Cognition in freshwater fish: effects of the environment

For animals that live in a reasonably variable environment the capacity for learning and memory allow them to adapt to the changes they experience. Ecological factors that vary between habitats can affect a range of learning behaviours. Less attention has been directed at how this variation may affect memory processes, or how different ecological variables might interact when shaping cognition and behaviour. Therefore one aim of this thesis was to investigate how different ecological variables shape memory abilities and to test whether those same variables affect other related behaviours such as learning. In order to test this, I selected natural populations of a temperate freshwater fish, the three-spined stickleback (Gasterosteus aculeatus) from pond and river habitats that were proposed to differ in predation pressure, and assayed their learning, memory and other behavioural traits. Pond and river populations differed in their memory and orientation behaviour. An interaction between pond/river habitat and predation pressure affected learning rate, and a similar interaction affected temperament behaviours. Two further studies were conducted to address how captive rearing environments and typical handling procedures affect behaviour in different species. Rearing environment affected memory, but not learning or temperament behaviours in three-spined sticklebacks. Handling caused stress responses in three-spined sticklebacks, Panamanian bishops (Brachyraphis episcopi) and Rainbow trout (Oncorhynchus mykiss), but handling with a water filled scoop compared to a traditional dip-net decreased these responses in three-spined sticklebacks and Panamanian bishops, and also affected behaviour in Panamanian bishops. The results presented in this thesis suggest that ecological variables play a substantial role in shaping learning, memory and other behavioural traits in fish, and highlight the utility of behavioural assays in answering welfare-based questions

A shortened protocol for assessing cognitive bias in rats

Reliable measurement of affective state in animals is a significant goal of animal welfare. Such measurements would also improve the validity of pre-clinical mental health research which relies on animal models. However, at present, affective states in animals are inaccessible to direct measurement. In humans, changes in cognitive processing can give reliable indications of emotional state. Therefore, similar techniques are increasingly being used to gain proxy measures of affective states in animals. In particular, the ‘cognitive bias’ assay has gained popularity in recent years. Major disadvantages of this technique include length of time taken for animals to acquire the task (typically several weeks), negative experiences associated with task training, and issues of motivation. New method Here we present a shortened cognitive bias protocol using only positive reinforcers which must actively be responded to. Results The protocol took an average of 4 days to complete, and produced similar results to previous, longer methods (minimum 30 days). Specifically, rats housed in standard laboratory conditions demonstrated negative cognitive biases when presented with ambiguous stimuli, and took longer to make a decision when faced with an ambiguous stimulus. Comparison with existing methods Compared to previous methods, this protocol is significantly shorter (average 4 days vs. minimum 30 days), utilises only positive reinforcers to avoid inducing negative affective states, and requires active responses to all cues, avoiding potential confounds of motivational state. Conclusions We have successfully developed a shortened cognitive bias protocol, suitable for use with laboratory rats. Keywords Cognitive bias; Emotion; Affect; Rat; Protoco

Enduring neuroimmunological consequences of developmental experiences: from vulnerability to resilience

The immune system is crucial for normal neuronal development and function (neuroimmune system). Both immune and neuronal systems undergo significant postnatal development and are sensitive to developmental programming by environmental experiences. Negative experiences from infection to psychological stress at a range of different time points (in utero to adolescence) can permanently alter the function of the neuroimmune system: given its prominent role in normal brain development and function this dysregulation may increase vulnerability to psychiatric illness. In contrast, positive experiences such as exercise and environmental enrichment are protective and can promote resilience, even restoring the detrimental effects of negative experiences on the neuroimmune system. This suggests the neuroimmune system is a viable therapeutic target for treatment and prevention of psychiatric illnesses, especially those related to stress. In this review we will summarise the main cells, molecules and functions of the immune system in general and with specific reference to central nervous system development and function. We will then discuss the effects of negative and positive environmental experiences, especially during development, in programming the long-term functioning of the neuroimmune system. Finally, we will review the sparse but growing literature on sex differences in neuroimmune development and response to environmental experiences

Female HPA axis displays heightened sensitivity to pre-pubertal stress

Early life stress (ELS) is a risk factor in the development of psychiatric disorders. The underlying biological mechanisms governing this phenomenon are not fully understood, but dysregulation of stress responses is likely to play a key role. Males and females differ in their propensity to develop psychiatric disorders, with far higher rates of anxiety, major depressive disorder, affective disorders and post-traumatic stress disorder found in women. We hypothesised that sex differences in response to ELS may play a crucial role in differential vulnerability between the sexes. To test this, we evaluated the consequences of pre-pubertal stress (PPS) on the HPA axis in adult female and male Lister Hooded rats. PPS animals were exposed to swim, restraint and elevated platform stress on postnatal days 25-27, controls remained in their home cage. Once adult, animals were either a) sacrificed directly and brains collected or b) sacrificed 20 minutes or 1 week after a social test and trunk blood collected. In the female hippocampal formation, PPS increased expression of FKBP5 and AVPR1a. In the female prefrontal cortex, PPS resulted in increased glucocorticoid receptor expression, increased glucocorticoid:mineralocorticoid (GR:MR) receptor expression ratio and decreased AVPR1a expression. Females exposed to PPS did not show the normal rise in blood corticosterone levels following a social interaction test. In contrast, PPS did not alter the expression of oxytocin or oxytocin receptors, and no effects of PPS were seen in males. However, striking sex differences were found. Females had higher oxytocin receptor expression in the prefrontal cortex and AVPR1a and oxytocin expression in the hypothalamus, whereas males demonstrated higher expression of GR, MR, GR:MR, FKBP5 and oxytocin receptor in the hypothalamus. These results demonstrate heightened reactivity of the female HPA axis to PPS and may help explain why in humans females display an increased susceptibility to certain stress-related psychopathologies. Lay Summary: Women are at greater risk of developing several psychiatric illnesses. Using a rodent model, we show that the female stress system is more reactive to the lasting effects of early life stress. This heightened reactivity of the female stress response may help explain why women are at a greater risk of developing psychiatric disorders

Environmental enrichment induces optimistic cognitive bias in rats

People’s affective or emotional state can alter their cognitive processing, biasing interpretation of ambiguous stimuli. Those in a more positive state interpret such stimuli in a more optimistic manner than those in a negative state. Recently this research has extended to animals, and has shown that manipulations associated with negative affect cause animals to interpret ambiguous stimuli more pessimistically. We investigated whether exposure to environmental enrichment engenders optimistic responses to ambiguous stimuli. Rats, Rattus norvegicus, were trained on a novel conditional discrimination task whereby they learned the correct response necessary to obtain a food reward given the stimulus present during approach (rough or smooth sandpaper). One stimulus was associated with a higher-value reward than the other. Once the rats were trained, cognitive bias was probed by exploring their responses to an ambiguous stimulus (intermediate grade of sandpaper); a rat was defined as optimistic if it chose the response appropriate to the stimulus associated with the better reward. Animals transferred from unenriched to enriched cages showed more optimistic responses following the change. A control group maintained in unenriched cages showed pessimistic responses throughout. These results demonstrate for the first time that environmental enrichment can induce an optimistic cognitive bias in rats previously housed in standard caging, possibly indicative of a more positive affective state. These results add support to the suggestion that measuring cognitive biases can give an insight into animal emotional states; this has implications for animal welfare and preclinical testing of potential therapeutics for mood disorders

Cacna1c hemizygosity results in aberrant fear conditioning to neutral stimuli

CACNA1C, a gene that encodes an alpha-1 subunit of L-type voltage-gated calcium channels, has been strongly associated with psychiatric disorders including schizophrenia and bipolar disorder. An important objective is to understand how variation in this gene can lead to an increased risk of psychopathology. Altered associative learning has also been implicated in the pathology of psychiatric disorders, particularly in the manifestation of psychotic symptoms. In this study, we utilize auditory-cued fear memory paradigms in order to investigate whether associative learning is altered in rats hemizygous for the Cacna1c gene. Cacna1c hemizygous (Cacna1c+/−) rats and their wild-type littermates were exposed to either delay, trace, or unpaired auditory fear conditioning. All rats received a Context Recall (24 h post-conditioning) and a Cue Recall (48 h post-conditioning) to test their fear responses. In the delay condition, which results in strong conditioning to the cue in wild-type animals, Cacna1c+/− rats showed increased fear responses to the context. In the trace condition, which results in strong conditioning to the context in wild-type animals, Cacna1c+/− rats showed increased fear responses to the cue. Finally, in the unpaired condition, Cacna1c+/− rats showed increased fear responses to both context and cue. These results indicate that Cacna1c heterozygous rats show aberrantly enhanced fear responses to inappropriate cues, consistent with key models of psychosis

Social interaction following prepubertal stress alters prefrontal gene expression associated with cell signalling and oligodendrocytes

Early-life adversity is associated with an increased risk of psychopathology, including mood disorders, later in life. Early-life stress affects several physiological systems, however, the exact mechanisms underlying pathological risk are not fully understood. This knowledge is crucial in developing appropriate therapeutic interventions. The prepubertal period is documented as a key developmental period for the maturation of the prefrontal cortex (PFC), a brain region involved in higher cognitive functions, including social function. In this study, we performed RNA sequencing on the PFC of adult rats who had experienced prepubertal stress (PPS) and controls to investigate the genome-wide consequences of this stress. PPS alters social behaviour in adulthood, therefore we also performed RNA sequencing on PPS and control rats following a social interaction test to determine social activity-dependent gene changes. At a baseline state (1 week following a social interaction test), no genes were differentially expressed in the PPS group. However, 1603 genes were differentially expressed in PPS rats compared to controls following a social interaction. These genes were enriched in biological pathways associated with cell signalling and axon myelination dynamics. Cell enrichment analysis showed these genes were associated with oligodendrocytes, and a comparison with an existing early-life stress sequencing dataset showed that pathways linked to oligodendrocyte morphology are impacted in a range of models of early-life stress in rodents. In conclusion, we identify pathways, including those involved in axon myelination, that are differentially activated in the adult in response to social stimulation following PPS. These differential responses may contribute to vulnerability to psychiatric pathology

Childhood stress impairs social function through AVP-dependent mechanisms

Impaired social function is a core feature of many psychiatric illnesses. Adverse experiences during childhood increase risk for mental illness, however it is currently unclear whether stress early in life plays a direct role in the development of social difficulties. Using a rat model of pre-pubertal stress (PPS), we investigated effects on social behaviour, oxytocin and arginine vasopressin (AVP) in the periphery (plasma) and centrally in the paraventricular and supraoptic hypothalamic nuclei. We also explored social performance and AVP expression (plasma) in participants with borderline personality disorder (BPD) who experienced a high incidence of childhood stress. Social behaviour was impaired and AVP expression increased in animals experiencing PPS and participants with BPD. Behavioural deficits in animals were rescued through administration of the AVPR1a antagonist Relcovaptan (SR49059). AVP levels and recognition of negative emotions were significantly correlated in BPD participants only. In conclusion, early life stress plays a role in the precipitation of social dysfunction, and AVP mediates at least part of this effect

Imaging learned fear circuitry in awake mice using fMRI

Functional magnetic resonance imaging (fMRI) of learned behaviour in ‘awake rodents’ provides the opportunity for translational preclinical studies into the influence of pharmacological and genetic manipulations on brain function. fMRI has recently been employed to investigate learned behaviour in awake rats. Here, this methodology is translated to mice, so that future fMRI studies may exploit the vast number of genetically modified mouse lines that are available. One group of mice was conditioned to associate a flashing light (conditioned stimulus, CS) with foot shock (PG; paired group), and another group of mice received foot shock and flashing light explicitly unpaired (UG; unpaired group). The blood oxygen level-dependent signal (proxy for neuronal activation) in response to the CS was measured 24 h later in awake mice from the PG and UG using fMRI. The amygdala, implicated in fear processing, was activated to a greater degree in the PG than in the UG in response to the CS. Additionally, the nucleus accumbens was activated in the UG in response to the CS. Because the CS signalled an absence of foot shock in the UG, it is possible that this region is involved in processing the safety aspect of the CS. To conclude, the first use of fMRI to visualise brain activation in awake mice that are completing a learned emotional task is reported. This work paves the way for future preclinical fMRI studies to investigate genetic and environmental influences on brain function in transgenic mouse models of disease and aging

The effects of juvenile stress on anxiety, cognitive bias and decision making in adulthood:a rat model

Stress experienced in childhood is associated with an increased risk of developing psychiatric disorders in adulthood. These disorders are particularly characterized by disturbances to emotional and cognitive processes, which are not currently fully modeled in animals. Assays of cognitive bias have recently been used with animals to give an indication of their emotional/cognitive state. We used a cognitive bias test, alongside a traditional measure of anxiety (elevated plus maze), to investigate the effects of juvenile stress (JS) on adulthood behaviour using a rodent model. During the cognitive bias test, animals were trained to discriminate between two reward bowls based on a stimulus (rough/smooth sandpaper) encountered before they reached the bowls. One stimulus (e.g. rough) was associated with a lower value reward than the other (e.g. smooth). Once rats were trained, their cognitive bias was explored through the presentation of an ambiguous stimulus (intermediate grade sandpaper): a rat was classed as optimistic if it chose the bowl ordinarily associated with the high value reward. JS animals were lighter than controls, exhibited increased anxiety-like behaviour in the elevated plus maze and were more optimistic in the cognitive bias test. This increased optimism may represent an optimal foraging strategy for these underweight animals. JS animals were also faster than controls to make a decision when presented with an ambiguous stimulus, suggesting altered decision making. These results demonstrate that stress in the juvenile phase can increase anxiety-like behaviour and alter cognitive bias and decision making in adulthood in a rat model