Matthew Parker

Housing conditions differentially affect physiological and behavioural stress responses of zebrafish, as well as the response to anxiolytics.

Zebrafish are a widely utilised animal model in developmental genetics, and owing to recent advances in our understanding of zebrafish behaviour, their utility as a comparative model in behavioural neuroscience is beginning to be realised. One widely reported behavioural measure is the novel tank-diving assay, which has been often cited as a test of anxiety and stress reactivity. Despite its wide utilisation, and various validations against anxiolytic drugs, reporting of pre-test housing has been sparse in the literature. As zebrafish are a shoaling species, we predicted that housing environment would affect their stress reactivity and, as such, their response in the tank-diving procedure. In our first experiment, we tested various aspects of housing (large groups, large groups with no contact, paired, visual contact only, olfactory contact only) and found that the tank diving response was mediated by visual contact with conspecifics. We also tested the basal cortisol levels of group and individually housed fish, and found that individually housed individuals have lower basal cortisol levels. In our second experiment we found ethanol appeared to have an anxiolytic effect with individually housed fish but not those that were group housed. In our final experiment, we examined the effects of changing the fishes water prior to tank diving as an additional acclimation procedure. We found that this had no effect on individually housed fish, but appeared to affect the typical tank diving responses of the group housed individuals. In conclusion, we demonstrate that housing represents an important factor in obtaining reliable data from this methodology, and should be considered by researchers interested in comparative models of anxiety in zebrafish in order to refine their approach and to increase the power in their experiments.

Discrimination reversal and attentional sets in zebrafish (Danio rerio)

The potential of zebrafish as a comparative model in behavioural neuroscience is currently hampered only by the lack of reliable and validated behavioural assays available to researchers. In the present experiment, we describe the performance of zebrafish in a test of attentional set formation. The fish were initially trained on a two-choice colour discrimination. Upon reaching acquisition criterion, the reinforced alternative was switched to the previously unreinforced alternative. Again, upon reaching criterion, the cues were replaced with a novel pair of colours (intra-dimensional shift) and reversed again on reaching criteria. We found that zebrafish show a steady decrease in trials-to-criteria over the four phases of the experiment, suggesting that they are forming and maintaining an attentional set, as has previously been demonstrated with mammals. Reversal learning deficits have been implicated in a variety of human psychological disorders (e.g., disorders of impulse control) and as such, we propose that performance of zebrafish in this procedure may represent a useful comparative model to complement existing rodent models.

Impaired instrumental choice in crib-biting horses (Equus caballus)

Horses displaying an oral stereotypy were tested on an instrumental choice paradigm to examine differences in learning from non-stereotypic counterparts. Stereotypic horses are known to have dysfunction of the dorsomedial striatum, and lesion studies have shown that this region may mediate response-outcome learning. The paradigm was specifically applied in order to examine learning that requires maintenance of response-outcome judgements. The non-stereotypic horses learned, over three sessions, to choose a more immediate reinforcer, whereas the stereotypic horses failed to do so. This suggests an initial behavioural correlate for dorsomedial striatum dysregulation in the stereotypy phenotype.

Development and implementation of a three-choice serial reaction time task for zebrafish (Danio rerio)

Zebrafish are an established and widely utilized developmental genetic model system, but limitations in developed behavioral assays have meant that their potential as a model in behavioral neuroscience has yet to be fully realized. Here, we describe the development of a novel operant behavioral assay to examine a variety of aspects of stimulus control in zebrafish using a 3 choice serial reaction time task (3 CSRTT). Fish were briefly exposed to three spatially distinct, but perceptually identical stimuli, presented in a random order after a fixed-time inter-trial interval (ITI). Entries to the correct response aperture either during the stimulus presentation, or within a brief limited hold period following presentation, were reinforced with illumination of the magazine light and delivery of a small food reward. Following training, premature responding was probed with a long-ITI session three times; once at baseline, once following a saline injection and once following an injection of a low dose of amphetamine (AMPH; 0.025 mg/kg). We predicted that if premature responding was related to impulsivity (as in rodents) it would be reduced following the AMPH injection. Results confirmed that zebrafish could learn to perform a complex operant task similar to tasks developed for rodents which are used to probe sustained attention and impulsivity, but the results from the AMPH trials were inconclusive. This study provides the foundations for development and further validation of this species as a model for some aspects of human attentional and impulse control disorders, such as substance abuse disorder.

Comparison of Polar ® heart rate interval data with simultaneously recorded ECG signals in horses

Human heart rate monitors (HRMs) are frequently used in equine studies to measure heart rate (HR) and interbeat intervals (IBIs). However, to date, the most commonly used HRM (the Polar® system) in horses has not been validated against simultaneously recorded electrocardiogram (ECG) signals during a range of ambulatory conditions. Polar® S810i and ECG IBIs were simultaneously recorded from six horses under three conditions commonly included in behavioural observation: standing at rest, loose in the stable and at liberty in a field. Following recording, Polar® IBI data were corrected for error processing in cardiac data. Corrected and uncorrected Polar® data were then compared with simultaneously recorded ECG data using a variety of commonly measured time and frequency domain parameters (e.g. HR variability (HRV)). Polar® data collected while horses were stabled or in the field were significantly different from ECG data, even following correction of the data, and therefore, it may not be possible for the two systems to be used interchangeably. This study indicates the need for caution while using Polar® S810i for collecting HRV data, unless horses are stationary, and even when the IBI data are corrected for measurement error.

Differential place and response learning in horses displaying an oral stereotypy

Significant similarities exist between the neural and behavioural features of environmentally and drug-induced stereotypy. For example, exposure to dopamine agonists, such as amphetamine, induces stereotypy and causes alterations in midbrain neurophysiology similar to those observed following chronic stress. An additional behavioural feature of these neural changes in the drug-induced phenotype is an enhanced rate of switching from response-outcome (R-O) to stimulus-response (S-R) learning. The aim of the current experiment was to examine R-O and S-R learning in horses displaying environmentally induced oral stereotypies. This was achieved by employing variations of the place/response paradigm. In Experiment 1, we found that crib-biting horses displayed response learning after 20 learning trials, whereas non-crib-biting controls tended to display place learning throughout the experiment. In Experiment 2, we used a modified version of the procedure, in which the subjects were introduced to the maze from different start points and forced always to turn the same way. We found that the crib-biters acquired the task at a faster rate suggesting again that this group was displaying response learning. Finally, in Experiment 3, we carried out an arena test to ensure that crib-biters were capable of place learning. These results are the first to show that horses displaying an oral stereotypy, a behavioural phenotype previously associated with stress-induced perturbations of the basal ganglia, preferentially use response learning. The findings are discussed in relation to the search for an aetiological model of stereotypy.

Zebrafish (Danio rerio) models of substance abuse:: harnessing the capabilities

Zebrafish are fast becoming one of the most widely used animal models in systems biology and developmental genetics, but their potential as models for behavioural neuroscience is only just beginning to be realised. This progress is dependent upon the rate at which behavioural assays for adult zebrafish are designed, reported and validated. As models for drug abuse and addiction, this species has been shown thus far to show high face validity, in that they show robust place preference for drug-related cues, as well as evidence for withdrawal, relapse and compulsive drug seeking. This review will outline the case for using zebrafish as models to study endophenotypes relating to drug addiction.

The impact of chronic environmental stressors on growing pigs, Sus scrofa (part 1):stress physiology, production and play behaviour

Commercially farmed animals are frequently housed in conditions that impose a number of concurrent environmental stressors. For pigs housed indoors, elevated levels of mechanical noise, atmospheric ammonia and low light intensities are commonplace. This experiment examined the effects on growing pigs of chronic exposure to combinations of commercially relevant levels of these potential stressors. Four-week-old hybrid female pigs (n = 224) were housed under experimentally manipulated conditions of nominally either <5 or 20 ppm atmospheric concentration of ammonia (24 h), a light intensity of 40 lux or 200 lux (12 h) and mechanical noise at either ⩽60 or 80 dB(A) (24 h) for 15 weeks in a fully factorial arrangement (23) of treatments. The response of pigs to these environmental factors was assessed using a suite of physiological, production and behavioural measures. These included indicators of hypothalamic-pituitary-adrenal (HPA) axis activation such as salivary cortisol and adrenal morphometry, as well as body weight, food conversion efficiency and general health scores. Play behaviour was recorded as it is thought to be inversely related to stress. Chronic exposure to ammonia produced the strongest effect, shown by lower concentrations of salivary cortisol and larger adrenal cortices in the pigs reared under 20 ppm ammonia, which may have been indicative of a period of HPA activation leading to a downregulation of cortisol production. The pigs in the ammoniated rooms also performed less play behaviour than pigs in non-ammoniated rooms. There was evidence for an interaction between high noise and ammonia on the health scores of pigs and for brighter light to ameliorate the effect of ammonia on salivary cortisol. However, there was no measurable impact of these potential stressors on the productivity of the pigs or any of the other physiological parameters measured. We conclude that there should be little concern in terms of performance about the physical stressors tested here, within current European Union legal limits. However, 20 ppm ammonia may have had an adverse influence on the well-being of growing pigs. In this study, all other aspects of the pigs husbandry were optimal; therefore, it is possible that under less favourable conditions, more pronounced effects of ammonia, noise and dim light would be observed.

The impact of chronic environmental stressors on growing pigs, Sus scrofa (Part 2): social behaviour

The effects of common and concurrent environmental stressors on the social behaviour of farm animals are poorly understood. Here, we report the results of a multifactorial experiment designed specifically to examine the individual, additive or interactive effects of elevated ammonia, noise and low light (LL) levels on the social behaviour of growing pigs. Social behaviour was measured in terms of the nature, frequency and duration of both initiated and response behaviours for 4 weeks following mixing of the groups. General activity patterns, group cohesion and social discrimination were also examined as a function of the environmental treatments. Elevated concentrations of atmospheric ammonia (∼20 v. <5 ppm) and LL intensity (∼40 v. 200 lux) had the most pronounced effects, particularly on the nature of social interactions, with pigs under these conditions showing more aggression in the early stages of the experiment. In addition, pigs exposed to a high level of mechanical noise representative of artificial ventilation (∼80 v. 40 dB [A]) were less submissive to aggressive acts, while pigs in ∼20 ppm ammonia showed more reciprocated aggression when in coincident LL (<40 lux). The results indicate that atmospheric ammonia at commonly experienced concentrations may undermine social stability, particularly in the presence of low lighting, though the mechanisms are currently unknown. These findings have implications for the welfare of growing pigs and hence policy makers and farmers alike, with respect to the improvement of welfare in intensive pig farming.

Effect of low light and high noise on behavioural activity, physiological indicators of stress and production in laying hens

1. Commercial laying hens are commonly housed in noisy and dim environments, yet relatively little is known about whether these conditions, particularly in combination, have any effect on welfare or egg production. 2. The study was designed to investigate whether chronic exposure to continuous noise (60u2009dB(A) vs. 80u2009dB(A)) and/or light intensity (150u2009lux vs. 5u2009lux) during the critical period of coming into lay (16–24 weeks of age) influenced behaviour (activity, resting and feather maintenance), physiological stress (plasma corticosterone and heterophil to lymphocyte ratio) and production (number and weight of eggs laid) in laying hens. 3. Hens in the low light pens were less active and preened and dust-bathed more than those housed in 150u2009lux; hens in the high noise pens rested more frequently than those in quieter pens. 4. There was no evidence that chronic exposure to low light or high noise caused appreciable physiological stress but egg production was affected by these conditions. Hens kept in pens with low light or high noise laid fewer eggs per day than those kept in high light or low noise pens. These effects were additive, so that the fewest eggs were laid by hens subject to both low light and high noise. 5. These results show that low light intensity and continual high background noise have a detrimental effect on egg production in the early laying phase as well as influencing the time allocated to different behaviours. However there was no strong evidence for a physiological stress response to either of these conditions or their combination.