Differential neurite growth on astrocyte substrates: interspecies facilitation in green fluorescent protein-transfected rat and human neurons.
In the present study, we used co-culture of astrocytes from one species with neurons from a different species to examine neuritic outgrowth. We include a focus on human cells. Three types of neuron were used, including rat hippocampal dentate granule cells, rat hypothalamic neurons and human cortical neurons. To visualize neuronal processes, neurons were either immunostained with GABA antiserum or transfected with the jellyfish green fluorescent protein gene. The entire axonal and dendritic fields of single neurons could be quantitatively analysed based on their strong green fluorescent protein label. Astrocytes were obtained from rat hippocampus or hypothalamus, chicken cortex, normal human cortex, human cortex lesion, and from the sclerotic human hippocampus after surgery for intractable temporal lobe epilepsy. In the absence of astrocytes, isolated neurons died within three to four days. In contrast, neurons from both rat and human brains survived and extended dendrites and axons on rat, chicken and human astrocytes or in their conditioned medium. Astrocytes from interspecies cultures were not only capable of enhancing the survival of neuron co-cultures, but neuronal neurite extension in some cases was even greater on heterospecific astrocytes than on homospecific astrocytes. To support the hypothesis that synaptogenesis of rat hippocampal neurons was accelerated by a substrate of human astrocytes, we used a functional assay based on time-lapse confocal laser or digital imaging of calcium responses to transmitter release; synaptic responses were found earlier when rat neurons were grown on rat or human astrocytes than in the absence of these astrocytes. These data indicate that rodent glial cells enhance human neurite extension, and that rat neurite outgrowth can be used as a type of bioassay for the neurite promoting capacity of different derivations of human glia.
van den Pol
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Department of Neurosurgery, Yale University School of Medicine, New Haven, CT 06520, USA.
Spencer
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D015763:2-Amino-5-phosphonovalerate / Q000494:pharmacology; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione / Q000494:pharmacology; D000818:Animals; D001253:Astrocytes / Q000166:cytology*; D001369:Axons / Q000502:physiology; D002450:Cell Communication / Q000502:physiology; D018929:Cell Culture Techniques / Q000379:methods*; D002540:Cerebral Cortex / Q000166:cytology; D018076:DNA, Complementary; D003712:Dendrites / Q000502:physiology; D018891:Dentate Gyrus / Q000166:cytology; D004594:Electrophysiology; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D020439:Growth Cones / Q000502:physiology; D006801:Humans; D007031:Hypothalamus / Q000166:cytology; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D016501:Neurites / Q000502:physiology*; D009474:Neurons / Q000648:ultrastructure*; D010957:Plasmids; D051381:Rats; D013045:Species Specificity; D013569:Synapses / Q000502:physiology; D014162:Transfection / Q000379:methods
D018076:DNA, Complementary; D018690:Excitatory Amino Acid Agonists; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D015763:2-Amino-5-phosphonovalerate