Early life adversity is a known risk factor for mood and anxiety disorders in adult
humans (Heim et al., 2010; Huh et al., 2014; Rehan et al., 2017). Given the prevalence
of both maltreatment in childhood and mental illness in adulthood, understanding the
neurobiological mechanisms of this connection is important as it may suggest targets
for new therapeutic interventions. Ethical constraints on conducting studies with
humans have highlighted the need for reliable and robust animal models that researchers
can utilize to identify relevant neurobiological processes (Guzman et al., 2016).
Since the work of Harlow and colleagues beginning in the 1940s, which involved raising
infant macaques with cloth and wire mothers (reviewed in van der Horst and van der
Veer, 2008), researchers have sought to develop useful animal models of early life
adversity. These and other more recent studies have shown obvious behavioral abnormalities
in monkeys subjected to early life stress (ELS) (Schino et al., 2001; Corcoran et
al., 2012; Howell et al., 2014). Despite the relevance of these models to humans,
nonhuman primates have practical and ethical limitations that are obstacles for their
use in high-throughput studies. By contrast, animal models of early life stress in
rodents, which were first used in the laboratory more than 50 years ago (Levine, 1957),
have gained in usage.
One of the most commonly used manipulations to produce a rodent model of ELS has been
maternal separation. Studies have shown that maternal separation in rats, as long
as it is of sufficient duration (typically 3 h/day during the first 2 postnatal weeks
of life) increases anxiety- and depressive-like behaviors in adulthood, suggesting
that it has translational validity (Janus, 1987; Huot et al., 2001; Kalinichev et
al., 2002; Romeo et al., 2003; Daniels et al., 2004; Lee et al., 2007; Wei et al.,
2010; Masrour et al., 2018). However, other studies in both rats and mice have shown
considerable variability in behavioral results from maternal separation, with several
reports showing no behavioral effect (Lehmann et al., 1999; Eklund and Arborelius,
2006; Slotten et al., 2006; Millstein and Holmes, 2007; Savignac et al., 2011). In
addition to inconsistent behavioral findings with this model, concerns have been raised
about whether maternal separation mimics neglect, abuse or a combination of both.
It has been reported that after prolonged separation, maternal behavior toward pups
differs and these differences may be as important, if not more, than the lack of contact
with the mother (Boccia and Pedersen, 2001; Huot et al., 2004). Some reports have
also observed that dams increase maternal care post-separation possibly attenuating
the effects of the separation itself (Millstein and Holmes, 2007). The type of human
maltreatment that rodent maternal separation reflects might be important for establishing
its translational validity, since human studies have separated early adverse experiences
into several categories, including emotional abuse, emotional neglect, physical abuse,
physical neglect and sexual abuse (Kendler et al., 2004; van Harmelen et al., 2010;
Young and Widom, 2014; Rehan et al., 2017; Gallo et al., 2018) and some studies suggest
that the type of maltreatment may be important for the adult outcome in terms of behavioral
dysfunction (Huh et al., 2014; Young and Widom, 2014).
To address concerns about the unspecified nature of the maternal separation manipulation,
researchers have developed another way to impair maternal care with the limited bedding/nesting
model (Brunson et al., 2005; Cui et al., 2006; Ivy et al., 2008; Rice et al., 2008).
The most extreme version of this model involves housing dams in a wire mesh floored
cage with no bedding and a scarcity of material with which to make a nest, while variations
involve just limiting nesting material (Walker et al., 2017). The result is an increase
in maternal anxiety and fractured caregiving where behavior toward the pups might
be interpreted as abusive (Rice et al., 2008). As with the earlier investigations
of the maternal separation model, some studies using this manipulation reported evidence
for increased anxiety- and depressive-like behavior in adulthood (Cui et al., 2006;
Dalle Molle et al., 2012; Raineki et al., 2012; Wang et al., 2012), supporting its
translational validity. However, other studies using this model failed to find an
increase in anxiety- or depressive-like behavior (Brunson et al., 2005; Rice et al.,
2008; van der Kooij et al., 2015; Johnson et al., 2018) raising questions about reliability
similar to those observed with the maternal separation model.
Contradictory results of studies using both of these rodent models are puzzling and
may be attributable to differences in experimental design. To fully understand these
discrepancies, many factors must be considered (Figure 1). First, the genetic background
of the experimental animal is important. Human studies have clearly shown genetic
predisposition to mood and anxiety disorders and it follows that this factor should
be considered in studies using experimental animals to model the human condition.
Studies have shown varying effects of maternal separation on anxiety- and depressive-like
behaviors in different strains of mice; the C57Bl/6 strain appears to be most resistant
to stress compared to other strains, such as the Balb/c strain, which is inherently
more anxious (Millstein and Holmes, 2007; Wei et al., 2010; Savignac et al., 2011).
However, different studies using the same strain have reported conflicting results
with seemingly identical ELS manipulations, so genetic strain differences cannot account
for all of the variance in the literature. It should be noted, however, that individual
subtler genetic differences within a specific rodent strain may be relevant to establishing
vulnerability to such manipulations. That is, ELS manipulations likely impact some
animals more than others and such variability may obscure overall group differences
in behavior. Second, the sex of the animal should be considered. Somewhat paradoxically
given that women exhibit greater prevalence of mood/anxiety disorders than do men
(Altemus et al., 2014), several rodent studies have shown that ELS produces either
no effect or a reduction in anxiety- and depressive-like behaviors in females (Lehmann
et al., 1999; McIntosh et al., 1999; Eklund and Arborelius, 2006; Slotten et al.,
2006). These unexpected results raise questions about whether the standard laboratory
tests of anxiety- and depressive-like behavior, which were developed for use in males
and typically involve measures of behavioral inhibition, are accurate measures of
these states in female rodents, given known estrous cycle variations in behavioral
activity levels. Clearly, the field would benefit from new sensitive behavioral assays
that are useful for both sexes, particularly given the need to correct the under-emphasis
of research on females (Clayton and Collins, 2014).
Figure 1
Factors influencing variability in ELS animal models. Cartoon depicting the two most
commonly used models of ELS and the different factors influencing behavioral outcomes
in adulthood. MS, Maternal separation stress; LB-LN, Limited bedding-Limited nesting.
The timing and duration of the stressful experience during the postnatal period may
also be important to consider. In rodent studies, differential effects of early vs.
late postnatal stress exposure on depressive-like behaviors have been demonstrated
(van der Kooij et al., 2015; Peña et al., 2017). By contrast, however, a recent human
study concluded that data on the link between childhood maltreatment and psychopathology
do not fit a sensitive period theoretical model (Dunn et al., 2018), again raising
questions about the direct translational validity of some ELS models in rodents. It
is likely relevant that the HPA axis response to stress is attenuated in pups during
the stress hyporesponsive period, a phenomenon that serves a protective effect on
the developing brain (Sapolsky and Meaney, 1986). A similar state has been reported
in humans up until about 1 year of age, but it does not extend throughout childhood
when the majority of reported maltreatment occurs (Gunnar and Donzella, 2002).
The duration of stress seems to be more definitively associated with worse outcomes
compared to the timing of stress, and data from human studies support a cumulative
and/or recency model of stress effects on vulnerability to psychopathology (Dunn et
al., 2018). To address the issue about the duration of stress as well as inconsistencies
in the ELS literature, researchers have developed “two-hit” models that incorporate
maternal separation followed by additional stress, either shortly thereafter or in
adulthood. The models are based on the assumption that the first stressful period
may create an internal vulnerability that is alone insufficient to manifest itself
behaviorally, but when aggravated by subsequent stress, produces detectable behavioral
changes. One set of such studies used longer periods of separation followed by early
weaning of pups (George et al., 2010). Early weaning by itself has been shown to increase
anxiety-like behavior in adulthood (Kikusui et al., 2004) and when combined with maternal
separation, it not only increases anxiety-like behaviors but also results in hyperactivity,
gene dysregulation and neuroanatomical changes to the brain; some of which have been
observed in humans with a history of early life abuse. Another set of such studies
used maternal separation and/or limited bedding followed by exposure to chronic stress
in adulthood (Vargas et al., 2016; Peña et al., 2017). Both of these approaches mimic
the “dose-response” or “cumulative” stress links to mental illness that have been
described in humans. However, like the other rodent models of ELS, data from these
two-hit models need to be interpreted with caution as null effects have also been
reported (Santarelli et al., 2017; Tan et al., 2017).
Notwithstanding the potential importance of strain, sex, timing, duration, type of
stress experience and other factors (Figure 1) across studies as reasons for variable
results, it is clear that variable results can emerge even in the face of virtually
identical experimental designs. What is the explanation for these differences? While
we do not know for certain, there are some important points to consider. First, baseline
housing and testing conditions may vary across laboratories in seemingly unspecified
ways (Cavigelli et al., 2006; Sorge et al., 2014), adding additional stress to both
control and experimental groups and potentially reducing the behavioral differences
between them. Second, evidence suggests that rodent maternal behavior varies considerably
even within control groups (Francis and Meaney, 1999). In other words, some rat and
mouse dams may be more capable of compensating for the effects of maternal separation
or limited bedding than others. This could be influenced by the early life experiences
of the dams themselves and the amount of stress they were exposed to before entering
breeding. This natural variation in maternal behavior may introduce additional variability
into ELS-induced long-term behavioral outcomes. Third, perhaps related to the second
point, rodent populations likely display considerable individual variability in response
to ELS, such that depending on the cohort examined, statistically significant differences
in anxiety- and depressive-like behavior may or may not be detectable. Thus, reproducible
significant differences may require larger numbers of animals than are often used
in such studies, consistent with what has been the norm for human studies (Collins
and Tabak, 2014). In addition, these studies might be more informative if the data
from rodents subjected to ELS manipulations were analyzed in ways that do not group
them together with the assumption that they comprise a homogenous group. In searching
for neurobiological mechanisms underlying behavioral signs of mental illness, it may
be more fruitful to separate out the experimental animals that show robust ELS-induced
increases in anxiety- and depressive-like behavior. This approach might reveal informative
correlations between brain changes and relevant behaviors. While this suggestion makes
experimental designs and statistical analyses more complicated than commonly used
methods of comparing means between groups, it may produce more reliable results across
laboratories.
Considering rodent populations as heterogeneous with regard to their susceptibility
to ELS-induced behavioral changes would address an interesting parallel with humans.
While the connection between early life adversity and mood/anxiety disorders in humans
has been widely accepted, it is perhaps less well-known that the majority of people
subjected to childhood maltreatment (>70%) do not show anxiety and depression symptoms
that are clinically significant (Rehan et al., 2017). Thus, as with rodents, humans
display a considerable amount of resilience and resistance to early life adversity,
a phenomenon that deserves scientific attention as it may provide clues about how
to encourage these characteristics in the entire population. Finally, it deserves
mention that many people develop anxiety and mood disorders that are not retrospectively
traceable to childhood maltreatment, so examining control rodents that score as more
anxious/depressed despite a lack of prior stress manipulation may be informative as
well. Here again, looking at individual differences within groups may be most informative
and also help to reduce the inconsistency across studies using rodent models of stress-induced
mental illness.
Author contributions
SM wrote the article, edited the article and made the figure. EG wrote the article,
edited the article and edited the figure.
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.