Specific targeting of layer 4 for direct cortical stimulation is not necessary to induce reliable perception

Abstract The primary somatosensory cortex (S1) processes tactile inputs through microcircuits involving connections between and within layers. In particular, layer 4 is the main first cortical target of tactile inputs. However, the need for the initial activation of layer 4 neurons in order to generate a tactile percept has not been studied yet. Here, we investigated whether optogenetic activation of layer 4 neurons first is necessary to induce perception, or whether stimulation of neurons in other layers may suffice. To achieve this, we generated mouse profiles expressing a light-sensitive opsin in excitatory neurons in different combinations of cortical layers, including or excluding layer 4. We generated these profiles by combining transgenic mouse lines and peripheral injections of PHP.eB adeno-associated viruses. After providing proof-of-concept for neuronal optogenetic control in vivo with PHP.eB viruses, we trained mice in a task involving the discrimination of moving optogenetic stimuli projected onto the whisker S1 area, thanks to patterned photostimulation. Mice learned to track the position of a light bar in order to obtain rewards. Direct activation of layer 4 was sufficient for the mice to learn the task, but not necessary. Indeed, activation of all layers but layer 4 induced learning. Moreover, the simultaneous direct activation of all layers accelerated learning. These findings contribute to a better understanding of the cortical mechanisms underlying somatosensory perception. They could also help to optimize artificial sensory stimuli to provide efficient feedback in cortical neuroprostheses.