AbstractWith the advent of two-photon imaging as a tool for systems neuroscience, the mouse has become a preeminent model system for studying sensory processing. One notable difference that has been found however, between mice and traditional model species like cats and primates is the responsiveness of the cortex. In the primary visual cortex of cats and primates, nearly all neurons respond to simple visual stimuli like drifting gratings. In contrast, imaging studies in mice consistently find that only around half of the neurons respond to such stimuli. Here we show that visual responsiveness is strongly dependent on the cortical depth of neurons. Moving from superficial layer 2 down to layer 4, the percentage of responsive neurons increases dramatically, ultimately reaching levels similar to what is seen in other species. Over this span of cortical depth, neuronal response amplitude also increases and orientation selectivity moderately decreases. These depth dependent response properties may be explained by the distribution of thalamic inputs in mouse V1. Unlike in cats and primates where thalamic projections to the granular layer are constrained to layer 4, in mice they spread up into layer 2/3, qualitatively matching the distribution of response properties we see. These results show that the analysis of neural response properties must take into consideration not only the overall cortical lamina boundaries but also the depth of recorded neurons within each cortical layer. Furthermore, the inability to drive the majority of neurons in superficial layer 2/3 of mouse V1 with grating stimuli indicates that there may be fundamental differences in the role of V1 between rodents and other mammals.