In vertical penetrations through the striate cortex in the tree shrew (Tupaia belangeri), we found regions where neural activity was evoked predominantly by light ON. These were followed by regions where responses were evoked predominantly by light OFF. Histological reconstructions indicated that the ON regions were correlated with layer IVa and the OFF regions were correlated with layer IVb. Local application of cobalt chloride produced a transient cessation of visually evoked activity, suggesting that the electrodes sampled cortical activity rather than lateral geniculate nucleus afferents. These data demonstrate that separate ON and OFF regions are present in the tree shrew striate cortex and suggest that spatially separate, parallel ON and OFF afferent channels extend, in this species, at least through the first synapse in the striate cortex.