USING TWO-TIMEFRAME RABIES TRACING TO INVESTIGATE LEARNING-RELATED INPUTS TO MOUSE MOTOR CORTEX

Primary motor cortex (M1) has been extensively studied as a hub for motor learning. Previous work has shown that learning a motor task induces the formation of new dendritic spines in M1 in rodents. However, the identity of the presynaptic partners of the newly formed spines remains largely unknown. To investigate the origin of learning-induced inputs to M1, we employ a novel two-timeframe rabies tracing (TTT) approach.
In timeframe 1, TTT labels existing monosynaptic inputs to the forepaw region of M1 in a tdTomato reporter mouse, figuratively taking a “snapshot” of pre-existing connections. In timeframe 2, mice are trained on a forepaw seed grasping task, inducing synaptic turnover in the respective region of M1. Novel inputs formed during the learning phase can be distinguished from pre-existing connections by the combination of fluorophores they express (timeframe 1: rabies-tdStayGold + tdTomato; timeframe 2: rabies-tdStayGold only). Brain-wide assessment of novel, putatively learning-relevant inputs to M1 is carried out using serial sectioning 2p imaging.
In pilot experiments, we have established a method of functionally targeting the M1 forepaw region using intrinsic optical imaging. Next, mice were trained on the grasping task while TTT was carried out. High-speed videos were acquired to quantify behavioral performance. Preliminary results indicate a larger fraction of putatively novel inputs in regions associated with motor learning (e.g. M1 and somatosensory cortices), in trained as compared to untrained animals.
Currently, we are expanding the dataset to assess learning-related changes in presynaptic connectivity to M1 and how they relate to behavioral performance.