Adult neurogenesis and neuronal regeneration in the brain of teleost fish

Until the late 1990s, the dogma that no new neurons are formed in the adult human brain dominated neurology. Although this dogma was overturned when the laboratories of Peter Eriksson and Fred Gage reported in 1998 the discovery of newborn neurons in the adult human brain, their work, as well as numerous other studies, have also revealed the limits of the mammalian brain: the generation of new neurons (‘adult neurogenesis’) is restricted to two brain regions, and the number of adult-born neurons is very low, compared to the total number of brain cells.

These limits of the mammalian brain contrast with the enormous potential of teleost fish to generate a large number of new neurons in numerous areas of the adult brain. Our laboratory entered this field in the early 1990s, at a time when only a handful of research groups in the world worked on the phenomenon of adult neurogenesis. Using the brown ghost knifefish (Apteronotus leptorhynchus) as a model system, we performed the first brain mapping of the proliferation zones in the adult brain of any vertebrate species. We analyzed in detail the development of newly generated neurons in the structure that exhibits the highest proliferative activity in the brain, the cerebellum. We were the first to show that the new cells are born in specific proliferation zones within the cerebellum, from where they migrate to specific target areas. During their migration, the young cells are guided by radial glial fibers. These fibers express glial fibrillary acidic protein (GFAP) and delineate the paths taken by the migrating cells (see video, below). Furthermore, we demonstrated long-term persistence of the adult-born cells, and we were the first to succeed in the isolation and cultivation of adult stem cells from the teleostean brain. Our mapping of the proliferation zones in the brain of zebrafish, and our analysis of the patterns of migration, differentiation, and long-term survival of the cells that are born in these zones, established the adult zebrafish brain as a new model system in the field of adult neurogenesis.

Complimentary to our investigations on adult neurogenesis in the intact brain, we have studied the extraordinary potential of teleost fish to regenerate neuronal tissue after brain injury through generation of new neurons (‘neuronal regeneration’). As we have shown, this ability is mediated by the elimination of injured cells through apoptotic cell death, followed by replacement of neurons lost to injury by newly generated ones. By employing proteome analysis, we succeeded in the identification of a large number of candidate proteins putatively involved in this regenerative potential.

Based on an examination of the chromosomal compliment of the cells born in the adult teleost brain, we discovered an unexpected degree of chromosomal variation. Only a fraction of the newborn cells display the euploid set of chromosomes. The majority of the adult-born cells show a loss or gain of chromosomes (‘aneuploidy’). This variability is caused by segregation defects during mitotic division. Despite the chromosomal aberrations, a significant number of the new cells differentiate into neurons and persist long-term. The functional significance of the aneuploidy among the population of adult-born cells is unknown. Perhaps, the numerical variation in chromosome number forms the basis for a mechanism to regulate gene expression at the level of whole chromosomes, rather than at a single-gene level.

Progeny of adult stem cells isolated from the brain of the teleost fish, Apteronotus leptorhynchus. Differentiation is induced by plating neurospheres onto a laminin‑coated surface and cultivating them in medium containing fetal bovine serum. Approximately 24 h after plating, the cells start to differentiate into various cell types. (A) GFAP‑immunoreactive astrocyte. (B) Vimentin‑positive cell. (C) Hu C/D‑expressing neurons. (D) Microtubule‑associated protein‑2 (2a + 2b)‑immunopositive neuron. Nuclei are stained with either propidium iodide (red) or 4’‑6‑diamidino‑2‑phenylindole dihydrochloride, DAPI (blue). Note the absence of immunolabeling in some cells in (C) and (D). (From Hinsch & Zupanc, 2006.)

Three-dimensional reconstruction of radial glial fibers in the cerebellum of the teleost fish, Apteronotus leptorhynchus. These fibers play an important role in the guidance of adult-born neurons migrating from proliferation zones to their target areas. (From: Zupanc et al., 2012.)

Selected Publications:

Zupanc, G.K.H., Zupanc, M.M.: Birth and migration of neurons in the central posterior/prepacemaker nucleus during adulthood in weakly electric knifefish, Eigenmannia sp. Proceedings of the National Academy of Sciences U.S.A. 89, 9539-9543 (1992), URL: http://www.pnas.org/content/89/20/9539

Zupanc, G.K.H., Horschke, I.: Proliferation zones in the brain of adult gymnotiform fish: a quantitative mapping study. Journal of Comparative Neurology 353, 213-233 (1995), doi:10.1002/cne.903530205

Zupanc, G.K.H., Horschke, I., Ott, R., Rascher, G.B.: Postembryonic development of the cerebellum in gymnotiform fish. Journal of Comparative Neurology 370, 443-464 (1996), doi:10.1002/(SICI)1096-9861(19960708)370:4<443::AID-CNE3>3.0.CO;2-4

Zupanc, G.K.H., Kompass, K.S., Horschke, I., Ott, R., Schwarz, H.: Apoptosis after injuries in the cerebellum of adult teleost fish. Experimental Neurology 152, 221-230 (1998), doi:10.1006/exnr.1998.6853

Zupanc, G.K.H., Ott, R.: Cell proliferation after lesions in the cerebellum of adult teleost fish: time course of generation, site of origin, and type of new cells produced. Experimental Neurology 160, 78-87 (1999), doi:10.1006/exnr.1999.7182

Zupanc, G.K.H., Hinsch, K., Gage, F.H.: Proliferation, migration, neuronal differentiation, and long-term survival of new cells in the adult brain of zebrafish. Journal of Comparative Neurology 488, 290-319 (2005), doi:10.1002/cne.20571

Zupanc, M.M., Wellbrock, U.M., Zupanc, G.K.H.: Proteome analysis identifies novel protein candidates involved in regeneration of the cerebellum of teleost fish. Proteomics 6, 677-696 (2006), doi:10.1002/pmic.200500167

Hinsch, K., Zupanc, G.K.H.: Isolation, cultivation, and differentiation of neural stem cells from adult fish brain. Journal of Neuroscience Methods 158, 75-88 (2006), doi:10.1016/j.jneumeth.2006.05.020

Hinsch, K., Zupanc, G.K.H.: Generation and long-term persistence of new neurons in the adult zebrafish brain: a quantitative analysis. Neuroscience 146, 679-696 (2007), doi:10.1016/j.neuroscience.2007.01.071

Rajendran, R.S., Zupanc, M.M., Lösche, A., Westra, J., Chun, J., Zupanc, G.K.H.: Numerical chromosome variation and mitotic segregation defects in the adult brain of teleost fish. Developmental Neurobiology 67, 1334-1347 (2007), doi:10.1002/dneu.20365

Rajendran, R.S., Wellbrock, U.M., Zupanc, G.K.H.: Apoptotic cell death, long-term persistence, and neuronal differentiation of aneuploid cells generated in the adult brain of teleost fish. Developmental Neurobiology 68, 1257-1268 (2008), doi:10.1002/dneu.20656

Zupanc, G.K.H., Wellbrock, U.M., Sîrbulescu, R.F., Rajendran, R.S.: Generation, long-term persistence, and neuronal differentiation of cells with nuclear aberrations in the adult zebrafish brain. Neuroscience 159, 1338-1348 (2009), doi:10.1016/j.neuroscience.2009.02.014

Zupanc, G.K.H., Sîrbulescu, R.F., Ilieş, I.: Radial glia in the cerebellum of adult teleost fish: Implications for the guidance of migrating new neurons. Neuroscience 210, 416-430 (2012), doi:10.1016/j.neuroscience.2012.03.012

Teles, M.C., Sîrbulescu, R.F., Wellbrock, U.M., Oliveira, R.F., Zupanc, G.K.H.: Adult neurogenesis in the brain of the Mozambique tilapia, Oreochromis mossambicus. Journal of Comparative Physiology A 198, 427-449 (2012), doi:10.1007/s00359-012-0721-6

Ilieş, I., Zupanc, M.M., Zupanc, G.K.H.: Proteome analysis reveals protein candidates involved in early stages of brain regeneration in teleost fish. Neuroscience 219, 302-313 (2012), doi:10.1016/j.neuroscience.2012.05.028

Sîrbulescu, R.F., Ilieş, I., Zupanc, G.K.H.: Quantitative analysis reveals dominance of gliogenesis over neurogenesis in an adult brainstem oscillator. Developmental Neurobiology 74, 932-952 (2014), doi:10.1002/dneu.22176

Sîrbulescu, R.F., Ilieş, I., Zupanc, G.K.H.: Matrix metalloproteinase-2 and -9 in the cerebellum of teleost fish: Functional implications for adult neurogenesis. Molecular and Cellular Neuroscience 68, 9-23 (2015), doi:10.1016/j.mcn.2015.03.015

Sîrbulescu, R.F., Ilieş, I., Vitalo, A.G., Trull, K., Zhu, J., Traniello, I.M., Zupanc, G.K.H.: Adult stem cells in the knifefish cerebellum. Developmental Neurobiology 75, 39-65 (2015), doi:10.1002/dneu.22210