Classically, the zebrafish model organism has been used to elucidate the genetic and
cellular mechanisms related to development since the embryo forms and grows externally
following fertilization. This provides insight into the genetic control of developmental
processes in humans because their genomes are similar. Also, unlike other animal models,
the genes of zebrafish can be manipulated quite easily by using reverse genetic screens
tools such as morpholinos, which transiently silence target genes of interest or systems
such as the transposon-mediated insertional mutagenesis or CRISPR-Cas9. Moreover,
one pair of fish will provide up to 300 offspring, which means that if there is a
gene of interest that is manipulated, then it can be transmitted to a large population
of fish. What is beginning to emerge is that similar to other mammals, adult zebrafish
have an integrated nervous system, which is proposed to contain homologous brain structures
to those found in humans, as well as equivalent cellular and synaptic structure and
function. Moreover, like humans, zebrafish exhibit age-related declines in cognitive
functions, and a convergence of evidence has indicated that subtle changes in cellular
and synaptic integrity underlie these changes. Therefore, the zebrafish is a powerful
model organism for studying the neurobiological consequences of aging-related behavioral
and biological changes, which offers the potential to identify possible interventions
that would promote healthy aging. In what follows, we present and discuss recent findings
and advances along these directions.
Behavioral tasks and abilities altered in aged zebrafish
The zebrafish is a promising model for studying age-related changes in cognition and
perception. Early behavioral studies date back to 1960s and the characterization of
zebrafish behavior has accelerated since 2000 (Kalueff et al., 2013). They have been
suggested to reflect the evolutionarily conserved nature of many behaviors and to
resemble those of other species (Kalueff et al., 2014; Stewart et al., 2014; Orger
and de Polavieja, 2017). A rich repertoire of behavioral phenotypes has been identified
for cognitive functioning, perceptual processes, and associated disorders (Stewart
and Kalueff, 2012). Using different behavioral assays (e.g., inter- and intra-trial
habituation, T-maze, conditioned place preference paradigms), previous studies indicated
that zebrafish have both simple and relatively complex forms of learning, and also
display good performance on cognitive tasks dependent on short-term and long-term
memory (Blaser and Vira, 2014; Gerlai, 2016). There is also growing interest in other
aspects of zebrafish behavior which significantly depend on perception, low-level
discrimination, and sensitivity (Neuhauss, 2010). For instance, the basic components
of the zebrafish visual system, the visual processing hierarchy, and pathways are
similar to those commonly found in other species (Bilotta and Saszik, 2001). In particular,
most of the previous research evaluated visual motion perception and sensitivity through
optomotor response and/or optokinetic reflexive eye movements. These behavioral studies
point to qualitatively similar visual acuity and contrast sensitivity functions for
zebrafish (Rinner et al., 2005; Haug et al., 2010; Tappeiner et al., 2012). It has
also been shown that zebrafish perceive first- and second-order motion. They also
experience motion illusions commonly used in studies on human vision such as reverse-phi
illusion, motion aftereffect, and rotating snakes illusion (Orger et al., 2000; Gori
et al., 2014; Najafian et al., 2014). Within the context of visual motion, these studies
provide behavioral evidence that mechanisms and principles similar to those of humans
and other species underlie zebrafish sensory processing and associated behavior.
Characterizing aging-related changes in zebrafish behavior has important implications
for our understanding of cognition and perception. First, aging-related changes in
cognition are a part of the normal aging process and common in all the species. Monitoring
age-dependent changes in cognition and perception is difficult to perform on the same
human subject throughout life. Due to their short lifespan, behavioral assays and
paradigms developed, zebrafish provides an ideal model to study cognitive and perceptual
performance during aging. Second, when these behavioral studies are combined with
already developed molecular and genetic tools on this aging model, we will also have
a deeper understanding on the functional links between key synaptic targets, cognition,
and perception during neural aging. Previous studies report significant declines in
learning and memory in aged zebrafish. Typically, old zebrafish have less performance
on tasks relevant with associative learning, avoidance, spatial learning and working
memory (Yu et al., 2006; Arey and Murphy, 2017; Brock et al., 2017). Compared to wild-types,
mutants with impaired acetylcholinesterase function had better performance in spatial
learning, entrainment and increased rate of learning (Yu et al., 2006; Parker et al.,
2015). These findings suggest that cholinergic signaling may also play a role in age-related
cognitive decline. In terms of perceptual performance, there are studies comparing
larvae and adult zebrafish. However, we have limited knowledge on how perceptual performance
(and thus perception and sensitivity) changes during neural aging. A challenge for
the future is to characterize aging-related changes in perceptual performance and
sensitivity of adult zebrafish. As mentioned above, we consider that such studies
can provide comprehensive information not only on perception and behavior in general
(Owsley, 2016) but also on the cellular and molecular mechanisms underlying specific
aspects (e.g., motion) of perception and sensitivity.
Aging-related neurobiological alterations
Understanding the cellular mechanisms that underlie cognitive decline is important
for determining sites of actions for possible interventions that could ameliorate
alterations in cognitive function. Early reports indicated that age-related cognitive
decline was due to significant cell (Brody, 1955; Devaney and Johnson, 1980; Henderson
et al., 1980) and synapse loss (Geinisman et al., 1977; Bondareff, 1979; Curcio and
Hinds, 1983; Haug and Eggers, 1991; Shi et al., 2005). However, it has become well
accepted that significant cell (Haug and Eggers, 1991; Rapp and Gallagher, 1996; Rasmussen
et al., 1996; Peters et al., 1998) and synapse loss does not occur in conjunction
with normal aging-related declines in cognitive capacities (Poe et al., 2001; Newton
et al., 2007; Shi et al., 2007). Therefore, research studies have been designed at
examining markers of cellular and synaptic integrity during the aging process, such
as altered neurogenesis rates (Kempermann et al., 1998, Luo et al., 2006) and the
levels of key excitatory and inhibitory pre- and post-synaptic proteins (Newton et
al., 2007; Shi et al., 2007; Adams et al., 2008), since subtle changes in cellular
and synaptic functions likely underlie the aging-related declines in cognitive abilities.
Moreover, examining key molecular targets that control these processes will increase
our understanding of the cellular and synaptic regulation of behavior across the lifespan.
While these aging-related changes in cellular and synaptic processes could be examined
in many different animal species, the zebrafish model organism is well-adapted to
studying the cellular and molecular changes with aging because they have similar patterns
as mammals with regards to the cellular aging process. Zebrafish on average live approximately
three to five years and share a similar genome with humans (Kishi et al., 2003; Howe
et al., 2013). Moreover, senescence-associated ß-galactosidase, which is a biomarker
of aging, increases with advancing age in zebrafish, and this cellular alteration
has been described in humans as well (Kishi et al., 2003; Arslan-Ergul et al., 2016).
Finally, zebrafish have continued neurogenesis even into late adulthood (Kizil et
al., 2012; Schmidt et al., 2013), they express key excitatory and inhibitory pre-
and post-synaptic proteins (Karoglu et al., 2017), and classical cellular synaptic
plasticity (i.e., long-term potentiation) is found in their brains (Nam et al., 2004).
Recent work in the zebrafish brain has demonstrated that there are age-related declines
in genes related to cellular and synaptic structure and growth (Arslan-Ergul and Adams,
2014), neurogenesis (Edelmann et al., 2013; Arslan-Ergul et al., 2016), and synaptic
alterations (Arslan-Ergul et al., 2016; Karoglu et al., 2017). Interestingly, as has
been shown in mammals, these changes depend on the gender of the animal (Arslan-Ergul
and Adams, 2014; Karoglu et al., 2017), and the data are in good agreement with those
showing sexually-dimorphic patterns published in young zebrafish brains (Ampatzis
et al., 2012). Taken together, these findings indicate that the zebrafish is an appropriate
model to study the effects of cellular and synaptic aging and its relationship to
cognitive decline.
Use of interventions to alter aging-related processes
A major goal of research related to elucidating the altered cellular and synaptic
processes that underlie cognitive aging is to determine possible interventions to
restore youthful cellular and synaptic function. As was mentioned previously, mutant
zebrafish with lower levels of acetylcholinesterase had better performance in spatial
learning, entrainment, and increased rate of learning (Yu et al., 2006; Parker et
al., 2015). Therefore, these animals likely have a more youthful cellular and synaptic
profile as compared to their wild-type counterparts. Currently, we are investigating
this possibility and our data suggest that genetic manipulation of the cholinergic
system alters the course of aging-related changes in the synaptic protein levels.
We have demonstrated that at old ages as compared to their wild-type siblings, mutants
have higher levels of synaptophysin, which is an indicator of presynaptic integrity,
and gephyrin, a component of post-synaptic inhibitory transmission, and interestingly
these changes are gender-dependent (Karoglu et al., 2018). If we can determine the
cellular and synaptic profile of these mutants and how they relate to cognitive aging,
it would provide potential targets for drug development to ameliorate the effects
of cognitive decline.
Another potential intervention with promise is dietary restriction (DR), which is
the only non-genetic intervention that reliably increases both lifespan and healthspan.
Numerous studies have shown that a lifelong reduction in caloric intake from ad libitum
levels increases lifespan (Roth et al., 2001; Lin et al., 2002; Colman et al., 2009).
Additionally, DR increases neuronal proliferation and survival (Lee et al., 2002;
Kitamura et al., 2006; Park and Lee, 2011; Park et al., 2013). We applied a short-term
DR of 10 weeks and observed that this treatment did not prevent an age-related decline
in cell proliferation but altered the telomere lengths of these neuronal cells (Arslan-Ergul
et al., 2016), thereby DR exerted positive effects by subtly altering the cell cycle
dynamics of these neurons. We have tested the timing and duration of short-term DR
and a potential DR-mimetic, rapamycin, as the positive effects of DR are thought to
be modulating the mammalian target of rapamycin signaling pathway. Our data indicate
that a longer duration of both DR and its mimetic is more effective on aging-related
changes in synaptic protein levels and transcripts, which might reflect a conserved
mechanism of the beneficial effects of DR and rapamycin on life- and healthspan (Celebi-Birand
et al., 2018). These studies also have the potential to provide suitable therapeutic
targets around which drug development can proceed for ameliorating the devastating
effects of cognitive decline.
Conclusions
The zebrafish is clearly a powerful model organism that can be used to understand
the aging-related changes in both cognition and the underlying cellular and molecular
processes. As previously mentioned, zebrafish exhibit characteristics that are similar
to humans, as well as other mammals, including the fact that these animals age gradually,
and they demonstrate aging-related changes across both cognitive and neurobiological
spectrums. It clear that both genetic and non-genetic interventions can be applied
to alter the course of the aging process and provide potential drug targets that could
be manipulated to ameliorate age-related cognitive declines. Therefore, this model
will help researchers elucidate the biological mechanisms that underlie aging-related
cognitive decline.
Author contributions
All authors listed have made a substantial, direct and intellectual contribution to
the work, and approved it for publication.
Conflict of interest statement
The authors declare that the research was conducted in the absence of any commercial
or financial relationships that could be construed as a potential conflict of interest.