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Nothobranchius annual killifishes
Full text linkAnnual fishes of the genus Nothobranchius inhabit ephemeral habitats in Eastern and Southeastern Africa. Their life cycle is characterized by very rapid maturation, a posthatch lifespan of a few weeks to months and embryonic diapause to survive the dry season. The species N. furzeri holds the record of the fastest-maturing vertebrate and of the vertebrate with the shortest captive lifespan and is emerging as model organism in biomedical research, evolutionary biology, and developmental biology. Extensive characterization of age-related phenotypes in the laboratory and of ecology, distribution, and demography in the wild are available. Species/populations from habitats differing in precipitation intensity show parallel evolution of lifespan and age-related traits that conform to the classical theories on aging. Genome sequencing and the establishment of CRISPR/Cas9 techniques made this species particularly attractive to investigate the effects genetic and non-genetic intervention on lifespan and aging-related phenotypes. At the same time, annual fishes are a very interesting subject for comparative approaches, including genomics, transcriptomics, and proteomics. The N. furzeri community is highly diverse and rapidly expanding and organizes a biannual meeting
Immunofluorescence and Aggresome Staining of Nothobranchius furzeri Cryosection
No full textProtein aggregation is a hallmark of many age-related diseases and, in particular, of neurodegenerative
diseases such as Parkinson’s and Alzheimer’s diseases. The teleost Nothobranchius furzeri shows the
shortest median life span among all vertebrate animal models and has recently gained popularity as a
convenient model for experimental approaches to aging. Immunofluorescence staining is the primary
technique used to visualize the distribution of proteins in fixed cells and tissues and it has proven to be a
powerful tool to study aggregates and proteins involved in neurodegenerative diseases. Specifically,
immunofluorescence staining allows for precise localization of aggregates in specific cell types and can
be used to identify the proteins constituting such aggregates. To facilitate the study of aggregate-related
pathologies in the context of aging using the new model N. furzeri, we report a protocol to visualize
general protein aggregates and specific proteins optimized for N. furzeri brain cryosections
EdU and immunofluorescence staining of nothobranchius furzeri organotypic cultures
No full text: Immunofluorescence is a widely used technique to visualize the localization of proteins of interest. Nucleoside analogs, such as 5-ethynyl-2'-deoxyuridine (EdU), are incorporated into newly synthesized DNA and enable permanent labeling of newly divided cells. Both these techniques can be applied to long-term organotypic culture of Nothobranchius furzeri in a fashion similar to that already described for tissue sections. We report here our optimized method for immunofluorescence and EdU staining of N. furzeri organotypic slices
The short-lived fish Nothobranchius furzeri as a new model system for aging studies
No full textGenetic and pharmacological research on aging is hampered by the lifespan of available vertebrate models. We recently initiated studies on Nothobranchius furzeri, a species with a maximum life expectancy in captivity of just three months which represents the shortest documented captive lifespan for a vertebrate. Further research on N. furzeri has demonstrated that1.Short lifespan is tied with explosive growth and accelerated sexual maturation.2.Short lifespan is correlated with expression of age-related behavioral and histological changes.3.Lifespan and expression of age-related markers can be modulated by water temperature.4.Resveratrol, a drug characterized for its life-extending action in Caenorhabditis elegans and Drosophila, increases lifespan and retards expression of age-related markers.5.Aging-related genes can be easily isolated by homology cloning. Finally, different populations or species of Nothobranchius show large-scale differences in captive lifespan. In the last three years, N. furzeri has moved from biological curiosity to a promising model system for drug validation. Furthermore, this species occupies a favorable position in the Teleost's "tree of life". It is very close to the Japanese Medaka, and close to the pufferfishes and stickleback and might represent a very useful model for comparative genomics of aging
Localization and Characterization of Major Neurogenic Niches in the Brain of the Lesser-Spotted Dogfish Scyliorhinus canicula
Full text linkAdult neurogenesis is defined as the ability of specialized cells in the postnatal brain to produce new functional neurons and to integrate them into the already-established neuronal network. This phenomenon is common in all vertebrates and has been found to be extremely relevant for numerous processes, such as long-term memory, learning, and anxiety responses, and it has been also found to be involved in neurodegenerative and psychiatric disorders. Adult neurogenesis has been studied extensively in many vertebrate models, from fish to human, and observed also in the more basal cartilaginous fish, such as the lesser-spotted dogfish, Scyliorhinus canicula, but a detailed description of neurogenic niches in this animal is, to date, limited to the telencephalic areas. With this article, we aim to extend the characterization of the neurogenic niches of S. canicula in other main areas of the brain: we analyzed via double immunofluorescence sections of telencephalon, optic tectum, and cerebellum with markers of proliferation (PCNA) and mitosis (pH3) in conjunction with glial cell (S100β) and stem cell (Msi1) markers, to identify the actively proliferating cells inside the neurogenic niches. We also labeled adult postmitotic neurons (NeuN) to exclude double labeling with actively proliferating cells (PCNA). Lastly, we observed the presence of the autofluorescent aging marker, lipofuscin, contained inside lysosomes in neurogenic areas
Distribution of Brain-Derived Neurotrophic Factor in the Brain of the Small-Spotted Catshark Scyliorhinus canicula, and Evolution of Neurotrophins in Basal Vertebrates
Full text linkNeurotrophins (NTFs) are structurally related neurotrophic factors essential for differentiation, survival, neurite outgrowth, and the plasticity of neurons. Abnormalities associated with neurotrophin-signaling (NTF-signaling) were associated with neuropathies, neurodegenerative disorders, and age-associated cognitive decline. Among the neurotrophins, brain-derived neurotrophic factor (BDNF) has the highest expression and is expressed in mammals by specific cells throughout the brain, with particularly high expression in the hippocampus and cerebral cortex. Whole genome sequencing efforts showed that NTF signaling evolved before the evolution of Vertebrates; thus, the shared ancestor of Protostomes, Cyclostomes, and Deuterostomes must have possessed a single ortholog of neurotrophins. After the first round of whole genome duplication that occurred in the last common ancestor of Vertebrates, the presence of two neurotrophins in Agnatha was hypothesized, while the monophyletic group of cartilaginous fishes, or Chondrichthyans, was situated immediately after the second whole genome duplication round that occurred in the last common ancestor of Gnathostomes. Chondrichthyans represent the outgroup of all other living jawed vertebrates (Gnathostomes) and the sister group of Osteichthyans (comprehensive of Actinopterygians and Sarcopterygians). We were able to first identify the second neurotrophin in Agnatha. Secondly, we expanded our analysis to include the Chondrichthyans, with their strategic phylogenetic position as the most basal extant Gnathostome taxon. Results from the phylogenetic analysis confirmed the presence of four neurotrophins in the Chondrichthyans, namely the orthologs of the four mammalian neurotrophins BDNF, NGF, NT-3, and NT-4. We then proceeded to study the expression of BDNF in the adult brain of the Chondrichthyan Scyliorhinus canicula. Our results showed that BDNF is highly expressed in the S. canicula brain and that its expression is highest in the Telencephalon, while the Mesencephalic and Diencephalic areas showed expression of BDNF in isolated and well-defined cell groups. NGF was expressed at much lower levels that could be detected by PCR but not by in situ hybridization. Our results warrant further investigations in Chondrichthyans to characterize the putative ancestral function of neurotrophins in Vertebrates
Effects of dietary restriction on mortality and age-related phenotypes in the short-lived fish Nothobranchius furzeri
Full text linkThe short-lived annual fish Nothobranchius furzeri shows extremely short captive life span and accelerated expression of age markers, making it an interesting model system to investigate the effects of experimental manipulations on longevity and age-related pathologies. Here, we tested the effects of dietary restriction (DR) on mortality and age-related markers in N. furzeri. DR was induced by every other day feeding and the treatment was performed both in an inbred laboratory line and a longer-lived wild-derived line. In the inbred laboratory line, DR reduced age-related risk and prolonged maximum life span. In the wild-derived line, DR induced early mortality, did not reduce general age-related risk and caused a small but significant extension of maximum life span. Analysis of age-dependent mortality revealed that DR reduced demographic rate of aging, but increased baseline mortality in the wild-derived strain. In both inbred- and wild-derived lines, DR prevented the expression of the age markers lipofuscin in the liver and Fluoro-Jade B (neurodegeneration) in the brain. DR also improved performance in a learning test based on conditioning (active avoidance in a shuttle box). Finally, DR induced a paradoxical up-regulation of glial fibrillary acidic protein in the brain
Nerve Growth Factor Receptor (NGFR/p75NTR) of the Small-Spotted Catshark (Scyliorhinus canicula): Evolutionary Conservation and Brain Function
Full text linkThe p75NTR receptor, a member of the tumor necrosis factor (TNF) receptor superfamily, can participate in signaling pathways either by forming heteromeric complexes with other receptors, such as the Trk family (tropomyosin receptor kinases), or by functioning independently. p75NTR was investigated prevalently in the brain and retina of mammals, whereas almost nothing is known about its conservation among species. Here, we reconstructed the phylogenetic arb of p75NTR and described for the first time the p75NTR expression in the brain of the basal vertebrate Chondrichthyan Scyliorhinus canicula (S. canicula), uncovering the existing parallelism between ancient vertebrates and mammals. p75NTR functional conservation among vertebrates was further investigated by cloning the S. canicula nerve growth factor (NGF) and performing the canonical posterior commissure (PC)-12 differentiation assay, which results in standard neurite-like production. We then investigated the S. canicula p75NTR, which proves to be capable of complementing a specific clone of PC-12 lacking p75NTR (PC-12 p75NTR-/-). All together, our results highlighted the expression and functional conservation of p75NTR among vertebrates during the evolution
New lessons on TDP‐43 from old N. furzeri killifish
Full text linkFrontotemporal dementia and amyotrophic lateral sclerosis are fatal and incurable neurodegenerative diseases linked to the pathological aggregation of the TDP-43 protein. This is an essential DNA/RNA-binding protein involved in transcription regulation, pre-RNA processing, and RNA transport. Having suitable animal models to study the mechanisms of TDP-43 aggregation is crucial to develop treatments against disease. We have previously demonstrated that the killifish Nothobranchius furzeri offers the advantage of being the shortest-lived vertebrate with a clear aging phenotype. Here, we show that the two N. furzeri paralogs of TDP-43 share high sequence homology with the human protein and recapitulate its cellular and biophysical behavior. During aging, N. furzeri TDP-43 spontaneously forms insoluble intracellular aggregates with amyloid characteristics and colocalizes with stress granules. Our results propose this organism as a valuable new model of TDP-43-related pathologies making it a powerful tool for the study of disease mechanism
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