The molecular identity of the membrane Cl- channels responsible for cell volume homeostasis has not been defined. Since previous studies have shown that Xenopus-expressed ClC-2 channels activate during hypotonic exposure, our purpose was to evaluate whether endogenous mammalian ClC-2 channels contribute to volume-sensitive changes in membrane Cl- permeability. A cDNA closely homologous with rat brain ClC-2 was isolated from a model liver cell line that demonstrates swelling-activated currents typical of most mammalian cell types. Abundant cellular expression of ClC-2 mRNA and membrane localization of ClC-2 protein was shown by in situ hybridization and immunocytochemistry. Intracellular delivery of antibodies to the cytoplasmic amino-terminal region of ClC-2 thought to be important for channel gating nearly abolished Cl- current activation by volume increases or exposure to ATP. lntracellular injection of ClC-2 antibodies also prevented recovery of cell volume following swelling. The characteristics of volume-and ATP-stimulated unitary currents include a slope conductance that averaged 7.6 pS in symmetrical Cl- containing solutions, reversal near ECl, and a permeability ratio of NO3- > Cl- > aspartate. These studies provide the first direct evidence that native mammalian ClC-2 channel proteins contribute to membrane Cl- permeability and volume homeostasis, and may have important implications for other cell types in light of the near ubiquitous expression of ClC-2.
|Original language||English (US)|
|State||Published - Mar 20 1998|
ASJC Scopus subject areas
- Molecular Biology