RESUMO
Contactin1a (Cntn1a) is a zebrafish homolog of contactin1 (F3/F11/contactin) in mammals, an immunoglobulin superfamily recognition molecule of neurons and oligodendrocytes. We describe conspicuous Cntn1a mRNA expression in oligodendrocytes in the developing optic pathway of zebrafish. In adults, this expression is only retained in glial cells in the intraretinal optic fiber layer, which contains 'loose' myelin. After optic nerve lesion, oligodendrocytes re-express Cntn1a mRNA independently of the presence of regenerating axons and retinal ganglion cells upregulate Cntn1a expression to levels that are significantly higher than those during development. After spinal cord lesion, expression of Cntn1a mRNA is similarly increased in axotomized brainstem neurons and white matter glial cells in the spinal cord. In addition, reduced mRNA expression in the trigeminal/anterior lateral line ganglion in erbb3-deficient mutant larvae implies Cntn1a in Schwann cell differentiation. These complex regulation patterns suggest roles for Cntn1a in myelinating cells and neurons particularly in successful CNS regeneration.
Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Diferenciação Celular/fisiologia , Sistema Nervoso Central/patologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Regeneração Nervosa/fisiologia , Oligodendroglia/fisiologia , Animais , Animais Geneticamente Modificados , Animais Recém-Nascidos , Moléculas de Adesão Celular Neuronais/genética , Sistema Nervoso Central/fisiopatologia , Contactina 1 , Contactinas , Embrião não Mamífero , Enucleação Ocular/métodos , Hibridização In Situ/métodos , Microscopia Eletrônica de Transmissão/métodos , Proteína P0 da Mielina/metabolismo , Glicoproteína Associada a Mielina/metabolismo , Neurônios/fisiologia , Neurônios/ultraestrutura , Oligodendroglia/ultraestrutura , Traumatismos do Nervo Óptico/patologia , Traumatismos do Nervo Óptico/fisiopatologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor ErbB-3/genética , Traumatismos da Medula Espinal/patologia , Traumatismos da Medula Espinal/fisiopatologia , Peixe-Zebra , Proteínas de Peixe-ZebraRESUMO
In zebrafish, the capacity to regenerate long axons varies among different populations of axotomized neurons after spinal cord transection. In specific brain nuclei, 84-92% of axotomized neurons upregulate expression of the growth-related genes GAP-43 and L1.1 and 32-51% of these neurons regrow their descending axons. In contrast, 16-31% of spinal neurons with axons ascending to the brainstem upregulate these genes and only 2-4% regrow their axons. Dorsal root ganglion (DRG) neurons were not observed to regrow their ascending axons or to increase expression of GAP-43 mRNA. Expression of L1.1 mRNA is high in unlesioned and axotomized DRG neurons. In the lesioned spinal cord, expression of growth-related molecules is increased in a substantial population of non-axotomized neurons, suggesting morphological plasticity in the spinal-intrinsic circuitry. We propose that locomotor recovery in spinal-transected adult zebrafish is influenced less by recovery of ascending pathways, but more by regrowth of descending tracts and rearrangement of intraspinal circuitry.
Assuntos
Gânglios Espinais/fisiologia , Regeneração Nervosa/fisiologia , Neurônios/fisiologia , Medula Espinal/fisiologia , Animais , Biomarcadores , Divisão Celular , Proteína GAP-43/análise , Proteína GAP-43/genética , Hibridização In Situ , Neurônios/citologia , RNA Mensageiro , Peixe-ZebraRESUMO
Adult zebrafish, in contrast to mammals, regrow axons descending from the brainstem after spinal cord transection. L1.1, a homolog of the mammalian recognition molecule L1, is upregulated by brainstem neurons during axon regrowth. However, its functional relevance for regeneration is unclear. Here, we show with a novel morpholino-based approach that reducing L1.1 protein expression leads to impaired locomotor recovery as well as reduced regrowth and synapse formation of axons of supraspinal origin after spinal cord transection. This indicates that L1.1 contributes to successful regrowth of axons from the brainstem and locomotor recovery after spinal cord transection in adult zebrafish.
Assuntos
Regeneração Nervosa/fisiologia , Traumatismos da Medula Espinal/fisiopatologia , Medula Espinal/fisiologia , Peixe-Zebra/fisiologia , Animais , Axônios/metabolismo , Tronco Encefálico/fisiopatologia , Cordotomia , Implantes de Medicamento , Esponja de Gelatina Absorvível , Morfolinas/farmacologia , Regeneração Nervosa/efeitos dos fármacos , Oligodesoxirribonucleotídeos/farmacologia , Recuperação de Função Fisiológica , Método Simples-Cego , NataçãoRESUMO
A number of genes affecting axonal projections are currently being identified in zebrafish mutant screens. Analyzing the expression of these genes in the adult brain in relation to specific neuronal populations could yield insights into new functional contexts, such as the successful axonal regeneration in adult zebrafish. Here, we provide a relatively simple procedure for non-radioactive in situ hybridization in sections of adult zebrafish brains in combination with retrograde axonal tracing using the fluorescent neuronal tracer rhodamine dextran amine (RDA). A lesion is inflicted on the spinal cord of adult zebrafish and a crystal of RDA is then applied to the lesion site resulting in retrograde labeling of neurons in the brain through their spinal axons. Six to eighteen days later fish are perfusion-fixed, and in situ hybridization is carried out on vibratome-cut floating sections using a protocol simplified from that used for whole-mounted zebrafish embryos. This procedure leads to robust double labeling of axotomized neurons with RDA and an in situ hybridization signal for the growth-associated protein 43 (GAP-43). This method can be used to identify gene expression in specific populations of projection neurons and to detect changes in gene expression in axotomized neurons in the CNS of adult zebrafish.
