Survival and death of mature avian motoneurons in organotypic slice culture: trophic requirements for survival and different types of degeneration |
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Authors: | Brunet Núria Tarabal Olga Portero-Otín Manel Oppenheim Ronald W Esquerda Josep E Calderó Jordi |
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Institution: | Unitat de Neurobiologia Cel.lular, Departament de Ciències Mèdiques Bàsiques, Facultat de Medicina, Universitat de Lleida and IRB Lleida, 25008 Lleida, Catalonia, Spain. |
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Abstract: | We have developed an organotypic culture technique that uses slices of chick embryo spinal cord, in which trophic requirements for long-term survival of mature motoneurons (MNs) were studied. Slices were obtained from E16 chick embryos and maintained for up to 28 days in vitro (DIV) in a basal medium. Under these conditions, most MNs died. To promote MN survival, 14 different trophic factors were assayed. Among these 14, glial cell line-derived neurotrophic factor (GDNF) and vascular endothelial growth factor were the most effective. GDNF was able to promote MN survival for at least 28 DIV. K(+) depolarization or caspase inhibition prevented MN death but also induced degenerative-like changes in rescued MNs. Agents that elevate cAMP levels promoted the survival of a proportion of MNs for at least 7 DIV. Examination of dying MNs revealed that, in addition to cells exhibiting a caspase-3-dependent apoptotic pattern, some MNs died by a caspase-3-independent mechanism and displayed autophagic vacuoles, an extremely convoluted nucleus, and a close association with microglia. This organotypic spinal cord slice culture may provide a convenient model for testing conditions that promote survival of mature-like MNs that are affected in late-onset MN disease such as amyotrophic lateral sclerosis. |
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Keywords: | motoneurons spinal cord neurotrophic factors cell death organotypic slice culture chick embryo |
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