Our understanding of the cellular and molecular mechanisms underlying the adult neural stem cell state remains fragmentary. To provide new models on this issue, we searched for stem cells in the adult brain of the zebrafish. Using BrdU tracing and immunodetection of cell-type-specific markers, we demonstrate that the adult zebrafish telencephalon contains self-renewing progenitors, which show features of adult mammalian neural stem cells but distribute along the entire dorso-ventral extent of the telencephalic ventricular zone. These progenitors give rise to newborn neurons settling close to the ventricular zone within the telencephalon proper. They have no equivalent in mammals and therefore constitute a new model of adult telencephalic neural stem cells. In addition, progenitors from the ventral subpallium generate rapidly dividing progenitors and neuroblasts that reach the olfactory bulb (OB) via a rostral migratory stream and differentiate into GABAergic and TH-positive neurons. These ventral progenitors are comparable to the mammalian neural stem cells of the subependymal zone. Interestingly, dorsal and ventral progenitors in the adult telencephalon express a different combination of transcription factors than their embryonic counterparts. In the case of neurogenin1, this is due to the usage of different enhancer elements. Together, our results highlight the conserved and unique phylogenic and ontogenic features of adult neurogenesis in the zebrafish telencephalon and open the way to the identification of adult neural stem cell characters in cross-species comparative studies.

The zebrafish has become a new model for adult neurogenesis, owing to its abundant neurogenic areas in most brain subdivisions. Radial glia-like cells, actively proliferating cells, and label-retaining progenitors have been described in these areas. In the telencephalon, this complexity is enhanced by an organization of the ventricular zone (VZ) in fast and slow-dividing domains, suggesting the existence of heterogeneous progenitor types. In this work, we studied the expression of various transgenic or immunocytochemical markers for glial cells (gfap:gfp, cyp19a1b:gfp, BLBP, and S100beta), progenitors (nestin:gfp and Sox2), and neuroblasts (PSA-NCAM) in cycling progenitors of the adult zebrafish telencephalon (identified by expression of proliferating cell nuclear antigen (PCNA), MCM5, or bromodeoxyuridine incorporation). We demonstrate the existence of distinct populations of dividing cells at the adult telencephalic VZ. Progenitors of the overall slow-cycling domains express high levels of Sox2 and nestin:gfp as well as all glial markers tested. In contrast, domains with an overall fast division rate are characterized by low or missing expression of glial markers. PCNA-positive cells in fast domains further display a morphology distinct from radial glia and co-express PSA-NCAM, suggesting that they are early neuronal precursors. In addition, the VZ contains cycling progenitors that express neither glial markers nor nestin:gfp, but are positive for Sox2 and PSA-NCAM, identifying them as committed neuroblasts. On the basis of the marker gene expression and distinct cell morphologies, we propose a classification for the dividing cell states at the zebrafish adult telencephalic VZ.

Adult neurogenesis arises from niches that harbor neural stem cells (NSC). Although holding great promise for regenerative medicine, the identity of NSC remains elusive. In mammals, a key attribute of NSC is the expression of the filamentous proteins glial fibrillary acidic protein (GFAP) and NESTIN. To assess whether these two markers are relevant in the fish model, two transgenic zebrafish lines for gfap and nestin were generated. Analysis of adult brains showed that the fusion GFAP-green fluorescent protein closely mimics endogenous GFAP, while the nestin transgene recapitulates nestin at the ventricular zones. Cells expressing the two reporters display radial glial morphology, colocalize with the NSC marker Sox2, undergo proliferation, and are capable of self-renewal within the matrix of distinct thickness in the telencephalon. Together, these two transgenic lines reveal a conserved feature of putative NSC in the adult zebrafish brain and provide a means for the identification and manipulation of these cells in vivo. Developmental Dynamics 238:475-486, 2009.

The neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces selective loss of dopaminergic neurons in the mammalian midbrain, eliciting symptoms characteristic of Parkinson's disease. By exploiting the advantages of zebrafish embryos, we report here that dopaminergic neurons in this species are specifically perturbed when exposed to MPTP. In contrast to mammals, the zebrafish does not possess a midbrain dopaminergic system. Instead, the main population of neurons expressing the dopamine transporter is located in the posterior tuberculum of the diencephalon. Exposure of embryos to MPTP led to a pronounced reduction in the number of dopaminergic cells in the diencephalon. This effect can be reversed by deprenyl, a specific inhibitor of monoamine oxidase B that catalyses the conversion of MPTP to its active metabolite, MPP+. Similarly, direct treatment of embryos with MPP+ abolished the diencephalic dopaminergic neurons. These larvae also demonstrated behavioural defects in swimming responses. Thus, dopaminergic neurons in the posterior tuberculum of the zebrafish may be homologous to the midbrain dopaminergic system of mammals. In addition, the mechanism behind the loss of dopaminergic neurons following pharmacological perturbation may be conserved among vertebrates and suggest that the zebrafish can be used as a convenient and economical system to study the pathogenesis of Parkinson's disease and for testing potential therapeutic strategies.

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