Parallel down-regulation of chloride channel CLC-K1 and barttin mRNA in the thin ascending limb of the rat nephron by furosemide
Tóm tắt
In the past few years the pivotal role of kidney Cl−channels (ClC-K) channels in maintaining salt and water homeostasis in the kidney has been established. The aim of the present study was to investigate the influence of the loop diuretic furosemide on the gene expression of the kidney chloride channel ClC-K1 and its recently described functional subunit barttin. Male Sprague Dawley rats received the loop diuretic furosemide (12 mg/kg/day) for 6 days. Rats had free access to 0.9% NaCl, 0.1%KCl solution to prevent volume depletion. Localisation and regulation of ClC-K1 and barttin mRNA was analysed by RNase protection and in situ hybridisation. Nephron-specific regulation was investigated by microdissection and real-time PCR quantification. In furosemide-treated rats ClC-K1 mRNA decreased to half in the inner medulla. In the renal cortex and outer medulla ClC-K1 mRNA levels were weak and did not change. Under furosemide treatment barttin mRNA was regulated in parallel with ClC-K1 mRNA. A significant mRNA decrease occurred after furosemide treatment in inner medulla (0.50 fold), whereas cortical and outer medulla levels remained unaffected. 35S in situ hybridisation confirmed the regulation and distribution seen in the RNase protection assay experiments. Microdissection of the inner medullary collecting duct and thin limb of Henle's loop followed by real-time PCR revealed that CLC-K1 and barttin mRNA regulation in inner medulla was limited to the thin limb; mRNA levels in collecting ducts were not affected by furosemide treatment. Our findings imply that during furosemide treatment selective down-regulation of ClC-K1 and barttin mRNAs in thin limb plays a role in maintaining salt and water homeostasis.
Tài liệu tham khảo
Jentsch TJ, Stein V, Weinreich F, Zdebik AA (2002) Molecular structure and physiological function of chloride channels. Physiol Rev 82:503–568
Adachi S, Uchida S, Ito H, Hata M, Hiroe M, Marumo F, Sasaki S (1994) Two isoforms of a chloride channel predominantly expressed in thick ascending limb of Henle's loop and collecting ducts of rat kidney. J Biol Chem 269:17677–17683
Kieferle S, Fong P, Bens M, Vandewalle A, Jentsch TJ (1994) Two highly homologous members of the ClC chloride channel family in both human and rat kidney. Proc Natl Acad Sci USA 91:6943–6947
Zimniak L, Winters CJ, Reeves WB, Andreoli TE (1996) Cl− channels in basolateral renal medullary vesicles. XI. rb-ClC-Ka cDNA encodes basolateral MTAL Cl− channels. Am J Physiol 270:F1066–F1072
Uchida S, Sasaki S, Furukawa T, Hiraoka M, Imai T, Hirata Y, Marumo F (1993) Molecular cloning of a chloride channel that is regulated by dehydration and expressed predominantly in kidney medulla. J Biol Chem 268:3821–3824
Kere J (1999) Kidney Genetics and chloride ion pumps. Nature Genet 21:67–68
Matsumara M, Uchida S, Kondo Y, Miyazaki H, Ko SBH, Hayama A, Morimoto T, Liu W, Arisawa M, Sasaki S, Marumo F (1999) Overt nephrogenic diabetes insipidus in mice lacking the CLC-K1 chloride channel. Nature Genet 21:95–98
Vandewalle A, Cluzeaud F, Bens M, Kieferle S, Steinmeyer K, Jentsch TJ (1997) Localization and induction by dehydration of CLC-K chloride channels in the rat kidney. Am J Physiol 272:F678–F688
Estevez R, Boettger T, Stein V, Birkenhager R, Otto E, Hildebrandt F, Jentsch TJ (2001) Barttin is a Cl− channel beta-subunit crucial for renal Cl− reabsorption and inner ear K+ secretion. Nature 414:558–561
Waldegger S, Jeck N, Barth P, Peters M, Vitzthum H, Wolf K, Kurtz A, Konrad M, Seyberth HW (2002) Barttin increases surface expression and changes current properties of ClC-K channels. Pflugers Arch 444:411–418
Waldegger S, Jentsch TJ (2000) Functional and structural analysis of ClC-K chloride channels involved in renal disease. J Biol Chem 275:24527–24533
Uchida S, Sasaki, S, Nitta K, Uchida K, Horita S, Nihei H, Marumo F (1995) Localization and functional characterization of rat kidney-specific chloride channel, CLC-K1. J Clin Invest 95:104–113
Vitzthum H, Castrop H, Meier-Meitinger M, Riegger GA, Kurtz A, Krämer BK, Wolf K (2002) Nephron specific regulation of chloride channel ClC-K2 mRNA in the rat. Kidney Int 61:547–554
Miyazaki H, Kaneko T, Uchida S, Sasaki S, Takei Y (2002) Kidney-specific chloride channel, OmClC-K, predominantly expressed in the diluting segment of freshwater-adapted tilapia kidney. Proc Natl Acad Sci USA 99:15782–15787
Chomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156–159
Schafer JA, Watkins ML, Li L, Herter P, Haxelmans S, Schlatter E (1997) A simplified method for isolation of large numbers of defined nephron segments. Am J Physiol 273:F650–F657
Simon DB, Bindra RS, Mansfield TA, Nelson-Williams C, Mendonca E, Stone R, Schurman S, Nayir A, Alpay H, Bakkaloglu A, Rodriguez-Soriano J, Morales JM, Sanjad SA, Taylor CM, Pilz D, Brem A, Trachtman H, Griswold W, Richard GA, John E, Lifton RP (1997) Mutations in the chloride channel gene, CLCNKB, cause Bartter's syndrome type III. Nature Genet 17:171–178
Birkenhager R, Otto E, Schurmann MJ, Vollmer M, Ruf EM, Maier-Lutz I, Beekmann F, Fekete A, Omran H, Feldmann D, Milford DV, Jeck N, Konrad M, Landau D, Knoers NV, Antignac C, Sudbrak R, Kispert A, Hildebrandt F (2001) Mutation of BSND causes Bartter syndrome with sensorineural deafness and kidney failure. Nat Genet. 29:310–314
Winters CJ, Zimniak L, Reeves WB, Andreoli TE (1997) Cl− channels in basolateral renal medullary membranes. XII. Anti-rbClC-Ka antibody blocks MTAL Cl− channels. Am J Physiol 273:F1030–F1038
Winters CJ, Reeves WB, Andreoli TE (1999) Cl− channels in basolateral membranes. XIV. Kinetic properties of a basolateral MTAL Cl− channel. Kidney Int 55:1444–1449
Kobayashi K, Uchida S, Mizutani S, Sasaki S, Marumo F (2001) Intrarenal and cellular localization of CLC-K2 protein in the mouse kidney. J Am Soc Nephrol 12:1327–1334
Castrop H, Kramer BK, Riegger GA, Kurtz A, Wolf K (2000) Overexpression of chloride channel CLC-K2 mRNA in the renal medulla of Dahl salt-sensitive rats. J Hypertens 18:1289–1295
Wolf K, Castrop H, Riegger GA, Kurtz A, Kramer BK (2001) Differential gene regulation of renal salt entry pathways by salt load in the distal nephron of the rat. Pflugers Arch 442:498–504
Sone M, Ohno A, Albrecht GJ, Thurau K, Beck FX (1995) Restoration of urine concentrating ability and accumulation of medullary osmolytes after chronic diuresis. Am J Physiol 269:F480–F490