in Row Trout (Oncorhynchus mykiss) !" " #$%#&' (" )%$$'%#*)+%$ ,, ,,-.%#*& !" # $ % &''( !')'' * * * + , - . , /0 1, &''(, / / 2 . - 3 4, # , !!", (& , , 5/ %!6((7689%:6(, # . , - * . . . * , + . . , * . *. * , ; * *, ; . . . 3<24 . 324 ** , <2 * . . 39' 4 2 *, = . ** . <2 2 * . . * , - * . . * . . , < . . * * ** . . . ** > , $ . * * . , $ * . ** , < . . . , ; * ** *, ! / * / / ? - ? / " #$ % & $ '( )* +$ $ ,-./012 $ @ 1 /0 &''( 5// !8(!68&!7 5/ %!6((7689%:6( ) ))) 68!9& 3 )AA ,>,A B C ) ))) 68!9&4 List of Papers This thesis is based on the following articles, which will be referred to by their roman numerals. I J. Schjolden, K.G.T. Pulman, T.G. Pottinger, N.B. Metcalfe & S. Winberg. Divergence in locomotor activity within established and stable social structures in two strains of rainbow trout (Oncorhynchus mykiss) with contrasting stress responsiveness. Journal of Fish Biology – in press. II J. Schjolden, T. Backström, K.G.T. Pulman, T.G. Pottinger & S. Winberg. Divergence in behavioural responses to stress in two strains of rainbow trout (Oncorhynchus mykiss) with contrasting stress responsiveness. Hormones and Behavior – in press. III J. Schjolden, K.G.T. Pulman, T.G. Pottinger, O. Tottmar & S. Winberg. Serotonergic characteristics of rainbow trout divergent in stress responsiveness. Physiology and Behavior – submitted. IV J. Schjolden, A. Stoskhus & S. Winberg. 2004. Does intraspecific variation in stress responses and agonistic behaviour reflect divergent stress coping strategies in juvenile rainbow trout? – Physiological and Biochemical Zoology 78 (5): 715-723. Reprints of these papers were made with kind permissions from the following publishers: Paper I: Journal of Fish Biology; Blackwell Publishing, Edinburgh. Paper II: Hormones and Behavior; Elsevier, San Diego. Paper III: Submitted, Physiology and Behavior. Paper IV: Physiological and Biochemical Zoology, The University of Chicago Press, Chicago. Table of Contents 1 Introduction..................................................................................................7 1.1 The stress response...............................................................................7 1.1.1 The cortisol response to stress in teleost fish................................8 1.1.2 The catecholamine response to stress in teleost fish...................10 1.1.3 Brain serotonergic activity..........................................................11 1.2 Stress coping strategies ......................................................................14 1.3 consistent behavioural patterns in fish ...............................................15 1.4 Does stress coping strategies exist in fish?.........................................16 1.5 Objectives...........................................................................................18 2 Materials and Methods...............................................................................19 2.1 Locations and experimental animals ..................................................19 2.2 Experimental conditions and procedures ...........................................20 2.2.1 The home environment...............................................................20 2.2.2 Plasma cortisol and catecholamine responses to confinement ...21 2.2.3 Behavioural response to a novel environment or object.............21 2.2.4 Locomotor activity in HR and LR fish interacting in groups .....22 2.2.5 Aggression – Intruder test and dyadic interaction ......................22 2.2.6 Resumption of food intake subsequent to transfer......................23 2.2.7 Assays.........................................................................................23 3 Results and Discussion ..............................................................................26 3.1 Physiological stress responses in HR and LR fish .............................26 3.1.1 The cortisol response ..................................................................26 3.1.2 The catecholamine response .......................................................26 3.1.3 The serotonergic activity ............................................................27 3.2 Behavioural stress responses in hr and lr fish ....................................30 3.2.1 Locomotor activity and social structure......................................30 3.2.2 Do HR and LR rainbow trout exhibit different stress coping strategies? ............................................................................................33 3.3 Stress responses in an unselected line of rainbow trout .....................36 3.4 Summary and conclusion ...................................................................40 Svensk sammanfattning ................................................................................41 Acknowledgements.......................................................................................42 References.....................................................................................................43 Abbreviations 5-HIAA – 5-Hydroxyindole-3-acetic acid 5-HT – 5-Hydroxytryptamine (Serotonin) 5-HTP – 5-Hydroxytryptophan D-MSH – D-Melanocyte stimulating hormone AAD – L-Amino acid decarboxylase ACTH – Adrenocorticotropic hormone ALDH – Aldehyd dehydrogenase AVT – Arginine vasotocin BST – Brain stem CA – Catecholamines CRH – Corticotropin releasing hormone DA – Dopamine DHBA – 3,4 Dihydroxybenzylamine ED – Electrochemical detection EDTA – Ethylenediaminetetraacetic acid DHBA – 3,4-Dihydroxybenzylamine E – Epinephrine EDTA – Ethylenediaminetetraacetic acid HPA – Hypothalamus-pituitary-adrenal HPI – Hypothalamus-pituitary-interrenal HPLC – High performance liquid chromatography HR – High responder or high responding HSC – Hypothalamic-sympathetic-chromaffin HYP – Hypothalamus IT – Isotocin LR – Low responder or low responding MAO – Monoamineoxidase MCH – Melanocyte concentrating hormone NAD+ – Nicotinamide adenine dinucleotide NE – Norepinephrine NPY – Neuropeptide Y OPT – Optic tectum PCA – Perchloric acid PCPA – Para chlorophenylalanine POMC – Proopiomelanocortin SOS – Sodium octyl sulphate TRH – Thyrotropin releasing hormone TEL – Telencephalon TH – Tryptophan-5-hydroxylase TRP – Tryptophan 1 Introduction 1.1 The stress response Stress is a concept which is widely used to describe an animal’s physiological response to an unfavourable change in its environment (Henry & Stephens, 1977; Von Holst, 1988; Ladewig et al., 1993). This change is often referred to as a stressor. The identification of stressors can be difficult because there is a continuum ranging from mild everyday events to directly life threatening situations that has to be considered. The question is at which point in this continuum a change in an animal's environment becomes a stressor. Furthermore, in addition to unfavourable stressors (distress) there are also stressors that can be positive (eustress) for the animal. Examples of these are the appearance of a predator or a food source. In both situations the animal experiences a change in its physiological status, which prepares the animal to cope with the new situation. One could argue that a situation can be considered stressful on the basis of the physiological response to that situation, but this argument is circular in that the stress response must be defined by the situation experienced. This has been the source of many controversies regarding the definition of stress and stressors. According to Selye (1952) stress can be defined as “a non-specific response of the body to any demand made on it”. This definition sets the lower limit for what is to be considered a stressor very low. But it doesn't say that all changes are stressors and hence the circular argument is not avoided. In addition to this Selye was wrong about the stress response being non-specific. The reason for this can be found in the following. When an animal is exposed to a challenge that has a negative effect on its fitness this will cause the animal to respond with a combination of behavioural, neuroendocrine and autonomic changes that aim to reduce the adverse effect of that challenge. The change in behaviour allows the animal to either escape or counter the challenge, while the autonomic and neuroendocrine response provides the animal with the resources needed to meet the demands of the altered behaviour as well as trying to maintain the physiological homeostasis during the aversive situation (Moberg, 1985). It is not difficult to imagine that the behavioural responses are as diverse as the stressors. At least a single behavioural pattern is not sufficient to meet the diversity of all possible challenges. The diversity of the behavioural response has to reflect diversity in the response of the neuroendocrine system. In fact, the neuroendocrine system responds in 7 patterns characteristic for each stressor (Mason, 1968). Ideally it would be better not to use the terms "stress"