FRINK Linked Data Fragments server

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2068448909> ?p ?o ?g. }

• W2068448909 endingPage "37880" @default.
• W2068448909 startingPage "37871" @default.
• W2068448909 abstract "ROMK is an apical K+channel expressed in the thick ascending limb of Henle (TALH) and throughout the distal nephron of the kidney. Null mutations in theROMK gene cause type II Bartter's syndrome, in which abnormalities of electrolyte, acid-base, and fluid-volume homeostasis occur because of defective NaCl reabsorption in the TALH. To understand better the pathogenesis of type II Bartter's syndrome, we developed a mouse lacking ROMK and examined its phenotype. Young null mutants had hydronephrosis, were severely dehydrated, and ∼95% died before 3 weeks of age. ROMK-deficient mice that survived beyond weaning grew to adulthood; however, they had metabolic acidosis, elevated blood concentrations of Na+ and Cl−, reduced blood pressure, polydipsia, polyuria, and poor urinary concentrating ability. Whole kidney glomerular filtration rate was sharply reduced, apparently as a result of hydronephrosis, and fractional excretion of electrolytes was elevated. Micropuncture analysis revealed that the single nephron glomerular filtration rate was relatively normal, absorption of NaCl in the TALH was reduced but not eliminated, and tubuloglomerular feedback was severely impaired. These data show that the loss of ROMK in the mouse causes perturbations of electrolyte, acid-base, and fluid-volume homeostasis, reduced absorption of NaCl in the TALH, and impaired tubuloglomerular feedback. ROMK is an apical K+channel expressed in the thick ascending limb of Henle (TALH) and throughout the distal nephron of the kidney. Null mutations in theROMK gene cause type II Bartter's syndrome, in which abnormalities of electrolyte, acid-base, and fluid-volume homeostasis occur because of defective NaCl reabsorption in the TALH. To understand better the pathogenesis of type II Bartter's syndrome, we developed a mouse lacking ROMK and examined its phenotype. Young null mutants had hydronephrosis, were severely dehydrated, and ∼95% died before 3 weeks of age. ROMK-deficient mice that survived beyond weaning grew to adulthood; however, they had metabolic acidosis, elevated blood concentrations of Na+ and Cl−, reduced blood pressure, polydipsia, polyuria, and poor urinary concentrating ability. Whole kidney glomerular filtration rate was sharply reduced, apparently as a result of hydronephrosis, and fractional excretion of electrolytes was elevated. Micropuncture analysis revealed that the single nephron glomerular filtration rate was relatively normal, absorption of NaCl in the TALH was reduced but not eliminated, and tubuloglomerular feedback was severely impaired. These data show that the loss of ROMK in the mouse causes perturbations of electrolyte, acid-base, and fluid-volume homeostasis, reduced absorption of NaCl in the TALH, and impaired tubuloglomerular feedback. thick ascending limb of Henle isoform 2 of the Na+-K+-2Cl− cotransporter single nephron glomerular filtration rate tubuloglomerular feedback. NKCC2 and Nkcc2 refer to human and mouse genes, respectively, encoding NKCC2. ROMK andRomk are human and mouse genes, respectively, encoding ROMK.Romk +/+, Romk +/−, andRomk −/− refer to wild-type, heterozygous, and homozygous mutant mice, respectively Bartter's syndrome, a hypokalemic alkalosis with dehydration, hypotension, and severe polyuria which develops before birth or during infancy (1Simon D.B. Lifton R.P. Curr. Opin. Cell Biol. 1998; 10: 450-454Crossref PubMed Scopus (37) Google Scholar), is caused by null mutations in any of four genes encoding proteins involved in NaCl absorption in the renal thick ascending limb of Henle (TALH).1 These are the NKCC2 Na+-K+-2Cl−cotransporter (2Simon D.B. Karet F.E. Hamdan J.M. DiPietro A. Sanjad S.A. Lifton R.P. Nat. Genet. 1996; 13: 183-188Crossref PubMed Scopus (797) Google Scholar), the ROMK potassium channel (3Simon D.B. Karet F.E. Rodriguez-Soriano J. Hamdan J.H. DiPietro A. Trachtman H. Sanjad S.A. Lifton R.P. Nat. Genet. 1996; 14: 152-156Crossref PubMed Scopus (731) Google Scholar), the CLC-KB chloride channel (4Simon D.B. Bindra R.S. Mansfield T.A. Nelson-Williams C. Mendonca E. Stone R. Schurman S. Nayir A. Alpay H. Bakkaloglu A. Rodriguez-Soriano J. Morales J.M. Sanjad S.A. Taylor C.M. Pilz D. Brem A. Trachtman H. Griswold W. Richard G.A. John E. Lifton R.P. Nat. Genet. 1997; 17: 171-178Crossref PubMed Scopus (770) Google Scholar), and barttin (5Birkenhager R. Otto E. Schurmann M.J. Vollmer M. Ruf E.M. Maier-Lutz I. Beekmann F. Fekete A. Omran H. Feldmann D. Milford D.V. Jeck N. Konrad M. Landau D. Knoers N.V. Antignac C. Sudbrak R. Kispert A. Hildebrandt F. Nat. Genet. 2001; 29: 310-314Crossref PubMed Scopus (430) Google Scholar), a β-subunit of the chloride channel (6Estevez R. Boettger T. Stein V. Birkenhager R. Otto E. Hildebrandt F. Jentsch T.J. Nature. 2001; 414: 558-561Crossref PubMed Scopus (482) Google Scholar). Na+ and Cl−, in a 1:2 ratio, are absorbed across the apical membrane of TALH cells by the coupled activities of NKCC2 and ROMK and extruded via the basolateral Na+,K+-ATPase and chloride channel (6Estevez R. Boettger T. Stein V. Birkenhager R. Otto E. Hildebrandt F. Jentsch T.J. Nature. 2001; 414: 558-561Crossref PubMed Scopus (482) Google Scholar, 7Hebert S.C. Am. J. Physiol. 1998; 275: F325-F327PubMed Google Scholar); additional Na+ is absorbed via the paracellular pathway. Although NKCC2 directly mediates uptake of Na+, K+, and Cl−, the activity of ROMK is critical because the K+ concentration in the luminal fluid is much lower than that of Na+ and Cl−. Thus, the continuous electroneutral uptake of Na+, K+, and Cl− requires that K+ be recycled to the lumen of the tubule. Apical K+ secretion via ROMK replenishes luminal K+ and also contributes, in concert with basolateral Cl− efflux via CLC-KB/barttin (5Birkenhager R. Otto E. Schurmann M.J. Vollmer M. Ruf E.M. Maier-Lutz I. Beekmann F. Fekete A. Omran H. Feldmann D. Milford D.V. Jeck N. Konrad M. Landau D. Knoers N.V. Antignac C. Sudbrak R. Kispert A. Hildebrandt F. Nat. Genet. 2001; 29: 310-314Crossref PubMed Scopus (430) Google Scholar, 6Estevez R. Boettger T. Stein V. Birkenhager R. Otto E. Hildebrandt F. Jentsch T.J. Nature. 2001; 414: 558-561Crossref PubMed Scopus (482) Google Scholar), to the transcellular electrical potential that is the driving force for Na+ absorption via the paracellular pathway (8Hebert S.C. Andreoli T.E. Am. J. Physiol. 1984; 246: F745-F756PubMed Google Scholar). The different types of Bartter's syndrome, caused by null mutations in NKCC2, ROMK, CLC-KB, and barttin, are referred to as types I–IV, respectively. The syndrome is thus heterogeneous, consistent with the variety in genetic mechanisms, and the physiological phenotypes overlap to some degree with those of Gitelman's syndrome, a milder hypokalemic alkalosis caused by null mutations in the thiazide-sensitive NaCl cotransporter of the distal convoluted tubule (9Simon D.B. Nelson-Williams C. Bia M.J. Ellison D. Karet F.E. Molina A.M. Vaara I. Iwata F. Cushner H.M. Koolen M. Gainza F.J. Gitelman H.J. Lifton R.P. Nat. Genet. 1996; 12: 24-30Crossref PubMed Scopus (1048) Google Scholar, 10Schultheis P.J. Lorenz J.N. Meneton P. Nieman M.L. Riddle T.M. Flagella M. Duffy J.J. Doetschman T. Miller M.L. Shull G.E. J. Biol. Chem. 1998; 273: 29150-29155Abstract Full Text Full Text PDF PubMed Scopus (264) Google Scholar). Detailed analysis of the physiological functions and relative importance of the transporters involved in each type of Bartter's syndrome would be facilitated by the development of knockout mouse models. A mouse model for Bartter's syndrome type I, involving NKCC2, has already been developed (11Takahashi N. Chernavvsky D.R. Gomez R.A. Igarashi P. Gitelman H.J. Smithies O. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 5434-5439Crossref PubMed Scopus (214) Google Scholar); null mutants exhibit severe hydronephrosis, dehydration, polydipsia, polyuria, and an inability to concentrate the urine, and they usually die before weaning. There are multiple N-terminal variants of ROMK (gene locusKcnj1) (12Shuck M.E. Bock J.H. Benjamin C.W. Tsai T.D. Lee K.S. Slightom J.L. Bienkowski M.J. J. Biol. Chem. 1994; 269: 24261-24270Abstract Full Text PDF PubMed Google Scholar, 13Ho K. Nichols C.G. Lederer W.J. Lytton J. Vassilev P.M. Kanazirska M.V. Hebert S.C. Nature. 1993; 362: 31-38Crossref PubMed Scopus (833) Google Scholar, 14Boim M.A., Ho, K. Shuck M.E. Bienkowski M.J. Block J.H. Slightom J.L. Yang Y. Brenner B.M. Hebert S.C. Am. J. Physiol. 1995; 268: F1132-F1140Crossref PubMed Google Scholar). One or more of these variants is expressed in the TALH, distal convoluted tubule, connecting tubule, collecting duct, and macula densa (15Lee W.S. Hebert S.C. Am. J. Physiol. 1995; 268: F1124-F1131Crossref PubMed Google Scholar, 16Xu J.Z. Hall A.E. Peterson L.N. Bienkowski M.J. Eessalu T.E. Hebert S.C. Am. J. Physiol. 1997; 273: F739-F748Crossref PubMed Google Scholar), consistent with functions in K+ recycling to facilitate Na+ reabsorption in the TALH, K+ secretion in the distal nephron, and tubuloglomerular feedback (17Wang W. Hebert S.C. Giebisch G. Annu. Rev. Physiol. 1997; 59: 413-436Crossref PubMed Scopus (176) Google Scholar, 18Hurst A.M. Lapointe J.Y. Laamarti A. Bell P.D. J. Gen. Physiol. 1994; 103: 1055-1070Crossref PubMed Scopus (37) Google Scholar). The broad distribution of ROMK in the renal nephron, in contrast to the restricted distribution of NKCC2, and the possibility that other apical K+ channels (17Wang W. Hebert S.C. Giebisch G. Annu. Rev. Physiol. 1997; 59: 413-436Crossref PubMed Scopus (176) Google Scholar) or K+-independent modes of NaCl transport for NKCC2 (19Plata C. Meade P. Hall A. Welch R.C. Vazquez N. Hebert S.C. Gamba G. Am. J. Physiol. 2001; 280: F574-F582Crossref PubMed Google Scholar) might provide some compensation for its absence, suggested that the loss of ROMK might lead to a different phenotype than that of theNkcc2 knockout. In this study we have developed a mouse model for Bartter's syndrome type II, involving ROMK. ROMK-deficient mice have hydronephrosis, polydipsia, polyuria, extracellular fluid volume depletion, and a urinary concentrating defect. This phenotype is similar to that of theNkcc2 knockout, although it is not as severe, suggesting that renal Na+ handling is perturbed less by the loss of ROMK than by the loss of NKCC2. To examine this possibility we performed micropuncture analysis of a single nephron function. These experiments revealed that tubuloglomerular feedback is severely impaired and that Na+ reabsorption is significantly reduced but not eliminated in the TALH of the Romk knockout. The latter observation supports the hypothesis that ROMK plays an essential role in K+ recycling in the TALH, which is required for maximum Na+ reabsorption via NKCC2, and also shows that a significant amount of Na+ reabsorption does take place in its absence. A phage library prepared using genomic DNA from a 129/SvJ mouse was screened with a RomkcDNA probe. Clones containing the Romk gene were isolated and analyzed by restriction endonuclease mapping. Two restriction fragments containing sequences from the large core exon were inserted into the MJK-KO targeting vector (20Meneton P. Schultheis P.J. Greeb J. Nieman M.L. Liu L.H. Clarke L.L. Duffy J.J. Doetschman T. Lorenz J.N. Shull G.E. J. Clin. Invest. 1998; 101: 536-542Crossref PubMed Scopus (138) Google Scholar), which allows a positive-negative selection strategy. A 3.8-kb BglII fragment terminating with codon 181 was inserted between the 3′-end of the neomycin resistance gene and the 5′-end of the herpes simplex virus-thymidine kinase gene, and a 2.8 kb BglII fragment beginning with codon 182 was inserted between the 5′-end of the neomycin resistance gene and vector sequences. The targeting construct was linearized and electroporated into embryonic stem cells derived from 129/SvJ mice, which were then cultured in the presence of G418 and gancyclovir as described previously (20Meneton P. Schultheis P.J. Greeb J. Nieman M.L. Liu L.H. Clarke L.L. Duffy J.J. Doetschman T. Lorenz J.N. Shull G.E. J. Clin. Invest. 1998; 101: 536-542Crossref PubMed Scopus (138) Google Scholar). DNA was isolated from cells that survived the selection procedure and analyzed by Southern blot analysis using a 2-kb HindIII-BglII fragment from the region just 5′ to fragments used to prepare the targeting construct. Blastocyst-mediated transgenesis was performed, chimeric mice were bred with Black Swiss mice, and a colony carrying the null allele was established by breeding heterozygous mutant mice. These studies and the experiments described below were performed in accordance with the guidelines established by the Institutional Animal Care and Use Committee at the University of Cincinnati College of Medicine. DNA from tail biopsies was analyzed by PCR using a three primer set. A 275-bp fragment from the wild-type gene was amplified using a forward primer (5′-GTGACAGAACAGTGTGCC-3′) corresponding to codons 149–154 and a reverse primer (5′-CTCCTTCAGGTGTGATGG-3′) corresponding to anticodons 240–234. A 385-bp product from the mutant gene was amplified using the reverse primer from the Romk gene and a primer (5′-CTGACTAGGGGAGGAGTAGAAGG-3′) complementary to sequences in the 5′-untranslated region of the neomycin resistance gene. Total RNA was isolated from pooled kidneys of 8-day-old Romk −/−(n = 10) and Romk +/+(n = 6) mice and from individual adultRomk −/− and Romk +/+mice using Tri-ReagentTM (Molecular Research Center, Inc., Cincinnati, OH). Blots were prepared, hybridized, and washed as described previously (21Schultheis P.J. Clarke L.L. Meneton P. Harline M. Boivin G.P. Stemmermann G. Duffy J.J. Doetschman T. Miller M.L. Shull G.E. J. Clin. Invest. 1998; 101: 1243-1253Crossref PubMed Scopus (222) Google Scholar), using rat ROMK cDNA probes corresponding to codons 13–158 and codons 197–372. At postnatal day 7–9, pups were sacrificed by decapitation, and trunk blood was collected into heparinized tubes for analysis on a Chiron model 448 (Chiron Diagnostics, Medfield, MA) blood gas analyzer. Urine was collected from the bladder, and electrolytes were analyzed by flame photometry (Corning model 480, Medfield, MA). Kidneys collected from 7–9-day-old and adult Romk +/+ andRomk −/− mice were prepared for routine light and electron microscopic histology and morphometry (22Flagella M. Clarke L.L. Miller M.L. Erway L.C. Giannella R.A. Andringa A. Gawenis L.R. Kramer J. Duffy J.J. Doetschman T. Lorenz J.N. Yamoah E.N. Cardell E.L. Shull G.E. J. Biol. Chem. 1999; 274: 26946-26955Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar). SevenRomk −/− and 11 Romk +/+mice were used, with approximately equal numbers of young, old, male, and female mice. Phase-contrast microscopy of toluidine blue-stained, 1.5-μm thick plastic sections was used to quantify the relative amounts of proximal and distal tubule, and of glomeruli, in the kidney cortex subadjacent to the capsule (to correlate with those nephrons analyzed by micropuncture). A grid of 75 intersections was visualized on the light microscopic image using a camera lucida, and the number of positive intersections lying over glomerular, proximal, and distal tubular cells was determined. Proximal tubular cells were identified as those with brush-border membranes and nuclei close to the basal membrane, whereas distal tubular cells were identified as those with no brush-border and more apically located nuclei. The volume density of each of these was determined by dividing the number of intersections found over each tubule type by the total number of intersections lying over all the tissue. In addition, a subjective appraisal of the degree of mitochondrial organization at the base of the tubular cells was made, with 1+ connoting no organization and 6+ being the highly organized and parallel alignment of mitochondria in a perpendicular position relative to the basement membrane. Means and S.E. values were generated using the General Linear model of SAS 6.1. Differences were considered significant when p < 0.05. AdultRomk −/− mice of either sex (n= 4) and matched Romk +/+ mice (n= 6) were housed individually in conventional shoebox cages for the duration of the experiment, and they were provided a diet of normal rodent chow. Blood pressure measurements were made by tail-cuff using a Visitech Systems BP-2000 blood pressure analyzer (Apex, NC) after 4 days of acclimation to the instrument. Urine concentrating ability was evaluated by measuring osmolality in urine samples obtained by bladder massage before and after 6 h of water deprivation. For the following 3 days, mice were given distilled water to drink, and water intake, body weight, and blood pressure were monitored daily. For the next 3 days, animals were provided with both water and isotonic saline to drink, and measurements were repeated. At the end of the tail-cuff protocol, animals resumed drinking water only and were made available for clearance and micropuncture experiments. Experiments were performed on adult Romk −/− mice of either sex (n = 5) and matched Romk +/+ mice (n = 5). Whole kidney clearance measurements were performed in all of the mice, and micropuncture measurements were made in three mutant and three wild-type mice. Animals were surgically prepared according to conventional techniques modified for use in the mouse as described previously (23Lorenz J.N. Schultheis P.J. Traynor T. Shull G.E. Schnermann J. Am. J. Physiol. 1999; 277: F447-F453PubMed Google Scholar). Mice were anesthetized with separate intraperitoneal injections of ketamine (50 μg/g of body weight) and thiobutabarbital (Inactin, Research Biochemicals International, Natick, MA; 100 μg/g of body weight) and placed on a thermally controlled surgical table. After tracheostomy, the right femoral artery and vein were cannulated with polyethylene tubing hand-drawn to a fine tip over a flame (OD 0.3–0.5 mm). The arterial catheter was connected to a COBE CDXIII fixed dome pressure transducer (COBE Cardiovascular, Arvada, CO) for measurement of arterial blood pressure, and the venous catheter was connected to a syringe pump for infusion. The bladder was also cannulated with flared PE-10 tubing for the collection of urine. Blood pressure and heart rate were monitored throughout the experiment using a PowerLab data acquisition system (AD Instruments, Boston) with a sampling rate of 200 samples/s. Body temperature was maintained at 37.5 °C, and animals were provided with a steady stream of 100% O2 to breathe. For micropuncture experiments, the left kidney was exposed via a flank incision, carefully dissected free of adherent fat and connective tissue, placed in a Lucite cup and covered with mineral oil. A 4-μl/g of body weight bolus infusion of isotonic saline containing 0.75 g of fluorescein isothiocyanate-inulin/100 ml (Sigma), 2.25 g of bovine serum albumin/100 ml, and 1.0 g of glucose/100 ml was then administered, followed by a maintenance infusion of the same solution at 0.2 μl/min/g of body weight. After a 30–45-min equilibration period, micropuncture collections and/or urine collections were begun. For micropuncture collections, surface convolutions of the same nephron were identified by injecting a small volume of saline containing 0.25% Fast Green dye (Sigma) into a random proximal segment. Late proximal puncture sites were identified as the last surface segment to fill with green dye before it disappeared into the loop of Henle. In a small population of nephrons (10–20%), an early distal puncture site could be identified when the green dye returned to the kidney surface. During two consecutive clearance periods lasting 30–60 min, at least five timed proximal collections were made, and usually two to three paired distal collections were made. In those nephrons having both proximal and distal collection sites, the distal tubule collection was performed before the proximal collection. Blood samples (5–10 μl) were taken in heparinized tubes before and after each clearance period. Sharpened glass micropipettes used for dye injection were 2–3 μm in diameter, and those used for fluid collection were 6–7 μm. At the end of each experiment, tubular fluid samples were transferred individually to 1-μl constant bore microcaps for determination of volume and inulin concentration as described previously (24Lorenz J.N. Gruenstein E. Am. J. Physiol. 1999; 276: F172-F177PubMed Google Scholar). The tubular fluid chloride concentration was determined by electrometric titration (25Ramsay J.A. Brown R.H. Croghan P.C. J. Exp. Biol. 1955; 32: 822-829Google Scholar). Blood samples were centrifuged, and aliquots of plasma were transferred into 1-μl microcaps for inulin determination. Urine samples were also evaluated for inulin for the determination of whole kidney glomerular filtration rate. SeparateRomk −/− (n = 3) and wild-type mice (n = 5) of either sex and weighing 20–30 g were prepared and proximal tubule segments identified for micropuncture as described above. Early proximal portions of the identified tubules were blocked with wax, and a micropipette attached to a nanoliter infusion pump was inserted into the last superficial proximal segment for loop of Henle perfusion. Another micropipette, attached to a Servo-null pressure device (World Precision Instruments, Sarasota, FL), was then inserted into an early proximal segment recognizable from the widening of the tubular lumen. When stop-flow pressure stabilized, the loop of Henle perfusion rate was altered from 0 to 40 nl/min, and maximal responses in stop-flow pressures were recorded. Perfusion fluid contained (in mm) 136 NaCl, 4 NaHCO3, 4 KCl, 2 CaCl2, 7.5 urea, and 1 mg/ml Fast Green. In a few tubules (see under “Results”), 36 mm NaCl in the perfusion fluid was replaced with 36 mm KCl to test the effect of perfusion with high K+ (100 mm Na+, 40 mm K+). Statistical analysis was performed by analysis of variance using a single factor design or a mixed factorial design with repeated measures on the second factor. Where necessary, individual comparisons of group means were accomplished using individual contrasts. Data are expressed as means ± S.E., and differences are regarded as significant at p < 0.05. TheRomk gene was disrupted in embryonic stem cells by insertion of the neomycin resistance gene into the large core exon (Fig.1 A). Targeted cells were identified by Southern blot analysis (Fig. 1 B) and used to generate a mutant line carrying the null allele. As shown by PCR analysis of tail DNA (Fig. 1 C), breeding of heterozygous mutant mice resulted in the birth of live pups of all three genotypes.Romk +/+, heterozygous, and null mutant mice were born in a normal Mendelian ratio (26.6% +/+, 49.8% +/−, and 23.6% −/− among the first 500 pups), demonstrating that ROMK is not required for survival of the embryo. Northern blot analysis of kidney mRNA from wild-type and Romk −/− mice showed that insertion of the neomycin resistance gene had virtually eliminated expression of Romk mRNA (Fig. 1 D), although trace levels of an mRNA that was larger than the wild-type mRNA was detected with the 3′-probe. Null mutants exhibited growth retardation, and by 1 week of age most of them could be identified because of poor turgor and wrinkled skin, probably reflecting fluid volume depletion. Mortality ofRomk −/− mice was high (Fig.2), with 85% dying by 12 days of age, and only 5% surviving to weaning at 21 days. Daily subcutaneous injections of either indomethacin, which was reported to improve the survival rate of NKCC2-deficient mice (11Takahashi N. Chernavvsky D.R. Gomez R.A. Igarashi P. Gitelman H.J. Smithies O. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 5434-5439Crossref PubMed Scopus (214) Google Scholar), or isotonic saline, which rescued mice lacking the mineralocorticoid receptor (26Bleich M. Warth R. Schmidt-Hieber M. Schulz-Baldes A. Hasselblatt P. Fisch D. Berger S. Kunzelmann K. Kriz W. Schutz G. Greger R. Pflügers Arch. Eur. J. Physiol. 1999; 438: 245-254Crossref PubMed Scopus (96) Google Scholar), did not improve the survival rate of null mutants.Romk −/− mice that survived to weaning exhibited no excess mortality and grew well. Adult mutants were slightly smaller than wild-type mice but appeared healthy. Both male and female null mutants were fertile. Grossly, mutant kidneys from young (7–9 days) and adult mice showed moderate to severe dilation of the renal pelvis, indicative of hydronephrosis (Fig.3, A and B). Histologically, it was readily apparent at the macro level that the renal cortex was considerably thinned, and an enormously dilated renal pelvis surrounded the renal papilla in mutant kidneys, which made it difficult to visualize thin loops in the mutant mice. This was accompanied by a loss of organization and structure within the medulla of the kidney. Unlike in wild-type mice (Fig. 3 C), abundant droplets of lipidic material were seen within the proximal tubule cells of adult Romk −/− mice (Fig. 3 D). This debris, possibly lysosomal, sometimes had a myelin character. Similar structures were found as casts in the tubular lumen (Fig.3 E). These structures may represent the lysis or loss of apical membranes of compromised cells but did not contain calcium crystalline deposits (27Ghadially F.N. Ultrastruct. Pathol. 2001; 25: 243-267Crossref PubMed Scopus (46) Google Scholar). In the proximal tubule mainly, but also in the distal tubule, mitochondria of adultRomk −/− mice contained significantly more dense, native intramitochondrial granules (Fig. 3 F) than those of wild-type mice, often accompanied by an increase in overall electron density of the cell. The results of morphometric analyses of the renal cortex are presented in Table I. There were no differences in volume density of the glomeruli between wild-type andRomk −/− mice of any age, although in young mutant mice there were often dark bodies within the glomeruli (perhaps apoptotic debris). This difference was not found in adults. Although glomeruli from both young and adult null mutants were usually histologically indistinguishable from those of wild-type mice, on occasion the glomerular tuft was dwarfed by a ballooning of the urinary space. The glomerular basement membrane in mutant mice was examined for thickening and for calcium deposits using electron microscopy, but neither was encountered. Distal and collecting tubules were often dilated in both young and adult Romk −/− mice. The volume density of proximal tubule cells was significantly decreased in the knockout compared with the wild-type, and at the same time the volume density of distal tubules was significantly increased. The ratio of distal to proximal tubules was significantly increased in mutant mice (Table I), indicating that a compensatory increase had occurred in the relative length of the distal tubule in the knockout. A loss of organization of mitochondria within the basal portion of individual proximal tubule cells was found in the mutant mice. With electron microscopy (data not shown), proximal tubule cells from mutant animals showed less organellar order; specifically, there were fewer basal membrane infoldings and mitochondrial interdigitation in knockout mice than in the wild-type mice. This was not true of the distal tubule, however, where the level of subcellular organization appeared to be similar in wild-type and null mice. The amount of lipid debris in the proximal tubule was significantly greater in the knockout.Table IMorphometric analysis of renal cortex in young and adult Romk+/+ and Romk−/− mice+/+ (n = 14)−/− (n = 17)p valueVd glomeruli7.4 ± 1.96.44 ± 1.3NSVd PT64.5 ± 3.5647.7 ± 2.20.0001Vd DT28.0 ± 2.650.3 ± 2.30.0001Distal: proximal tubule ratio0.47 ± 0.071.36 ± 0.170.0001PT mitochondrial organization3.36 ± 0.242.35 ± 0.250.0075DT mitochondrial organization2.24 ± 0.262.2 ± 0.27NSVd lipid debris0.22 ± 0.051.28 ± 0.180.0001Vd, volume density; PT, proximal convoluted tubule; DT, distal convoluted tubule. p value compared with +/+. NS, not significant. Open table in a new tab Vd, volume density; PT, proximal convoluted tubule; DT, distal convoluted tubule. p value compared with +/+. NS, not significant. Prior to weaning, pups were evaluated for blood and urine chemistries (Table II). Wild-type and heterozygous pups were indistinguishable based on the measured variables. Compared with Romk +/+ mice, most of the null mutants had markedly reduced body weight and failed to thrive. Wet kidney weight did not differ between wild-type and mutant mice, so the kidney weight:body weight ratio was significantly greater in Romk −/− mice. There were no significant differences in blood pCO2 orpO2 or between wild-type and mutant mice; however, pH and HCO 3− were significantly reduced, and hematocrit and plasma Na+ and Cl− concentrations were significantly elevated in the mutants, consistent with metabolic acidosis, volume depletion, and dehydration.Table IIBlood and urine data from 7–9-day-old Romk+/+, Romk+/− and Romk−/− pups+/+ (n = 6)+/− (n = 12)−/− (n = 7)BW (g)5.9 ± 0.65.0 ± 0.33.3 ± 0.6*KW (g)0.08 ± 0.010.08 ± 0.010.08 ± 0.01KW/BW1.26 ± 0.071.34 ± 0.022.36 ± 0.19*Hct (%)37 ± 136 ± 147 ± 2*pCO2 (mm Hg)51.5 ± 2.750.6 ± 1.853.3 ± 3.7pO2 (mm Hg)43.8 ± 5.347.0 ± 5.136.6 ± 3.8pH7.38 ± 0.017.40 ± 0.017.26 ± 0.04*PHCO−3(mm)30.0 ± 1.630.3 ± 1.024.7 ± 1.1*PNa+(mm)131 ± 2132 ± 1170 ± 6*PK+(mm)6.4 ± 0.36.3 ± 0.47.2 ± 0.4PCL−(mm)98 ± 298 ± 1119 ± 5*UNa+(mm)35 ± 826 ± 641 ± 19UK+(mm)55 ± 1938 ± 755 ± 11Uosm (mosm/liter)685 ± 134582 ± 39566 ± 57BW, body weight; KW, kidney weight; Px, plasma concentration of x; Ux, urine concentration of x; Hct, hematocrit; *p < 0.05 compared with +/+. Open table in a new tab BW, body weight; KW, kidney weight; Px, plasma concentration of x; Ux, urine concentration of x; Hct, hematocrit; *p < 0.05 compared with +/+. A small" @default.
• W2068448909 created "2016-06-24" @default.
• W2068448909 creator A5004540545 @default.
• W2068448909 creator A5005072769 @default.
• W2068448909 creator A5010967815 @default.
• W2068448909 creator A5017319962 @default.
• W2068448909 creator A5024979827 @default.
• W2068448909 creator A5032778941 @default.
• W2068448909 creator A5054487742 @default.
• W2068448909 creator A5058751203 @default.
• W2068448909 creator A5061383127 @default.
• W2068448909 creator A5081451576 @default.
• W2068448909 date "2002-10-01" @default.
• W2068448909 modified "2023-10-15" @default.
• W2068448909 title "Impaired Renal NaCl Absorption in Mice Lacking the ROMK Potassium Channel, a Model for Type II Bartter's Syndrome" @default.
• W2068448909 cites W1608336039 @default.
• W2068448909 cites W1643229297 @default.
• W2068448909 cites W1969244514 @default.
• W2068448909 cites W1983469458 @default.
• W2068448909 cites W1985793100 @default.
• W2068448909 cites W1995826486 @default.
• W2068448909 cites W1997610195 @default.
• W2068448909 cites W2005140628 @default.
• W2068448909 cites W2007259410 @default.
• W2068448909 cites W2010368222 @default.
• W2068448909 cites W2015344956 @default.
• W2068448909 cites W2030840085 @default.
• W2068448909 cites W2036212752 @default.
• W2068448909 cites W2047640546 @default.
• W2068448909 cites W2051231609 @default.
• W2068448909 cites W2058633925 @default.
• W2068448909 cites W2064094026 @default.
• W2068448909 cites W2065978737 @default.
• W2068448909 cites W2066031126 @default.
• W2068448909 cites W2081620339 @default.
• W2068448909 cites W2083818654 @default.
• W2068448909 cites W2090362914 @default.
• W2068448909 cites W2092552227 @default.
• W2068448909 cites W2092976088 @default.
• W2068448909 cites W2096293547 @default.
• W2068448909 cites W2107609623 @default.
• W2068448909 cites W2110261346 @default.
• W2068448909 cites W2119375870 @default.
• W2068448909 cites W2131297348 @default.
• W2068448909 cites W2151206192 @default.
• W2068448909 cites W2395135404 @default.
• W2068448909 cites W2403464624 @default.
• W2068448909 doi "https://doi.org/10.1074/jbc.m205627200" @default.
• W2068448909 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/12122007" @default.
• W2068448909 hasPublicationYear "2002" @default.
• W2068448909 type Work @default.
• W2068448909 sameAs 2068448909 @default.
• W2068448909 citedByCount "163" @default.
• W2068448909 countsByYear W20684489092012 @default.
• W2068448909 countsByYear W20684489092013 @default.
• W2068448909 countsByYear W20684489092014 @default.
• W2068448909 countsByYear W20684489092015 @default.
• W2068448909 countsByYear W20684489092016 @default.
• W2068448909 countsByYear W20684489092017 @default.
• W2068448909 countsByYear W20684489092018 @default.
• W2068448909 countsByYear W20684489092019 @default.
• W2068448909 countsByYear W20684489092020 @default.
• W2068448909 countsByYear W20684489092021 @default.
• W2068448909 countsByYear W20684489092022 @default.
• W2068448909 countsByYear W20684489092023 @default.
• W2068448909 crossrefType "journal-article" @default.
• W2068448909 hasAuthorship W2068448909A5004540545 @default.
• W2068448909 hasAuthorship W2068448909A5005072769 @default.
• W2068448909 hasAuthorship W2068448909A5010967815 @default.
• W2068448909 hasAuthorship W2068448909A5017319962 @default.
• W2068448909 hasAuthorship W2068448909A5024979827 @default.
• W2068448909 hasAuthorship W2068448909A5032778941 @default.
• W2068448909 hasAuthorship W2068448909A5054487742 @default.
• W2068448909 hasAuthorship W2068448909A5058751203 @default.
• W2068448909 hasAuthorship W2068448909A5061383127 @default.
• W2068448909 hasAuthorship W2068448909A5081451576 @default.
• W2068448909 hasBestOaLocation W20684489091 @default.
• W2068448909 hasConcept C126322002 @default.
• W2068448909 hasConcept C134018914 @default.
• W2068448909 hasConcept C178790620 @default.
• W2068448909 hasConcept C185592680 @default.
• W2068448909 hasConcept C2777616141 @default.
• W2068448909 hasConcept C2778242168 @default.
• W2068448909 hasConcept C2910219414 @default.
• W2068448909 hasConcept C517785266 @default.
• W2068448909 hasConcept C537181965 @default.
• W2068448909 hasConcept C71924100 @default.
• W2068448909 hasConcept C83743174 @default.
• W2068448909 hasConceptScore W2068448909C126322002 @default.
• W2068448909 hasConceptScore W2068448909C134018914 @default.
• W2068448909 hasConceptScore W2068448909C178790620 @default.
• W2068448909 hasConceptScore W2068448909C185592680 @default.
• W2068448909 hasConceptScore W2068448909C2777616141 @default.
• W2068448909 hasConceptScore W2068448909C2778242168 @default.
• W2068448909 hasConceptScore W2068448909C2910219414 @default.
• W2068448909 hasConceptScore W2068448909C517785266 @default.
• W2068448909 hasConceptScore W2068448909C537181965 @default.
• W2068448909 hasConceptScore W2068448909C71924100 @default.

• next