The gray squirrel (Sciurus carolinensis) is a diurnal highly visual rodent with a cone-rich retina. In order to determine which features of visual cortex are common to highly visual mammals and which are restricted to non-rodent species, we studied the laminar organization of response properties in primary visual area V1 of isoflurane-anesthetized squirrels using extra-cellular single-unit recording and sinusoidal grating stimuli. Of the responsive cells, 75% were tuned for orientation. Only 10% were directionally selective, almost all in layer 6, a layer receiving direct input from the dorsal lateral geniculate nucleus (LGN). Cone opponency was widespread, but almost absent from layer 6. Median optimal spatial frequency tuning was 0.21 cpd. Median optimal temporal frequency a high 5.3 Hz. Layer 4 had the highest percentage of simple cells and shortest latency (26 ms). Layers 2/3 had the lowest spontaneous activity and highest temporal frequency tuning. Layer 5 had the broadest spatial frequency tuning and most spontaneous activity. At the layer 4/5 border were sustained cells with high cone opponency. Simple cells, determined by modulation to drifting sinusoidal gratings, responded with shorter latencies, were more selective for orientation and direction, and were tuned to lower spatial frequencies. A comparison with other mammals shows that while the laminar organization of orientation selectivity is variable, the cortical input layers contain more linear cells in most mammals. Nocturnal mammals appear to have more orientation-selective neurons in V1 than diurnal mammals of similar size.