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Differential effects of growth factors in bulbospinal neuron survival and neurite outgrowth in vitro Pataky, David Michael

Abstract

During development, the central nervous system (CNS) is capable of not only growth, but regeneration as well. There are two possible reasons why this ability is lost with maturation: 1) the CNS environment no longer supports growth, and 2) the capacity of the neurons for growth diminishes. There is good evidence that treating injured neurons with the appropriate small polypeptide growth factors can reverse their diminished growth capacity and overcome the inhibition of the mature CNS environment. Neurons in the brainstem with axonal projections to the spinal cord (bulbospinal neurons) are necessary for the initiation and control of many motor behaviours, notably locomotion. After spinal cord injury, these motor behaviours are permanently compromised due to the ineffectiveness of CNS repair. I hypothesized that treating bulbospinal neurons with the appropriate trophic factor(s) will enhance their growth. To address this hypothesis, I designed a novel assay to examine trophic effects on bulbospinal neurons specifically. The assay is based on retrogradely labeling the bulbospinal neurons as they develop axonal projections to the spinal cord in the chick embryo. Subsequently, the brainstem tissue is dissociated (whole or vestibulospinal) or explanted (vestibulospinal or reticulospinal) into culture, creating survival or neurite outgrowth assays respectively. Two families of growth factors, neurotrophins and Fibroblast Growth Factors (FGFs) were examined for trophic effects on bulbospinal neurons. The neurotrophins were largely ineffective, but NT-3 significantly increased neurite outgrowth from reticulospinal (not vestibulospinal) explants. Four FGFs (FGF-1, FGF-2, FGF-5 and FGF-9) were also tested. FGF-2 was the most effective, stimulating survival and neurite outgrowth for all populations examined. FGF-1 only enhanced bulbospinal neurite outgrowth, FGF-9 only survival, and FGF-5 was largely ineffective. At this stage of development, bulbospinal neurons did not express FGF receptors, but non-neuronal cells in situ and in vitro did. Astrocyte-conditioned medium also increased bulbospinal neuron survival, suggesting that perhaps the FGFs act by stimulating the production of the appropriate growth factor from non-neuronal cells. In conclusion, indirect effects may be an important mechanism for growth factor action on bulbospinal neurons. Furthermore, NT-3, FGF-1 and FGF-2 are good candidates for promoting regeneration from injured bulbospinal neurons in vivo.

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