RESUMO
Studying the development of mesodiencephalic dopaminergic (mdDA) neurons provides an important basis for better understanding dopamine-associated brain functions and disorders and is critical for establishing cell replacement therapy for Parkinson's disease. The transcription factors Otx2 and Lmx1b play a key role in the development of mdDA neurons. However, little is known about the genes downstream of Otx2 and Lmx1b in the pathways controlling the formation of mdDA neurons in vivo. Here we report on our investigation of Lmx1b as downstream target of Otx2 in the formation of mdDA neurons. Mouse mutants expressing Otx2 under the control of the En1 promoter (En1+/Otx2) showed increased Otx2 expression in the mid-hindbrain region, resulting in upregulation of Lmx1b and expansion of mdDA neurons there. In contrast, Lmx1b-/- mice showed decreased expression of Otx2 and impairments in several aspects of mdDA neuronal formation. To study the functional interaction between Otx2 and Lmx1b, we generated compound mutants in which Otx2 expression was restored in mice lacking Lmx1b (En1+/Otx2;Lmx1b-/-). In these animals Otx2 was not sufficient to rescue any of the aberrations in the formation of mdDA neurons caused by the loss of Lmx1b, but rescued the loss of ocular motor neurons. Gene expression studies in Lmx1b-/- embryos indicated that in these mutants Wnt1, En1 and Fgf8 expression are induced but subsequently lost in the mdDA precursor domain and the mid-hindbrain organizer in a specific, spatio-temporal manner. In summary, we demonstrate that Otx2 critically depends on Lmx1b for the formation of mdDA neurons, but not for the generation of ocular motor neurons. Moreover, our data suggest that Lmx1b precisely maintains the expression pattern of Wnt1, Fgf8 and En1, which are essential for mid-hindbrain organizer function and the formation of mdDA neurons.
Assuntos
Neurônios Dopaminérgicos/fisiologia , Proteínas com Homeodomínio LIM/genética , Mesencéfalo/fisiologia , Fatores de Transcrição Otx/genética , Fatores de Transcrição/genética , Animais , Padronização Corporal/genética , Diferenciação Celular/genética , Dopamina/genética , Embrião de Mamíferos/fisiologia , Feminino , Fator 8 de Crescimento de Fibroblasto/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Homeodomínio/genética , Camundongos , Proteínas do Tecido Nervoso/genética , Rombencéfalo/fisiologia , Proteína Wnt1/genéticaRESUMO
The locus coeruleus (LC) which is the major noradrenergic nucleus in the brain develops under the influence of Bmps secreted by the roof plate and Fgf8 emitted from the mid-hindbrain organizer. We studied the development of the LC in different Bmp mouse mutants and report the absence of this nucleus in Bmp5(-/-);Bmp7(-/-) double knockouts. Notably, genes marking organizers and neuronal populations adjacent to the LC precursor field are unperturbed in Bmp5(-/-);Bmp7(-/-) animals. In addition, we found that in En1(+/Otx2) mutants in which the caudal Otx2 expression domain and thereby the mid-hindbrain organizer are shifted caudally, LC neurons are concomitantly reduced along with Bmp5/7. Complementing these results, Otx1(-/-);Otx2(+/-) mutants, in which the mid-hinbrain organizer is shifted rostrally, show a rostrally extended Bmp5 expression area and an increase in LC neurons. Taken together, our data indicate that LC development requires either Bmp5 or Bmp7, and one is able to compensate for the loss of the other. In addition, we conclude that the position of the mid-hindbrain organizer determines the size of the LC and propose that Bmp5/7 play an important role in mediating this organizer function.
Assuntos
Proteína Morfogenética Óssea 5/metabolismo , Proteína Morfogenética Óssea 7/metabolismo , Locus Cerúleo/citologia , Locus Cerúleo/embriologia , Mesencéfalo/fisiologia , Norepinefrina/metabolismo , Rombencéfalo/fisiologia , Animais , Apoptose , Fator 8 de Crescimento de Fibroblasto/genética , Fator 8 de Crescimento de Fibroblasto/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Locus Cerúleo/metabolismo , Mesencéfalo/citologia , Camundongos , Camundongos Knockout , Neurônios/citologia , Neurônios/fisiologia , Fatores de Transcrição Otx/genética , Fatores de Transcrição Otx/metabolismo , Rombencéfalo/citologia , Células-Tronco/citologia , Células-Tronco/fisiologiaRESUMO
During late developmental phases individual sympathetic neurons undergo a switch from noradrenergic to cholinergic neurotransmission. This phenomenon of plasticity depends on target-derived signals in vivo and is triggered by neurotrophic factors in neuronal cultures. To analyze genome-wide expression differences between the two transmitter phenotypes we employed DNA microarrays. RNA expression profiles were obtained from chick paravertebral sympathetic ganglia, treated with neurotrophin 3, glial cell line-derived neurotrophic factor or ciliary neurotrophic factor, all of which stimulate cholinergic differentiation. Results were compared with the effect of nerve growth factor, which functions as a pro-noradrenergic stimulus. The gene set common to all three comparisons defined the noradrenergic and cholinergic synexpression groups. Several functional categories, such as signal transduction, G-protein-coupled signaling, cation transport, neurogenesis and synaptic transmission, were enriched in these groups. Experiments based on the prediction that some of the identified genes play a role in the neurotransmitter switch identified bone morphogenetic protein signaling as an inhibitor of cholinergic differentiation.
