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Parkinson's disease (PD) is characterized by phosphorylation and aggregation of the protein alpha-Synuclein and ensuing neuronal death progressing from the noradrenergic locus coeruleus to midbrain dopaminergic neurons. In 2019, Matsui and colleagues reported a spontaneous age-dependent degeneration of dopaminergic neurons and an even greater neurodegeneration of the noradrenergic neurons in the short-lived killifish Nothobranchius furzeri. Given the great possible relevance of a spontaneous model for PD, we assessed neurodegeneration of noradrenergic and dopaminergic neurons in two further laboratory strains of N. furzeri. We implemented, for the first time in N. furzeri, a whole-brain clarification technique and proceeded to entire 3D nuclei reconstruction to quantify total cell numbers in two different stains of N. furzeri. In both strains, we observed that age-dependent neurodegeneration is limited to the locus coeruleus and does not involve the posterior tuberculum. We also applied 3D counting to the optic tectum, an area of active adult neurogenesis, and detected an increase of neurons with age. Our results confirm age-dependent neurodegeneration of noradrenergic neurons, a condition reminiscent of the presymptomatic stage of PD indicating that N. furzeri could be used in the future to identify modifying factors for age-dependent neurodegeneration and open the intriguing possibility that natural genetic variation may influence the susceptibility of dopaminergic neurons.

Summary The brains of teleost fish show extensive adult neurogenesis and neuronal regeneration. The patterns of gene regulation during fish brain aging are unknown. The short-lived teleost fish Nothobranchius furzeri shows markers of brain aging including reduced learning performances, gliosis, and reduced adult neurogenesis. We used RNA-seq to quantify genome-wide transcript regulation and sampled five different time points to characterize whole-genome transcript regulation during brain aging of N. furzeri. Comparison with human datasets revealed conserved up-regulation of ribosome, lysosome, and complement activation and conserved down-regulation of synapse, mitochondrion, proteasome, and spliceosome. Down-regulated genes differ in their temporal profiles: neurogenesis and extracellular matrix genes showed rapid decay, synaptic and axonal genes a progressive decay. A substantial proportion of differentially expressed genes (~40%) showed inversion of their temporal profiles in the last time point: spliceosome and proteasome showed initial down-regulation and stress-response genes initial up-regulation. Extensive regulation was detected for chromatin remodelers of the DNMT and CBX families as well as members of the polycomb complex and was mirrored by an up-regulation of the H3K27me3 epigenetic mark. Network analysis showed extensive coregulation of cell cycle/DNA synthesis genes with the uncharacterized zinc-finger protein ZNF367 as central hub. In situ hybridization showed that ZNF367 is expressed in neuronal stem cell niches of both embryonic zebrafish and adult N. furzeri. Other genes down-regulated with age, not previously associated with adult neurogenesis and with similar patterns of expression are AGR2, DNMT3A, KRCP, MEX3A, SCML4, and CBX1. CBX7, on the other hand, was up-regulated with age.

Summary The current molecular understanding of the aging process derives almost exclusively from the study of random or targeted single‐gene mutations in highly inbred laboratory species, mostly invertebrates. Little information is available as to the genetic mechanisms responsible for natural lifespan variation and the evolution of lifespan, especially in vertebrates. Here, we investigated the pattern of positive selection in annual (i.e., short‐lived) and nonannual (i.e., longer‐lived) African killifishes to identify a genomic substrate for evolution of annual life history (and reduced lifespan). We identified genes under positive selection in all steps of mitochondrial biogenesis: mitochondrial (mt) DNA replication, transcription from mt promoters, processing and stabilization of mt RNAs, mt translation, assembly of respiratory chain complexes, and electron transport chain. Signs of paralleled evolution (i.e., evolution in more than one branch of Nothobranchius phylogeny) are observed in four out of five steps. Moreover, some genes under positive selection in Nothobranchius are under positive selection also in long‐lived mammals such as bats and mole‐rats. Complexes of the respiratory chain are formed in a coordinates multistep process where nuclearly and mitochondrially encoded components are assembled and inserted into the inner mitochondrial membrane. The coordination of this process is named mitonuclear balance, and experimental manipulations of mitonuclear balance can increase longevity of laboratory species. Our data strongly indicate that these genes are also casually linked to evolution lifespan in vertebrates.

Summary Aging research in vertebrates is hampered by the lack of short- lived models. Annual fishes of the genus Nothobranchius live in East African seasonal ponds. Their life expectancy in the wild is limited by the duration of the wet season and their lifespan in captivity is also short. Nothobranchius are popular aquarium fishes and many different species are kept as captive strains, providing rich material for comparative studies. The present paper aims at reviving the interest in these fishes by reporting that: (1) Nothobranchius can be cultured, and their eggs stored dry at room temperature for months or years, offering inexpensive methods of embryo storage; (2) Nothobranchius show accelerated growth and expression of aging biomarkers at the level of histology and behaviour; (3) the species Nothobranchius furzeri has a maximum lifespan of only 3 months and offers the possibility to perform investigations thus far unthinkable in a vertebrate, such as drug screening with life-long pharmacological treatments and experimental evolution; (4) when the lifespan of different species is compared, a general correlation is found between wet season duration in their natural habitat and longevity in captivity; and (5) vertebrate aging-related genes, such as p66Shc and MTP , can be easily isolated in Nothobranchius by homology cloning. These fishes can become excellent models for aging studies. They can be employed to test the effects of experimental manipulation on aging at a pace com- parable with that of Drosophila and to probe the effects of natural selection on the evolution of aging-related genes.

The African annual fish Nothobranchius furzeri emerged as a new model for age research over recent years. Nothobranchius furzeri show an exceptionally short lifespan, age-dependent cognitive/behavioral decline, expression of age-related biomarkers, and susceptibility to lifespan manipulation. In addition, laboratory strains differ largely in lifespan. Here, we set out to study the genetics of lifespan determination. We crossed a short- to a long-lived strain, recorded lifespan, and established polymorphic markers. On the basis of genotypes of 411 marker loci in 404 F(2) progeny, we built a genetic map comprising 355 markers at an average spacing of 5.5 cM, 22 linkage groups (LGs) and 1965 cM. By combining marker data with lifespan values, we identified one genome-wide highly significant quantitative trait locus (QTL) on LG 9 (P0.01), which explained 11.3% of the F(2) lifespan variance, and three suggestive QTLs on LG 11, 14, and 17. We characterized the highly significant QTL by synteny analysis, because a genome sequence of N. furzeri was not available. We located the syntenic region on medaka chromosome 5, identified candidate genes, and performed fine mapping, resulting in a c. 40% reduction of the initial 95% confidence interval. We show both that lifespan determination in N. furzeri is polygenic, and that candidate gene detection is easily feasible by cross-species analysis. Our work provides first results on the way to identify loci controlling lifespan in N. furzeri and illustrates the potential of this vertebrate species as a genetic model for age research.

Among vertebrates that can be kept in captivity, the annual fish Nothobranchius furzeri possesses the shortest known lifespan. It also shows typical signs of aging and is therefore an ideal model to assess the role of different physiological and environmental parameters on aging and lifespan determination. Here, we used Nothobranchius furzeri to study whether aging is associated with mitochondrial DNA (mtDNA) alterations and changes in mitochondrial function. We sequenced the complete mitochondrial genome of N. furzeri and found an extended control region. Large-scale mtDNA deletions have been frequently described to accumulate in other organisms with age, but there was no evidence for the presence of detectable age-related mtDNA deletions in N. furzeri. However, mtDNA copy number significantly decreased with age in skeletal muscle, brain, liver, skin and dorsal fin. Consistent with this finding, expression of Pgc-1α that encodes a transcriptional coactivator of mitochondrial biogenesis and expression of Tfam and mtSsbp both encoding mtDNA binding factors was downregulated with age. The investigation of possible changes in mitochondrial function revealed that the content of respiratory chain complexes III and IV was reduced in skeletal muscle with age. In addition, ADP-stimulated and succinate-dependent respiration was decreased in mitochondria of old fish. These findings suggest that despite the short lifespan, aging in N. furzeri is associated with a decline in mtDNA copy number, the downregulation of mtDNA-associated genes and an impairment of mitochondrial function.

We studied adult neurogenesis in the short-lived annual fish Nothobranchius furzeri and quantified the effects of aging on the mitotic activity of the neuronal progenitors and the expression of glial fibrillary acid protein (GFAP) in the radial glia. The distribution of neurogenic niches is substantially similar to that of zebrafish and adult stem cells generate neurons, which persist in the adult brain. As opposed to zebrafish, however, the N. furzeri genome contains a doublecortin (DCX) gene. Doublecortin is transiently expressed by newly generated neurons in the telencephalon and optic tectum (OT). We also analyzed the expression of the microRNA miR-9 and miR-124 and found that they have complementary expression domains: miR-9 is expressed in the neurogenic niches of the telencephalon and the radial glia of the OT, while miR-124 is expressed in differentiated neurons. The main finding of this paper is the demonstration of an age-dependent decay in adult neurogenesis. Using unbiased stereological estimates of cell numbers, we detected an almost fivefold decrease in the number of mitotically active cells in the OT between young and old age. This reduced mitotic activity is paralleled by a reduction in DCX labeling. Finally, we detected a dramatic up-regulation of GFAP in the radial glia of the aged brain. This up-regulation is not paralleled by a similar up-regulation of S100B and Musashi-1, two other markers of the radial glia. In summary, the brain of N. furzeri replicates two typical hallmarks of mammalian aging: gliosis and reduced adult neurogenesis.

Temperature variations are known to modulate aging and life-history traits in poikilotherms as different as worms, flies and fish. In invertebrates, temperature affects lifespan by modulating the slope of age-dependent acceleration in death rate, which is thought to reflect the rate of age-related damage accumulation. Here, we studied the effects of temperature on aging kinetics, aging-related behavioural deficits, and age-associated histological markers of senescence in the short-lived fish Nothobranchius furzeri. This species shows a maximum captive lifespan of only 3 months, which is tied with acceleration in growth and expression of aging biomarkers. These biological peculiarities make it a very convenient animal model for testing the effects of experimental manipulations on life-history traits in vertebrates. Here, we show that (i) lowering temperature from 25 degrees C to 22 degrees C increases both median and maximum lifespan; (ii) life extension is due to reduction in the slope of the age-dependent acceleration in death rate; (iii) lowering temperature from 25 degrees C to 22 degrees C retards the onset of age-related locomotor and learning deficits; and (iv) lowering temperature from 25 degrees C to 22 degrees C reduces the accumulation of the age-related marker lipofuscin. We conclude that lowering water temperature is a simple experimental manipulation which retards the rate of age-related damage accumulation in this short-lived species.

Among vertebrates that can be kept in captivity, the annual fish Nothobranchius furzeri possesses the shortest known lifespan. It also shows typical signs of aging and is therefore an ideal model to assess the role of different physiological and environmental parameters on aging and lifespan determination. Here, we used Nothobranchius furzeri to study whether aging is associated with mitochondrial DNA (mtDNA) alterations and changes in mitochondrial function. We sequenced the complete mitochondrial genome of N. furzeri and found an extended control region. Large-scale mtDNA deletions have been frequently described to accumulate in other organisms with age, but there was no evidence for the presence of detectable age-related mtDNA deletions in N. furzeri. However, mtDNA copy number significantly decreased with age in skeletal muscle, brain, liver, skin and dorsal fin. Consistent with this finding, expression of Pgc-1α that encodes a transcriptional coactivator of mitochondrial biogenesis and expression of Tfam and mtSsbp both encoding mtDNA binding factors was downregulated with age. The investigation of possible changes in mitochondrial function revealed that the content of respiratory chain complexes III and IV was reduced in skeletal muscle with age. In addition, ADP-stimulated and succinate-dependent respiration was decreased in mitochondria of old fish. These findings suggest that despite the short lifespan, aging in N. furzeri is associated with a decline in mtDNA copy number, the downregulation of mtDNA-associated genes and an impairment of mitochondrial function.