Alessandro Cellerino

Resveratrol Prolongs Lifespan and Retards the Onset of Age-Related Markers in a Short-Lived Vertebrate

Resveratrol, a natural phytoalexin found in grapes and red wine, increases longevity in the short-lived invertebrates Caenorhabditis elegans and Drosophila and exerts a variety of biological effects in vertebrates, including protection from ischemia and neurotoxicity. Its effects on vertebrate lifespan were not yet known. The relatively long lifespan of mice, which live at least 2.5 years, is a hurdle for life-long pharmacological trials. Here, the authors used the short-lived seasonal fish Nothobranchius furzeri with a maximum recorded lifespan of 13 weeks in captivity. Short lifespan in this species is not the result of spontaneous or targeted genetic mutations, but a natural trait correlated with the necessity to breed in an ephemeral habitat and tied with accelerated development and expression of ageing biomarkers at a cellular level. Resveratrol was added to the food starting in early adulthood and caused a dose-dependent increase of median and maximum lifespan. In addition, resveratrol delays the age-dependent decay of locomotor activity and cognitive performances and reduces the expression of neurofibrillary degeneration in the brain. These results demonstrate that food supplementation with resveratrol prolongs lifespan and retards the expression of age-dependent traits in a short-lived vertebrate.

Molecular determinants of retinal ganglion cell development, survival, and regeneration.

The retina is an easily accessible part of the CNS with a well-defined cytological architecture. It allows for detailed study of the regulation of neurogenesis, determinants of cell fate specification, and signals for cell survival versus programmed cellular death during development. Within the retina, retinal ganglion cells (RGCs) are the only neurons connecting to the brain. Their axonal projection to the midbrain targets, the superior colliculus (SC), and the lateral geniculate nucleus (LGN) has been subject of a number of investigations, and led to the identification of molecular signals directing topographic information for precise wiring during development. Transcription factors, guidance molecules, extracellular matrix proteins, neurotrophic factors, and cell death-regulating factors of the Bcl-2 family and caspases, have all been reported to be involved in the processes of formation of a precise retino-collicular map, and regulation of developmental cell death.During adulthood, RGCs and their projection have to be maintained, since-to our current knowledge-they cannot be replaced following injury. On the other hand, insults of various kinds can be potentially hazardous to RGCs. Therefore, much work has been directed towards understanding of the molecular regulation of RGC degeneration following insults such as retinal ischaemia, axonal lesion, or in optic neuropathy. Experimental strategies are being devised towards protection of lesioned RGCs. Since following axonal lesion, these cells not only need to survive, but also have to reconnect in order to be functionally relevant, efforts are directed towards not only survival, but also axonal regeneration and proper rewiring of injured RGCs. This paper reviews the molecular determinants of RGC fate determination and the development of the retino-tectal projection. We summarize what is known (and hypothesized) on the determinants of RGC survival during normal adulthood, and the mechanisms of RGC degeneration in the injured retina. We also try to develop perspectives towards neuroprotection and regeneration of adult lesioned RGCs that may be applicable to lesioned CNS neurons in vertebrates in a broader sense.

Annual fishes of the genus Nothobranchius as a model system for aging research

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 comparable with that of Drosophila and to probe the effects of natural selection on the evolution of aging‐related genes.

Brain-derived neurotrophic factor/neurotrophin-4 receptor TrkB is localized on ganglion cells and dopaminergic amacrine cells in the vertebrate retina

The tyrosine kinase TrkB is a receptor for the neurotrophic factors brain‐derived neurotrophic factor (BDNF) and neurotrophin‐4 (NT‐4). Retinal ganglion cells are responsive to BDNF, and TrkB has been localized in ganglion cells as well as in a subpopulation of amacrine cells in the retina of the chicken and the rat. In the present paper, we analyzed the distribution of TrkB immunoreactivity in the retina of marmoset monkeys, ferrets, rabbits, rats, mice, chickens, pigeons, barn owls, Pseudemys turtles, Xenopus frogs, goldfishes, and carps. TrkB antibodies gave a positive reaction in all of these vertebrates. TrkB immunoreactivity was detected in the majority of retinal ganglion cells. Some amacrine cells also contained TrkB immunoreactivity; they were located mainly at the vitreal border of the inner nuclear layer, and their relative abundance varied in the different species. Until now, no information has been available concerning the neurochemical identity of the amacrine neurons containing TrkB. In some species (marmoset monkeys, rats, pigeons), we observed that the morphology and location of TrkB‐immunoreactive amacrine cells was reminiscent of that of the well‐described dopaminergic cells. To determine whether dopaminergic amacrine cells contained TrkB immunoreactivity, we therefore performed double‐labelling immunohistochemistry by using tyrosine hydroxylase (TH) antibodies in combination with TrkB antibodies in marmoset monkeys, rats, pigeons, Pseudemys turtles, and goldfishes. The most novel finding of the present paper is that, in all of these species, the majority of dopaminergic neurons were found to contain TrkB immunoreactivity. Dopaminergic neurons, on the other hand, represented only a fraction of the TrkB+ amacrine cells.

THE ACTION OF NEUROTROPHINS IN THE DEVELOPMENT AND PLASTICITY OF THE VISUAL CORTEX

Nerve growth factor (NGF) and the other members of the NGF gene family have been extensively characterized as neurotrophic factors. Recently a modulatory action of these neurotrophic factors on synapse efficacy has emerged. The developing visual system has provided a convenient model to test the role of neurotrophins on neural plasticity in vivo.

The Distribution of Brain-derived Neurotrophic Factor and its Receptor trkB in Parvlbumin-containing Neurons of the Rat Visual Cortex

We analysed the distribution of brain‐derived neurotrophic factor (BDNF) and its receptor trkB in the adult rat visual cortex, paying particular attention to a GABAergic neuronal subpopulation—the parvalburnin‐positive cells. We found expression of trkB in the cell body and apical dendrite of pyramidal neurons and in the cell body of non‐pyramidal neurons. Double labelling experiments revealed extensive colocalization of parvalbumin and trkB immunoreactivity in non‐pyramidal neurons. Interestingly, the trkB‐positive pyramidal neurons appeared surrounded by parvalbumin‐labelled boutons. The use of double immunohistochemistry and in situ hybridization histochemistry showed that parvalbumin‐positive neurons express trkB mRNA. BDNF rnRNA was found in several cells. Coexpression of BDNF mRNA and parvalbumin immunoreactivity was extremely rare. These data strongly suggest that BDNF synthesized by cortical neurons acts as a postsynaptically derived factor for parvalbumin‐positive neurons in the adult rat visual cortex.

Large Differences in Aging Phenotype between Strains of the Short-Lived Annual Fish Nothobranchius furzeri

Background A laboratory inbred strain of the annual fish Nothobranchius furzeri shows exceptionally short life expectancy and accelerated expression of age markers. In this study, we analyze new wild-derived lines of this short-lived species. Methodology/Principal Findings We characterized captive survival and age-related traits in F1 and F2 offspring of wild-caught N. furzeri. Wild-derived N. furzeri lines showed expression of lipofuscin and neurodegeneration at age 21 weeks. Median lifespan in the laboratory varied from to 20 to 23 weeks and maximum lifespan from 25 to 32 weeks. These data demonstrate that rapid age-dependent decline and short lifespan are natural characteristics of this species. The N. furzeri distribution range overlaps with gradients in altitude and aridity. Fish from more arid habitats are expected to experience a shorter survival window in the wild. We tested whether captive lines stemming from semi-arid and sub-humid habitats differ in longevity and expression of age-related traits. We detected a clear difference in age-dependent cognitive decline and a slight difference in lifespan (16% for median, 15% for maximum lifespan) between these lines. Finally, we observed shorter lifespan and accelerated expression of age-related markers in the inbred laboratory strain compared to these wild-derived lines. Conclusions/Significance Owing to large differences in aging phenotypes in different lines, N. furzeri could represent a model system for studying the genetic control of life-history traits in natural populations.

Temperature affects longevity and age-related locomotor and cognitive decay in the short-lived fish Nothobranchius furzeri

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 °C to 22 °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 °C to 22 °C retards the onset of age‐related locomotor and learning deficits; and (iv) lowering temperature from 25 °C to 22 °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.

Extremely short lifespan in the annual fish Nothobranchius furzeri

Evolutionary theories of senescence postulate that lifespan is determined by the age-dependent decrease in the effects of natural selection. Factors that influence survival and reproduction at early life stages have a larger impact on fitness than factors that influence later life stages. According to these views, selection for rapid sexual maturation and a steep age-dependent decrease in fitness drive the evolution of short lifespans. Here, we report on the survival trajectory of Nothobranchius furzeri (Pisces: Ciprinodontidae): a member of a group of annual species found in temporary bodies of water whose life expectancy in the wild is limited to a few months. We find that maximum survival of N. furzeri in the laboratory is less than 12 weeks. The temporal trajectory of survival shows an age-dependent increase in the mortality rate that is typical of organisms with defined lifespans. The lifespan of N. furzeri is exceptionally short for a vertebrate: owing to its small size and the possibility of propagation in captivity, N. furzeri could be used as a convenient model for ageing research.

Brain-Derived Neurotrophic Factor Modulates the Development of the Dopaminergic Network in the Rodent Retina

Dopaminergic cells in the retina express the receptor for brain-derived neurotrophic factor (BDNF) (Cellerino and Kohler, 1997). To investigate whether BDNF can influence the development of the retinal dopaminergic pathway, we performed intraocular injections of BDNF during the second or third postnatal week and visualized the dopaminergic system with tyrosine hydroxylase (TH) immunohistochemistry. Both regimens of BDNF treatment caused an increase in TH immunoreactivity in stratum 1 and stratum 3 of the inner plexiform layer (IPL). D2 dopamine receptor immunoreactivity, a presynaptic marker of dopaminergic cells (Veruki, 1996), was also increased in stratum 1 and stratum 3 of the inner plexiform layer. These data suggest that BDNF causes sprouting of dopaminergic fibers in the inner plexiform layer. Other neurochemical systems, for example, the cholinergic amacrine cells, remained unaffected. Similar effects were observed after injections of neurotrophin-3 and neurotrophin-4, but not nerve growth factor. Analysis of whole-mounted TH-immunolabeled retinae revealed hypertrophy of dopaminergic cells (+41% in soma areas; p < 0.01) and an increase of labeled dopaminergic varicosities in stratum 1 of the IPL (+51%;p < 0.01) after BDNF treatment. The opposite was observed in mice homozygous for a null mutation of thebdnf gene: dopaminergic cells were atrophic (−22.5% in soma areas; p < 0.05), and the density of TH-positive varicosities in stratum 1 was reduced (57%;p < 0.01). We conclude that BDNF controls the development of the retinal dopaminergic network and may be particularly important in determining the density of dopaminergic innervation in the retina.