
Mrna Levels ◽  
Wild Type ◽  
Western Blots ◽  
Α Subunit ◽  
Exon 1 ◽  
Serum Pth

Abstract XLαs, a variant of the stimulatory G protein α-subunit (Gsα), can mediate receptor-activated cAMP generation and, thus, mimic the actions of Gsα in transfected cells. However, it remains unknown whether XLαs can act in a similar manner in vivo. We have now generated mice with ectopic transgenic expression of rat XLαs in the renal proximal tubule (rptXLαs mice), where Gsα mediates most actions of PTH. Western blots and quantitative RT-PCR showed that, while Gsα and type-1 PTH receptor levels were unaltered, protein kinase A activity and 25-hydroxyvitamin D 1-α-hydroxylase (Cyp27b1) mRNA levels were significantly higher in renal proximal tubules of rptXLαs mice than wild-type littermates. Immunohistochemical analysis of kidney sections showed that the sodium-phosphate cotransporter type 2a was modestly reduced in brush border membranes of male rptXLαs mice compared to gender-matched controls. Serum calcium, phosphorus, and 1,25 dihydroxyvitamin D were within the normal range, but serum PTH was ∼30% lower in rptXLαs mice than in controls (152 ± 16 vs. 222 ± 41 pg/ml; P &lt; 0.05). After crossing the rptXLαs mice to mice with ablation of maternal Gnas exon 1 (E1m−/+), male offspring carrying both the XLαs transgene and maternal Gnas exon 1 ablation (rptXLαs/E1m−/+) were significantly less hypocalcemic than gender-matched E1m−/+ littermates. Both E1m−/+ and rptXLαs/E1m−/+ offspring had higher serum PTH than wild-type littermates, but the degree of secondary hyperparathyroidism tended to be lower in rptXLαs/E1m−/+ mice. Hence, transgenic XLαs expression in the proximal tubule enhanced Gsα-mediated responses, indicating that XLαs can mimic Gsα in vivo.

scholarly journals On the Mechanism of Inhibition in Fluid Reabsorption by the Renal Proximal Tubule of the Volume-Expanded Rat

1971 ◽  
Vol 50 (8) ◽  
pp. 1596-1602 ◽  
Author(s):  
Barry M. Brenner ◽  
Julia L. Troy ◽  
Terrance M. Daugharty ◽  
I. F. Ueki ◽  
D. P. Nicholas ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Renal Proximal Tubule ◽  
Fluid Reabsorption ◽  
Mechanism Of Inhibition

Defective PTH regulation of sodium-dependent phosphate transport in NHERF-1−/− renal proximal tubule cells and wild-type cells adapted to low-phosphate media

2005 ◽  
Vol 289 (4) ◽  
pp. F933-F938 ◽  
Author(s):  
Rochelle Cunningham ◽  
Xiaofei E ◽  
Deborah Steplock ◽  
Shirish Shenolikar ◽  
Edward J. Weinman
Keyword(s):  
Proximal Tubule ◽  
Primary Cultures ◽  
Phosphate Transport ◽  
Inhibitory Response ◽  
Renal Proximal Tubule ◽  
Wild Type ◽  
Proximal Tubule Cells ◽  
Sodium Dependent ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells

The present experiments using primary cultures from renal proximal tubule cells examine two aspects of the regulation of sodium-dependent phosphate transport and membrane sodium-dependent phosphate transporter (Npt2a) expression by parathyroid hormone (PTH). Sodium-dependent phosphate transport in proximal tubule cells from wild-type mice grown in normal-phosphate media averaged 4.4 ± 0.5 nmol·mg protein−1·10 min−1 and was inhibited by 30.5 ± 8.6% by PTH (10−7 M). This was associated with a 32.7 ± 5.2% decrease in Npt2a expression in the plasma membrane. Proximal tubule cells from Na+/H+ exchanger regulatory factor-1 (NHERF-1)−/− mice had a lower rate of phosphate transport compared with wild-type cells and a significantly reduced inhibitory response to PTH. Wild-type cells incubated in low-phosphate media for 24 h had a higher rate of phosphate transport compared with wild-type cells grown in normal-phosphate media but a significantly blunted inhibitory response to PTH. These data indicate a role for NHERF-1 in mediating the membrane retrieval of Npt2a and the subsequent inhibition of phosphate transport in renal proximal tubules. These studies also suggest that there is a blunted phosphaturic effect of PTH in cells adapted to low-phosphate media.

scholarly journals Expression of NHE-3 in the apical membrane of rat renal proximal tubule and thick ascending limb

1995 ◽  
Vol 48 (4) ◽  
pp. 1206-1215 ◽  
Author(s):  
Morimasa Amemiya ◽  
Jan Loffing ◽  
Marius Lötscher ◽  
Brigitte Kaissling ◽  
Robert J. Alpern ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Apical Membrane ◽  
Renal Proximal Tubule ◽  
Thick Ascending Limb ◽  
Rat Renal Proximal Tubule

scholarly journals Transcriptomic alterations induced by Monuron in rat and human renal proximal tubule cells in vitro and comparison to rat renal-cortex in vivo

2015 ◽  
Vol 4 (2) ◽  
pp. 423-431 ◽  
Author(s):  
Katarzyna M. Bloch ◽  
Noreen Yaqoob ◽  
Sikander Sharma ◽  
Andrew Evans ◽  
Lydia Aschauer ◽  
...  
Keyword(s):  
Developing Countries ◽  
Proximal Tubule ◽  
Renal Cortex ◽  
Renal Proximal Tubule ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells

Monuron (1,1-dimethyl-3-(4-chlorophenyl)urea) is a widely used herbicide in developing countries although concerns have been raised about its toxicity and carcinogenicity.

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