Studing the role of Lmx1a during midbrain dopamine neuron specification using NesE-Lmx1a gain of function cell line

The midbrain dopaminergic (mDA) neurons constitute about 75% of dopaminergic neurons in the adult brain. These neurons are involved in the control of voluntary movements and in the regulation of emotion - related behaviour. Loss of mDA neurons can cause Parkinson’s disease and other neurological disorders as well. In order to understand the selective degeneration of these neurons, insight in the pathways and factors involved in the development and maintenance of this subset of dopaminergic neurons is needed. On the other hand, the prospect of using stem–cell derived DA neurons has emphasized the requirement of understanding the normal pathway of DA neuron development. Among the factors implicated in mDA neuron development are Wnt1, Wnt5a, En1/2, Otx2, Foxa1, Foxa2, Ngn2, Nurr1, Pitx3, Msx1 and LIM homeodomain transcription factors Lmx1a and Lmx1b. Several studies suggested a role for Lmx1a in establishing a mDA neuronal phenotype (Andersson et al., 2006; Chung et al.,2009). Gain and loss of function studies in chick revealed that Lmx1a is required for the specification of mDA neurons. Furthermore Lmx1a can induce mouse embryonic stem (mES) cells into DA neurons. Together these experiments in chick and in mES cells suggest an essential role for Lmx1a in determination of mDA neurons. Beside several studies suggesting a role for Lmx1a in proliferation and neurogenesis, the precise role of Lmx1a in the mouse mDA is still not fully understood. Therefore, to understand the Lmx1a phenotype in depth we studied the gain of function of Lmx1a in NesE-Lmx1a gain of function cell line. In this cell line Lmx1a is under the control of the enhancer of Nestin. Using NesE-Lmx1a cell line Andersson and colleagues (Andersson et al., 2006) showed that Lmx1a functions as DA neuron determinant for midbrain dopamine neurons in mES cells. We were interested to know the molecular network that Lmx1a activates to do that. To elucidate the molecular programming we performed qPCR analysis on d5 and d9 NesE-Lmx1a cells. RT-qPCR analysis at d5 showed that several genes involed in midbrain dopamine neuron specification, differentiation and migration such as Ngn2, Foxa2, Nurr1, Rspo2, Slit2, Aldh1a1 were upregulated. At d9 qPCR results showed that Tuj1, which is used as marker for postmitotic neuronal cells was upregulated in NesE-Lmx1a compared with control cells and so Nurr1. Altogether these data indicate that Lmx1a is essential for the correct development of mDA neurons and it does that by regulating the expression of of several genes involved in dopamine neuron differentiation and migration

The Surface Proteome of Adult Neural Stem Cells in Zebrafish Unveils Long-Range Cell-Cell Connections and Age-Related Changes in Responsiveness to IGF

Summary: In adult stem cell populations, recruitment into division is parsimonious and most cells maintain a quiescent state. How individual cells decide to enter the cell cycle and how they coordinate their activity remains an essential problem to be resolved. It is thus important to develop methods to elucidate the mechanisms of cell communication and recruitment into the cell cycle. We made use of the advantageous architecture of the adult zebrafish telencephalon to isolate the surface proteins of an intact neural stem cell (NSC) population. We identified the proteome of NSCs in young and old brains. The data revealed a group of proteins involved in filopodia, which we validated by a morphological analysis of single cells, showing apically located cellular extensions. We further identified an age-related decrease in insulin-like growth factor (IGF) receptors. Expressing IGF2b induced divisions in young brains but resulted in incomplete divisions in old brains, stressing the role of cell-intrinsic processes in stem cell behavior. : In this article, Chapouton and colleagues use the brain of the adult zebrafish to identify communication pathways in a native population of neural stem cells. They identify proteins expressed on the apical surfaces by a biotinylation technique and document the presence of filopodial extensions between cells. They further show the appearance of an abnormal mitotic response to IGF with age. Keywords: telencephalon, pallium, GFAP, radial glia, filopodia, lamellipodia, biotinylation, mass spectrometry, aging, neurogenesis, quiescenc

The Aryl Hydrocarbon Receptor Pathway Defines the Time Frame for Restorative Neurogenesis

Summary: Zebrafish have a high capacity to replace lost neurons after brain injury. New neurons involved in repair are generated by a specific set of glial cells, known as ependymoglial cells. We analyze changes in the transcriptome of ependymoglial cells and their progeny after injury to infer the molecular pathways governing restorative neurogenesis. We identify the aryl hydrocarbon receptor (AhR) as a regulator of ependymoglia differentiation toward post-mitotic neurons. In vivo imaging shows that high AhR signaling promotes the direct conversion of a specific subset of ependymoglia into post-mitotic neurons, while low AhR signaling promotes ependymoglial proliferation. Interestingly, we observe the inactivation of AhR signaling shortly after injury followed by a return to the basal levels 7 days post injury. Interference with timely AhR regulation after injury leads to aberrant restorative neurogenesis. Taken together, we identify AhR signaling as a crucial regulator of restorative neurogenesis timing in the zebrafish brain. : Zebrafish have a high capacity to replace lost neurons after brain injury. Di Giaimo et al. identify the aryl hydrocarbon receptor (AhR) as a crucial regulator of restorative neurogenesis timing in the zebrafish brain. Interference with timely AhR regulation after injury leads to aberrant restorative neurogenesis. Keywords: neurogenesis, regeneration, aryl hydrocarbon receptor, direct conversion, zebrafish, live imagin