Roles of brain monoamine neurotransmitters in agonistic behaviour and stress reactions, with particular reference to fish

0742-8413/93 $6.00 + 0.00 0 1993 Pergamon Press Ltd

Camp. Biochem. Physiol.Vol. 106C, No. 3, pp. 597-614, 1993

Printed in Great

Britain

MINI REVIEW

ROLES OF BRAIN MONOAMINE NEUROTRANSMITTERS IN AGONISTIC BEHAVIOUR AND STRESS REACTIONS, WITH PARTICULAR REFERENCE TO FISH SVANTEWINBERG and G&AN E. NILSSON Department of Zoophysiology, Uppsala University, NorbyvIgen (Fax 46-185-188-43) (Received 21 June 1993; acceptedfor

18 A, S-752 36 Uppsala, Sweden

publication 30 July 1993)

Abstract-1. Experimental results on the involvement of brain monoamines in agonistic behaviour and stress in fish are reviewed and discussed in relation to available data from other vertebrates. 2. In fish as well as mammals, stress induces increased brain serotonergic activity, and a similar increase in serotonergic activity is seen in subordinate individuals in a dominance hierarchy. 3. The brain serotonergic system appears to inhibit aggression and spontaneous locomotor activity in both fish and mammals. 4. Subordinate fish show several behavioural characteristics, notably inhibition of aggressive behaviour, low spontaneous locomotor activity and decreased food intake, that are likely to be related to their increased brain serotonergic activity. 5. By contrast, the brain dopaminergic system appears to stimulate aggressive behaviour in both fish and mammals, and dominant fish show signs of elevated dopaminergic activity in telencephalon.

6. The similarities between fish and mammalian monoaminergic functions suggest that these are phylogenetically very old mechanisms that have been conserved during the last 400 million years of vertebrate evolution.

INTRODUCTION Monoaminergic neurons compose a very small fraction of the neurons in the vertebrate brain (e.g. Cooper et al., 1986). In fact, monoaminergic neurons number in the thousands whereas the total quantity of neurons in the vertebrate brain numbers in the hundreds of millions or more. However, the influence of monoaminergic neurons on their target sites appears to go far beyond their numbers. In the mammalian brain, where the monoaminergic systems have been most extensively studied, monoaminergic neurotransmitters are believed to be involved in the control of several behavioural patterns, notably aggression (e.g. Mason, 1984; Miczek and Donat, 1989; Olivier et al., 1989), mating (e.g. Meyerson and Malmnb, 1978) and feeding (e.g. Leibowitz, 1992). Monoaminergic systems in the brain have also been connected to stress reactions (e.g. Dunn, 1989) as well as to the central regulation of autonomic and neuroendocrine functions (e.g. Tuomisto and Mlnnistii, 1985). Moreover, human diseases, including schizophrenia, depression and Parkinsons disease, seem to be more or less directly related to unbalance or malfunction of brain monoaminergic systems (e.g. Mason, 1984). In comparison to the wealth of information, although sometimes inconclusive, available on brain monoaminergic functions in mammals, very little is known about the function of monoamine neurotransmitter systems in non-mammalian vertebrates.

The present paper focusses on the function of brain monoamines in fish behaviour. Studies of behavioural roles of brain monoamines in fish will not only increase our knowledge of how fishes function. Fishes deviated from other vertebrates during the Devonian period, 400 million years ago, so comparing monoaminergic mechanisms in the behaviour of fish and mammals will give new perspectives on the age of monoamine function in vertebrates. Indeed, several similarities between monoaminergic mechanisms in fish and mammalian behaviour, discussed in this paper, suggest that these functions have been conserved during vertebrate evolution and are phylogenetically very old. It should also be pointed out that a better understanding of monoaminergic mechanisms involved in fish behaviour could be of particular importance for fish culture. The formation of dominance hierarchies, which will be in focus of the present review, involves both stress and aggression, and is probably responsible for considerable production losses in many fish culturing systems.

THE MONOAMINERGICSYSTEMS OF THE BRAIN

The monoamine neurotransmitters include the catecholamines, dopamine (DA), norepinephrine (NE) and epinephrine (E), and the indoleamine serotonin (S-hydroxytryptamine, S-HT). 597

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this enzyme, little or no 5-HTP can normally be detected in 5-HT neurones. Catecholamines are synthesized from the amino acid tyrosine which is taken up from the blood by the same carrier as for tryptophan. The first and rate limiting step in catecholamine synthesis is the hydroxylation of tyrosine to 3,4_dihydroxyphenylalanine (L-DOPA) by tyrosine hydroxylase (TYH), a soluble enzyme found in the cytosol of all catecholamine synthesizing cells (Fig. 1). L-DOPA is subsequently decarboxylated by AAD to form DA. The conversion of L-DOPA to DA is very efficient and only low levels of L-DOPA occur in catecholaminergic neurons. Like 5-HT, DA is concentrated in vesicles. In NE neurons, these vesicles contain dopamine-/I-hydroxylase (DBH) which synthesizes NE from DA (Fig. 1). In E containing cells, NE is methylated by the enzyme phenylethanolamine-N-methyltransferase (PNMT) to form E (Fig. 1). Monoamines, like neurotransmitters in general, are concentrated and stored in vesicles which release their content into the synaptic cleft by exocytosis when the presynaptic membrane depolarizes (Fig. 2). The

We will start this review by providing some basic information on monoamine neurochemistry and distribution, since some knowledge in this field is helpful for the interpretation of the experimental data on monoamines and behaviour that will subsequently be discussed. Metabolism

Monoamine metabolism appears to be. qualitatively identical in all vertebrates. The indoleamine 5-HT is synthesized from the amino acid tryptophan which is taken up from the blood by means of a non-specific carrier that transports large neutral amino acids across the blood-brain barrier. The initial and rate limiting step in 5-HT synthesis is the hydroxylation of tryptophan to S-hydroxytryptophan (5-HTP), catalyzed by tryptophan-Shydroxylase (TH) (Fig. 1). TH is only found in 5-HT synthesizing cells, and its activity is restricted by tryptophan availability. 5-HTP is decarboxylated to 5-HT by aromatic L-amino acid decarboxylase (AAD), a nonspecific decarboxylase with a wide distribution throughout the body. Because of the high activity of

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Fig. 1. Pathways of monoamine metabolism. See the text for further details. TH: tryptophan hydroxylase; AAD: aromatic L-amino acid decarboxylase; MAO: monoamine oxidase; ALDH: aldehyde dehydrogenase; ALRD: aldehyde reductase; TYH: tyrosine hydroxylase; DBH: dopamine-/?-hydroxylase; PNMT: phenylethanolamine-IV-methyltransferase; COMT: catechol-O-methyl transferase.

Monoamine neurotransmitters

Fig. 2. A monoaminergic synapse. Monoamine neurotransmitters are concentrated in vesicles (1) which are emptied by exocytosis (2) when the presynaptic membrane depolarizes. Following release into the synaptic cleft, the monoamine can activate post- (3) and pre-synaptic (4) receptors. The actions of a released monoamine are terminated by uptake (5) into the presynaptic nerve terminal. Following uptake, monoamines either enter vesicles (6) or are metabolized, e.g. deaminated by MAO, localized on the outer membrane of the mitochondria (7).

effects of monoamine neurotransmitters are terrninated by uptake into presynaptic nerve terminals and possibly also glial cells (Hansson, 1983; Katz and Kimelberg, 1985). Following uptake, monoamines are deaminated to their corresponding aldehydes by monoamine oxidase (MAO, Fig. 1), an enzyme located on the outer membrane of mitochondria (Fig. 2). Aldehydes originating from the deamination of catecholamines are efficiently converted by aldehyde dehydrogenase (ALDH) to the corresponding acids or by aldehyde reductase (ALRD) to form alcohols (Fig. 1). Similarly, ALDH rapidly converts 5-hydroxyindoleacetaldehyde, the aldehyde formed by deamination of 5-HT, to S-hydroxyindoleacetic acid (5-HIAA, the main metabolite of 5-HT). The alcohol 5-hydroxytryptophol (5-HTOH) formed by ALRD appears to be a minor 5-HT metabolite in vertebrates (Fig. 1). Alternatively, catecholamines may be inactivated by methylation of the m-hydroxyl group, a reaction catalyzed by catechol-O-methyl transferase (COMT). This relatively non-specific enzyme, which has been suggested to be localized mainly extraneuronally, methylates catecholamines and various catechol compounds, including DA and 3,4_dihydroxyphenylacetic acid (DOPAC), a primary DA metabolite (Fig. 1). Methylated catecholamines may be deaminated and converted to corresponding acids or alcohols by MAO and ALDH or ALRD, respectively (Fig. 1). There seems to be no single dominating DA metabolite in fishes (Dulka et al., 1992). Nilsson (1989, 1990a) reported that homovanillic acid (HVA) was the major DA metabolite in crucian carp (Carassius carassius), whereas DOPAC seems to be the predominant DA metabolite in goldfish (Sloley et al., 1992; Dulka et al., 1992). Saligaut et al. (1990) suggested that 3-methoxytyramine (3-MT) was

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the major DA metabolite in rainbow trout (Oncorhynchus mykiss), while Saligout et al. (1992) reported that the concentration of DOPAC exceeded that of 3-MT in rainbow trout hypothalamus and pituitary. The reason for the apparent discrepancies between studies on DA metabolism in fish is unclear, even if it could, at least in part, be related to species-specific differences in DA catabolism. Differences in the analytical techniques used to measure DA and its metabolites are other possible explanations. It is also likely that the rate of conjugation and/or clearance of DA metabolites from the brain differ between, or even within, species. For instance, in Arctic chart-, HVA was the only DA metabolite detected by Winberg et al. (1991) whereas only DOPAC was detected in another study on this species (Winberg and Nilsson, 1992). The Artic charr used in these two studies were from the same population, although the fish used in the study by Winberg et al. (1991) were older and larger (mean weight 130 f 40 g) than the ones used by Winberg and Nilsson (1992) (mean weight 18 + 4 g). For a more detailed discussion on monoamine metabolism, the reviews by Boadle-Biber (1982), Westerink (1985), Green (1989), Weiner and Molinoff (1989) and Fillenz (1990) are recommended. Distribution The organization of monoaminergic systems, especially that of 5-HT, seems to be remarkably constant throughout the vertebrate subphylum (Parent et al., 1984). For example, a typical feature of vertebrate monoaminergic systems is a concentration of 5-HT cellbodies to the midline raphe region (Parent, 1981; Parent et al., 1984; Takeuchi, 1988; T&k, 1990; Jacobs and Azmitia, 1992). Catecholaminergic cells, on the other hand, are more abundant in the lateral portion of the brain stem (Parent et al., 1984). The 5-HT (Takeuchi, 1988; T&k, 1990; Jacobs and Azmitia, 1992) and NE (Fillenz, 1990) systems of the brain could be described as diffuse systems with extremely divergent projection patterns. Furthermore, not all serotonergic (T&k, 1990; Jacobs and Azmitia, 1992) and noradrenergic fibers (Fillenz, 1990) make classical synaptic connections. NE (Fillenz, 1990) and 5-HT (Jacobs and Azmitia, 1992) released from such non-junctional fibers might function as neuromodulators by diffusing in the extracellular space to act on distant cellular targets. In teleosts, DBH immuno-reactive (DBH-IR) cell bodies, presumptive NE synthesizing neurons, occur in three areas of the hind brain, the dorsomedial medulla, the medullary tegmentum and the isthmal tegmentum, whereas DBH-IR cell bodies appear to be absent in all other brain areas (Hornby and Peikut, 1990; Ekstriim et al., 1992a). Similarly, in the mammalian brain noradrenergic cell bodies are localized in the brain stem, where they occur in several separate groups (Fillenz, 1990). However, in fish as well as in

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mammals, the brain stem noradrenergic neurons project to most parts of the neuroaxis, with a particular dense innervation of hypothalamus and telencephalon. In mammals, serotonergic cell bodies are mainly confined to brain stem raphe nuclei and have axonal terminals reaching virtually all regions of the central nervous system (Takeuchi, 1988; Tork, 1990; Jacobs and Azmitia, 1992). The distribution of serotonergic cell bodies appears to be similar in the teleost brain (Kah and Chambolle, 1983; Yoshida et al., 1983; Parent ef al., 1978, 1981, 1984; Frankenhuis-van den Heuvel and Nieuwenhuys, 1984; Bonn and Konig, 1990; Bolliet and Ali, 1992; Ekstriim et al., 1992b). However, in fish, numerous 5-HT immunoreactive (5-HT-IR) cells are also found outside the raphe region, especially in ventral thalamic and hypothalamic areas. In the hypothalamus, a population of these 5-HT-IR cells is in direct contact with the cerebrospinal fluid (CSF) in the third ventricle (Parent, 1981; Parent et al., 1984). The brain dopaminergic systems seem to be more complex in their organization. DA containing cell bodies are found in larger numbers and are gathered in several major nuclei with a more topographic projection (Shepherd, 1988). Furthermore, some specialized DA neurons, for example in the olfactory bulb, make only local connections (Shepherd, 1988). In the fish brain, TYH immuno-reactive (TYH-IR) cell bodies, presumptive DA synthesizing neurons, occur together with DBH-IR cell bodies in the hind brain (Hornby and Peikut, 1990; Sas et al., 1990; Ekstriim et al., 1992a). In addition, presumptive dopaminergic cell bodies are also found in the olfactory bulbs as well as in several areas of the telencephalon and diencephalon (Hornby et al., 1987; Sas et al., 1990; Ekstriim et al., 1992a). In the goldfish (Carassius auratus) telencephalon, the largest group of TYH-IR cell bodies is found in the central zone of area dorsalis, and in diencephalon a large group of TYH-IR cell bodies occurs in the preoptic area (Hornby et al., 1987). Furthermore, a considerable number of catecholaminergic cell bodies is found in the hypothalamus (Parent et al., 1984). Some of these cells are small in size and have club like apical processes reaching into the CSF in the third ventricle (Yoshida et al., 1983; Parent et al., 1978, 1984). Like in many othsr vertebrates, E is only found at very low concentrations in the teleost brain (Pennypacker et al., 1985; Hornby and Peikut, 1988; Nilsson, 1989), and has, as of yet, no known functions there. Receptors As with many neurotransmitters, it is becoming increasingly clear that monoamines activate a great variety of receptor subtypes. Monoamine function may gain specificity by differences in the distribution of receptor subtypes in brain. Furthermore,

monoaminergic neurotransmission is probably modulated by heterogeneity of receptor distribution on the post- and pre-synaptic surfaces (Fig. 2). However, very little is known about monoamine receptor subtypes in the teleost brain. With regard to NE four receptor subtypes, ai, tag, /I, and p2 have been described in the mammalian brain (Fillenz, 1990). Pharmacological evidence suggests that the stimulatory effect of NE on gonadotropin (GtH) release from goldfish pituitary cells is mediated by a,-like adrenergic receptors (Chang et al., 1991). The effects of DA in the mammalian brain are mediated by at least two different DA receptors types, D, and Dr (Clark and White, 1987; Creese, 1987). Concerning DA receptors in fish, Chang et al. (1984) found that the inhibitory effect of DA on GtH secretion in goldfish is mediated by receptors resembling the D, receptor in mammals. The serotonergic system shows the most considerable receptor divergence. At least three major 5-HT receptors families, 5-HT,, 5-HT, and 5-HT,, are found in the mammalian brain (Leff and Martin, 1988; Peroutka, 1988, 1990; Frazer et al., 1990; Giithert, 1992). The 5-HT, receptor family presently consists of at least three receptor subtypes, 5-HT,,, 5-HT,, and 5-HT,,. The 5-HT, receptor family includes two receptor subtypes, 5-HT, and 5-HT,, (Leff and Martin, 1988; Peroutka, 1988, 1990; Frazer et al., 1990; Gothert, 1992). Consequently, postsynaptic effects of monoamine release will depend on the distribution of individual receptor subtypes in different projection areas. In a comparative study, Palacios and Diet1 (1988) found presumptive 5-HT,, sites, but no 5-HT,, or 5-HT,, sites, in the fish brain. Furthermore, Hensley and Cohen (1992) obtained pharmacological evidence for the presence of 5-HT,, receptors in the goldfish retina. Burka et al. (1988) reported that the non-selective 5-HT, receptor antagonist methysergide inhibited 5-HT induced contraction of rainbow trout intestine, indicating the presence of peripheral 5-HT, receptors in fish. Recently, Bakker et al. (1993) obtained results indicating the presence of a previously unknown 5-HT receptor subtype in the goldfish intestine. This receptor stimulates adenylate cyclase activity and seems not to be identical with any previously described 5-HT receptor. 5-HT,, and 5-HT,, receptors appear to be missing in amphibian and reptile brains but occur in birds (Palacios and Dietl, 1988). BRAIN MONOAMINES

AND AGONISTIC

BEHAVIOUR

Roles of central catecholaminergic systems in agonistic behaviour In dominance hierarchies, which will be discussed in more detail in subsequent sections, agonistic behaviour plays a central role. The DA as well as the

monoamine neurotransmitters

NE system of the mammalian brain are believed to be involved in the regulation of agonistic behaviour (Mason, 1984), and it was early postulated that some DA systems may function as excitatory systems for intraspecies aggression (Avis, 1974). Central catecholaminergic activity has been related to agonistic behaviour also in fish, Munro (1986a) suggested a role of DA in the regulation of aggressive behaviour in blue acara (Aequidens p&her), although he found that both a DA agonist (apomorphine) and a DA antagonist (chlorpromazine) caused reduced aggression, Maler and Ellis (1987) reported that intraventricular injections of DA had powerful effects on inter-male aggressive signaling in the weakly electric fish, Apteronotus leptorhy~chus. However, both enhancing and suppressing effects of intraventricular DA injections on aggressive signaling were recorded in their study. By contrast, Tiersch and Griffith (1988) observed increased aggressive behaviour in rainbow trout given apomorphine, although no quantitative data were presented. The discrepancy between the results obtained by Tiersh and Griffith (1988) and the results by Munro (1986a) might be related to the dose of apomorphine used. Munro (1P&a) added apomorphine to the aquarium water, giving a final concentration of 0.3-3 mg/l, whereas Tiersch and Griffith administered apomorphine by i.p. injections in doses of 3&120mg/kg. Apomorphine is a nonselective DA receptor agonist, acting on both D, and D, receptors, although it shows higher affinity for the D, receptor. Presynaptic DA autoreceptors are believed to be of the D, type (Clark and White, 1987) and low doses of apomorphine (0.1-0.5 mg/kg) have been reported to decrease DA release in the mammalian brain (Cabib and Puglisi-Allegra, 1991). However, it is now becoming increasingly clear that D, and D, receptors can interact in both opposing and synergistic fashions (reviewed by Clark and White, 1987; Blackburn et al., 1992). In cats, apomorphine facilitates predatory attack behaviour (Shaikh et al., 1991) as well as effective defense behaviour (Sweidan et al., 1991). In both these studies, the effect of apomorphine appears to be mediated by Dz receptor stimulation (Shaikh et al., 1991; Sweidan et al., 1991). It has also been shown that spontaneously aggressive rats have higher DA levels in hypothalamus than non-aggressive conspecifics (Barr et af., 1979). Furthermore, fighting increases the concentration of HVA in the brain of mice, without affecting DA levels (Modigh, 1973). However, it has been questioned if the fighting related increase in DA metabolites is related to aggression or just reflects a stress induced increase in brain dopaminergic activity (Hutchins et al., 1975; PuglisiAllegra and Cabib, 1990). The central NE system has also been connected with agonistic ~haviour, although in mammals, increased brain norepinephric activity seems to relate mainlv to stress (Mason. 1984‘1. I However. in the

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weakly electric fish, Apteronotus leptorhynchus, intracranial injections of NE cause strong and consistent enhancement of the inter-male aggressive signaling (Fig. 3) (Maler and Ellis, 1987). Roles of the central serotonergic system in agonistic behav~our The serotonergic system of the brain is believed to be involved in the regulation of agonistic behaviour among diverse animal groups, and in most species increased serotonergic activity appears to have an inhibitory effect on aggressive behaviour (Avis, 1974; Huntingford and Turner, 1987; Miczek and Donat, 1989; Olivier et ai., 1989). For example, Munro (1986b) found that intracranial injections of S-HT reduced aggression in the cichlid, Aequidens p&her. However, the effect was abolished if S-HT was administered together with S-adenosyl homocysteine, which inhibitsjhe conversion of 5-HT to melatonin, indicating melatonin as the anti-aggressive agent. Maler and Ellis (1987) reported that intra-cranial injections of S-HT greatly reduced aggressive signaling in electric fish (A~teronotus ~eptorhynch~s) males (Fig. 3), an effect which they found likely to be caused by 5-HT itself as the S-HT up-take inhibitor, fluoxetine, had a similar effect. Pharmacological stimulation of the serotonergic system of the mammalian brain has repeatedly been found to inhibit aggression (see Olivier et al., 1989 for a review). For instance, pharmacological studies have shown that several 5-HT receptor agonists, such as eltoprazine, RU 24949 and TFMPP, specitically inhibit offensive aggression in rodents (Olivier et at., 1989; Mos et ai., 1992). Furthermore, pharmacological stimulation of the brain serotonergic system also decreases aggressive reactions in pigeons (Fachinelli 6

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Time (mid Fig. 3. Time course of monoamine effect on the fresuencv of the inter-male aggressive signal (chirping) in the weakly electric fish Apteronotus leptorhynchus. NE, S-HT or vehicle was injected at time 0. Values are normalized with respect to the mean standard deviation of the six preinjection values. Thus, normalized chirps are given as units of stan_.... ..~ dard deviation (preuqection) from the preinjection mean.

Modified from Mafer and Ellis (1987).

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et al., 1989). In mammals, stimulation of the central serotonergic system also inhibits predatory aggression. In rats, electrical stimulation of raphe nucleus as well as administration of the S-HT receptor agonist, quipazine, or the selective S-HT re-uptake inhibitor, CGP 6085-A, inhibit predatory aggression (Kostowski et al., 1980). Similarly, in the mink (Mute/a uison), stimulation of brain serotonergic activity, by administration of S-HTP, the endogenous precursor of 5-HT, decreases predatory attacks (Nikulina and Popova, 1988). Inhibition of brain serotonergic activity, on the other hand, seems to produce variable effects on aggressive behaviour in mammals. Treatment with p-CPA, a S-HT synthesis inhibitor, increases aggressive behaviour in rats @heard, 1969; Matte and Tornow, 1978; Vergnes et al., 1986). Similarly, rats with surgical (Yamamoto and Ueki, 1977) or chemical (Ellison, 1976; Vergnes et al., 1988) lesions of the brain 5-HT system show increased aggressive behaviour. However, in several mammalian studies, pharmacological inhibition of serotonergic activity has been found to be ineffective or reduce various forms of intra-specific aggressive behaviour (reviewed by Miczek and Donat, 1989; Miczek et al., 1989). By contrast, predatory aggression seems to be more consistently increased by inhibition of serotonergic activity (Miczek and Donat, 1989; Miczek et al., 1989). It must be pointed out that agonistic behaviour is a complex collection of behavioural acts which are unlikely to be controlled by any single neurotransmitter system. Thus, contradictory results on the role of 5-HT, and other neurotransmitters, in the control of agonistic behaviour might be related to the diverse forms of aggression studied (predatory aggression, social aggression, isolation induced aggression, etc.) and the different behavioural variables measured. Behavioural roles of 5-HT are further complicated by the considerable receptor diversity displayed by the serotonergic system. Activation of different 5-HT receptor subtypes may have opposing effects on various behavioural acts. This makes many studies hard to interpret as the 5-HT agonists and antagonists most widely used are relatively unselective. DOMINANCE HIERARCHIES

In many fish species, the social organization is characterized by the development of a dominance based social hierarchy, a social structure where agonistic behaviour is an important component. The hierarchy, which is established through pairwise encounters, often tends to be essentially linear (i.e. each individual dominates all others with ranks lower than itself), at least in small groups of fish. This kind of relationship is likely to occur when the outcome of an encounter between two individuals mainly depends on their fighting ability (Huntinford and Turner, 1987). In fish, size is likely to be a major factor

determining individual fighting ability. Furthermore, in a group of fish, high ranking individuals constantly consolidate their social position by a much faster growth rate than subordinate fish (Abbott and Dill, 1989; Winberg et al., 1992a). However, previous social experience also affects the competitive ability of a fish. Experience of a low social position seems to inhibit aggressive behaviour, whereas fish previously occupying high social positions show enhanced aggression (Abbott et al., 1985; Wallen and Wojciechowski-Metzlar, 1985; Nelissen and Andries, 1988; Winberg et al., 1992a). Abbott et al. (1985) showed that the established dominance relationships in pairs of juvenile steelhead trout (Oncohynchus mykiss) could not be reversed by providing the subordinate fish with more food than the corresponding dominant individual. Indeed, in no case did the subordinate fish gain dominance, even if, at the end of the experiment, it had grown to become considerably larger than its opponent. This kind of social modulation of agonistic behaviour, and conservation of hierarchic positions, could be mediated by the brain monoaminergic systems. Social rank, stress and brain monoaminergic

activity

Social experience has been found to greatly affect brain monoaminergic activity in fish. For example, subordinate fish display a general increase in brain 5-HT activity, a stress related effect, which might at least in part explain the inhibition of competitive behaviour in these individuals. Dominant fish, on the other hand, show increased brain DA activity and it is tempting to interpret this as a reflection of the enhanced aggressive behaviour displayed by these individuals. Catecholamines

Effects of social rank on catecholamine levels in the fish brain were first reported by McIntyre et al. (1979). They found that dominant rainbow trout had lower NE and higher DA concentrations in the brain than subordinate fish. Furthermore, they reported that the subordinate fish most frequently attacked showed the greatest reduction in DA levels. Increased brain DA levels in dominant fish could be related to increased aggression in these individuals, while higher concentrations of NE in the brains of subordinate fish might indicate a stress related increase in noradrenergic activity. Prolonged exposure to stressful situations has been reported to coincide with increased brain levels of NE in mammals (Adell et al., 1988). However, the fact that McIntyre et al. (1979) did not measure catecholamine metabolite levels makes their results hard to interpret, as catecholamine levels per se are poor sources of information on the activity of the catecholaminergic systems. For example, a decreased steady state level of a monoamine can be interpreted in two diametrically different ways: (1) that the neurotransmitter is being more extensively used; or (2) that the neurotransmitter synthesis is

Monoamine neurotransmitters being down regulated due to a decreased utilization of the system. The effect of social rank on brain monoamine metabolite levels has been studied in Arctic charr (Saluelinus alpinus). In this species, socially dominant individuals have been found to have higher concentrations of the DA metabolite HVA in telencephalon than subordinate fish (Winberg et al., 1991). The difference in telencephalic HVA concentration between dominant and subordinate individuals, might be related to differences in aggressive behaviour, since telencephalon has been implicated in the regulation and integration of agonistic behaviour in fish (de Bruin, 1980). This hypothesis was further supported by the finding that pre-treatment with L-DOPA (levodopa, 10 mg/g) greatly increased the chance (from 50 to 80%) of a juvenile Arctic charr to become dominant over a size-matched opponent during their first encounter (Winberg and Nilsson, 1992). LDOPA is the immediate precursor of dopamine and L-DOPA treatment will increase the rate of dopamine synthesis and thereby stimulate the activity of dopaminergic systems in the brain. Indeed, brain levels of DA and DOPAC showed a dose dependent increase in L-DOPA treated fish (Winberg and Nilsson, 1992). It should be mentioned that DA is the precursor also of NE, so L-DOPA treatment may indirectly elevate the synthesis of NE. However, brain levels of NE were unaffected by L-DOPA treatment in this experiment. The blood volume of the fish studied was too small to allow catecholamine analysis. Thus, an involvement of peripheral catecholamines could not be excluded. Nechayev and Musatov (1992) showed that peripheral administration of catecholamines increased aggressive behaviour in dominant rainbow trout, and induced threatening poses in subdominant individuals. In these experiments, DA turned out to be the most effective catecholamine, while E was least effective (Nechayev and Musatov, 1992). Furthermore, when DA injections were given prior to hierarchy formation, the DA treated fish always became dominant (Nechayev and Musatov, 1992). These results seem to indicate that peripheral catecholamines affect aggressive behaviour in rainbow trout. However, it appears likely that behavioural effects observed after peripheral administration of catecholamines are exerted by the central nervous system. In fish, the blood-brain barrier has been found to be permeable to NE (Busacker and Chavin, 1977; Nekvasil and Olson, 1986). Unfortunately, DA has not been examined in this respect. 5-HT Social experience has striking effects on the level of 5-HIAA in the fish brain. Juvenile Arctic charr occupying low positions in a dominance hierarchy display increased concentrations of 5-HIAA in telencephalon, hypothalamus and brain stem (Winberg et al., 1991, 1992a; Winberg and Nilsson, 1993). The 5-HIAA/S-HT ratio, an index of brain serotonergic

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activity (Shannon et al., 1986), is also increased in these brain areas of subordinate individuals (Fig. 4). Brain 5-HT levels, on the other hand, are not affected by subordinate experience (Winberg et al., 1991, 1992a, 1993a; Winberg and Nilsson, 1993). Thus, brain 5-HIAA/S-HT ratios (Winberg et al., 1992a) and 5-HIAA concentrations (Winberg et al., 1991)

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Fig. 4. A-C 5-HIAA/S-HT ratios in (A) telencephalon, (B) hypothalamus and (C) brain stem of Arctic charr having 1, 3, 7 or 21 days experience of a dominant or subordinate position in a pair. Fish that had been isolated for one week were used as controls. Values are means and SEM from 7-10 individuals. Asterisks denote significant differences between dominant and subordinate individuals whereas asterisks in parenthesis denote significant differences from controls. *P < 0.05, **P -c 0.01, ***P < 0.001 MannWhitney U-test (two tailed). From Winberg and Nilsson (1993).

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S. WINBERGand G. E. NLS~ON

are, in this species, inversely correlated with social rank. This relationship between brain 5-HT utilization and social rank in Arctic charr seems to be socially induced and not caused by innate differences in brain serotonergic activity predisposing fish for dominant or subordinate positions in a dominance hierarchy (Winberg et al., 1992a). Furthermore, the increase in brain serotonergic activity indicated in subordinate fish has a rapid onset and is clearly apparent already after 1 day of exposure to a dominant conspecific (Fig. 4) (Winberg and Nilsson, 1993). Both stress (Bliss et al., 1972; Curzon et al., 1972; Morgan et al., 1975; Morgan and Rudeen, 1976; Adell et al., 1988; Mitchell and Thomas, 1988) and food deprivation (Curzon et at., 1972; Kantak et al., 1978; Fuenmayor and Garcia, 1984) are known to activate the serotonergic system of the mammalian brain, as indicated by increased brain 5-HIAA levels. Fish occupying low positions in a social hierarchy are likely to be subjected to both food deprivation and stress, caused by repeated attacks from high ranking fish as well as by the constant threat imposed by the sheer presence of dominant individuals. Thus, not surprisingly, subordinate fish show many of the physiological signs of stress (Noakes and Leatherland, 1977; Ejike and Schreck, 1980; Peters et al., 1980; Scott and Currie, 1980) as well as reduced food intake (Abbott and Dill, 1989; McCarthy et al., 1992a, 1992b). So how do stress and starvation per se affect the 5-HT system in fish? In Arctic charr, a 4 week period of daily artificial stress increases brain S-HIAA concentrations and 5-HIAA/S-HT ratios (Fig. 5), while starvation per se has no effect on brain 5-HT utilization (Winberg et al., 1992b). Consequently, stress is likely to be a factor underlying the signs of increased brain serotonergic activity seen in subordinate fish while food deprivation, on the other hand, probabIy can be ruled out as a contributing factor in this context. In mammals, exposure to stressful episodes is known to increase brain tryptophan levels (Curzon et al., 1972; Neckers and Sze, 1975; Dunn, 1988; Dunn and Welch, 1991). This may affect the function of serotonergic systems, since tryptophan is the amino acid precursor of 5-HT and the rate of 5-HT synthesis is normally restricted by tryptophan availability in both mammals (Boadle-Biber, 1982) and fish (Johnston et al., 1990, 1992). Furthermore, tryptophan administration has been found to increase brain 5-HIAA levels in mammals (Fernstrom, 1987) although the physiological significance of this increase has been questioned (Grahame-Smith, 1971; Lookingland et al., 1986; De Simoni et al., 1987). The increase in brain 5-HIAA levels observed after tryptophan administration could, for example, reflect an increase in intra-neuronal 5-HT catabolism and not an increase in functional 5-HT release (GrahameSmith, 1971; Lookingland et al., 1986; De Simoni et al., 1987).

Al

0.5

Bl 0.5

Fig. 5. A,B. Effect of stress or starvation during 4 weeks on 5-HIAA/S-HT ratios in telencephalon (A) and brain stem (B) of Arctic charr. Stress was imposed on the fish (3 x 15 mitt daily) by forcing the fish close to the surface by lifting up a net pIaced on the bottom of the aquaria. Neither stressed fish nor controls were fed during the stress experiment. Values are mean and SEM from 8 individuals. *P <: 0.05, ***P < 0.001 Mann-Whitney U-test (two tailed). Modified from Winberg et al. (1992b).

At least in the Arctic charr, it appears clear that the indications of increased serotonergic activity in subordinate individuals are not directly related to increased tryptophan levels. In pairs of Arctic charr, 1 day of social interaction causes an increase in tryptophan levels in telencephalon, hypothalamus and brain stem of both the dominant and subordinate individual (Fig. 6) (Winberg and Nilsson, 1993). However, after longer periods of pair rearing, tryptophan levels fall, and brain tryptophan concentrations were, after 3 days of social interaction, found to be back to control levels in all three brain areas of dominant and subordinate fish (Fig. 6). In fact, the decrease in telencephalon and hypothalamus tryptophan concentrations were more pronounced in subordinate individuals than dominant fish (Winberg and Nilsson, 1993). By contrast, brain 5-HIAA/S-HI’ ratios, which were increased in telencephalon, hypothalamus and brain stem of subordinate, but not dominant, individuals already after I day, remained high also after 21 days of exposure to a dominant conspecific (Fig. 4). Furthermore, in this and other experiments, 5-HIAA leveis or 5-HIAA/S-HT ratios

Monoamine neurotransmitters TELENCEPHALON o control n dominant q subordinate

Al

71

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B’

10

5

15

20

HYPOTHALAMUS 16

1

14 12 10 II64-

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02

0

5

10

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BRAIN STEM

Cl 6

0

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10

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days together

Fig. 6. AC Concentrations of tryptophan (TRP) in (A) telencephalon, (B) hypothalamus and (C) brain stem of Arctic charr having 1, 3, 7 or 21 days experience of a dominant or subordinate position in a pair. Fish that had been isolated for one week were used as controls. Values are means and SEM from 610 individuals, except for hypothalamus of fish being subordinate for 21 days where n = 4. Asterisks denote significant differences between dominant and subordinate individuals whereas asterisks in parenthesis denote significant differences from controls. *P -c 0.05 Mann-Whitney U-test (two tailed). From Winberg and Nilsson (1993).

did not correlate with tryptophan levels in any of the brain areas examined (Winberg and Nilsson, 1993; Winberg et al., 1993b). Consequently, the increase in brain S-HIAA concentrations and brain S-HIAA/ 5-HT ratios observed in subordinate Arctic charr are

and behaviour

605

likely to indicate an increase in functional release of 5-HT and not just a tryptophan related increase in intraneuronal 5-HT catabolism. Dunn and Welch (1991) argued that the increase in brain tryptophan concentration observed in connection with stress could counteract depletion of existing 5-HT stores during a stress induced elevation of 5-HT release. However, the rate of 5-HT synthesis might be rapidly adjusted by mechanisms (such as phosphorylation) acting on TH activity per se, in order to maintain 5-HT levels during periods of increased 5-HT use. In fact, a rapid increase in brain TH activity has been observed in rats exposed to sound stress (Azmitia and McEwen, 1974; Beadle-Biber et al., 1989). A stress-induced elevation of brain tryptophan levels could, of course, further facilitate such an increase in 5-HT synthesis in response to a rise in 5-HT release. We recently obtained indirect evidence for a stress induced increase in TH activity in Arctic charr. In this experiment, the effect of stressful experiences on whole brain 5-HT turnover was studied in Arctic charr by measuring the accumulation of 5-HTP after inhibition of AAD by N-m-hydroxybenzylhydrazine (NSD 1015) (Curzon, 1981). This is a widely used technique to study brain 5-HT turnover that has the advantage that the increase in 5-HTP is measured against a very low background level of 5-HTP. To avoid effects of social interaction, fish were isolated in individual compartments and stress was imposed by lifting the fish to the water surface (i.e. by mechanically elevating the substrate as described by Winberg et al., 1992b) three times a day during 5 days. Neither stressed fish nor controls, which were isolated in equivalent compartments, were fed during the experiment. The fish (held at 10°C) were treated with NSD 1015 (100 mg/kg, i.p.) 1 hr prior to sacrifice. Individual brains were analyzed for 5-HT, 5-HTP and tryptophan. Individual 5-HTP synthesis rates were calculated by subtracting the amount of 5-HTP found in whole brain of non NSD 1015 treated Arctic charr (0.22 + 0.13 nmol/g, mean f SD, n = 4) and a theoretical value for the whole brain 5-HT turnover time was subsequently obtained, for each individual, by dividing 5-HT concentrations with 5-HTP synthesis rates. The results from this study showed that exposure to stressful episodes significantly decreased the whole brain 5-HT turnover time (Fig. 7) i.e. increased the turnover rate. Brain tryptophan levels, on the other hand, did not differ between stressed fish and controls (Fig. 7). Thus, the elevation in 5-HT turnover rate observed in stressed Arctic charr is likely to reflect a stress induced increase in TH activity. The 5-HT synthesis rate in unstressed controls was 0.49 f 0.22 nmol/g hr (mean + SD, n = 8) corresponding to a theoretical whole brain turnover time for 5-HT of 8.5 hr. Brain 5-HT turnover time in winter-acclimatized crucian carp (held at 8°C) estimated by the same method, ranged between 1 and 3

S. WINBER~ and

606

G. E. N~t.sso~

1 0

5-HT turnover time (A), estimated from S-HTP accumulation after NSD 1015 injection, and tryptophan concentrations (B) in Arctic charr (held at 10°C). Values are means & SEM from eight fish. + P < 0.05 Mann-Whitney U-test (two-tailed).

3

I

5

Fig. 7. The effect of stress on whole brain

days in different 5-HT turnover

MSM

12

15

1

18

(xl

Bl

I l +&_

r- -0.577 tr--0.517. S-0.0061 .

brain areas (Nilsson, 1990b). Thus, seems to be faster in Arctic charr than

in crucian carp, but it is still about S-6 times slower than in mammals (Curzon, 1981). (2i0 for brain metabolic rate has been estimated to be 2.1, a value that seems to be similar in ali vertebrates studied (Mink et ai., 1981). Thus, the apparent difference in 5-HT turnover rate between Arctic charr and mammals might be fully explained by the temperature difference. In the studies on Arctic charr discussed so far, small groups consisting of 24 individuals were used in order to make individual social rank assessable by behavioural observations. Thus, an interesting question was if a similar relationship between social rank and brain SHIAA/S-HT ratios existed in larger groups where the hierarchial structure might be more complicated. In rainbow trout, feeding hierarchies have been demonstrated in larger groups of fish using radiography to measure the individual food consumption rates (McCarthy et al., 1992a, 1992b). Thus, individual food intake, expressed as an individual’s mean share of group meals, seems to provide an index of individual social rank (McCarthy ef al., 1992a,b). Recently, the relationship between social position, assessed by mean share of group meal, and brain 5-HT utilization, has been studied in larger groups (15 individuals) of rainbow trout (Winberg et al., 1993b). The results from that study show that individual food intake and brain serotonergic activity, measured as brain 5-HIAA levels or 5HIAA/S-HT ratios, inversely correlate with each other, suggesting that both methods can be used as indicators of the positions of individual fish in a dominance heirarchy (Fig. 8). Furthermore, these results suggest that the relationship between brain serotonergic activity and social rank holds also in larger groups of salmonids. Is this relationship specific for salmonid fish or is it a general phenomenon occurring also in other species? Recently, the effect of social experience on brain 5-HT utilization was studied in bicolor

I

a

.

0.07

1. 0

I

3

6

9 MSM

1

12

15

18

(X)

Fig. 8. A,B. The relationship between mean share of meal (MSM) and 5-HIAA/S-HT ratios in telencephalon (A, n = 26) and brain stem (B, n = 28) of rainbow trout. Values and least square regression lines are original non-transformed data. Before correlation analysis the MSM data were subjected to arcsin transformation and r- and p-values for the correlation analysis on transformed data are given within parenthesis. Modified from Winberg et al. (1993b).

damselfish (Pomatocentms partitus). This is a common species on the coral reefs throughout the western Atlantic in which the formation of social hierarchies has been studied extensively (Myrberg. 1972a, 1972b). Interestingly, subordinate bicolor damselfish also seem to show a stress mediated increase of brain serotonergic activity. In fact, brain 5-HIAA/S-HT ratios in subordinate individuals correlated positively with the number of aggressive acts received (Winberg, My&erg and Nilsson, unpublished results). Furthermore, the effect of predator induced stress on brain 5-HIAA levels and brain 5-HIAA/5-HT ratios has also been studied in the bicolor damselfish (Winberg et al., 1993c). This study showed that brain S-HIAA concentrations and SHIAA/S-HT ratios were doubled in bicolor damselfish which, for 2 hr had been exposed to a graysby (Epinephelus cruentatus), a (Fig.

common

predator

in the damselfish

habitat

9). The same effect was seen in telencephalon,

hypothalamus

and brain stem. Thus,

predator-induced activity

seems

stress to

have

on

the

a wide

the effect of this

brain

serotonergic

distribution

in the

Monoamine neurotransmitters

cl

Control

q Exposed

to predator

l*

telencephrlon

hypothalamus

brain stem

Fig. 9. The effect of predator exposure on 5-HIAA/S-HT ratios in telencephalon, hypothalamus and brain stem of bicolor damselfish. Values are mean f SEM from eight fish. ** = P i 0.01, Mann-Whitney U-test (two tailed). Modified from Winberg et al. (1993~).

damselfish brain and very much resembles the effect of artificial stress on brain serotonergic activity in Arctic charr. In fact, the effect of social experience on brain serotonergic activity is not restricted to fishes. Increase in serotonergic activity in response to subordinate experience seems to be a general phenomenon occurring in several vertebrate species, including primates. For example, 5-HIAA levels are increased in limbic brain regions and the spinal cord of rats occupying low social positions in a colony (Blanchard et al., 1991). Furthermore, in talapoin monkeys, Yodyingyuad et al. (1985) found a relationship between social rank and brain serotonergic activity. In monkeys acquiring social dominance, 5-HIAA levels in cerebra spinal fluid (CSF) decreased, while they increased in individuals that became subordinate in the group. The differences in 5-HIAA levels were stable as long as the hierarchy remained unchanged, and short term fluctuations in S-HIAA concentrations did not correlate with day to day variations in agonistic behaviour. Consequently, Yodyingyuad et al. (1985) postulated that the increased 5-HIAA concentration in low ranking monkeys was, at least in part, a consequence of their social position, and not merely an effect of intercurrent aggressive interactions. In mammals, effects of social interactions and stress on brain serotonergic activity have also been studied by monitoring the spontaneous firing rate of 5-HT neurones in freely moving animals (reviewed by Jacobs and Fornal, 1991). For instance, by using telemetry, Walletschek and Raab (1982) were able to measure the spontaneous firing rate of 5-HT neurons in dorsal raphe nuclei (DRN) of freely moving treeshrews. In their study, they found that stress, induced by confrontation with the experimenter, as well as defensive encounters with a conspecific, increased the firing rate of DRN neurons, while offensive encoun-

and behaviour

607

ters with the opponent, on the other hand, decreased DRN neural activity. However, the effect of stress on DRN neuronal activity in cats is ambiguous. Heym et al. (1982) reported that strong acoustic stimuli enhanced DRN activity, while Wilkinson and Jacobs (1988), reported that neither loud white noise (100 dB) nor restraint or confrontation with a dog affected DRN activity in cats, despite the fact that all of these stressors induced typical behavioural and physiological stress responses. In rats, sound stress (Azmitia and McEwen 1974; Boadle-Biber et al., 1989) as well as electrical stimulation of DRN 5-HT neurons (Boadle-Biber et al., 1986) increase biochemical indices of 5-HT neuronal activity, TH activity and 5-HIAA/S-HT ratios. In a recent study Corely et al. (1992) obtained indirect evidence of a sound stress-induced elevation of DRN neuronal activity in rats. They showed that, infusion of gepirone, a specific 5-HT,, agonist, into the DRN blocks the sound stress-induced increase in TH activity, as well as the elevation of 5-HIAA/S-HT ratios. The 5-HT,, receptors in the DRN are believed to be somatodendritic autoreceptors (Giithert and Schlicker, 1990; Giithert, 1992) and the results by Corely et al. (1992) indicate that gepirone blocks the sound stress-induced increase in DRN activity by activating these autoreceptors. Possible functions of increased serotonergic activity in subordinate and stressed individuals The fact that stress and subordinate experience seem to increase brain serotonergic activity, as indexed by elevated brain 5-HIAA levels, in both mammals and fish, suggests that this is a phylogenetically very old response to stress that has been conserved during the vertebrate evolution. Of course, the conservation of this mechanism also suggests that it is functionally important, although its role remains largely unknown. We will point out here the possibility that one or more of the behavioural characteristics of subordinate (and stressed) fish could depend upon increased serotonergic activity. The central serotonergic system is believed to be involved in endocrine regulation, and 5-HT has been found to stimulate corticotropin (ACTH) release from the anterior pituitary (Tuomisto and Mannisto, 1985). Thus, the stress-induced increase in brain serotonergic activity might form a link between stressful experiences and elevated plasma cortisol levels. However, the wide distribution of the stress related increase in the brain serotonergic activity indicates that it serves a multitude of functions, apart from stimulating cortisol production. This is supported by results from experiments on rats showing differences in the responses of the ACTH system and 5-HT system. The increase in brain 5-HIAA levels, induced by subjecting rats to electric shock, shows no adaptation to repeated exposure to the same electric shock, while the corresponding ACTH response, on

608

S. WINBERGand G. E. NIL.%~N

the other hand, is reduced in rats with previous experience of chronic exposure to this stressor (Armario et al., 1989). In fact, chronic exposure to a stressful situation has been found to sensitize the serotonergic system of the mammalian brain (Adell et al., 1988). Armario et al. (1989) made the interesting suggestion that elevated brain serotonergic activity might be an important mechanism allowing the animal to cope with stress. It was early hypothesized that behavioural arousal is a joint function of serotonergic inhibition and catecholaminergic excitation (Brodie and Shore, 1957), and the idea that the serotonergic system plays a role in behavioural inhibition has thereafter gradually emerged (reviewed by Sourbie, 1986). In mammals, pharmacological blockade of serotonergic transmission produces a shift of behaviour towards facilitation of responding (Sourbie, 1986). For instance, lesions of median and dorsal raphe nuclei, as well as pharmacological depletion of 5-HT, cause a pronounced increase in startle response in mammals, whereas intraventricular infusion of 5-HT depresses startle response (reviewed by Davis, 1979). Moreover, rodents subjected to prolonged isolation show decreased brain serotonergic activity (Valzelli and Garrantini, 1972; Valzelli, 1973; Popova and Petkov, 1990; Jones et al., 1992), as well as enhanced aggressive behaviour, hyperactivity and impairment of response inhibition (Valzelli and Garratini, 1972; Valzelli, 1973). Behavioural inhibition might be an adaptive response in a subordinate fish, since it will reduce the risk of initiating attacks from superior individuals. In other words, a subordinate individual will have to do the best out of a bad situation while waiting for better times to come. Thus, the reversible inhibition of aggressive behaviour observed in subordinate Arctic charr (Winberg et al., 1992a) might be related to such a behavioural inhibition induced by a stress mediated increase in brain 5-HT activity. Interestingly, boldness of sticklebacks (Gusterosteus aculeatus) towards pike (Esox lucius) covaries with the intensity by which they attack conspecifics (Huntingford, 1976; Tulley and Huntingford, 1988). This covariance suggests that some factors within individual sticklebacks that influence their readiness to show intraspecific aggression also modulate their behaviour in encounters with a predator (Tulley and Huntingford, 1988). As discussed above, bicolor damselfish exposed to a natural predator show a rapid increase in brain serotonergic activity (Winberg et al., 1993~). Thus, increased brain serotonergic activity might form part of a common causal system producing timidity. Interestingly, Kostowski et al. (1984) found that pharmacological inhibition of brain 5-HT activity provoked a shift from subordination to dominance in rats competing for water, whereas stimulation of the 5-HT system in dominant individuals had the opposite effect. Moreover, in the same study it was re-

ported that rats with surgical lesions of the 5-HT system (midbrain raphe nucleus) always became dominant when paired with a sham lesioned opponent (Kostowski et al., 1984). Treatment with 5-HT receptor antagonists has also been found to affect aggresive behaviour and social positions in rodents. For example, Gao and Cutler (1992) reported that the 5-HT, receptor antagonist, BRL 46470, increased aggressive behaviour of mice during social encounters in an unfamiliar cage. Furthermore, Mitchell and Redfern (1992) found that chronic treatment with the antidepressant drugs clomipramine, a 5-HT re-uptake inhibitor, or mianserine, a selective 5-HT, receptor antagonist, increased the relative rank position of subdominant rats. The effect of clomipramine could at first look like evidence for a stimulating effect of 5-HT on aggression. However, in addition to the acute effects of these drugs, chronic treatment with 5-HT, receptor antagonists as well as with tricyclic antidepressants, like clomipramine, might cause a down regulation of 5-HT, receptors (Frazer et al., 1988; Sleight et al., 1991). In the study by Mitchell and Redfern (1992), the effect of both clomipramine and mianserine was dependent on chronic treatment, and no effects were observed after 1 day of drug treatment. Furthermore, the effect of drug treatment was still apparent 1 day after the cessation of treatment (Mitchell and Redfern, 1992). Thus, it is tempting to speculate that the increase in relative rank position of subdominant rats that Mitchell and Redfern (1992) observed was caused by a down regulation of 5-HT, receptors, and, therefore, a decrease in serotonergic neurotransmission. In addition to an inhibition of aggressive behaviour, subordinate fish also show reduced food intake (Abbott et al., 1985; McCarthy et al., 1992a, 1992b; Winberg et al., 1993b) and lower spontaneous locomotor activity than dominant conspecifics. These effects of subordinate experience could also be related to a behavioural inhibition caused by a stress induced increase in brain serotonergic activity. Increased brain serotonergic activity appears to have an inhibitory effect on food intake in mammals (Samanin, 1989; Angel, 1990) and there are indications of a similar effect of increased serotonergic activity in fish (Johnston et al., 1992). There are a few studies on fish that indicate an inhibitory role of 5-HT on spontaneous locomotor activity. Fenwick (1970) reported higher swimming activity in blinded goldfish, which had depressed levels of brain 5-HT. Furthermore, he found a negative correlation between brain 5-HT concentration and locomotor activity in the goldfish. Similarly, the results by Fingerman (1976) indicate that 5-HT has an inhibitory effect on locomotor activity in Gulf killifish (Fundulus grandis). As mentioned above, the dominant fish in a group is the most active individual, while subordinate fish spend most of the time standing still close to the surface. Furthermore, artificially stressed Arctic charr

Monoamine neurotransmitters display a reduction in motor activity as well as increased 5-HIAA/S-HT ratios (Winberg et al., 1992b). Thus, the decreased locomotor activity in subordinate fish could, at least partly, be an effect of the increased serotonergic activity. A computerized video-image analysis system was recently used to study the effects of 5-HT and social rank on spontaneous locomotor activity in Arctic charr. The fish studied had just experienced dominant or subordinate positions in a pair. These fish were compared to fish where the S-HT system was inhibited by a S-HT synthesis inhibitor @-CPA) or stimulated by a specific 5-HT re-uptake blocker (zimeldine) (Winberg et al., 1993a). The results showed that the subordinate individuals had significantly lower spontaneous locomotor activity than dominant fish. Similarly, stimulation of 5-HT activity by treatment with zimeldine resulted in a significant reduction in locomotor activity (Fig. IOA), whereas inhibition of brain serotonergic activity by treatment with p-CPA signifi-

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cantly increased spontaneous locomotor activity in the fish (Fig. IOB) (Winberg et al., 1993a). In the same experiment it was found that neither social experience nor p-CPA or zimeldine had any striking effects on the diurnal activity rhythm of Arctic charr (Fig. lOA,B) (Winberg ef al., 1993a). The highest locomotor activity was always recorded during the first hours of the experiment, a high activity period that might reflect exploratory behaviour or anxiety related to disorientation, as the test area was a novel environment for the fish (Winberg et al., 1993a). Interestingly, both the effect of p-CPA (Fig. IOB), and the difference in locomotor activity between dominant and subordinate individuals, were most pronounced during the first 6 hr of the experiment. Thus, these experiments also indicate an inhibitory effect of brain 5-HT on exploratory behaviour and responsiveness to environmental stimuli. The central serotonergic system seems to be involved in the regulation of spontaneous locomotor activity also in mammals (reviewed by Gerson and Baldessarini, 1980). In rodents, it has generally been found that inhibition of the brain serotonergic system increases locomotor activity, whereas activation of serotonergic system of the brain has the opposite effect (Gerson and Baldessarini, 1980).

SUMMARY B g

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609

AND CONCLUSIONS

The monoaminergic systems are phylogenetically very old, and their distribution within the brain seems to be .well conserved between different vertebrate classes. Both the NE and 5-HT system have their cellbodies localized in a few major nuclei with extremely wide and diffuse projection patterns, indicating neuromodulatory functions. The dopaminergic system, on the other hand, consists of several major with different topographic projections, nuclei, suggesting more specialized functions. In the mammalian brain, monoamine neurotransmitters act through a variety of receptor subtypes. The receptor divergence might be less pronounced in lower vertebrates; however, the information about monoamine receptor subtypes in the brain of these animals is presently very restricted. Several similarities with regard to functions of central monoamine neurotransmitters in fish and mammals have been discussed in the present paper. Thus, brain DA systems seem to act in a stimulatory manner on aggressive or competitive behaviour in fish, and a similar relationship between DA and aggressive behaviour has been reported in mammalian studies. Increased dopaminergic activity has been indicated in telencephalon of dominant Arctic charr in a dominance hierarchy, and by pharmacological stimulation of the dopaminergic system, it has been possible to induce social dominance in this species.

610

S. WINBERG and G. E. NIIZXIN

The brain serotonergic system has, on the other hand, been suggested to act in an inhibitory manner on aggressive behaviour in both fish and mammals. Stimulation of the brain serotonergic system has also been found to decrease locomotor activity in fish and mammals. Furthermore. an inhibitorv role of the brain serotonergic system in the control of food intake has been suggested in mammals, a function which has not yet been confirmed in fish. A general increase in brain serotonergic activity has been found in fish occupying low positions in dominance hierarchies, and again similar-results have been reported ing primates. sive behaviour,

from

some

mammalian

Subordinate locomotor

fish, behavioural

species,

experience

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activity

characteristics

and food that

could,

includaggresintake

in

at least in

part, be connected with the increased brain serotonergic activity displayed by these individuals. Taken functions with

together,

available

in fish behaviour

mammals,

indicating

results suggest -_ that

on many

these

monoamine similarities mechanisms

have been conserved during the last 400 million years of vertebrate evolution. Acknowledgemenrs-This work was financially supported by the Swedish Council for Forestry and kg&cultural Research, the Helge Ax: son Johnson Foundation and the Carl Trygger Foundation.

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