In the last few years, optogenetic tools and optical functional indicators are increasingly used together to perform simultaneous manipulation and recording of neuronal activity. Nevertheless, this method has still some limitations mainly due to the spectral cross-talk between optogenetic actuators and functional sensors [1;2]. To address this issue, red variants of genetically encoded calcium indicators (red-GECIs) have been recently developed [3;4]. The main goal of this project is to develop a full-optical system that allows effective interrogation of brain circuits. To this aim, we combined a red-shifted calcium indicator (jRCaMP1a), with the most common blue-light activated opsin, Channelrhodopsin II (ChR2). The results presented here show: (I) extended expression of the full-optical system that covers all the motor areas, (II) functional correlation between the laser power and the evoked neuronal activity, (III) segregation of the cortical functional areas of two different forelimb evoked movements. The future perspective of this project concerns the study of the functional areas correlation during optogenetically-evoked forelimb complex movements.
Neuro-rehabilitative research is developing novel strategies to enhance the effectiveness of therapies after stroke by using a combination of physical and plasticizing treatments <sup>1-3</sup>. Previous studies have shown that repeated optogenetics stimulation of neurons in the peri-lesioned area induces a significant improvement in cerebral blood flow and neurovascular coupling response <sup>4-6</sup>. Up to now the mechanisms underneath the reshaping of brain circuitry induced by rehabilitation after stroke are widely unknown. To investigate how rehabilitative therapies shape new cortical maps in the peri-infarct region, we induce a photothrombotic stroke in the primary motor cortex and the expression of Channelrhodopsine 2 (ChR2) in the peri-infarct area on Thy1-GCaMP6f mice. To promote functional recovery after stroke we use both an optogenetic strategy to stimulate targeted excitatory neurons in the peri-lesional region and motor training on a robotic platform (M-Platform) <sup>7</sup>. A 473 nm laser repeatedly stimulates ChR2-transfected neurons; the optostimulation is performed five days a week. The motor rehabilitation consists in a pulling task: after the forelimb is passively extended by the linear actuator of the M-platform, the animal has to pull back up to the resting position. By analysing the spatio-temporal calcium dynamic and the reshaping of cortical activation area during the movement throughout the treatment period, we found that the combined treatment restores cortical activation profiles during the forelimb movement. Through behavioural experiments, using Schallert test, we also evaluate changes of forelimb functionality during rehabilitation. Our combination of techniques allows obtaining unprecedented views on cortical plasticity induced by rehabilitative therapies.