This study examined whether substitution of chromosome 5 containing the CYP4A genes from Brown Norway rat onto the Dahl S salt-sensitive (SS) genetic background upregulates the renal production of 20-HETE and attenuates the development of hypertension. The expression of CYP4A protein and the production of 20-HETE were significantly higher in the renal cortex and outer medulla of SS.5 BN (chromosome 5-substituted Brown Norway rat) consomic rats fed either a low-salt (LS) or high-salt (HS) diet than that seen in SS rats. The increase in the renal production of 20-HETE in SS.5 BN rats was associated with elevated expression of CYP4A2 mRNA. MAP measured by telemetry rose from 117 Ϯ 1 to 183 Ϯ 5 mmHg in SS rats fed a HS diet for 21 days, but only increased to 151 Ϯ 5 mmHg in SS.5 BN rats. The pressure-natriuretic and diuretic responses were twofold higher in SS.5 BN rats compared with SS rats. Protein excretion rose to 354 Ϯ 17 mg/day in SS rats fed a HS diet for 21 days compared with 205 Ϯ 13 mg/day in the SS.5 BN rats, and the degree of glomerular injury was reduced. Baseline glomerular capillary pressure (Pgc) was similar in SS.5 BN rats (43 Ϯ 1 mmHg) and Dahl S (44 Ϯ 2 mmHg) rats. However, Pgc increased to 59 Ϯ 3 mmHg in SS rats fed a HS diet for 7 days, while it remained unaltered in SS.5 BN rats (43 Ϯ 2 mmHg). Chronic administration of an inhibitor of the synthesis of 20-HETE (HET0016, 10 mg·kg Ϫ1 ·day Ϫ1 iv) reversed the antihypertensive phenotype seen in the SS.5 BN rats. These findings indicate that the transfer of chromosome 5 from the BN rat onto the SS genetic background increases the renal expression of CYP4A protein and the production of 20-HETE and that 20-HETE contributes to the antihypertensive and renoprotective effects seen in the SS.5 BN consomic strain.hypertension; glomerulosclerosis; chromosome 5; Dahl S rats; pressure natriuresis; renal hemodynamics; kidney THE DAHL SALT-SENSITIVE (SS) rat is an inbred genetic model that rapidly develops severe hypertension, proteinuria, glomerulosclerosis, and renal interstitial fibrosis when fed a high salt (HS) diet (4, 7-8, 23, 25, 27, 30, 38, 43). However, the genes and pathways that contribute to the development of hypertension and renal disease have yet to be identified. Previous studies from our laboratory have demonstrated that the pressure natriuretic relationship is impaired in SS rats and that this is associated with increased Cl Ϫ transport in the thick ascending limb of Henle (TALH) (13, 15-16, 29, 44). They also exhibit a deficiency in the renal production of 20-HETE that contributes to the increase in loop Cl Ϫ transport (13,35,44).More recently, Mattson et al. (20) demonstrated that substitution of chromosome 5 from the Brown Norway (BN) rat onto the SS genetic background (SS.5 BN strain) attenuates the development of hypertension and proteinuria in SS.5 BN rats fed a high-salt (HS) diet for 21 days, but the mechanism is unknown. Because the CYP4A genes that produce 20-HETE are located on chromosome 5 and this region has been found to cosegre...

A fundamental requirement for cellular vitality is the maintenance of plasma ion concentration within strict ranges. It is the function of the kidney to match urinary excretion of ions with daily ion intake and nonrenal losses to maintain a stable ionic milieu. NADPH oxidase is a source of reactive oxygen species (ROS) within many cell types, including the transporting renal epithelia. The focus of this review is to describe the role of NADPH oxidase-derived ROS toward local renal tubular ion transport in each nephron segment and to discuss how NADPH oxidase-derived ROS signaling within the nephron may mediate ion homeostasis. In each case, we will attempt to identify the various subunits of NADPH oxidase and reactive oxygen species involved and the ion transporters, which these affect. We will first review the role of NADPH oxidase on renal Na ϩ and K ϩ transport. Finally, we will review the relationship between tubular H ϩ efflux and NADPH oxidase activity.

TR, Satlin LM. Role of NKCC in BK channel-mediated net K ϩ secretion in the CCD. Am J Physiol Renal Physiol 301: F1088 -F1097, 2011. First published August 3, 2011 doi:10.1152/ajprenal.00347.2011.-Apical SK/ROMK and BK channels mediate baseline and flow-induced K secretion (FIKS), respectively, in the cortical collecting duct (CCD). BK channels are detected in acid-base transporting intercalated (IC) and Na-absorbing principal (PC) cells. Although the density of BK channels is greater in IC than PC, Na-K-ATPase activity in IC is considered inadequate to sustain high rates of urinary K secretion. To test the hypothesis that basolateral NKCC in the CCD contributes to BK channel-mediated FIKS, we measured net K secretion (JK) and Na absorption (JNa) at slow (ϳ1) and fast (ϳ5 nl·min Ϫ1 ·mm Ϫ1 ) flow rates in rabbit CCDs microperfused in vitro in the absence and presence of bumetanide, an inhibitor of NKCC, added to the bath. Bumetanide inhibited FIKS but not basal JK, JNa, or the flow-induced [Ca 2ϩ ]i transient necessary for BK channel activation. Addition of luminal iberiotoxin, a BK channel inhibitor, to bumetanide-treated CCDs did not further reduce JK. Basolateral Cl removal reversibly inhibited FIKS but not basal JK or JNa. Quantitative PCR performed on single CCD samples using NKCC1-and 18S-specific primers and probes and the TaqMan assay confirmed the presence of the transcript in this nephron segment. To identify the specific cell type to which basolateral NKCC is localized, we exploited the ability of NKCC to accept NH4 ϩ at its K-binding site to monitor the rate of bumetanide-sensitive cytosolic acidification after NH4 ϩ addition to the bath in CCDs loaded with the pH indicator dye BCECF. Both IC and PC were found to have a basolateral bumetanide-sensitive NH4 ϩ entry step and NKCC1-specific antibodies labeled the basolateral surfaces of both cell types in CCDs. These results suggest that BK channel-mediated FIKS is dependent on a basolateral bumetanide-sensitive, Cl-dependent transport pathway, proposed to be NKCC1, in both IC and PC in the CCD. apical membrane voltage; tubular flow rates; bumetanide; intercalated cell; principal cell

The aims of the present study were to determine whether superoxide (O(2)(-)) production is enhanced in medullary thick ascending limb (mTAL) of Dahl salt-sensitive (SS) rats compared with a salt-resistant consomic control strain (SS.13(BN)) and to elucidate the cellular pathways responsible for augmented O(2)(-) production. Studies were carried out in 7- to 10-wk-old male SS and SS.13(BN) rats fed either a 0.4% NaCl diet or a 4.0% NaCl diet for 3 days before tissue harvest. Tissue strips containing mTAL were isolated from the left kidney, loaded with the O(2)(-)-sensitive fluorescent dye dihydroethidium, superfused with modified Hanks' solution, and imaged at x60 magnification on a heated microscope stage. O(2)(-) production was stimulated in mTAL by incrementing superfusate NaCl concentration from 154 to 254 to 500 mM. O(2)(-) production was enhanced in mTAL of SS rats compared with SS.13(BN) rats in response to incrementing bath NaCl. Addition of N-methyl-amiloride (100 muM) or inhibition of NAD(P)H oxidase reduced O(2)(-) production in SS mTAL to levels observed in SS.13(BN) rats. Both amiloride- and ouabain-sensitive pathways of O(2)(-) production were elevated following 3 days of high (4.0%) NaCl feeding in mTAL of SS and SS.13(BN) rats. We conclude that mTAL from SS rats exhibit enhanced amiloride-sensitive O(2)(-) production. The amiloride-sensitive O(2)(-) response in mTAL is independent of active Na(+) transport and appears to be mediated by NAD(P)H oxidase. Amiloride-sensitive O(2)(-) production is likely to contribute to augmented outer medullary O(2)(-) production observed in SS rats during both normal and high NaCl diets.

This study describes a high-throughput fluorescence dilution technique to measure the albumin reflection coefficient (σAlb) of isolated glomeruli. Rats were injected with FITC-dextran 250 (75 mg/kg), and the glomeruli were isolated in a 6% BSA solution. Changes in the fluorescence of the glomerulus due to water influx in response to an imposed oncotic gradient was used to determine σAlb. Adjustment of the albumin concentration of the bath from 6 to 5, 4, 3, and 2% produced a 10, 25, 35, and 50% decrease in the fluorescence of the glomeruli. Pretreatment of glomeruli with protamine sulfate (2 mg/ml) or TGF-β1 (10 ng/ml) decreased σAlb from 1 to 0.54 and 0.48, respectively. Water and solute movement were modeled using Kedem-Katchalsky equations, and the measured responses closely fit the predicted behavior, indicating that loss of albumin by solvent drag or diffusion is negligible compared with the movement of water. We also found that σAlb was reduced by 17% in fawn hooded hypertensive rats, 33% in hypertensive Dahl salt-sensitive (SS) rats, 26% in streptozotocin-treated diabetic Dahl SS rats, and 21% in 6-mo old type II diabetic nephropathy rats relative to control Sprague-Dawley rats. The changes in glomerular permeability to albumin were correlated with the degree of proteinuria in these strains. These findings indicate that the fluorescence dilution technique can be used to measure σAlb in populations of isolated glomeruli and provides a means to assess the development of glomerular injury in hypertensive and diabetic models.