During postnatal development, neural circuits undergo extensive refinement to establish efficient and mature connectivity. A key mechanism underlying this process is axonal pruning, whereby redundant projections are selectively eliminated in an activity-dependent manner. The corticospinal tract (CST), essential for voluntary motor control, undergoes large-scale pruning during early postnatal life, a process in which glia—particularly microglia, the brain’s resident immune cells—play an active role through the recognition and phagocytosis of supernumerary axons and synapses.

Neuronal activity is a critical signal for glial engagement during circuit refinement. Snap25, a presynaptic protein required for synaptic vesicle fusion and regulated neurotransmitter release, is a central regulator of activity-dependent signalling. Previous work from our laboratory demonstrated that selective loss of Snap25 in cortical layer 5 neurons reduce synaptic transmission, alters glia–neuron interactions, and leads to sex-specific differences in microglial activity.

Here, we investigate whether Snap25-dependent neuronal activity is required for CST axonal pruning and whether glia contribute to this process. Using Cre-dependent AAV-mediated labelling, we targeted primary motor cortex (M1) neurons in male and female Snap25 conditional knockout and wild-type mice at postnatal day 8. Brains were collected four weeks later and analysed using fluorescence imaging to assess CST projections. All animal experiments were conducted in accordance with the UK Animals (Scientific Procedures) Act, 1986 (ASPA), and approved by the university’s ethical review committee.

Preliminary findings indicate that, in the absence of Snap25, the normally eliminated M1 projection to the superior colliculus persists, suggesting a failure of large-scale CST pruning. These results support a role for Snap25-dependent neuronal activity in driving glia-mediated circuit refinement. Ongoing quantitative and glial analyses will further elucidate the mechanisms by which activity-dependent signals shape CST development and whether these processes differ between sexes.