Identification of Intrinsic Axon Growth Modulators for Intact CNS Neurons after Injury.

Functional deficits persist after spinal cord injury (SCI) because axons in the adult mammalian central nervous system (CNS) fail to regenerate. However, modest levels of spontaneous functional recovery are typically observed after trauma and are thought to be mediated by the plasticity of intact circuitry. The mechanisms underlying intact circuit plasticity are not delineated. Here, we characterize the in vivo transcriptome of sprouting intact neurons from Ngr1 null mice after partial SCI. We identify the lysophosphatidic acid signaling modulators LPPR1 and LPAR1 as intrinsic axon growth modulators for intact corticospinal motor neurons after adjacent injury. Furthermore, in vivo LPAR1 inhibition or LPPR1 overexpression enhances sprouting of intact corticospinal tract axons and yields greater functional recovery after unilateral brainstem lesion in wild-type mice. Thus, the transcriptional profile of injury-induced sprouting of intact neurons reveals targets for therapeutic enhancement of axon growth initiation and new synapse formation.

Reorganization of Intact Descending Motor Circuits to Replace Lost Connections After Injury.

Neurons have a limited capacity to regenerate in the adult central nervous system (CNS). The inability of damaged axons to re-establish original circuits results in permanent functional impairment after spinal cord injury (SCI). Despite abortive regeneration of axotomized CNS neurons, limited spontaneous recovery of motor function emerges after partial SCI in humans and experimental rodent models of SCI. It is hypothesized that this spontaneous functional recovery is the result of the reorganization of descending motor pathways spared by the injury, suggesting that plasticity of intact circuits is a potent alternative conduit to enhance functional recovery after SCI. In support of this hypothesis, several studies have shown that after unilateral corticospinal tract (CST) lesion (unilateral pyramidotomy), the intact CST functionally sprouts into the denervated side of the spinal cord. Furthermore, pharmacologic and genetic methods that enhance the intrinsic growth capacity of adult neurons or block extracellular growth inhibitors are effective at significantly enhancing intact CST reorganization and recovery of motor function. Owing to its importance in controlling fine motor behavior in primates, the CST is the most widely studied descending motor pathway; however, additional studies in rodents have shown that plasticity within other spared descending motor pathways, including the rubrospinal tract, raphespinal tract, and reticulospinal tract, can also result in restoration of function after incomplete SCI. Identifying the molecular mechanisms that drive plasticity within intact circuits is crucial in developing novel, potent, and specific therapeutics to restore function after SCI. In this review we discuss the evidence supporting a focus on exploring the capacity of intact motor circuits to functionally repair the damaged CNS after SCI.

Comprehensive Corticospinal Labeling with mu-crystallin Transgene Reveals Axon Regeneration after Spinal Cord Trauma in ngr1-/- Mice.

Spinal cord injury interrupts descending motor tracts and creates persistent functional deficits due to the absence of spontaneous axon regeneration. Of descending pathways, the corticospinal tract (CST) is thought to be the most critical for voluntary function in primates. Even with multiple tracer injections and genetic tools, the CST is visualized to only a minor degree in experimental studies. Here, we identify and validate the mu-crystallin (crym) gene as a high-fidelity marker of the CST. In transgenic mice expressing green fluorescent protein (GFP) under crym regulatory elements (crym-GFP), comprehensive and near complete CST labeling is achieved throughout the spinal cord. Bilateral pyramidotomy eliminated the 17,000 GFP-positive CST axons that were reproducibly labeled in brainstem from the spinal cord. We show that CST tracing with crym-GFP is 10-fold more efficient than tracing with biotinylated dextran amine (BDA). Using crym-GFP, we reevaluated the CST in mice lacking nogo receptor 1 (NgR1), a protein implicated in limiting neural repair. The number and trajectory of CST axons in ngr1(-/-) mice without injury was indistinguishable from ngr1(+/+) mice. After dorsal hemisection in the midthoracic cord, CST axons did not significantly regenerate in ngr1(+/+) mice, but an average of 162 of the 6000 labeled thoracic CST axons (2.68%) regenerated >100 µm past the lesion site in crym-GFP ngr1(-/-) mice. Although traditional BDA tracing cannot reliably visualize regenerating ngr1(-/-) CST axons, their regenerative course is clear with crym-GFP. Therefore the crym-GFP transgenic mouse is a useful tool for studies of CST anatomy in experimental studies of motor pathways. SIGNIFICANCE STATEMENT: Axon regeneration fails in the adult CNS, resulting in permanent functional deficits. Traditionally, inefficient extrinsic tracers such a biotinylated dextran amine (BDA) are used to label regenerating fibers after therapeutic intervention. We introduce crym-green fluorescent protein (GFP) transgenic mice as a comprehensive and specific tool with which to study the primary descending motor tract, the corticospinal tract (CST). CST labeling with crym-GFP is 10 times more efficient compared with BDA. The enhanced sensitivity afforded by crym-GFP revealed significant CST regeneration in NgR1 knock-out mice. Therefore, crym-GFP can be used as a standardized tool for future CST spinal cord injury studies.

Plasticity of intact rubral projections mediates spontaneous recovery of function after corticospinal tract injury.

Axons in the adult CNS fail to regenerate after injury, and therefore recovery from spinal cord injury (SCI) is limited. Although full recovery is rare, a modest degree of spontaneous recovery is observed consistently in a broad range of clinical and nonclinical situations. To define the mechanisms mediating spontaneous recovery of function after incomplete SCI, we created bilaterally complete medullary corticospinal tract lesions in adult mice, eliminating a crucial pathway for voluntary skilled movement. Anatomic and pharmacogenetic tools were used to identify the pathways driving spontaneous functional recovery in wild-type and plasticity-sensitized mice lacking Nogo receptor 1. We found that plasticity-sensitized mice recovered 50% of normal skilled locomotor function within 5 weeks of lesion. This significant, yet incomplete, spontaneous recovery was accompanied by extensive sprouting of intact rubrofugal and rubrospinal projections with the emergence of a de novo circuit between the red nucleus and the nucleus raphe magnus. Transient silencing of this rubro-raphe circuit in vivo via activation of the inhibitory DREADD (designer receptor exclusively activated by designer drugs) receptor hM4di abrogated spontaneous functional recovery. These data highlight the pivotal role of uninjured motor circuit plasticity in supporting functional recovery after trauma, and support a focus of experimental strategies on enhancing intact circuit rearrangement to promote functional recovery after SCI.

Local connection patterns of parvalbumin-positive inhibitory interneurons in rat primary auditory cortex.

In the auditory cortex (AC), GABAergic neurons constitute approximately 15-25% of all neurons. GABAergic cells are present in all sensory modalities and essential for modulating sensory receptive fields. Parvalbumin (PV) positive cells represent the largest sub-group of the GABAergic population in auditory neocortex. We investigated the projection pattern of PV cells in rat primary auditory cortex (AI) with a retrograde tracer (wheat germ apo-HRP conjugated to gold [WAHG]) and immunocytochemistry for PV. All AC layers except layer I contained cells double-labeled for PV and WAHG. All co-localized PV+ cells were within 2 mm of the injection site, regardless of laminar origin. Most (ca. 90%) of the co-localized PV cells were within 500 µm of the injection site in both dorsal-ventral and rostral-caudal dimension of the auditory core region. WAHG-only cells declined less rapidly with distance and were found up to 6 mm from the deposit sites. WAHG-only labeled cells in the medial geniculate body were in ventral division loci compatible with an injection in AI. Differences in the range and direction of the distribution pattern of co-localized PV+ cells and WAHG-only cells in AI express distinct functional convergence patterns for the two cell populations.