Tissue stretching is a confounding factor for the evaluation of neurodegeneration in the ever-growing killifish

Abstract

Abstract The fast-ageing killifish, Nothobranchius furzeri, has gained increasing attention as a promising gerontology model to study age-related processes and neurodegeneration. Interestingly, it is the first vertebrate model organism that shows physiological neurodegeneration at old age in its central nervous system (CNS), including its brain and retina. However, the fact that the killifish brain and retina are ever-growing tissues complicates studying neurodegenerative events. Indeed, recent studies showed that the method of tissue sampling, either using sections or whole-organs, has a large effect on the observed cell densities in the fast-growing CNS. In this study, we elaborated on how these two sampling methods alter neuronal frequency within the retina and how this tissue grows throughout life. Analysis of cellular density across the different retinal layers in cryosections revealed age-dependent cell loss, that was not observed in retinal whole-mounts, as a result of an extremely fast retinal expansion with age. Using BrdU pulse-chase experiments, we were able to show that the young adult killifish retina mainly grows by cell addition, largely facilitated by the ciliary marginal zone. However, with increasing age, the neurogenic potency of this zone declines while the tissue keeps on growing. Further histological analyses revealed tissue stretching, including cell size increase, as the main driver of retinal growth in older fish. Indeed, both cell size and distance between retinal neurons augments with ageing, thereby decreasing neuronal density. All in all, these findings highlight the need for tissue-wide counting methods to reliably quantify cell numbers in the fast-growing killifish.