Nucleotides affect neurogenesis and dopaminergic differentiation of mouse fetal midbrain-derived neural precursor...

The fetal midbrain is a preferred source for isolating and producing dopaminergic neurons for subsequent grafting and replacement of damaged or lost dopaminergic midbrain neurons. We analysed the potential of a variety of nucleotides and of adenosine to support dopaminergic neuron formation from primary mouse fetal midbrain-derived cells, harvested at E10.5 and at E13.5 and subjected to adherent cell culture. In contrast to cells derived at E13.5, cells derived at E10.5 have the potential to produce dopaminergic neurons in culture. These neurons express tyrosine hydroxylase and the dopamine transporter. The fetal ventral midbrain contained mRNA encoding almost all P2X and P2Y receptors, all adenosine receptors as well as the ectonucleotidases nucleoside triphosphate diphosphohydrolase 2 and tissue nonspecific alkaline phosphatase. Essentially, all components of the purinergic signalling pathway were also expressed by the cultured cells. ATP, ADPβS, 2MeSATP, 2ClATP and adenosine increased neuron formation. There was, however, no preference for the formation of dopaminergic neurons-with the exception of 2ClATP that increased the relative contribution of tyrosine hydroxylase-positive neurons. In cells isolated at E13.5 UTP promoted neuron survival but ADPβS and ATPγS essentially eliminated neurons. These data showed that the outcome of nucleotide application was different even though cells isolated at E10.5 and E13.5 expressed very similar receptor mRNA profiles. They suggest that purinergic agonists carry potential for stimulating neurogenesis and enriching the contribution of dopaminergic neurons in vitro. Nucleotide receptor agonists may be of value for contributing to the formation and survival of dopaminergic neurons in vivo.