Organotypic slice cultures of embryonic ventral midbrain: a system to study dopaminergic neuronal development in vitro.
The  mouse is an excellent model organism to study mammalian brain development due  to the abundance of molecular and genetic data. However, the developing mouse  brain is not suitable for easy manipulation and imaging in vivo since the mouse embryo is inaccessible and opaque.  Organotypic slice cultures of embryonic brains are therefore widely used to  study murine brain development in vitro. Ex-vivo manipulation or the use of  transgenic mice allows the modification of gene expression so that subpopulations of neuronal or glial cells can be labeled with  fluorescent proteins. The behavior of labeled cells can then be observed using  time-lapse imaging. Time-lapse imaging  has been particularly successful for studying cell behaviors that underlie the  development of the cerebral cortex at late embryonic stages (1-2).  Embryonic organotypic slice culture systems in brain regions outside of the  forebrain are less well established. Therefore, the wealth of time-lapse  imaging data describing neuronal cell migration is restricted to the forebrain (3,4). It is still not known, whether the principles  discovered for the dorsal brain hold true for ventral brain areas. In the  ventral brain, neurons are organized in neuronal clusters rather than layers  and they often have to undergo complicated migratory trajectories to reach  their final position. The ventral midbrain is not only a good model system for  ventral brain development, but also contains neuronal populations such as  dopaminergic neurons that are relevant in disease processes. While the function  and degeneration of dopaminergic neurons has been investigated in great detail  in the adult and ageing brain, little is known about the behavior of these neurons  during their differentiation and migration phase (5). We describe here the generation of slice  cultures from the embryonic day (E) 12.5 mouse ventral midbrain. These slice  cultures are potentially suitable for monitoring dopaminergic neuron  development over several days in vitro.  We highlight the critical steps in generating brain slices at these early  stages of embryonic development and discuss the conditions necessary for  maintaining normal development of dopaminergic neurons in vitro. We also present results from time lapse imaging  experiments. In these experiments, ventral midbrain precursors (including  dopaminergic precursors) and their descendants were labeled in a mosaic manner  using a Cre/loxP based inducible fate mapping system (6).
Institute of Reconstructive Neurobiology, University of Bonn.
Gabriela Oana
GO
Bodea
Sandra
S
Blaess
D000818:Animals; D059290:Dopaminergic Neurons / Q000166:cytology*; D004622:Embryo, Mammalian; D005260:Female; D008636:Mesencephalon / Q000166:cytology* / Q000196:embryology*; D051379:Mice; D008867:Microtomy / Q000379:methods; D009924:Organ Culture Techniques / Q000379:methods*; D011247:Pregnancy; D013234:Stem Cells / Q000166:cytology