Calcitonin gene-related peptide promotes differentiation, but not survival, of rat mesencephalic dopaminergic neurons in vitro.
The aim of this study was to investigate putative effects of calcitonin gene-related peptide on developing dopaminergic neurons in the ventral mesencephalon. To determine a time-point for a physiological role of calcitonin gene-related peptide in the development of this system, we first investigated calcitonin gene-related peptide messenger RNA expression in the ventral mesencephalon of Wistar rats at embryonic days (E) 11-19. Calcitonin gene-related peptide messenger RNA was not detectable at E11, i.e. prior to the appearance of dopaminergic neurons in this area. From E14 to E19, calcitonin gene-related peptide messenger RNA was expressed in increasing amounts. We therefore investigated the effects of calcitonin gene-related peptide on serum-free cell cultures established from the E14 midbrain floor. Addition of calcitonin gene-related peptide (200 ng/ml) every other day significantly increased neuronal differentiation, including longer tyrosine hydroxylase-positive neurites, enhanced immunoreactivity for growth-associated protein-43 and increased dopaminergic uptake per neuron. These effects were maximal after seven to eight days. Calcitonin gene-related peptide acted synergistically with fibroblast growth factor-2 on these parameters. In contrast to fibroblast growth factor-2, however, calcitonin gene-related peptide did not promote survival of tyrosine hydroxylase-immunoreactive neurons. Lack of calcitonin gene-related peptide expression in the mesencephalon at E11 was paralleled by a lack of effect of calcitonin gene-related peptide on early presumptive dopaminergic neurons in terms of eliciting this phenotype. Our data suggest that calcitonin gene-related peptide may act physiologically as a differentiation-promoting factor for phenotypically defined dopaminergic neurons during a time period when dopaminergic neurons assemble in the ventral mesencephalon and grow axons towards their targets.
Department of Anatomy & Cell Biology-Neuroanatomy, University of Heidelberg, Germany.
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Bürvenich
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Unsicker
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Krieglstein
D000818:Animals; D015740:Calcitonin Gene-Related Peptide / Q000378:metabolism / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004298:Dopamine / Q000378:metabolism / Q000502:physiology*; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008636:Mesencephalon / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012333:RNA, Messenger; D016222:Fibroblast Growth Factor 2; D015740:Calcitonin Gene-Related Peptide; D004298:Dopamine