Volatile anesthetics depress neurotransmitter release in a brain region- and neurotransmitter-selective manner by unclear mechanisms. Voltage-gated sodium channels (Nas), which are coupled to synaptic vesicle exocytosis, are inhibited by volatile anesthetics through reduction of peak current and modulation of gating. Subtype-selective effects of anesthetics on Na might contribute to observed neurotransmitter-selective anesthetic effects on release. We analyzed anesthetic effects on Na currents mediated by the principal neuronal Na subtypes Na1.1, Na1.2, and Na1.6 heterologously expressed in ND7/23 neuroblastoma cells using whole-cell patch-clamp electrophysiology. Isoflurane at clinically relevant concentrations induced a hyperpolarizing shift in the voltage dependence of steady-state inactivation and slowed recovery from fast inactivation in all three Na subtypes, with the voltage of half-maximal steady-state inactivation significantly more positive for Na1.1 (-49.7 ± 3.9 mV) than for Na1.2 (-57.5 ± 1.2 mV) or Na1.6 (-58.0 ± 3.8 mV). Isoflurane significantly inhibited peak Na current () in a voltage-dependent manner: at a physiologically relevant holding potential of -70 mV, isoflurane inhibited peak of Na1.2 (16.5% ± 5.5%) and Na1.6 (18.0% ± 7.8%), but not of Na1.1 (1.2% ± 0.8%). Since Na subtypes are differentially expressed both between neuronal types and within neurons, greater inhibition of Na1.2 and Na1.6 compared with Na1.1 could contribute to neurotransmitter-selective effects of isoflurane on synaptic transmission.