Simultaneous all-optical stimulation and readout of neuronal activity during optogenetically-evoked motor task

The use of light to study neuronal networks has several advantages, such as the noninvasiveness and the possibility to target with high precision specific population of cells. In the past decade, there was a revolution in the use of light for both recording and stimulation of neuronal activity. The development of genetically encoded activity sensors brought us close to single-action-potential sensitivity and, on the other side, optogenetic allows to stimulate or inactivate defined populations of neurons with single-cell precision on millisecond time-scale. However, achieving an all-optical interrogation of neural circuits by combining these tools is still arduous, mainly due to the spectral overlap between actuators and indicators. To this aim we developed a new all-optical system based on a red-shifted GECI (RCaMP1a) combined with channelrhodopsin II (ChR2). Our preliminary results shows that we could induce the expression of RCaMP1a on most of the targeted hemisphere, including motor-associated areas. The coupling of RCaMP1a and ChR2 allowed simultaneous stimulation and readout from the same functional areas. By performing single pulse irradiation we observed that evoked calcium signals rise at increasing laser power, reaching a plateau around 5 mW laser power. We also observed that the stimulated calcium response did not change 3 or 4 weeks after the transfection, indicating the stability of the all-optical system in time and allowing longitudinal experiments. In order to study the cortical activation underlying a specific motor behavior we performed optical stimulation of the Rostral Forelimb Area (RFA) in awake mice. By using a stimulus train we could activate selective movements, like licking and grasping with the contralateral forelimb. Cortical dynamic recorded during the optogenetically-evoked motor task showed correlated activity in the RFA and nearby motor areas. The all-optical system developed here will allow full integration of stimulation and readout of cortical activity of the sensorimotor circuit and the correlated animal behavior, thus shedding light on the neuronal patterns responsible for selected behaviors. Finally, the development of an entirely optical device for interrogation and modulation of motor cortex activity would be a key point in the study of optogenetic-guided rehabilitation after stroke.