Recent results indicate that nitric oxide (NO) can play an important role in neuronal excitability by modifying the strength of activated synapses and regulating local cerebral blood flow. We sought to determine whether the level of NO synthase (NOS) could, in turn, also be regulated by neural activity. Results using a polyclonal anti-NOS antibody showed that, in cortical area V1 of monocular monkeys, NOS-immunoreactivity is diminished in lamina 4C neuropil of the deprived ocular dominance columns relative to the immediately adjacent non-deprived columns. Closer examination of lamina 4C indicated that the intercolumnar difference in NOS-immunoreactivity does not reflect differences in the distribution of NOS-labeled perikarya, since relatively few neurons were immunoreactive for NOS in lamina 4C of either monocular or normal binocular monkeys. Electron microscopy revealed that the majority (> 80%) of NOS-immunoreactive profiles in lamina 4C are axon terminals. NOS-immunoreactive spines and dendritic shafts also are present but these are more prevalent in the superficial laminae. In order to determine whether the intercolumnar differences in lamina 4C neuropil correspond to altered densities of NOS cells in the superficial laminae, we performed a series of quantitative analyses. In the superficial laminae, NOS-cells occur as two distinguishable classes: a few that are large and intensely NOS-immunoreactive and many more (ca. 24-fold) that are small and lightly immunoreactive. Analysis of the distribution of 559 small and 105 large NOS-immunoreactive cells within 40-microns-thick tangential sections spanning laminae 2-3 showed that the number of cells (large and small together) associated with each blob is approximately 14 for both deprived (lighter) and non-deprived (darker) blobs. These cells are distributed evenly from the center to periphery of columns. Analysis of the distribution of NOS-cells in the infragranular laminae also did not reveal any columnar differences. These observations suggest that local neural activity may be coupled to NO release via alteration of NOS protein levels specifically within distal axonal processes of neurons. This mechanism could operate in conjunction with the more instantaneous catalytic activation of NOS. Ultrastructural analyses further suggest that NO may act as an anterograde and retrograde messenger arising from terminals in addition to its proposed role as a retrograde messenger arising from dendrites.