Changes in the distribution, complement and/or activity of voltage-gated K+ channels can influence the stability of neural networks through homeostatic effects on neuronal excitability and may be neuroprotective during hyperexcitability (1). In auditory brainstem, NO generated by neuronal NOS in response to synaptic stimulation switches postsynaptic action potential repolarisation from Kv3 to Kv2 K+ channel dominance (2, 3). This switch decreases neuronal excitability and increases transmission fidelity, thereby adapting neuronal output to synaptic input. To determine whether similar modulation occurs at other levels of the auditory system, whole-cell patch recordings were made from morphologically identified pyramidal neurones in slices of mouse auditory cortex. Studies focused in cortical layer 2/3 since immunofluorescence for neuronal NOS was predominant in this area (n = 3 mice). Data are means ± SEM. Under current clamp, and consistent with past reports (4), layer 2/3 neurones fired action potentials with a rheobase around 50 pA and a peak frequency of 39 ± 1 Hz (n = 12 cells).The NOS antagonist L-nitroarginine (L-NNA, 100 µM) significantly reduced the peak frequency (29 ± 3 Hz; n = 14; unpaired t test, p < 0.05). Under voltage clamp, NOS inhibition by L-NNA or 7-nitroindazole (10 µM) significantly increased the amplitude of the whole-cell outward current (at 50 mV, currents ± 1 nA were 11 nA, 19 nA and 16 nA for control, L-NNA and 7-nitroindazole, respectively; n = 5-20 cells. ANOVA with Tukey test, control vs. either inhibitor, p < 0.05; L-NNA vs. 7-nitroindazole, p > 0.05). Immunofluorohistochemistry revealed protein for Kv2 K+ channel subunits throughout auditory cortex, including in micron-sized clusters in layer 2/3 pyramidal neuron soma and proximal dendrites (n = 3 mice). Under voltage clamp the Kv2 channel gating modifier guangxitoxin-1E (100 nM) reversed the effect of NOS inhibition (100 µM L-NNA) on the amplitude of the whole-cell outward current (currents ± 1 nA were 18 nA, 14 nA and 12 nA at 50 mV for L-NNA, L-NNA plus guangxitoxin-1E, and control respectively. ANOVA with Tukey test, L-NNA vs. L-NNA plus toxin, p < 0.05; L-NNA plus toxin vs. control, p > 0.05; n = 6-7 cells), but had no effect on the amplitude of the control current (control ± toxin, p > 0.05; n = 6-7 cells). These data suggest that NO, produced tonically or in response to spontaneous synaptic activity, persistently augments pyramidal neuron excitability in layer 2/3 of the auditory cortex, possibly by inhibiting Kv2-mediated voltage-gated K+ currents. NO therefore appears to control cell excitability at multiple levels of the auditory system. Important differences exist in the effect of NO between auditory cortex and brainstem (3), but Kv2 K+ channels appear to be a common downstream target.