In slices of visual cortex, long-term potentiation (LTP) of synaptic responses in layer III can be evoked by high-frequency stimulation of a site in the middle of the cortical thickness, corresponding mainly to layer IV. In contrast, stimulation of the white matter-layer VI border typically fails to evoke LTP in adult visual cortex unless GABAA receptors are partially blocked. We performed current-source density (CSD) analysis to determine how the patterns of cortical activation compare under these different stimulation conditions. Single-pulse stimulation of the middle layers (corresponding to layer IV and superficial V) and the deep layers (corresponding to white matter and deep layer VI) yielded very similar CSD patterns. The major current sinks were located within 500 mu m of the pia, corresponding to layers II and III, regardless of the stimulation site. The amplitude of all current sinks was diminished, and the latency was increased, in the presence of high concentrations of divalent cations (12 mM Ca2+ and 12 mM mg2+). Nonetheless, the major synaptic current sink was still present at a depth of approximately 400 microns regardless of the site of stimulation, indicating that stimulation of either site leads to monosynaptic EPSCs in layer III. However, superficial sinks, at a depth of approximately 200 microns, were virtually eliminated by high concentrations of divalent cations after deep layer stimulation, but not after middle layer stimulation, suggesting that stimulation at the two sites recruits different monosynaptic circuits. This conclusion was supported by experiments using paired-pulse stimulation of the two sites (12.5 ms interstimulus interval). While there was little evidence of a paired-pulse interaction after stimulation of the middle layers, there was marked paired-pulse suppression of superficial layer III current sinks after stimulation of the deep layers. Taken together, the data suggest a model in which deep layer stimulation activates the dendrites of layer III cells by a monosynaptic route and by a disynaptic route. The disynaptic input originates in the middle cortical layers and is controlled by inhibition. Differences in synaptic plasticity evoked from the different sites could be explained if the recruitment of middle layer inputs were required for the generation of LTP in layer III.