• Kevin E Conway and Joseph F Cotten.
• Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Boston, Massachusetts 02114, USA.
• Mol. Pharmacol. 2012 Mar 1;81(3):393-400.

AbstractTASK-3 (KCNK9) tandem-pore potassium channels provide a volatile anesthetic-activated and Gα(q) protein- and acidic pH-inhibited potassium conductance important in neuronal excitability. Met-159 of TASK-3 is essential for anesthetic activation and may contribute to the TASK-3 anesthetic binding site(s). We hypothesized that covalent occupancy of an anesthetic binding site would irreversibly activate TASK-3. We introduced a cysteine at residue 159 (M159C) and studied the rate and effect of Cys-159 modification by N-ethylmaleimide (NEM), a cysteine-selective alkylating agent. TASK-3 channels were transiently expressed in Fischer rat thyroid cells, and their function was studied in an Ussing chamber. NEM irreversibly activated M159C TASK-3, with minimal effects on wild-type TASK-3. NEM-modified M159C channels were resistant to inhibition by both acidic pH and active Gα(q) protein. M159C channels that were first inhibited by Gα(q) protein were more-slowly activated by NEM, which suggests protection of Cys-159, and similar results were observed with isoflurane activation of wild-type TASK-3. M159W and M159F TASK-3 mutants behaved like NEM-modified M159C channels, with increased basal currents and resistance to inhibition by active Gα(q) protein or acidic pH. TASK-3 wild-type/M159C dimers expressed as a single polypeptide demonstrated that modification of a single Cys-159 was sufficient for TASK-3 activation, and M159F/M159C and M159W/M159C dimers provided evidence for cross-talk between subunits. The data are consistent with residue 159 contributing to an anesthetic regulatory site or sites, and they suggest that volatile anesthetics, through perturbations at a single site, increase TASK-3 channel activity and disrupt its regulation by active Gα(q) protein, a determinant of central nervous system arousal and consciousness.

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