Functional Optic Tract Rewiring via Subtype- and Target-specific Axonal Regeneration and Presynaptic Activity Enhancement (scRNA-Seq)

Mechanisms underlying functional axonal rewiring after adult mammalian central nervous system (CNS) injuries remain unclear partially due to limited models. Using a mouse intracranial pre–olivary pretectal nucleus (OPN) optic tract injury model, we demonstrate that Pten/Socs3 knockout and CNTF expression in retinal ganglion cells (RGCs) promotes optic tract regeneration and reinnervation into OPN. Revealed by transmission electron microscopy, trans-synaptic labeling and electrophysiology, functional synapses are formed in OPN mainly by intrinsically photosensitive RGCs, thereby partially restoring the pupillary light reflex (PLR). Furthermore, combining with Lipin-1 knockdown accelerates the recovery and achieves functional reconnection in the chronic injury paradigm. PLR can be further boosted by increasing either RGC photosensitivity with melanopsin or optic tract presynaptic release with a voltage-gated calcium channel modulator, the latter of which also improves corticospinal-tract-sprouting-induced functional recovery after stroke. These findings highlight the importance of neuronal types and presynaptic activity for functional reconnection after CNS injuries. Overall design: We showed that with genetic manipulation as Pten/Socs3 knockout (KO), CNTF overexpression and Lipin1 knockdown (PSCL), RGCs could regenerate and reinnervate into OPN nuclei after pre-OPN optic tract injury. To probe the regenerative transcriptome dynamics, we isolated RGCs labelled with CTB-555 from PSCL regeneration and injury control mice at various time points, conducting single-cell RNA sequencing mainly targeting on OPN-projecting RGCs. We had total 8 groups: GFP-intact, GFP-5dpi (5 days post injury), GFP-1.5mpi (1.5 months post injury), GFP-3mpi, PSCL-intact, PSCL-5dpi, PSCL-1.5mpi, PSCL-3mpi.