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Keywords:

  • nerve growth factors;
  • neuronal plasticity;
  • mesencephalon;
  • tyrosine hydroxylase;
  • image reconstruction

Abstract

The effect of the various neurotrophin family members on the morphological structure of dopaminergic neurons was compared in dissociated cultures of embryonic rat ventral mesencephalon. Cultures were maintained in vitro in the presence of brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), neurotrophin-4/5 (NT-4/5), nerve growth factor (NGF) or no added growth factors. Three-dimensional reconstructions of 48 neurons were made in each of the experimental groups following immunocytochemical staining for tyrosine hydroxylase to detect dopaminergic neurons. In addition [3H]mazindol binding analyses were carried out in replicate cultures in order to quantify the effects of the neurotrophins on the number of dopamine uptake sites. Among the neurotrophins tested, NT-4/5 influenced the proximal morphological parameters most, as determined by a 36% increase in the soma profile area and 35% in the number of stem neurites. Analysis of neuritic size and complexity in these cultures revealed that combined neuritic length and number of segments/cell were increased by 45 and 40% respectively. A change in neurite complexity in the NT-4/5 treated cultures was further confirmed using Scholl's concentric sphere analysis. In addition, relative to the control, NT-4/5 increased the neuronal differentiation as evidenced by increases in varicosity density and [3H]mazindol binding by 114 and 101% respectively. BDNF and, to a lesser extent, NT-3 also increased both proximal parameters and parameters of differentiation, but were without effect on parameters of neuritic size and Complexity. No effects on neuronal structure were observed in NGF treated cultures. These findings demonstrate that BDNF, NT-3 and NT-4/5 influence the morphological differentiation of dopaminergic neurons in vitro, suggesting they may play a role in the structural development and plasticity of these neurons in the mesencephalon.