Repulsive guidance molecule A (RGMA) is widely expressed in the CNS of the sea lamprey Petromyzon marinus during early development.

RGM interactions with its receptor Neogenin play an important role in the regulation of axonal guidance or cell death in the developing central nervous system. The sea lamprey RGMA transcript has been recently identified. However, its expression has been only studied in the spinal cord of mature (premetamorphic) larval sea lampreys. Here, we report the expression of the sea lamprey RGMA transcript in developing embryos and prolarvae by means of in situ hybridization. Our data show that the RGMA transcript is broadly expressed in the central nervous system of embryos and prolarvae and with a rostro-caudal gradient of expression.

Spontaneous regeneration of cholecystokinergic reticulospinal axons after a complete spinal cord injury in sea lampreys.

In contrast to humans, lampreys spontaneously recover their swimming capacity after a complete spinal cord injury (SCI). This recovery process involves the regeneration of descending axons. Spontaneous axon regeneration in lampreys has been mainly studied in giant descending neurons. However, the regeneration of neurochemically distinct descending neuronal populations with small-caliber axons, as those found in mammals, has been less studied. Cholecystokinin (CCK) is a regulatory neuropeptide found in the brain and spinal cord that modulates several processes such as satiety, or locomotion. CCK shows high evolutionary conservation and is present in all vertebrate species. Work in lampreys has shown that all CCKergic spinal cord axons originate in a single neuronal population located in the caudal rhombencephalon. Here, we investigate the spontaneous regeneration of CCKergic descending axons in larval lampreys following a complete SCI. Using anti-CCK-8 immunofluorescence, confocal microscopy and lightning adaptive deconvolution, we demonstrate the partial regeneration of CCKergic axons (81% of the number of axonal profiles seen in controls) 10 weeks after the injury. Our data also revealed a preference for regeneration of CCKergic axons in lateral spinal cord regions. Regenerated CCKergic axons exhibit colocalization with synaptic vesicle marker SV2, indicative of functional synaptic connections. We also extracted swimming dynamics in injured animals by using DeepLabCut. Interestingly, the degree of CCKergic reinnervation correlated with improved swimming performance in injured animals, suggesting a potential role in locomotor recovery. These findings open avenues for further exploration into the role of specific neuropeptidergic systems in post-SCI spinal locomotor networks.

An in vivo drug screen in zebrafish reveals that cyclooxygenase 2-derived prostaglandin D2 promotes spinal cord neurogenesis.

The study of neurogenesis is essential to understanding fundamental developmental processes and for the development of cell replacement therapies for central nervous system disorders. Here, we designed an in vivo drug screening protocol in developing zebrafish to find new molecules and signalling pathways regulating neurogenesis in the ventral spinal cord. This unbiased drug screen revealed that 4 cyclooxygenase (COX) inhibitors reduced the generation of serotonergic interneurons in the developing spinal cord. These results fitted very nicely with available single-cell RNAseq data revealing that floor plate cells show differential expression of 1 of the 2 COX2 zebrafish genes (ptgs2a). Indeed, several selective COX2 inhibitors and two different morpholinos against ptgs2a reduced the number of serotonergic neurons in the ventral spinal cord and led to locomotor deficits. Single-cell RNAseq data and different pharmacological manipulations further revealed that COX2-floor plate-derived prostaglandin D2 promotes neurogenesis in the developing spinal cord by promoting mitotic activity in progenitor cells. Rescue experiments using a phosphodiesterase-4 inhibitor suggest that intracellular changes in cAMP levels underlie the effects of COX inhibitors on neurogenesis and locomotion. Our study provides compelling in vivo evidence showing that prostaglandin signalling promotes neurogenesis in the ventral spinal cord.

Neogenin expression in ependymo-radial glia of the larval sea lamprey Petromyzon marinus spinal cord.

Neogenin is a receptor mainly known for its roles during axon pathfinding. However, neogenin is expressed in neuronal precursors of ventricular and subventricular zones of the nervous system and recent work has shown that it regulates adult neurogenesis. Here, we generated an antibody against the sea lamprey neogenin to study its expression in the larval spinal cord. Immunofluorescence experiments show that neogenin is expressed in ependymo-radial glial cells (ERGs) located in the ependymal region of the central canal of mature larval sea lampreys. Our results provide a basis for the future study of the role of neogenin in lamprey ERGs.

Spinal cord RNA-seq data after a baclofen treatment in mice with a spinal cord injury.

Spinal cord injury (SCI) leads to severe functional deficits. Currently, there are no available pharmacological treatments to promote neurological recovery in SCI patients. Recent work from our group has shown that a baclofen treatment can promote functional recovery after a compression SCI in mice [1]. Here, we provide transcriptomic (RNA-seq) data from adult mouse spinal cords collected 7 days after a compression SCI and baclofen (vs vehicle) administration. The Illumina NovaSeq 6000 platform was used to generate the raw transcriptomic data. In addition, we also present bioinformatic analyses including differential gene expression analysis, enrichment analyses for various functional annotations (gene ontology, KEGG and BioCarta pathways or InterPro domains) and transcription factor targets. The raw RNA-seq data has been uploaded to the NCBI Sequence Read Archive (SRA) database (Bioproject ID PRJNA886048). The data generated from the bioinformatic analyses is contained within the article.

Organization of the corticotropin-releasing hormone and corticotropin-releasing hormone-binding protein systems in the central nervous system of the sea lamprey Petromyzon marinus.

The expression of the corticotropin-releasing hormone (PmCRH) and the CRH-binding protein (PmCRHBP) mRNAs was studied by in situ hybridization in the brain of prolarvae, larvae, and adults of the sea lamprey Petromyzon marinus. We also generated an antibody against the PmCRH mature peptide to study the distribution of PmCRH-immunoreactive cells and fibers. PmCRH immunohistochemistry was combined with antityrosine hydroxylase immunohistochemistry, PmCRHBP in situ hybridization, or neurobiotin transport from the spinal cord. The most numerous PmCRH-expressing cells were observed in the magnocellular preoptic nucleus-paraventricular nucleus and in the superior and medial rhombencephalic reticular formation. PmCRH expression was more extended in adults than in larvae, and some cell populations were mainly (olfactory bulb) or only (striatum, ventral hypothalamus, prethalamus) observed in adults. The preopto-paraventricular fibers form conspicuous tracts coursing toward the neurohypophysis, but many immunoreactive fibers were also observed coursing in many other brain regions. Brain descending fibers in the spinal cord mainly come from cells located in the isthmus and in the medial rhombencephalic reticular nucleus. The distribution of PmCRHBP-expressing neurons was different from that of PmCRH cells, with cells mainly present in the septum, striatum, preoptic region, tuberal hypothalamus, pretectum, pineal complex, isthmus, reticular formation, and spinal cord. Again, expression in adults was more extended than in larvae. PmCRH- and PmCRHBP-expressing cells are different, excluding colocalization of these substances in the same neuron. Present findings reveal a complex CRH/CRHBP system in the brain of the oldest extant vertebrate group, the agnathans, which shows similarities but important divergences with that of mammals.

Full regeneration of descending corticotropin-releasing hormone axons after a complete spinal cord injury in lampreys.

Sea lampreys are a vertebrate model of interest for the study of spontaneous axon regeneration after spinal cord injury (SCI). Axon regeneration research in lampreys has focused on the study of giant descending neurons, but less so on neurochemically-distinct descending neuronal populations with small caliber axons. Corticotropin-releasing hormone (CRH) is a neuropeptide that regulates the stress response or locomotion. CRH is also a neuropeptide of interest in the SCI context because descending CRHergic projections from the Barrington's nucleus control micturition behavior in mammals. Recent work from our group revealed that in sea lampreys the CRHergic innervation of the spinal cord is only of descending origin. Thus, the lack of intrinsic CRH spinal cord neurons provides the opportunity to analyze the regeneration of this descending system by using immunofluorescence methods. Here, we used an antibody against the sea lamprey mature CRH peptide, confocal microscopy, lightning adaptive deconvolution, and ImageJ to analyze the regenerative capacity of the descending CRH-immunoreactive (-ir) axons of larval sea lampreys after a complete SCI at the level of the fifth gill. At 10 weeks post-lesion, when behavioral analyses showed that injured animals had recovered normal appearing locomotion, our results revealed a full recovery of the number of CRH-ir profiles (axons) at the level of the sixth gill. Thus, the CRH descending axons of lampreys fully regenerate after a complete SCI. Our study provides a new model to study spontaneous and successful axonal regeneration in a specific neuronal type with small caliber axons by using simple immunohistochemical methods.