|Title||Isoflurane modulates activation and inactivation gating of the prokaryotic Na+ channel NaChBac|
|Publication Type||Journal Article|
|Year of Publication||2017|
|Authors||Sand RM, Gingrich KJ, Macharadze T, Herold KF, Hemmings HC|
|Journal||The Journal of General Physiology|
Voltage-gated Na+ channels (Nav) have emerged as important presynaptic targets for volatile anesthetic (VA) effects on synaptic transmission. However, the detailed biophysical mechanisms by which VAs modulate Nav function remain unclear. VAs alter macroscopic activation and inactivation of the prokaryotic Na+ channel, NaChBac, which provides a useful structural and functional model of mammalian Nav. Here, we study the effects of the common general anesthetic isoflurane on NaChBac function by analyzing macroscopic Na+ currents (INa) in wild-type (WT) channels and mutants with impaired (G229A) or enhanced (G219A) inactivation. We use a previously described six-state Markov model to analyze empirical WT and mutant NaChBac channel gating data. The model reproduces the mean empirical gating manifest in INa time courses and optimally estimates microscopic rate constants, valences (z), and fractional electrical distances (x) of forward and backward transitions. The model also reproduces gating observed for all three channels in the absence or presence of isoflurane, providing further validation. We show using this model that isoflurane increases forward activation and inactivation rate constants at 0 mV, which are associated with estimated chemical free energy changes of approximately −0.2 and −0.7 kcal/mol, respectively. Activation is voltage dependent (z ≈ 2e0, x ≈ 0.3), inactivation shows little voltage dependence, and isoflurane has no significant effect on either. Forward inactivation rate constants are more than 20-fold greater than backward rate constants in the absence or presence of isoflurane. These results indicate that isoflurane modulates NaChBac gating primarily by increasing forward activation and inactivation rate constants. These findings support accumulating evidence for multiple sites of anesthetic interaction with the channel.