Spontaneously Regenerative Corticospinal Neurons in Mice
The mammalian spinal cord receives input from the motor cortex via the corticospinal tract (CST). The CST comprises corticospinal neuron (CSN) axons that begin in Layer 5b of motor cortex and can extend up to one meter to reach targets in the caudal spinal cord in humans. CSNs regulate the spinal cord's sensorimotor function; however, the extent of their impact on motor control differs between species. In macaques, lesion of the pyramidal tract produces immediate limb paralysis. In rodents, their genetic ablation profoundly impacts skilled-reaching ability but does not produce paralysis. In almost all examined mammals, corticospinal axons primarily terminate contralateral to their cell bodies of origin. Despite the predominance of contralateral CSN termination, a minority of CSN axons terminate ipsilaterally. There has been considerable examination of ipsilateral axons in unilateral cortical injury because of evidence that they are a potential circuit for the uninjured cortex to compensate for the damaged one. However, little is known about their anatomy and function in healthy animals. Though sparse, these ipsilateral axons are a remarkable deviation from the norm. Brains have evolved only in bilaterally symmetric animals, so the midline is a foundational architectural principle of all central nervous systems. Accordingly, an intricate and redundant ensemble of molecular signals exerts tight control over midline crossing. The ipsilateral axons represent a consistent anomaly across many species at an evolutionarily ancient midline boundary that is under strict developmental control. To better understand these ipsilateral axons, we first sought to characterize their anatomy with a diverse array of emerging methods in mouse molecular neuroscience. Using anterograde tracing methods, tissue clearing, and Smart-seq3 single-nucleus RNA-sequencing (snRNA-seq), we found that ipsilateral CSN axons project to regions of the ventral horn, including directly to motor neurons. Barcode-based Multiplexed Analysis of Projections by Sequencing (MAPseq) of the CST revealed that the neurons contributing these axons primarily comprise a class of bilaterally projecting CSNs and represent a more substantial population of total CSNs than their sparse ipsilateral axons suggest. With retrograde tracing and tissue clearing, we found that the cell bodies of this neuronal population reside in distinct cortical regions, almost entirely absent from the caudo-lateral cortex. We deeply profiled their molecular characteristics using the viral implementation of Translating Ribosome Affinity Purification (vTRAP) and discovered a striking similarity to the embryonic-like molecular signature of regenerating corticospinal neurons. We noticed that the anatomical characteristics of IP-CSNs we had documented were shared with regenerating CSNs: in addition to molecular similarity, both share bilaterality, projection to ventral spinal cord regions, and motor neuronal connectivity. Given these similarities, we hypothesized that IP-CSNs might themselves have regenerative properties. Finally, we show that IP-CSNs are spontaneously regenerative. The discovery of a class of spontaneously regenerative CSNs may prove valuable to the study of spinal cord injury. Additionally, this work suggests that the retention of juvenile-like characteristics may be a widespread phenomenon in adult nervous systems.
corticospinal neurons, ipsilateral projections, motor cortex, spinal cord, axon regeneration, molecular profiling, Life Sciences
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