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The chloride channel ClC-2 contributes to the inwardly rectifying Cl^- conductance in cultured porcine choroid plexus epithelial cells

Hidetoshi Kajita, Koichiro Omori and Hiroko Matsuda

1. The contribution of ClC-2 protein to the inwardly rectifying Cl^- conductance in cultured porcine choroid plexus epithelial cells was investigated using Western analysis and whole-cell current recordings.

2. Inwardly rectifying currents were elicited by hyperpolarizing voltage at a potential more negative than -50 mV in the presence of intracellular protein kinase A (PKA). The relative halide selectivity estimated from the shift in the reversal potential (E_rev) was I^- > Br^- > Cl^- > F^-.

3. Extracellular vasoactive intestinal peptide (VIP) activated the same currents in a dose-dependent manner with a half-maximal concentration of 167·3 nM. H-89 (a PKA inhibitor) interfered with the current activation by VIP.

4. The Cl^- channel was inhibited by external Cd²⁺, Ba²⁺or H⁺, but only weakly inhibited by known Cl^- channel blockers including glibenclamide, NPPB, DIDS and anthracene-9-carboxylic acid (9AC).

5. A specific antibody to ClC-2 detected a 79 kDa protein in porcine choroid plexus cells, which was reduced in cells treated with antisense oligodeoxynucleotide for ClC-2. Both PKA and VIP failed to activate the inwardly rectifying Cl^- currents in cells transfected with the antisense oligodeoxynucleotide, while they activated the currents in cells transfected with GFP alone or the control oligodeoxynucleotide randomized from antisense oligonucleotide.

6. It is concluded that ClC-2 protein contributes to the inwardly rectifying Cl^- conductance in porcine choroid plexus epithelial cells.

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