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Does the Stress of Laboratory Life and Experimentation on Animals Adversely Affect Research Data? A Critical Review

  • Animal Free Research UK


Recurrent acute and/or chronic stress can affect all vertebrate species, and can have serious consequences. It is increasingly and widely appreciated that laboratory animals experience significant and repeated stress, which is unavoidable and is caused by many aspects of laboratory life, such as captivity, transport, noise, handling, restraint and other procedures, as well as the experimental procedures applied to them. Such stress is difficult to mitigate, and lack of significant desensitisation/habituation can result in considerable psychological and physiological welfare problems, which are mediated by the activation of various neuroendocrine networks that have numerous and pervasive effects. Psychological damage can be reflected in stereotypical behaviours, including repetitive pacing and circling, and even self-harm. Physical consequences include adverse effects on immune function, inflammatory responses, metabolism, and disease susceptibility and progression. Further, some of these effects are epigenetic, and are therefore potentially transgenerational: the biology of animals whose parents/grandparents were wild-caught and/or have experienced chronic stress in laboratories could be altered, as compared to free- living individuals. It is argued that these effects must have consequences for the reliability of experimental data and their extrapolation to humans, and this may not be recognised sufficiently among those who use animals in experiments.
In the December 2017 issue of ATLA, I published an
Editorial, in which I suggested that stress in ani-
mals in laboratories, resulting from their environ-
ment and experimental procedures, was not limited
to welfare concerns (1). It also adversely affects, sig-
nificantly and unavoidably, multiple biological sys-
tems and therefore affects the resultant
experimental data. This exacerbates inter-species
differences and makes extrapolation to humans even
more difficult and unreliable. In an effort to encour-
age and promote discussion of this issue, which is
underappreciated, I am expanding on that Editorial.
Stress may be thought of as “the sum of the biolog-
ical reactions to any adverse stimulus, physical,
mental, or emotional, internal or external, that
tends to disturb the homeostasis of an organism” (2).
It is increasingly acknowledged that diverse species
experience pain, stress and distress (see below), and
experience depression and anxiety disorders (see
Ferdowsian et al. [3]). This includes fish — notably
the ubiquitous laboratory zebrafish — which show
emotional fever/stress-induced hyperthermia in
response to a variety of stressors, including simple
handling (4, 5). Stress in fish can result in increased
aggressive behaviour (6), elevated anxiety, dimin-
ished weight-gain, and altered levels of dopamine
and serotonin metabolites in the brain (7), as well as
increased brain levels of extracellular adenosine,
which has neuromodulatory effects (8).
While all species experience stressors and stress
in their natural environments, psychological and
physiological problems can arise with exposure to
recurrent stressors, and/or when stress becomes
chronic and too difficult to cope with. This leads to
allostatic overload (excessive wear and tear on the
body), which may manifest in altered physiological
responses, some of which can be harmful (9) — gen-
erally thought of as a state of distress. How ever, the
US National Research Council (NRC) accepts that
there is “confusion in the scientific, regulatory, and
animal welfare communities” concerning the distinc-
tion between stress and distress; and that these
terms are often used interchangeably in animal wel-
Does the Stress of Laboratory Life and Experimentation on
Animals Adversely Affect Research Data? A Critical Review
Jarrod Bailey
Cruelty Free International, London, UK
Summary — Recurrent acute and/or chronic stress can affect all vertebrate species, and can have serious
consequences. It is increasingly and widely appreciated that laboratory animals experience significant and
repeated stress, which is unavoidable and is caused by many aspects of laboratory life, such as captivity,
transport, noise, handling, restraint and other procedures, as well as the experimental procedures
applied to them. Such stress is difficult to mitigate, and lack of significant desensitisation/habituation can
result in considerable psychological and physiological welfare problems, which are mediated by the
activation of various neuroendocrine networks that have numerous and pervasive effects. Psychological
damage can be reflected in stereotypical behaviours, including repetitive pacing and circling, and even
self-harm. Physical consequences include adverse effects on immune function, inflammatory responses,
metabolism, and disease susceptibility and progression. Further, some of these effects are epigenetic,
and are therefore potentially transgenerational: the biology of animals whose parents/grandparents were
wild-caught and/or have experienced chronic stress in laboratories could be altered, as compared to free-
living individuals. It is argued that these effects must have consequences for the reliability of
experimental data and their extrapolation to humans, and this may not be recognised sufficiently among
those who use animals in experiments.
Key words: animal welfare, cost–benefit analysis, data accuracy, glucocorticoids, psychological, stress,
translational medical research.
Address for correspondence: Jarrod Bailey, Cruelty Free International, 16a Crane Grove, London,
N7 8NN, UK.
ATLA 46, 291–305, 2018 291
fare literature and that relevant available informa-
tion is “far from complete”, with distress remaining
“a complex and still poorly understood phenomenon”
(10). Nevertheless, it is generally understood that
distress manifests when an individual is unable to
cope and adapt successfully to one or more stressors,
resulting in compromised well-being due to the
inability to return to physiological and psychological
homeostasis (11). Notably, an individual can remain
in distress, even when a stressor is removed.
While different species and individuals have dif-
ferent stressors, variable ranges of stress to which
they can adapt, diverse spectra of tolerance, and
dissimilar manifestations and sequelae of exces-
sive stress, they share the same biological path-
ways and mechanisms that are adversely affected
by stress. Briefly, stressors stimulate the hypotha-
lamic–pituitary–adrenal (HPA) axis, the sympa-
thetic adrenal medullary axis, and the sympathetic
and parasympathetic nerve projections that
directly innervate secondary lymphoid organs (12–
14). Among other things, this results in the eleva-
tion of the ‘stress hormones’, cortisol and
corticosterone (CORT). Raised glucocorticoid (GC)
levels, alongside significant elevations of heart
rate, blood pressure and other hormone levels, are
acknowledged indicators of fear, stress and dis-
tress, and are used as biomarkers for stress in ter-
restrial vertebrates in laboratories (e.g. 15–17).
Direct consequences of these neuroendocrine
changes include deleterious effects on innate and
adaptive immunity, central nervous system
pathology, and cardiovascular and reproductive
perturbations (18, 19), leading to extensive and
diverse adverse health effects. Psychological
trauma, for instance, may result in altered health
or damaging patterns of behaviour. In humans,
immune perturbations manifest in poor responses
to vaccines, increased susceptibility to infections,
and accelerated disease progression (see Gurfein et
al. [18]). Further, the various mechanisms under-
lying such effects mean that stress has ramifica-
tions beyond the individuals experiencing it.
Successive generations, and individuals who have
experienced prenatal and/or early-life stress, may
be destined to suffer the consequences in adult-
Caveats and Framing the Argument
Chronic or long-term stress is not unique to ani-
mals in laboratories — it is part and parcel of life
for all species in whatever environment. I do not
intend to suggest otherwise, and it is accepted that
animals in the wild experience acute stressors reg-
ularly (such as lack of food and shelter, predation,
disease, and so on); not only is this entirely normal,
but it can be beneficial in terms of building allo-
static capacity. This Comment, however, argues
that stresses resulting from life in the laboratory
often have negative consequences — given the
opportunity, the animals would avoid many labo-
ratory stressors, and so they can hardly be consid-
ered benign. In addition, the degree, type,
frequency and duration of laboratory stressors are
different to those in the wild, and together could be
more chronic. This may result in greater adverse
consequences for animal welfare and for scientific
data quality. Experiencing repeated, unnatural
stressors, such as blood draws and gavage, cannot
be compared to the brief elevations in CORT
resulting from the natural diurnal/nocturnal
adrenal cycle. For instance, the former stressors
are very different from simply waking up, or being
hungry. In well-designed experiments, increases in
stress biomarkers are relative to baseline levels in
any case (see, for example, 16, 20).
That laboratory stress can — arguably, frequently
and unavoidably — have effects that compromise
animal welfare and experimental results is accepted
by the US NRC’s Committee on Recognition and
Alleviation of Distress in Laboratory Animals (10):
“In the longer term… a breakdown in an animal’s
ability to cope with its environment is likely to lead
to adverse emotional states and poor welfare. Some
of these cases may be quite minor and not give rise
to significant ethical concerns; but prolonged or
intense circumstances would compromise the ani-
mal’s welfare enough to warrant concern and also
significantly affect the research results.” Further:
“Strong evidence in rodents has shown that mild
stress of 2–3 months duration — a regimen that pro-
duces no signs of overt distress — alters the animals’
performance in tests of anxiety, depression, and
memory… Other findings indicate that rats’ habitu-
ation to a test environment can dramatically affect
their response to a toxic substance”. Stress can also
be beneficial: “Over a longer time frame, glucocorti-
coid production in response to infection helps restrict
the immune system, thus preventing deleterious
effects of inflammatory factors on tissues” (10).
While this latter statement is true, it also supports
the argument made here, that stress — even when
not ‘bad’ — adversely affects biological systems, and
consequently confounds research data.
Therefore, it is not only distress that leads to bio-
logical consequences affecting data reliability,
quality and relevance. It seems clear that the con-
sequences of stress are also an issue for welfare
and data quality. To quote again the US NRC:
“The impact of distress on both animal welfare and
research results is likely even more pronounced
than that of stress. Animals exposed to prolonged
severe stress experience underlying changes in
physiological functions (e.g. gastric lesions or
immunosuppression) that can interfere with exper-
imental manipulations; alter experimental vari-
ables such as behaviour, drug dosing and
clearance; change the progress of a disease; and
292 Comment
contribute to morbidity and mortality. A variety of
stressors can contribute to unintended distress,
from postoperative pain or infection to barren
housing conditions or the solitary confinement of
an individual of a social species. Stereotypies,
abnormal repetitive behaviours indicative of poor
well-being that are often observed in distressed
animals, are thought to reflect defective brain
function and to be a result of poor animal welfare.
Stereotypies are thus likely to interfere with
behavioural, neuroscience, and pharmacological
studies” (10); and “The impact of stress and dis-
tress on the quality of scientific research can result
in the generation of compromised data, which in
turn necessitates the use of more animals… Both
stress and distress represent potential complica-
tions in a wide range of experiments, and should be
proactively addressed by good experimental
design” (10). While I (and others) disagree that the
use of more animals and the careful design of
experiments can have an acceptably positive
impact on welfare, and on the human-relevance of
the data generated, the main points in this
Comment stand. Efforts to address this issue over
many years are acknowledged, but I argue that,
due to fundamental biology and inter-species dif-
ferences, the improvement of husbandry, veteri-
nary care, regulation, oversight, training, etc. can
never be sufficient to significantly address and
overcome these issues.
Stress Resulting from Laboratory Life
and Research, and its Biological
The issue for animals in laboratories is that life
can be inherently and excessively stressful — per-
haps much more than in their natural environ-
ments, from which laboratory conditions differ
greatly, even when enriched and accounting for
efforts to mitigate stressors. Laboratory conditions
preclude or limit many natural behaviours, hous-
ing tends to be much smaller than the animals’
natural ranges, and the animals are subjected to
frequent manipulations and handling, as well as
other alien factors that they try to resist and avoid
(16, 20). Stressful procedures and environmental
factors are numerous and varied. Briefly, they
include, but are not limited to: general handling
and manipulations, such as weighing and saline
injections (15, 16, 21–23); anaesthesia (15, 24–31);
restraint (21, 32–36); gavage (37–42); blood sam-
pling (15–17, 43–50); food and water restriction
(51, 52); non-natural environment (53–55) and
associated factors, such as noise and light (56–62);
social crowding and/or isolation (63–71); cage con-
ditions/cleaning/changing (18, 58, 72–78); trans-
port (19, 58, 79–83); observing procedures on, and
killing of, other animals (31, 84–87); and even
enrichment itself (17, 88, 89). It has been sug-
gested that captive-bred animals may not know
that these stressors are different to those in the
wild, though there might be some perception that
they are not similar to ‘natural’ stressors such as
limited food, inadequate shelter, predation and so
on. In any case, the point is that they are different,
but more importantly, they are also frequent, reg-
ular and inescapable.
Naturally, this has animal welfare and scientific
implications, acknowledged at least in some quar-
ters. For instance, stress from handling is accepted
as a source of “unexplained variation within and
between animal studies”, as it influences “both the
behaviour and physiology of animals” (23; see also
Balcombe et al. [16] and Meijer et al. [17]), and rel-
atively poor caging conditions “may contribute to
problems in translating murine research into
human studies” (18, 77, 78). However, it is acc -
epted by some that these factors are probably
widely underappreciated (90). It should be noted
that some consider enriched environments simply
as ‘less bad’ rather than ‘better’ than those that are
non-enriched. To illustrate, significant numbers of
animals experiencing enrichment still go on to
develop stereotypies (e.g. 91–98).
The Nature of Stress: Biological
Basis/Mechanisms of Stress and its
Adverse Effects
The physiological consequences of stress are varied
and powerful. This, in itself, is of concern for the
translation of animal data to humans, as they com-
pound and confound existing difficulties in transla-
tion due to species differences. First, however, a
consideration of the underlying mechanisms is
important, to demonstrate their fundamental nature
and potency.
Primary mediators of stress, such as GCs and cat-
echolamines, are released in response to stressors,
with various biological consequences. Such biological
effects generally go beyond species boundaries/limits
for mammalian species, though the mechanisms and
specific effects differ to varying degrees. These pri-
mary mediators are extremely powerful, because
they ultimately modulate the expression of many
genes. Secondary outcomes have been documented
“…in every physiological system, including the car-
diovascular system, metabolism, the central nervous
system, and the immune system”, and are con-
founded by the characteristics of the stressor(s), as
well as the attributes of the affected individual, such
as age, health, status, genetic background, past
experience, etc. (99).
The potency and ubiquity of the stress response
has been demonstrated in a number of in vitro stud-
ies, revealing that it generally blocks every impor-
tant cellular process, including DNA replication,
Comment 293
transcription, pre-mRNA processing, mRNA export,
and translation, until the cells recover (100).
Therefore, stress exerts its effects via varied molec-
ular mechanisms, with far-reaching consequences.
The principal ones are highlighted below.
Epigenetic mechanisms (histone acetylation and
DNA methylation) (101–107): Psychological
stress alters gene expression via histone acety-
lation and DNA methylation. Much occurs in
response to environmental triggers, e.g. diet,
drugs, toxins, and psychological stress, e.g. fear
conditioning and maternal care (103). Genes
involved in HPA axis function are especially
susceptible (108), e.g. in suicide victims with a
history of child abuse (109), and post-traumatic
stress disorder (PTSD) is strongly associated
with the epigenetic modification of genes
involved in immune function and inflammation
Alternative splicing/expression of regulatory
microRNAs: Alternative splicing of gene tran-
scripts and microRNAs (miRNAs), is a powerful
means of altering gene expression that can be
significantly affected by stress (111, 112). For
example, acute stress in humans altered the
splicing of 27 genes in peripheral leukocytes
Oxidative damage and ageing: Mental stress con-
tributes to oxidative stress in the body, and
therefore to oxidative damage (113). This effect
has been identified in students undergoing aca-
demic examinations (114), and in the lympho-
cytes of psychologically stressed individuals
(115). Oxidative stress is also associated with
PTSD and depression (116), contributes to the
ageing process (117), and is associated with neu-
rodegenerative disease, ophthalmologic disease,
cancer and cardiovascular disease (including
atherosclerosis, hypertension, cardiomyopathy,
chronic heart failure, myocardial ischaemia and
ventricular arrhythmias; 117, 118). There may be
confounding data on the effects of oxidative stress
from different species/strains of mice, which fur-
ther challenges the translation of data across
species (119).
Direct Physiological Consequences of
The physiological consequences of stress are
numerous and varied. Table 1 shows how promis-
cuous these consequences, effects and manifesta-
tions are in many species, including humans.
Given that these effects involve so many biological
pathways and systems, the effects on experimental
data must be significant.
Habituation/Desensitisation to Stress
Some argue that animals become habituated (120)
and/or desensitised (121) to stress, so implications
for welfare and experimental results may be over-
come (23, 122, 123). For instance, non-human
primates can be trained to approach test environ-
ments, and to present their arms for blood with-
drawal and so on, seemingly voluntarily. However,
it has been shown that repeat exposure to homo-
typic stressors of greater intensity and/or severity
does not result in habituation, and could actually
result in sensitisation of the CORT response (122).
Furthermore, CORT levels might only decrease for
certain types of stressor, persisting for other types
(35); where desensitisation has been shown, it is
only to a modest degree, in a small proportion of
the animals in the studies, and in small sample
sizes (70). While some studies have suggested that
enrichment might decrease stress, others have
shown a paradoxical increase in stress via CORT
levels (18). Similarly, some studies have suggested
that transferring scent-marked materials from old
to new cages reduces stress-related aggression,
while others found that it increases aggression
(74). Furthermore, mice do not habituate to stress
associated with simple handling, and indeed seem
to become sensitised to it (124–126). In instances
where CORT levels decrease, there is increasing
evidence that this does not necessarily mean an
attendant decrease in stress; other indicators, such
as the neutrophil–lymphocyte ratio, might be ‘bet-
ter’ indicators of chronic stress, which can occur in
the absence of increased serum CORT (127).
Adverse Physiological Sequelae of
Psychological Stress are Initiated
Prenatally or in Early Life, and are
Captive-born animals can be exposed to stress pre-
natally via their wild-trapped mothers (128–131),
and as infants in laboratory environments, often
experiencing inadequate maternal contact and
care (132–134; cited by Camus et al. [135], and
Detter et al. [136]). If an animal’s parents or grand-
parents lived in laboratories, and/or were born of
parents that lived in laboratories and/or endured
being wild-caught, then their ancestors experi-
enced highly stressful lives and would have been
affected by the adverse consequences described
herein. Even if such an individual was subse-
quently afforded as stress-free a life as possible
(difficult, if not impossible, in a laboratory), the
consequences of their early lives, and the lives of
their ancestors, would lead to the same adverse
effects as if they had continued to experience
excessive stress as adults.
294 Comment
That early-life experiences affect adult psy-
chopathology is widely accepted. As Jean-Paul
Sartre put it, “Childhood decides” (see Murgatroyd
[102]). “A large body of data shows that stress dur-
ing pregnancy causes an increase of GCs in the
blood of the dams and the foetus, leading to alter-
ations of the structure and function of the develop-
ing brain… These alterations result in the
disturbance of the function of the neuroendocrine
system and different kinds of behaviour through-
out life” (137). Early-life/prenatal exposure to
stress leads to altered adrenocorticotrophic hor-
mone responsiveness, dysfunction of the HPA axis
(138, 139), and altered autonomic modulation of
immune function that may begin in utero (103).
Social isolation in several species leads to neuroen-
docrine changes, increased cortisol, and ensuing
behavioural problems (for references, see Cham -
pagne [101]). Maternal inflammation during preg-
nancy (from infection, or possibly stress) may
increase the risk of neurodevelopmental disorders
such as schizophrenia and cerebral palsy (140).
Physiological sequelae include cardiovascular dis-
ease and metabolic disorders such as diabetes
(141), compromised immune function, including
poor lymphocyte proliferation upon infection and
reduced placental transfer of antibodies during
pregnancy (140), autoimmune disorders, chronic
obstructive lung disease, asthma and obesity (see
Chang [142]). Pivotal to these adverse outcomes
are the aforementioned stress-related epigenetic
processes and oxidative damage. Methylation of
gene regulatory regions is partly involved (143),
the extent of which may be set during prenatal
development (141). Cord blood samples of infants
of mothers with late-pregnancy depression show
altered methylation of the GC receptor promoter,
which also predicts elevated salivary cortisol in
early life (144). Other modifications are inherited
and transgenerational in nature (145); for exam-
ple, poor prenatal nutrition affects GC receptor
methylation, affecting the growth and metabolism
of first and second-generation offspring (146), and
matrilineal transmission of the effects of diethyl-
stilbestrol (DES) occurs via hypomethylation, lead-
ing to increased cancer risk over two generations
The inherent, multi-faceted stress of laboratory life
— often excessive, relative to the more transient,
acute and ‘natural’ stresses experienced in the wild
— is evidenced by its often negative impact on the
well-being of the animals involved, both psycholog-
ically and physiologically. Because this harm is
mediated via established trans-species biological
mechanisms involving the HPA axis and the sym-
pathetic nervous system, and effected via oxidative
stress and epigenetic mechanisms, which affect
multiple biological pathways and systems, it can
have adverse and confounding effects on experi-
mental results. This modulation of many biochem-
ical pathways and gene expression can result in
downstream effects such as organ damage, cardio-
vascular diseases, attenuated immune function
and autoimmune disorders, premature ageing and
mortality, developmental abnormalities, elevated
tumour initiation and progression, and muscu-
loskeletal atrophy (16).
The absolute degree of translation of animal stud-
ies to humans is debatable, but undoubtedly,
“Studies using animal models are more translatable
to human disease when the animals’ welfare is
maximised” (127). Arguably, it is difficult to “max-
imise” welfare significantly, given the in herent,
widespread, substantial and largely intractable
nature of the stresses involved in animal research
and laboratory life. Notably, habituation and/or
desensitisation to many of the stressors is often not
possible, or at least not significant, and the impact
of these effects on experimental data — and their
extrapolation to humans — is likely to be signifi-
cant. The many sources of stress have been sum-
marised here, along with their effects on multiple
biological and physiological systems. The literature
warns that: “…animals subjected to the environ-
mental changes that occur during transportation…
react with changes in their physiology, such as body
weight, plasma hormonal levels, heart rate and
blood pressure changes… When measurements of
physiological parameters are performed using con-
ventional measurement techniques, the results
must be interpreted with caution as these conven-
tional techniques also have effects on the animals”
(148). Most importantly, “Suffering in animals can
result in physiological changes which may increase
the variability of experimental data” (149). Many
scientists are well aware of these effects and consid-
erations, and have cautioned against disregarding
them (150–152). Yet, while accepting the negative
effects of pain, stress and distress, and their influ-
ence on study outcome, such effects are often not
reported or are under-reported in scientific publica-
tions (90).
The impact of stress on immunological and
inflammatory responses seems particularly preva-
lent, and might be especially critical, seeing as
much animal experimentation involves infectious
agents and/or immune function (for a discussion
and references, see Bailey [153] and Bailey [154]).
Crucially, this impact exacerbates and compounds
existing immune differences between humans and
non-humans due to genetic differences. To illus-
trate, genomic duplications — one of the most sig-
nificant causes of genetic variation among
primates (155) and at the root of many aspects of
intra-species and inter-species diversity — differ-
entially affect many genes involved in immune and
Comment 295
Table 1: A summary of the physiological consequences and manifestations of stress, in many species, including humans
Consequences of general stress Notes
Stereotypies (abnormal, repetitive, invariant These correlate with basal plasma cortisol and corticotrophin-releasing hormone (CRH) in cerebrospinal fluid (166).
behaviours with no obvious function; 160, 161)
and self-harm (e.g. 162–165) in non-humans
Wide-ranging physiological symptoms in humans Anger, depression, anxiety, behavioural changes, food cravings, lack of appetite, frequent crying, difficulty sleeping, tiredness, lack of
concentration, chest pains, constipation, diarrhoea, cramps and muscle spasms, dizziness, fainting, nervous twitches, restlessness, sexual
dysfunctions, breathlessness, and a host of diseases and illnesses with probable associated psychogenic (as well as biological) causes (167).
Long-lasting neurophysiological changes These could “…have direct implications for electrophysiological, behavioural, and molecular studies” (see 69).
Isolated rats show “…structural and functional changes in the mesocorticolimbic dopaminergic system, exhibited hyperlocomotor
activity and impaired sensorimotor gating” (168).
Isolated pigs show “…sustained changes in behavioural, neuroendocrine and immune regulation” (169).
Socially isolated humans show increased risk of death, “…genome-wide transcriptional activity of impaired GC response genes and
increased activity of pro-inflammatory transcription control pathways”, and higher risk of developing “…conduct disorders,
personality disorders, major depression, PTSD, schizophrenia, and anxiety disorders” (see 170).
Multi-faceted modulation of the immune system This occurs in many, if not all, mammalian species (140, 171, 172). It is especially problematic for research involving immune function,
infectious diseases, etc. The expression of several hundred genes (many related to immune function) may be perturbed by simple
handling of animals (173; and see 33), which may be affected by the acute or chronic nature of the stressor (see 173). Chronic stress can:
shift neutrophil–lymphocyte ratios (127, 174);
affect expression of cytokines and cytokine receptors (175, 176), such as IL-2 and IL-6 (18);
alter gene splicing in peripheral leukocytes (10);
alter global and immune gene specific methylation (177); and
differentially affect brain activity and neurotransmitter release, macrophage activity and antibody production (33).
The same stressor in acute or chronic forms may have different effects:
In rodents, acute restraint increases delayed-type hypersensitivity (DTH) and leukocyte redeployment, but can increase or decrease
them when chronic (33, 178).
In humans, chronic, though not acute, stress increases susceptibility to colds (179).
These inconsistencies show that stress is mediated not only by GCs (180), and that accounting for the effects of stress in animal
research must be difficult, if not impossible.
Observed alterations in human immune function These include:
“attenuated responses to vaccination, poorer wound healing, exaggerated release of inflammatory mediators, & premature aging of
the immune system” (181);
increased plasma and CNS cytokine levels, impaired natural killer cell activity, lower T-lymphocyte counts (in PTSD/complex
PTSD patients; 182);
epigenetic changes exerting lifelong impact on immune and inflammatory function (in PTSD/complex PTSD patients; 182); and
greater inflammatory responses to vaccinations (in depressed humans; 183).
Neuropeptides involved in stress responses may accentuate pathophysiological sequelae in critically ill individuals (184). In healthy
humans, 49 different genetic pathways are affected by stress, including genes associated with immune function (185). Stressed students
undergoing examinations have significantly increased pro-inflammatory cytokines (186).
Activation of the HPA axis This accelerates ageing generally, with adverse effects on brain/central nervous system, immune system, skeletal muscle and bone tissue
(see 187–189). HPA dysregulation may lead to excessive inflammation via increases in the levels of circulatory inflammatory cytokines,
decreases in anti-inflammatory cytokines, and alterations in the expression of genes involved in immune activation of peripheral blood
cells (see 177).
296 Comment
Table 1: continued
Effects of specific stressors Notes
Disease susceptibility An increase in susceptibility has been noted across several species to:
general disease and somatic disorders (see 100, 103);
various cancers (190, 191);
pancreatitis and pancreatic tumours (192, 193);
gastrointestinal disorders (194);
thyroid pathology (195);
multiple sclerosis (171);
inflammatory bowel disease (142, 196);
cardiovascular disease (197–199);
accelerated ageing and age-related disorders (187); and
musculoskeletal injury (200).
Stressors can increase oxidative stress, triggering inflammatory pathways associated with type-2 diabetes, cardiovascular disease,
osteoporosis, arthritis, some cancers, and susceptibility to some infections (e.g. 201–205).
Handling Causes biological changes, affecting wellbeing and/or experimental results.
Enrichment intended to mitigate stress can substantially alter brain structure, function and physiology.
Light, noise, cage position/changing etc. all affect physiology, behaviour, anxiety and experimental data (206).
Effects of handling stress may often be “missed” by researchers (23).
Early-life stress in various species Results in:
abnormal brain development, leading to early-life psychopathologies and adult chronic mental illnesses (e.g. 207, 208);
epigenetic modifications associated with depression and suicidal behaviour in later life (e.g. 102, 209–211); and
elevated morbidity and mortality from chronic diseases of ageing, including vascular disease, autoimmune disorders, and premature
mortality (e.g. 141, 212).
Stress from maternal deprivation has negative effects on growth rates in rats, and adversely affects circadian clock and stress
responses (72).
Traumatic stress in humans Studies on PTSD patients have shown that:
acute stress affects glucose metabolism, inflammation and components of the immune system associated with type-2 diabetes (201);
serious long-term consequences include hypertension, heart attacks and stroke, as well as increased risk of obesity, Alzheimer’s
disease, and AIDS dementia complex (213).
Comment 297
inflammatory responses (156). Indels (genomic
insertions and deletions) also affect major histo-
compatibility complex (MHC) genes, which are
critical to immune responses, and are associated
with differences in response to infections, as well
as susceptibility to autoimmune diseases. These
are further confounded by sex-related and strain-
related differences. It has also been shown that the
susceptibility and responsiveness of mice to stres-
sors varies with the strain (157, 158). Stress also
affects sleep, and conversely sleep perturbations
exacerbate and sensitise individuals to stress, with
all the attendant consequences. These conse-
quences are also strain-dependent, and intimately
linked to the CRH/HPA system (159).
Additionally, the adverse consequences of stress
are multigenerational, as the associated epigenetic
mechanisms affect the germline. This is likely to
have significant consequences for animals, and
their offspring, in laboratories: if an animal’s par-
ents or grandparents experienced a stressful labo-
ratory life and experimental procedures, and/or if
the offspring experienced significant stress in early
life, then this will compound any further stress
that they experience as adults, in turn compound-
ing species differences and the translation of data
to humans.
Overall, these observations of detrimental phys-
iological effects and the general mechanisms
behind them have been detailed in many species
(including humans), and throughout the evolution-
ary scale from monkeys to rodents. The minutiae of
the genes and biochemical pathways responsible,
and their manifestations, may differ to some
degree, but there are common mechanisms and
adverse effects in all species examined to date. It
must be concluded that laboratory life for animals
used in experiments has serious and intractable
consequences for their welfare, and for the quality
and human relevance of the experimental data
obtained (which, in any case, are already of debat-
able applicability to humans, due to species differ-
Finally, I believe that this issue should be taken
much more seriously by legislators, regulators,
funders, practitioners and advocates of animal
experiments, and urge all involved to do so. The
information presented here could, and should, be a
valuable resource for project licence applicants,
ethics committees and the Home Office Inspect -
orate, for use in experimental design and harm–
benefit analyses, and to aid data interpretation.
Though it argues that relatively little can be done
to minimise many, if not all, stressors and stress,
it could inform attempts to do so — as well as con-
trolling variable factors and mitigating negative
consequences, etc. The information could also be
factored into existing guidance for the strategic
planning of animal experiments, since stress
impacts all areas of this planning, including study
objectives, species/strain selection, experimental
procedures, analgesia, training of staff, and so on.
This review was funded by Cruelty Free
International Trust, London, UK. It was based on
previous (not published) work conducted by the
same author in 2011, funded by the New England
Anti-Vivisection Society (NEAVS), Boston, USA, in
his role as its Science Director.
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Comment 305
... However, in natural settings, there are many external uncontrollable factors that may influence the observed outcomes. For these reasons, plethodontids are often tested in laboratories to facilitate the avoidance of external confounding factors, while simultaneously controlling biological variables such as age, sex or reproductive status [17][18][19]. ...
... Researchers mostly explored the trophic spectrum of Speleomantes with the analysis of stomach contents, identifying diet composition in multiple conspecific populations and assessing how seasonality contributes to defining the type and amount of consumed prey [42,43]. In some instances, assessments of individuals' food specialization were also performed [19,44]. On the other hand, many other aspects of Speleomantes foraging behavior are still unexplored or just hypothesized [45]. ...
... In fact, the experimental hypothesis-testing approach seems very informative in the study of behavioral ecology, since it allows researchers to infer causality [3,17,18]. Although, caveats should be always considered when extrapolating results observed in simplified, and possibly stressing, settings compared to natural habitats [19]. This type of approach was performed mostly on the continental species S. strinatii, with only one exception concerning S. italicus (see Table S1). ...
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There is a recent growing interest in the study of evolutionary and behavioral ecology of amphibians. Among salamanders, Plethodontidae is the most speciose family, with more than 500 species, while in Europe, there are only 8 species, all belonging to the genus Speleomantes. European plethodontids recently received increasing attention with regard to the study of their natural history, ecology and behavior; however, the lack of standardized data, especially for the latter, hampers comparative analysis with the species from the New World. We here synthetized the recent advances in Speleomantes behavioral ecology, considering as a starting point the comprehensive monography of Lanza and colleagues published in 2006. We identified the behavioral categories that were investigated the most, but we also highlighted knowledge gaps and provided directions for future studies. By reviewing the scientific literature published within the period 2006–2022, we observed a significant increase in the number of published articles on Speleomantes behavior, overall obtaining 36 articles. Behavioral studies on Speleomantes focused mainly on trophic behavior (42%), and on intraspecific behavior (33%), while studies on pheromonal communication and interspecific behavioral interactions were lacking. In addition, most of the studies were observational (83%), while the experimental method was rarely used. After providing a synthesis of the current knowledge, we suggest some relevant topics that need to be considered in future research on the behavioral ecology of European plethodontids, highlighting the importance of a more integrative approach in which both field observations and planned experiments are used.
... Despite the unquestioned importance of laboratory animals to scientific progress over 200 years, suffering has become institutionalized. Not only is suffering bad for welfare, but stress within the laboratory causes data-distortion and reduces the justification of such studies (Bailey, 2018). Fortunately, we are now far enough along in advancements and technology, that we can raise the standard of justification for a few historic assays that have limited usefulness. ...
... Furthermore, we understand that non-welfare-friendly designs may create uninterpretable data. This occurs when data is compromised after being collected from stressed animals (e.g., data distortion; Bailey, 2018). Yet the FEP, as we describe below, could improve research outcomes to address each of these crises. ...
... This would satisfy animal welfare concerns and at the same time, address issues about translatability of findings (Drucker, 2016;Oppenheim, 2019). Laboratory FEP may be designed to improve outcomes and welfare in three ways: (1) by increasing heterozygosity when wild animals are brought into controlled settings and allowed to freely enter the designed apparatus; (2) when naturalistic contexts such as availability of conspecifics and shelter are incorporated into laboratory FEP settings; and (3) by minimizing animal handling, we decrease animal stress which is known to cause data distortion (Bailey, 2018). In short, improved welfare also increases data quality. ...
... Even though the need for post-operative analgesia should be considered at least the same as for humans undergoing surgery, the direct translation of medicines and dosages from other species to pigs could result in failure to fulfil analgesia, and thus a breach of the refinement principle. Pain will negatively impact animal welfare and the physiological and psychological effects of pain can bias experimental data by adversely affecting multiple biological systems [12]. This highlights the importance of appropriate use and scientific reporting of analgesic agents [7]. ...
... Respiratory frequency increased in response to fentanyl ( Figure 10). Vocalization remained stable within the groups and the median (range) for the vocal- (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16) at each timepoint (baseline, injection 1-3). Repetitive behaviors were rooting behavior, circling, backward locomotion, frequency of being in contact with the water nipple and jumping. ...
... The marker assigned to each pig is shown on the right panel(Single column fitted figure). Animals 2023, 13, x FOR PEER REVIEW 13 of Repetitive behaviors displayed by each pig(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16) at each timepoint (baseline, injection 1-3). Repetitive behaviors were rooting behavior, circling, backward locomotion, frequency of being in contact with the water nipple and jumping. ...
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Analgesic effects of fentanyl have been investigated using behavior. The behavioral effects of fentanyl and possible serotonergic influence are largely unknown. We therefore investigated behavioral effects of fentanyl, with or without the serotonin antagonist ketanserin, in pigs. Fourteen mixed-breed pigs, weighing 17–25 kg were included in a randomised blinded prospective, balanced three-group study. Ten pigs received first 5 and then 10 µg/kg of fentanyl intravenously. Ketanserin at 1 mg/kg or saline was given intravenously as a third injection. Four control pigs received three injections of saline. Behavior was video-recorded. The distance moved was automatically measured by commercially available software, and behaviors manually scored in retrospect. Fentanyl inhibited resting and playing, and induced different repetitive behaviors. The mean (SD) distance moved in the control group and fentanyl group was 21.3 (13.0) and 57.8 (20.8) metres respectively (p < 0.05 for pairwise comparison). A stiff gait pattern was seen after fentanyl injection for median (range) 4.2 (2.8–5.1) minutes per 10 min, which was reduced to 0 (0–4) s after ketanserin administration. Conclusion: fentanyl-induced motor and behavioral effects, and serotonergic transmission may be involved in some of them. The psychomotor side effects of fentanyl could potentially interfere with post-operative pain evaluation in pigs.
... Animal models are used to study infectious diseases in both human and veterinary medicine, but the results of these studies are vulnerable to a series of variables such as handling, cage environment, and technical procedures, which can generate varying degrees of stress (3). The physiological and immunological consequences of stress, in addition to other factors such as the induction of general anesthesia, have the potential to alter scientific outcomes resulting in less applicable science (4). Additional consequences of these uncontrolled variables in infectious disease research are poorer animal welfare outcomes (5). ...
... Stress is a complex and multi-faceted process (chronic vs. acute, beneficial vs. adverse effects) and consequently, there are inherent difficulties in identifying what causes stress in different species under diverse study conditions (9). Stress experienced by animals in disease research can be caused by the disease itself and accompanying inflammatory responses (10,11), as well as regular animal handling and repeated procedures and interventions (3)(4)(5)12). The factors that cause stress also promote the organism's response as a homeostasis-related compensatory mechanism or returning to homeostasis, by modifying the physiological parameters and generating compensatory metabolic, hormonal or neurological responses that can alter study results (13)(14)(15)(16). ...
... This definition is in line with the founding principles of humane animal research developed by Russel and Burch (27), which defines distress in laboratory animals as a central nervous state of a certain rank on a scale, in the direction of the mass autonomic response which if protracted, would lead to the physiologic stress syndrome (27). Animals maintained in laboratory conditions are often far removed from their evolved or natural environment, and this can predispose these animals to experiencing greater levels of stress (4). In addition, keeping animals in controlled environments away from stress factors may predispose animals to experience a greater degree of stress, resulting in neurobiological, hormonal, and metabolic compensatory responses that result in the development of chronic stress (16,28). ...
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Stress and general anesthesia have an impact on the functional response of the organism due to the detrimental effects on cardiovascular, immunological, and metabolic function, which could limit the organism's response to an infectious event. Animal studies have formed an essential step in understanding and mitigating infectious diseases, as the complexities of physiology and immunity cannot yet be replicated in vivo. Using animals in research continues to come under increasing societal scrutiny, and it is therefore crucial that the welfare of animals used in disease research is optimized to meet both societal expectations and improve scientific outcomes. Everyday management and procedures in animal studies are known to cause stress, which can not only cause poorer welfare outcomes, but also introduces variables in disease studies. Whilst general anesthesia is necessary at times to reduce stress and enhance animal welfare in disease research, evidence of physiological and immunological disruption caused by general anesthesia is increasing. To better understand and quantify the effects of stress and anesthesia on disease study and welfare outcomes, utilizing the most appropriate animal monitoring strategies is imperative. This article aims to analyze recent scientific evidence about the impact of stress and anesthesia as uncontrolled variables, as well as reviewing monitoring strategies and technologies in animal models during infectious diseases.
... Animal handling and housing in the laboratory inevitably impacts the data acquired through experimentation, which could result in stress-induced differences in behaviour and other measurable traits (Bailey, 2018;Ferdowsian & Beck, 2011;Sensini et al., 2020). If researchers aim to study naturally occurring behaviours (whether impacted by parasites or not) in the laboratory, reducing artificial stress must be of prime importance. ...
... Identifying endoparasitism in individuals from natural populations represents one of the biggest methodological challenges for live animal research, which is important since any given trait is strongly associated with overall health and can thus be impacted by inter- (Bailey, 2018;Ferdowsian & Beck, 2011). In this study, we were able to amplify parasite DNA using a minimally invasive method that required optimized parasitespecific qPCRs and extracellular eDNA extracted from the frass of individual hosts, allowing us to identify infected individuals from two populations of evolutionary distinct host-parasite systems. ...
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Almost every animal trait is strongly associated with parasitic infection or the potential exposure to parasites. Despite this importance, one of the greatest challenges that researchers still face is to accurately determine the status and severity of the endoparasitic infection without killing and dissecting the host. Thus, the precise detection of infection with minimal handling of the individual will improve experimental designs in live animal research. Here, we quantified extracellular DNA from two species of endoparasitic worm that grow within the host body cavity, hairworms (phylum Nematomorpha) and mermithids (phylum Nematoda), from the frass of their insect host, a cave wētā (Orthoptera: Rhaphidophoridae) and an earwig (Dermaptera: Forficulidae), respectively. Frass collection was done at two successive time periods, to test if parasitic growth correlated with relative DNA quantity in the frass. We developed and optimised two highly specific TaqMan assays, one for each parasite‐specific DNA amplification. We were able to detect infection prevalence with 100% accuracy in individuals identified as infected through post‐study dissections. An additional infection in earwigs was detected with the TaqMan assay alone, likely because some worms were either too small or degraded to observe during dissection. No difference in DNA quantity was detected between sampling periods, although future protocols could be refined to support such a trend. This study demonstrates that a non‐invasive and minimally stressful method can be used to detect endoparasitic infection with greater accuracy than dissection alone, helping improve protocols for live animal studies.
... Various studies have shown that routine procedures, for instance separation, blood sampling or restraining techniques can cause stress in different species of animals (Balcombe et al. 2004;Yardimci et al. 2013;do Vale et al. 2020). Stress, in turn, affects the welfare of the animals involved and may also affect the reliability of the results in those studies (Poole 1997;Bailey 2018). ...
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Experimental procedures involving farm animals are often associated with stress due to restraining. Stress can be reduced through use of positive reinforcement training, which then serves as refinement according to the 3Rs principles. Trainer skills, however, may influence the feasibility and success of animal training. The potential influence of trainer skills as well as the education of animal trainers are rarely described in literature but are necessary information for the implementation of positive reinforcement training as a refinement measure. To investigate the effect of educational programmes on animal trainers, we compared the training success of two groups of participants in training goats to elicit a behaviour that would allow simulated venipuncture. One group was educated in a two-day workshop while the other was provided with specific literature for self-instructed learning. Training success was evaluated using an assessment protocol developed for this study. A greater training success in the WORKSHOP GROUP, reflected by objective and subjective measures, was clearly supported statistically. In addition, 73 versus only 13% of the participants of the WORKSHOP GROUP and the self-instructed BOOK GROUP, respectively, stated that they could completely implement the knowledge gained in the course of this study. Our results indicate that more intensively educated trainers can train animals more successfully. In conclusion, if animal training is implemented as refinement, animal caretakers should receive instruction for positive reinforcement training.
Laboratory-based research dominates the fields of comparative physiology and biomechanics. The power of lab work has long been recognized by experimental biologists. For example, in 1932, Georgy Gause published an influential paper in Journal of Experimental Biology describing a series of clever lab experiments that provided the first empirical test of competitive exclusion theory, laying the foundation for a field that remains active today. At the time, Gause wrestled with the dilemma of conducting experiments in the lab or the field, ultimately deciding that progress could be best achieved by taking advantage of the high level of control offered by lab experiments. However, physiological experiments often yield different, and even contradictory, results when conducted in lab versus field settings. This is especially concerning in the Anthropocene, as standard laboratory techniques are increasingly relied upon to predict how wild animals will respond to environmental disturbances to inform decisions in conservation and management. In this Commentary, we discuss several hypothesized mechanisms that could explain disparities between experimental biology in the lab and in the field. We propose strategies for understanding why these differences occur and how we can use these results to improve our understanding of the physiology of wild animals. Nearly a century beyond Gause's work, we still know remarkably little about what makes captive animals different from wild ones. Discovering these mechanisms should be an important goal for experimental biologists in the future.
The gut microbiome is important for host fitness and is influenced by many factors including the host's environment. Captive environments could potentially influence the richness and composition of the microbiome and understanding these effects could be useful information for the care and study of millions of animals in captivity. While previous studies have found that the microbiome often changes due to captivity, they have not examined how quickly these changes can occur. We predicted that the richness of the gut microbiome of wild-caught birds would decrease with brief exposure to captivity and that their microbiome communities would become more homogeneous. To test these predictions, we captured wild house sparrows (Passer domesticus) and collected fecal samples to measure their gut microbiomes immediately after capture ('wild sample') and again 5-10 days after capture ('captive sample'). There were significant differences in beta diversity between the wild and captive samples, and captive microbiome communities were more homogenous but only when using non-phylogenetic measures. Alpha diversity of the birds' microbiomes also decreased in captivity. The functional profiles of the microbiome changed, possibly reflecting differences in stress or the birds' diets before and during captivity. Overall, we found significant changes in the richness and composition of the microbiome after only a short exposure to captivity. These findings highlight the necessity of considering microbiome changes in captive animals for research and conservation purposes.
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Drug administration in preclinical rodent models is essential for research and development of novel therapies. Compassionate administration methods have been developed, but these are mostly incompatible with water-insoluble drugs such as tamoxifen or do not allow for precise timing or dosing of the drugs. For more than two decades, tamoxifen has been administered by oral gavage or injection to CreERT2/loxP gene-modified mouse models to spatiotemporally control gene expression, with the numbers of such models steadily increasing in recent years. Animal-friendly procedures for accurately administering tamoxifen or other water-insoluble drugs would therefore have an important impact on animal welfare. Based on a previously published micropipette feeding protocol, we developed palatable formulations to encourage voluntary consumption of tamoxifen. We evaluated the acceptance of the new formulations by mice during training and treatment and assessed the efficacy of tamoxifen-mediated induction of CreERT2/loxP dependent reporter genes. Both sweetened milk and syrup-based formulations encouraged mice to consume tamoxifen voluntarily, but only sweetened milk formulations were statistically non-inferior to oral gavage in inducing CreERT2-mediated gene expression. Serum concentrations of tamoxifen metabolites, quantified using an in-house developed cell assay, confirmed the lower efficacies of syrup- as compared to sweetened milk-based formulations. We found dosing with a micropipette to be more accurate, with the added advantage that the method requires little training for the experimenter. The new palatable solutions encourage voluntary consumption of tamoxifen without loss of efficacy compared to oral gavage and thus represent a refined administration method.
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Recurrent acute and/or chronic stress can affect all vertebrate species, and can have serious consequences. It is increasingly and widely appreciated that laboratory animals experience significant and repeated stress, which is unavoidable and is caused by many aspects of laboratory life, such as captivity, transport, noise, handling, restraint and other procedures, as well as the experimental procedures applied to them. Such stress is difficult to mitigate, and lack of significant desensitisation/habituation can result in considerable psychological and physiological welfare problems, which are mediated by the activation of various neuroendocrine networks that have numerous and pervasive effects. Psychological damage can be reflected in stereotypical behaviours, including repetitive pacing and circling, and even self-harm. Physical consequences include adverse effects on immune function, inflammatory responses, metabolism, and disease susceptibility and progression. Further, some of these effects are epigenetic, and are therefore potentially transgenerational: the biology of animals whose parents/grandparents were wild-caught and/or have experienced chronic stress in laboratories could be altered, as compared to free- living individuals. It is argued that these effects must have consequences for the reliability of experimental data and their extrapolation to humans, and this may not be recognised sufficiently among those who use animals in experiments.
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Assertions that the use of monkeys to investigate human diseases is valid scientifically are fre- quently based on a reported 90–93% genetic similarity between the species. Critical analyses of the rele- vance of monkey studies to human biology, however, indicate that this genetic similarity does not result in sufficient physiological similarity for monkeys to constitute good models for research, and that monkey data do not translate well to progress in clinical practice for humans. Salient examples include the failure of new drugs in clinical trials, the highly different infectivity and pathology of SIV/HIV, and poor extrapo- lation of research on Alzheimer’s disease, Parkinson’s disease and stroke. The major molecular differences underlying these inter-species phenotypic disparities have been revealed by comparative genomics and molecular biology — there are key differences in all aspects of gene expression and protein function, from chromosome and chromatin structure to post-translational modification. The collective effects of these dif- ferences are striking, extensive and widespread, and they show that the superficial similarity between human and monkey genetic sequences is of little benefit for biomedical research. The extrapolation of bio- medical data from monkeys to humans is therefore highly unreliable, and the use of monkeys must be con- sidered of questionable value, particularly given the breadth and potential of alternative methods of enquiry that are currently available to scientists.
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Unpredictable chronic mild stress (UCMS) and developmental social isolation are often utilized in laboratory animals to mimic unpredictable life stressors and early life adversity that may contribute to the development of major depressive disorder in humans. Zebrafish (Danio rerio) have been used to examine the effects of both developmental social isolation and UCMS. However, anxiety-like behavioral responses, social behavior, and neurochemical changes induced by stressors have not been well characterized. Furthermore, the possible interaction between UCMS and developmental isolation remains unexplored. In this study, we analyzed the effect of UCMS on developmentally isolated and socially reared zebrafish. The UCMS procedure entailed delivering unpredictably varying mild stressors twice a day for 15 consecutive days. To quantify social and anxiety-like behaviors, we measured the zebrafish's behavioral and neurochemical (dopaminergic and serotonergic) responses to an animated image of conspecifics in a novel tank. Our results suggest that UCMS increased anxiety-like behavioral responses, whereas developmental isolation altered motor responses during stimulus presentation. We also found that UCMS diminished weight gain and reduced whole-brain levels of dopamine and serotonin's metabolite 5-HIAA in developmentally isolated, but not socially reared zebrafish. Our findings reinforce the utility of combining developmental isolation with UCMS in zebrafish to model depressive-like behavior in humans.
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Assertions that the use of chimpanzees to investigate human diseases is valid scientifically are frequently based on a reported 98-99% genetic similarity between the species. Critical analyses of the relevance of chimpanzee studies to human biology, however, indicate that this genetic similarity does not result in sufficient physiological similarity for the chimpanzee to constitute a good model for research, and furthermore, that chimpanzee data do not translate well to progress in clinical practice for humans. Leading examples include the minimal citations of chimpanzee research that is relevant to human medicine, the highly different pathology of HIV/AIDS and hepatitis C virus infection in the two species, the lack of correlation in the efficacy of vaccines and treatments between chimpanzees and humans, and the fact that chimpanzees are not useful for research on human cancer. The major molecular differences underlying these inter-species phenotypic disparities have been revealed by comparative genomics and molecular biology - there are key differences in all aspects of gene expression and protein function, from chromosome and chromatin structure to post-translational modification. The collective effects of these differences are striking, extensive and widespread, and they show that the superficial similarity between human and chimpanzee genetic sequences is of little consequence for biomedical research. The extrapolation of biomedical data from the chimpanzee to the human is therefore highly unreliable, and the use of the chimpanzee must be considered of little value, particularly given the breadth and potential of alternative methods of enquiry that are currently available to science.
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Whether fishes are sentient beings remains an unresolved and controversial question. Among characteristics thought to reflect a low level of sentience in fishes is an inability to show stress-induced hyperthermia (SIH), a transient rise in body temperature shown in response to a variety of stressors. This is a real fever response, so is often referred to as 'emotional fever'. It has been suggested that the capacity for emotional fever evolved only in amniotes (mammals, birds and reptiles), in association with the evolution of consciousness in these groups. According to this view, lack of emotional fever in fishes reflects a lack of consciousness. We report here on a study in which six zebrafish groups with access to a temperature gradient were either left as undisturbed controls or subjected to a short period of confinement. The results were striking: compared to controls, stressed zebrafish spent significantly more time at higher temperatures, achieving an estimated rise in body temperature of about 2-4°C. Thus, zebrafish clearly have the capacity to show emotional fever. While the link between emotion and consciousness is still debated, this finding removes a key argument for lack of consciousness in fishes.