A variety of molecular targets for volatile anesthetics have been suggested including the anesthetic-sensitive K+ leak channel, TREK-1. Knockout of TREK-1 is reported to render mice resistant to volatile anesthetics, making TREK-1 channels compelling targets for anesthetic action. Spinal cord slices from mice, either wildtype or an anesthetic-hypersensitive mutant, Ndufs4, display an isoflurane-induced outward K+ leak that correlates with their minimum alveolar concentrations (MAC) and is blocked by norfluoxetine. We hypothesized that TREK-1 channels conveyed this current and contribute to the anesthetic hypersensitivity of Ndufs4. Our results led to evaluation of a second TREK channel, TREK-2, in control of anesthetic sensitivity.
We measured anesthetic sensitivities of mice carrying knockout alleles of Trek-1 and Trek-2, the double knockout Trek-1;Trek-2, and Ndufs4;Trek-1. Neurons from spinal cord slices from each mutant were patch clamped to characterize isoflurane-sensitive currents. Norfluoxetine was used to identify TREK-dependent currents.
We compared mean values for MAC (+/- SD) between wildtype and two Trek-1 knockout alleles in mice (p-values, Trek-1 compared to wildtype). Wildtype: (MAC(Hal), 1.30%(0.10); MAC(Iso), 1.40%(0.11): Trek-1tm1Lex (MAC(Hal), 1.27%(0.11); p=0.387; MAC(Iso), 1.38%(0.09); p=0.268): Trek-1tm1Lzd (MAC(Hal); 1.27%(0.11); p=0.482: MAC(Iso); 1.41%(0.12); p=0.188). Neither allele was resistant for loss of righting reflex. The EC50s of Ndufs4;Trek-1tm1Lex did not differ from Ndufs4. Ndufs4: (EC50(Hal), 0.65%(0.05); EC50 (Iso), 0.63%(0.05): Ndufs4;Trek-1tm1Lex (EC50(Hal), 0.58%(0.07); p=0.004; EC50(Iso); 0.61%(.06); p=0.442). Loss of TREK-2 did not alter anesthetic sensitivity in a wildtype or Trek-1 genetic background. Loss of TREK-1 or TREK-2, or both, did not alter the isoflurane-induced currents in wildtype cells but did cause them to be norfluoxetine-insensitive.
Loss of TREK channels did not alter anesthetic sensitivity in mice, nor did it eliminate isoflurane-induced transmembrane currents. However, the isoflurane-induced currents are norfluoxetine-resistant in Trek mutants indicating that other channels may function in this role when TREK channels are deleted.