During neurogenesis nerve cells are generated according to a precise spatio-temporal schedule, so that cohorts of newly-generated neurons settle in defined locations and establish appropriate connections with specific partners. At precise ontogenetic stages, the specification of neural progenitors is subject to complex regulatory mechanisms, needed to determine the neuronal types, the required quantities, and the specific location of their implant. In spite of our expanding knowledge of these phenomena, we miss an ensemble view of the process. To better understand the interplay of the different factors controlling this process, we have studied the variation in the expression of over 40 genes involved in the neuronal differentiation of dopaminergic (DA) neurones during adult neurogenesis in the olfactory bulb at three different stages of maturation. To this end, we have exploited a peculiar property of DA neurones in a transgenic line of animals expressing eGFP under the TH promoter, which is a direct relationship between the intensity of fluorescence and the degree of maturation. Using the fluorescence-activated cell sorting after enzymatic dissociation of the olfactory bulb we have separated the fluorescent cells as a function of their level of fluorescence in three groups, corresponding to immature, intermediate and fully mature DA neurones, and we have performed semiquantitative RT-PCRs. The results provide, for the first time, a synoptic view of how a large number of different factors controlling the dopaminergic differentiation vary their expression during the maturation of DA neurones. We have divided these factors in three groups: 1) Key regulators of the dopaminergic phenotype, 2) Phenotype-independent regulators, and 3) Regulators of OB differentiation through olfactory neuron innervations of the olfactory bulb. The first group includes AP-1, CREB, Dlx1 and 2, Engrailed-1, ER-81, Gsh-2, basic helix-loop-helix transcription factors (Mash1, Id2, and Hes1), Meis-2, Nurr-1, Pax-6, Zic 1 and 3. The second group includes DCX, Ephrin A2, Myst-4Notch-1, Olig-2, Prok-2, PSA- NCAM, genes involved in the reelin cascade (Reelin, APOEr2, VLDLR and Dab1), Shh, Slit 1 and2, Tenascin r, Vax-1. The third group includes ARX, CNG2, Dlx5, FezF1, Big2-Contactin4. For all these genes we provide a quantitative description of the level of expression in three different phases of maturation, a picture that becomes particularly instructive when the variations are looked synoptically. We believe that this will contribute to a better understanding of the complex signaling system required to make dopaminergic neurons in the adult olfactory bulb.
A synoptic view of the expression of genes controlling adult neurogenesis of dopaminergic neurones in the olfactory bulb
PIGNATELLI, Angela;GAMBARDELLA, Cristina;BELLUZZI, Ottorino
2010
Abstract
During neurogenesis nerve cells are generated according to a precise spatio-temporal schedule, so that cohorts of newly-generated neurons settle in defined locations and establish appropriate connections with specific partners. At precise ontogenetic stages, the specification of neural progenitors is subject to complex regulatory mechanisms, needed to determine the neuronal types, the required quantities, and the specific location of their implant. In spite of our expanding knowledge of these phenomena, we miss an ensemble view of the process. To better understand the interplay of the different factors controlling this process, we have studied the variation in the expression of over 40 genes involved in the neuronal differentiation of dopaminergic (DA) neurones during adult neurogenesis in the olfactory bulb at three different stages of maturation. To this end, we have exploited a peculiar property of DA neurones in a transgenic line of animals expressing eGFP under the TH promoter, which is a direct relationship between the intensity of fluorescence and the degree of maturation. Using the fluorescence-activated cell sorting after enzymatic dissociation of the olfactory bulb we have separated the fluorescent cells as a function of their level of fluorescence in three groups, corresponding to immature, intermediate and fully mature DA neurones, and we have performed semiquantitative RT-PCRs. The results provide, for the first time, a synoptic view of how a large number of different factors controlling the dopaminergic differentiation vary their expression during the maturation of DA neurones. We have divided these factors in three groups: 1) Key regulators of the dopaminergic phenotype, 2) Phenotype-independent regulators, and 3) Regulators of OB differentiation through olfactory neuron innervations of the olfactory bulb. The first group includes AP-1, CREB, Dlx1 and 2, Engrailed-1, ER-81, Gsh-2, basic helix-loop-helix transcription factors (Mash1, Id2, and Hes1), Meis-2, Nurr-1, Pax-6, Zic 1 and 3. The second group includes DCX, Ephrin A2, Myst-4Notch-1, Olig-2, Prok-2, PSA- NCAM, genes involved in the reelin cascade (Reelin, APOEr2, VLDLR and Dab1), Shh, Slit 1 and2, Tenascin r, Vax-1. The third group includes ARX, CNG2, Dlx5, FezF1, Big2-Contactin4. For all these genes we provide a quantitative description of the level of expression in three different phases of maturation, a picture that becomes particularly instructive when the variations are looked synoptically. We believe that this will contribute to a better understanding of the complex signaling system required to make dopaminergic neurons in the adult olfactory bulb.I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
