Measuring endocrine (cortisol) responses of zebrafish to stress (original) (raw)
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Chapter 11 Measuring Endocrine (Cortisol) Responses of Zebrafish to Stress
The zebrafish (Danio rerio) is rapidly becoming a popular model species in stress and neuroscience research. Their behavior, robustly affected by environmental and pharmacological manipulations, can be paralleled by physiological (endocrine) analysis. Zebrafish have a hypothalamic-pituitary-interrenal (HPI) axis, which is homologous to the human hypothalamic-pituitary-adrenal (HPA) axis. While mice and rats use corticosterone as their main stress hormone, both humans and zebrafish utilize cortisol. This protocol explains the whole-body cortisol extraction procedure and the use of the human salivary cortisol ELISA kit to measure the amount of cortisol in each zebrafish sample. The ability to correlate physiological data from individual fish with behavioral data provides researchers with a valuable tool for investigating stress and anxiety, and contributes to the utility of zebrafish neurobehavioral models of stress.
PeerJ, 2019
In recent years, the zebrafish (Danio rerio) has become a popular model to study the mechanisms of physiological and behavioral effects of stress, due to the similarity in neural structures and biochemical pathways between zebrafish and mammals. Previous research in this vertebrate animal model has demonstrated an increase in whole-body cortisol resulting from an acute (30-second) net handling stress, but it remains unclear whether such a stressor will concomitantly increase anxiety-like behavior. In addition, as the previous study examined the effects of this acute stressor in adult zebrafish after a brief period of isolation, it is unclear whether this stressor would be effective in eliciting cortisol increases in younger aged subjects without isolation. In the current study, young adult zebrafish (approximately 90 days post-fertilization) were briefly exposed to a net handling stressor and were subsequently subjected to either the novel tank test or the light/dark preference test...
Understanding behavioral and physiological phenotypes of stress and anxiety in zebrafish
Behavioural brain …, 2009
The zebrafish (Danio rerio) is emerging as a promising model organism for experimental studies of stress and anxiety. Here we further validate zebrafish models of stress by analyzing how environmental and pharmacological manipulations affect their behavioral and physiological phenotypes. Experimental manipulations included exposure to alarm pheromone, chronic exposure to fluoxetine, acute exposure to caffeine, as well as acute and chronic exposure to ethanol. Acute (but not chronic) alarm pheromone and acute caffeine produced robust anxiogenic effects, including reduced exploration, increased erratic movements and freezing behavior in zebrafish tested in the novel tank diving test. In contrast, ethanol and fluoxetine had robust anxiolytic effects, including increased exploration and reduced erratic movements. The behavior of several zebrafish strains was also quantified to ascertain differences in their behavioral profiles, revealing high-anxiety (leopard, albino) and low-anxiety (wild type) strains. We also used LocoScan (CleverSys Inc.) video-tracking tool to quantify anxiety-related behaviors in zebrafish, and dissect anxiety-related phenotypes from locomotor activity. Finally, we developed a simple and effective method of measuring zebrafish physiological stress responses (based on a human salivary cortisol assay), and showed that alterations in whole-body cortisol levels in zebrafish parallel behavioral indices of anxiety. Collectively, our results confirm zebrafish as a valid, reliable, and highthroughput model of stress and affective disorders.
Measuring behavioral and endocrine responses to novelty stress in adult zebrafish
Nature Protocols, 2010
several behavioral assays are currently used for high-throughput neurophenotyping and screening of genetic mutations and psychotropic drugs in zebrafish (Danio rerio). In this protocol, we describe a battery of two assays to characterize anxiety-related behavioral and endocrine phenotypes in adult zebrafish. Here, we detail how to use the 'novel tank' test to assess behavioral indices of anxiety (including reduced exploration, increased freezing behavior and erratic movement), which are quantifiable using manual registration and computer-aided video-tracking analyses. In addition, we describe how to analyze whole-body zebrafish cortisol concentrations that correspond to their behavior in the novel tank test. this protocol is an easy, inexpensive and effective alternative to other methods of measuring stress responses in zebrafish, thus enabling the rapid acquisition and analysis of large amounts of data. as will be shown here, fish anxiety-like behavior can be either attenuated or exaggerated depending on stress or drug exposure, with cortisol levels generally expected to parallel anxiety behaviors. this protocol can be completed over the course of 2 d, with a variable testing duration depending on the number of fish used.
Progress in Neuro-Psychopharmacology and Biological Psychiatry, 2011
Zebrafish (Danio rerio) have emerged as a promising model organism to study development, toxicology, pharmacology, and neuroscience, among other areas. Despite the increasing number of studies using zebrafish, behavioral studies with this species are still elementary when compared to rodents. The aim of this study was to develop a model of unpredictable chronic stress (UCS) in zebrafish. We evaluated the effects of UCS protocol during 7 or 14 days on behavioral and physiological parameters. The effects of stress were evaluated in relation to anxiety and exploratory behavior, memory, expression of corticotrophin-releasing factor (CRF) and glucocorticoid receptor (GR), and cortisol levels. As expected, UCS protocol increased the anxiety levels, impaired cognitive function, and increased CRF while decreased GR expression. Moreover, zebrafish submitted to 7 or 14 days of UCS protocol presented increased cortisol levels. The protocol developed here is a complementary model for studying the neurobiology and the effects of chronic stress in behavioral and physiological parameters. In addition, this protocol is less time consuming than standard rodent models commonly used to study chronic stress. These results confirm UCS in zebrafish as an adequate model to preclinical studies of stress, although further studies are warranted to determine its predictive validity.
Modeling Stress and Anxiety in Zebrafish
Neuromethods, 2010
While zebrafish (Danio rerio) are widely utilized as a model species for neuroscience research, they also possess several qualities that make them particularly useful for studying stress and anxiety-related behaviors. Zebrafish neuroendocrine responses are robust, and correlate strongly with behavioral endpoints. These fish are also highly sensitive to various environmental challenges, including novelty stress, exposure to predators, alarm pheromone, anxiogenic drugs, and drug withdrawal. In addition, varying levels of baseline anxiety can be observed in different strains of zebrafish. Collectively, this supports the validity and efficacy of the adult zebrafish model for studying both acute and chronic anxiety.
Chapter 3 Modeling Stress and Anxiety in Zebrafish
2014
While zebrafish (Danio rerio) are widely utilized as a model species for neuroscience research, they also possess several qualities that make them particularly useful for studying stress and anxiety-related behav-iors. Zebrafish neuroendocrine responses are robust, and correlate strongly with behavioral endpoints. These fish are also highly sensitive to various environmental challenges, including novelty stress, exposure to predators, alarm pheromone, anxiogenic drugs, and drug withdrawal. In addition, varying levels of baseline anxiety can be observed in different strains of zebrafish. Collectively, this supports the validity and efficacy of the adult zebrafish model for studying both acute and chronic anxiety.
Whole-body cortisol is an indicator of crowding stress in adult zebrafish, Danio rerio
Aquaculture, 2006
Plasma cortisol levels have been used to evaluate the stress response in a variety of cultured fish species. However, little is known about the stress response of zebrafish, Danio rerio, despite its extensive use as a laboratory research organism. Due to its small size, evaluation of whole-body cortisol has provided a means to assess zebrafish stress levels Understanding the role of crowding on zebrafish whole-body cortisol would allow researchers to optimize various rearing procedures. The aim of this study was to determine the zebrafish whole-body cortisol response to crowding stress. Baseline cortisol values have not been established for zebrafish; therefore, we compared specific treatment groups acclimated to similar conditions. We crowded zebrafish at a high density (40 fish/L) for 3 h (acute stress), or 5 days (chronic stress). Crowding resulted in a four-fold increase in mean whole-body cortisol level in both groups compared to zebrafish maintained at a much lower density (0.25 fish/L). Additional experiments demonstrated that the cortisol response to crowding was modulated by fasting, feeding and density. In large glass aquaria (76 L), fasted, crowded fish (40 fish/L) had significantly higher cortisol compared to fasted, control fish (0.2 fish/L). Furthermore, weight was inversely related to cortisol level in fasted, crowded fish held in large glass aquaria. For fed fish, crowding did not significantly increase cortisol level, suggesting an interaction between feeding and crowding. In small tanks (4 L), crowding (40 fish/L) did not significantly increase cortisol compared to control fish held at 4 fish/L. Our results suggest that whole-body cortisol is a useful indicator of crowding stress in fasted, adult zebrafish. Understanding how crowding and other environmental conditions affect zebrafish fitness could aid in optimizing zebrafish growth, health and reproduction as well as improving the consistency and reproducibility of in vivo studies that use this popular vertebrate model.
Behaviour, 2012
Animals encounter stressful situations multiple times throughout their lives and often successfully cope with them. Individuals vary in the nature and intensity of their behavioral and physiological response to stressors, often representing correlated and qualitatively distinct coping styles (e.g., proactive and reactive). These alternative coping styles are ways an animal can overcome a variety of stressful situations, which ultimately can have important fitness consequences. Here we use zebrafish (Danio rerio) recently wild-derived and selectively bred for amount of stationary behavior (High and Low lines) and a classic domesticated strain (AB) to document the utility of these zebrafish strains in understanding coping mechanisms. The Low Stationary Behavior (LSB) line of zebrafish displayed significantly lower stress and anxiety-related behaviors than the High Stationary Behavior (HSB) across six stress and anxiety-related behavioral assays. In some assays, we observed strain differences in behavior within three minutes of the start of the trial. Males also showed reduced levels of anxiety-related behaviors relative to females in two assays. Comparing wild-derived and domesticated strains, the AB line displayed significantly lower levels of anxietyrelated behavior in half of the assays. This study demonstrates that our selectively bred lines from wild-caught zebrafish (HSB, LSB) exhibit consistent and divergent behavioral stress responses across multiple distinct assays. Hence these lines may prove useful in understanding the proximate and ultimate mechanisms of coping with stress and anxiety.
A zebrafish model of glucocorticoid resistance shows serotonergic modulation of the stress response
Frontiers in behavioral neuroscience, 2012
One function of glucocorticoids is to restore homeostasis after an acute stress response by providing negative feedback to stress circuits in the brain. Loss of this negative feedback leads to elevated physiological stress and may contribute to depression, anxiety, and post-traumatic stress disorder. We investigated the early, developmental effects of glucocorticoid signaling deficits on stress physiology and related behaviors using a mutant zebrafish, gr(s357), with non-functional glucocorticoid receptors (GRs). These mutants are morphologically inconspicuous and adult-viable. A previous study of adult gr(s357) mutants showed loss of glucocorticoid-mediated negative feedback and elevated physiological and behavioral stress markers. Already at 5 days post-fertilization, mutant larvae had elevated whole body cortisol, increased expression of pro-opiomelanocortin (POMC), the precursor of adrenocorticotropic hormone (ACTH), and failed to show normal suppression of stress markers after ...