Impact of stress, fear and anxiety on the nociceptive responses of larval zebrafish

Both adult and larval zebrafish have been demonstrated to show behavioural responses to noxious stimulation but also to potentially stress- and fear or anxiety- eliciting situations. The pain or nociceptive response can be altered and modulated by these situations in adult fish through a mechanism called stress-induced analgesia. However, this phenomenon has not been described in larval fish yet. Therefore, this study explores the behavioural changes in larval zebrafish after noxious stimulation and exposure to challenges that can trigger a stress, fear or anxiety reaction. Five-day post fertilization zebrafish were exposed to either a stressor (air emersion), a predatory fear cue (alarm substance) or an anxiogenic (caffeine) alone or prior to immersion in acetic acid 0.1%. Pre- and post-stimulation behaviour (swimming velocity and time spent active) was recorded using a novel tracking software in 25 fish at once. Results show that larvae reduced both velocity and activity after exposure to the air emersion and alarm substance challenges and that these changes were attenuated using etomidate and diazepam, respectively. Exposure to acetic acid decreased velocity and activity as well, whereas air emersion and alarm substance inhibited these responses, showing no differences between pre- and post-stimulation. Therefore, we hypothesize that an antinociceptive mechanism, activated by stress and/or fear, occur in 5dpf zebrafish, which could have prevented the larvae to display the characteristic responses to pain

Efficient individual identification of zebrafish using Hue/Saturation/Value color model

Automated fish species recognition is widely investigated in research but it is not explored for the individuals with the same fish species. A new classifying method for zebrafish individuals that is based on statistical texture and Hue/Saturation/Value (HSV) color features are presented in this paper. Post image acquisition, pre-processing stages and features of sub-images are extracted, using statistical texture and HSV color space domain, and grouped into HSV and statistical sets of features. An artificial neural network (ANN) and K-Nearest Neighbors (KNN) are then used to identify the subjects under test. The impact of using statistical and HSV features on the prediction accuracy and average processing time is then assessed experimentally. An improved performance for the HSV over the statistical model is clearly demonstrated. The combination of HSV model and KNN classifier has also demonstrated a superior performance over the combination of HSV and ANN classifier in terms of the accuracy (KNN = 99.0%; ANN = 97.8%) and average processing time (KNN = 4.1 ms; ANN = 24.2 ms). Such promising findings encourage further testing of the HSV model towards developing a highly-efficient and fully-automated identification system for small species individual like zebrafish. Keywords: Fish identification, Statistical texture features, HSV color features, KNN classifier, ANN classifie

Impact of stress, fear and anxiety on the nociceptive responses of larval zebrafish

Both adult and larval zebrafish have been demonstrated to show behavioural responses to noxious stimulation but also to potentially stress- and fear or anxiety- eliciting situations. The pain or nociceptive response can be altered and modulated by these situations in adult fish through a mechanism called stress-induced analgesia. However, this phenomenon has not been described in larval fish yet. Therefore, this study explores the behavioural changes in larval zebrafish after noxious stimulation and exposure to challenges that can trigger a stress, fear or anxiety reaction. Five-day post fertilization zebrafish were exposed to either a stressor (air emersion), a predatory fear cue (alarm substance) or an anxiogenic (caffeine) alone or prior to immersion in acetic acid 0.1%. Pre- and post-stimulation behaviour (swimming velocity and time spent active) was recorded using a novel tracking software in 25 fish at once. Results show that larvae reduced both velocity and activity after exposure to the air emersion and alarm substance challenges and that these changes were attenuated using etomidate and diazepam, respectively. Exposure to acetic acid decreased velocity and activity as well, whereas air emersion and alarm substance inhibited these responses, showing no differences between pre- and post-stimulation. Therefore, we hypothesize that an antinociceptive mechanism, activated by stress and/or fear, occur in 5dpf zebrafish, which could have prevented the larvae to display the characteristic responses to pain

Reduction in activity by noxious chemical stimulation is ameliorated by immersion in analgesic drugs in zebrafish

Research has recently demonstrated that larval zebrafish show similar molecular responses to nociception to those of adults. Our study explored whether unprotected larval zebrafish exhibited altered behaviour after exposure to noxious chemicals and screened a range of analgesic drugs to determine their efficacy to reduce these responses. This approach aimed to validate larval zebrafish as a reliable replacement for adults as well as providing a high-throughput means of analysing behavioural responses. Zebrafish at 5 days postfertilization were exposed to known noxious stimuli: acetic acid (0.01%, 0.1% and 0.25%) and citric acid (0.1%, 1% and 5%). The behavioural response of each was recorded and analysed using novel tracking software that measures time spent active in 25 larvae at one time. Subsequently, the efficacy of aspirin, lidocaine, morphine and flunixin as analgesics after exposure to 0.1% acetic acid was tested. Larvae exposed to 0.1% and 0.25% acetic acid spent less time active, whereas those exposed to 0.01% acetic acid and 0.1–5% citric acid showed an increase in swimming activity. Administration of 2.5 mg l−1 aspirin, 5 mg l−1 lidocaine and 48 mg l−1 morphine prevented the behavioural changes induced by acetic acid. These results suggest that larvae respond to a noxious challenge in a similar way to adult zebrafish and other vertebrates and that the effect of nociception on activity can be ameliorated by using analgesics. Therefore, adopting larval zebrafish could represent a direct replacement of a protected adult fish with a non-protected form in pain- and nociception-related research

Impact of stress, fear and anxiety on the nociceptive responses of larval zebrafish

Both adult and larval zebrafish have been demonstrated to show behavioural responses to noxious stimulation but also to potentially stress- and fear or anxiety- eliciting situations. The pain or nociceptive response can be altered and modulated by these situations in adult fish through a mechanism called stress-induced analgesia. However, this phenomenon has not been described in larval fish yet. Therefore, this study explores the behavioural changes in larval zebrafish after noxious stimulation and exposure to challenges that can trigger a stress, fear or anxiety reaction. Five-day post fertilization zebrafish were exposed to either a stressor (air emersion), a predatory fear cue (alarm substance) or an anxiogenic (caffeine) alone or prior to immersion in acetic acid 0.1%. Pre- and post-stimulation behaviour (swimming velocity and time spent active) was recorded using a novel tracking software in 25 fish at once. Results show that larvae reduced both velocity and activity after exposure to the air emersion and alarm substance challenges and that these changes were attenuated using etomidate and diazepam, respectively. Exposure to acetic acid decreased velocity and activity as well, whereas air emersion and alarm substance inhibited these responses, showing no differences between pre- and post-stimulation. Therefore, we hypothesize that an antinociceptive mechanism, activated by stress and/or fear, occur in 5dpf zebrafish, which could have prevented the larvae to display the characteristic responses to pain

Swimming activity of larval zebrafish after exposure to irritant chemicals and analgesics

Swimming activity of larval zebrafish after exposure to irritant chemicals and analgesi

Change in time spent active (%) shown by 5dpf zebrafish exposed to different treatments.

<p>Each box shows the lower and upper quartile values and the central horizontal black line indicates the median value. The error bars indicate the variation for the rest of the data and outliers are indicated as white dots. Significant differences between groups and the control groups are indicated with an asterisk (control in the air emersion challenge: CO/AE), a hashtag (control in the alarm pheromone challenge: CO/AP) or a cross (control in the caffeine challenge: CO/CF) (Wilcoxon signed-rank test, P<0.0031).</p