A Novel voltage-dependent chloride current activated by extracellular acidic pH in cultured rat Sertoli cells

J Biol Chem. 2003 May 23;278(21):19230-6. doi: 10.1074/jbc.M301096200. Epub 2003 Mar 11.

Abstract

Sertoli cells from mammalian testis are key cells involved in development and maintenance of stem cell spermatogonia as well as secretion of a chloride- and potassium-rich fluid into the lumen of seminiferous tubules. Using whole-cell patch clamp experiments, a novel chloride current was identified. It is activated only in the presence of an extracellular acidic pH, with an estimated half-maximal activation at pH 5.5. The current is strongly outwardly rectifying, activated with a fast time-dependent onset of activation but a slow time-dependent kinetic at depolarization pulses. The pH-activated chloride current was not detected at physiological or basic pH and is not sensitive to intracellular or extracellular Ca2+ variation. Diphenylamine-2-carboxylic acid and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid blocked the induced currents, and its anionic selectivity sequence was Cl- > Br- > I-> gluconate. We have performed a reverse transcription-PCR analysis to search for voltage-dependent chloride rClC channels in cultured rat Sertoli cells. Among the nine members of the family only rClC-2, rClC-3, rClC-6, and rClC-7 have been identified. The inwardly rectifying rClC-2 chloride current was activated by hyperpolarization but not by pH variation. A different depolarization-activated outwardly rectifying chloride current was activated only by hypotonic challenge and may correspond either to rClC-3 or rClC-6. Immunolocalization experiments demonstrate that rClC-7 resides in the intracellular compartment of Sertoli cells. This study provides the first functional identification of a native acid-activated chloride current. Based on our molecular analysis of rClC proteins, this new chloride current does not correspond to rClC-2, rClC-3, rClC-6, or rClC-7 channels. The potential physiological role of this native current in an epithelial cell from the reproductive system is discussed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / pharmacology
  • Animals
  • CLC-2 Chloride Channels
  • Calcium / metabolism
  • Cells, Cultured
  • Chloride Channels / analysis
  • Chloride Channels / genetics
  • Chloride Channels / physiology*
  • Chlorides / metabolism
  • Electric Conductivity
  • Gene Expression
  • Hydrogen-Ion Concentration
  • Hypotonic Solutions
  • Male
  • Oocytes / metabolism
  • Patch-Clamp Techniques
  • RNA, Messenger / analysis
  • Rats
  • Rats, Wistar
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sertoli Cells / chemistry
  • Sertoli Cells / physiology*
  • Transfection
  • Xenopus laevis
  • ortho-Aminobenzoates / pharmacology

Substances

  • CLC-2 Chloride Channels
  • Chloride Channels
  • Chlorides
  • ClC-3 channel
  • Clcn6 protein, rat
  • Clcn7 protein, rat
  • Hypotonic Solutions
  • RNA, Messenger
  • ortho-Aminobenzoates
  • fenamic acid
  • 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid
  • Calcium