The effects of quisqualate and nocodazole on the organization of MAP2 and neurofilaments in spinal cord neurons in vitro.
The relationship between microtubules, neurofilaments and microtubule-associated protein (MAP)2 was investigated in spinal cord neurons grown for up to 14 days in vitro. Neurons were labelled using antibodies against MAP2, neurofilaments and tubulin, and immunofluorescence analyzed by confocal microscopy. A well-structured network of neurofilaments and microtubules was observed in unstimulated cultures. MAP2 staining was poorly structured but became more filamentous following depolymerization of microtubules with nocodazole. Double-staining experiments suggested that MAP2 was now closely associated with neurofilaments in cell bodies and dendrites. Stimulation of cultures with excitatory amino acids increased the resistance of the microtubular cytoskeleton to depolymerization by nocodazole. Again double-labelling experiments demonstrated an increased association between neurofilaments and MAP2 immunofluorescence. Previous results suggested that the stability of the neuronal cytoskeleton could be modulated by glutamate receptors acting through an increased binding of MAP2 to microtubules. From the results presented here, we further suggest that cross-linking of neurofilaments to microtubules may also play a role in this process.
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MRC Neurobiology Unit, MRC Centre, Cambridge, U.K.
Hunt
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D000818:Animals; D000911:Antibodies, Monoclonal; D002478:Cells, Cultured; D004622:Embryo, Mammalian; D005455:Fluorescent Antibody Technique; D007382:Intermediate Filaments / Q000187:drug effects / Q000648:ultrastructure*; D008869:Microtubule-Associated Proteins / Q000187:drug effects / Q000502:physiology*; D009474:Neurons / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D015739:Nocodazole / Q000494:pharmacology*; D016318:Quisqualic Acid / Q000494:pharmacology*; D051381:Rats; D013116:Spinal Cord / Q000502:physiology* / Q000648:ultrastructure; D014404:Tubulin / Q000187:drug effects / Q000502:physiology
D000911:Antibodies, Monoclonal; D008869:Microtubule-Associated Proteins; D014404:Tubulin; D016318:Quisqualic Acid; D015739:Nocodazole