Functional characterization and cell immunolocalization of AQP-CD water channel in kidney collecting duct

Kiyohide Fushimi, Sei Sasaki, Tadashi Yamamoto, Matsuhiko Hayashi, Tetsuji Furukawa, Shinichi Uchida, Michio Kuwahara, Kenichi Ishibashi, Masanobu Kawasaki, Itaru Kihara, Fumiaki Marumo

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Abstract

Vasopressin-regulated water permeability of the kidney collecting duct is a key component of the urine concentration machinery. Recently, a cDNA for AQP-CD, the vasopressin-regulated water channel, initially reported as WCH-CD, has been isolated (K. Fushimi, S. Uchida, Y. Hara, Y. Hirata, F. Marumo, and S. Sasaki. Nature Lond. 361: 549-552, 1993). AQP-CD was expressed in oocyte membrane using a Xenopus expression vector, and functional characteristics of AQP-CD were examined. Osmotic water permeability (Pf) of oocytes expressing AQP-CD was 138 ± 19 μm/s (mean ± SE), 12 times greater than the control (11 ± 3 μm/s), 90% inhibited by 0.3 mM HgCl2, and weakly temperature dependent (energy of activation for Pf was 4.0 kcal/mol). Urea influx measured from 15-min [14C]urea uptake by oocytes injected with AQP-CD/expression vector 1 cRNA was 86 ± 17% of the control. Two-electrode voltage-clamp experiments revealed insignificant ion conductance of AQP-CD. Immunoblots of membranes from rat kidney medulla and oocytes expressing AQP-CD using anti-AQP-CD COOH-terminal antibody showed a 29-kDa protein and 35- to 50-kDa high-molecular-mass forms. Immunohistochemistry showed apical and subapical localization of AQP-CD in the collecting duct principal cells. Our results indicated that AQP-CD is a 29-kDa protein, a selective water channel, distinct from a urea channel, and localized to the membranes of vasopressin-sensitive components in kidney collecting duct principal cells.

Original languageEnglish
JournalAmerican Journal of Physiology - Renal Fluid and Electrolyte Physiology
Volume267
Issue number4 36-4
Publication statusPublished - 1994 Oct
Externally publishedYes

Fingerprint

Collecting Kidney Tubules
Aquaporins
Oocytes
Vasopressins
Urea
Membranes
Permeability
Kidney Medulla
Complementary RNA
Mercuric Chloride
Water
Xenopus
Electrodes
Proteins
Complementary DNA
Immunohistochemistry
Urine
Ions
Temperature
Antibodies

Keywords

  • Osmotic water permeability
  • Urine concentration
  • Vasopressin
  • Water channel of collecting duct

ASJC Scopus subject areas

  • Physiology

Cite this

Fushimi, K., Sasaki, S., Yamamoto, T., Hayashi, M., Furukawa, T., Uchida, S., ... Marumo, F. (1994). Functional characterization and cell immunolocalization of AQP-CD water channel in kidney collecting duct. American Journal of Physiology - Renal Fluid and Electrolyte Physiology, 267(4 36-4).

Functional characterization and cell immunolocalization of AQP-CD water channel in kidney collecting duct. / Fushimi, Kiyohide; Sasaki, Sei; Yamamoto, Tadashi; Hayashi, Matsuhiko; Furukawa, Tetsuji; Uchida, Shinichi; Kuwahara, Michio; Ishibashi, Kenichi; Kawasaki, Masanobu; Kihara, Itaru; Marumo, Fumiaki.

In: American Journal of Physiology - Renal Fluid and Electrolyte Physiology, Vol. 267, No. 4 36-4, 10.1994.

Research output: Contribution to journalArticle

Fushimi, K, Sasaki, S, Yamamoto, T, Hayashi, M, Furukawa, T, Uchida, S, Kuwahara, M, Ishibashi, K, Kawasaki, M, Kihara, I & Marumo, F 1994, 'Functional characterization and cell immunolocalization of AQP-CD water channel in kidney collecting duct', American Journal of Physiology - Renal Fluid and Electrolyte Physiology, vol. 267, no. 4 36-4.
Fushimi, Kiyohide ; Sasaki, Sei ; Yamamoto, Tadashi ; Hayashi, Matsuhiko ; Furukawa, Tetsuji ; Uchida, Shinichi ; Kuwahara, Michio ; Ishibashi, Kenichi ; Kawasaki, Masanobu ; Kihara, Itaru ; Marumo, Fumiaki. / Functional characterization and cell immunolocalization of AQP-CD water channel in kidney collecting duct. In: American Journal of Physiology - Renal Fluid and Electrolyte Physiology. 1994 ; Vol. 267, No. 4 36-4.
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abstract = "Vasopressin-regulated water permeability of the kidney collecting duct is a key component of the urine concentration machinery. Recently, a cDNA for AQP-CD, the vasopressin-regulated water channel, initially reported as WCH-CD, has been isolated (K. Fushimi, S. Uchida, Y. Hara, Y. Hirata, F. Marumo, and S. Sasaki. Nature Lond. 361: 549-552, 1993). AQP-CD was expressed in oocyte membrane using a Xenopus expression vector, and functional characteristics of AQP-CD were examined. Osmotic water permeability (Pf) of oocytes expressing AQP-CD was 138 ± 19 μm/s (mean ± SE), 12 times greater than the control (11 ± 3 μm/s), 90{\%} inhibited by 0.3 mM HgCl2, and weakly temperature dependent (energy of activation for Pf was 4.0 kcal/mol). Urea influx measured from 15-min [14C]urea uptake by oocytes injected with AQP-CD/expression vector 1 cRNA was 86 ± 17{\%} of the control. Two-electrode voltage-clamp experiments revealed insignificant ion conductance of AQP-CD. Immunoblots of membranes from rat kidney medulla and oocytes expressing AQP-CD using anti-AQP-CD COOH-terminal antibody showed a 29-kDa protein and 35- to 50-kDa high-molecular-mass forms. Immunohistochemistry showed apical and subapical localization of AQP-CD in the collecting duct principal cells. Our results indicated that AQP-CD is a 29-kDa protein, a selective water channel, distinct from a urea channel, and localized to the membranes of vasopressin-sensitive components in kidney collecting duct principal cells.",
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