Tangi R. Summers

Does Serotonin Influence Aggression? Comparing Regional Activity before and during Social Interaction*

Serotonin is widely believed to exert inhibitory control over aggressive behavior and intent. In addition, a number of studies of fish, reptiles, and mammals, including the lizard Anolis carolinensis, have demonstrated that serotonergic activity is stimulated by aggressive social interaction in both dominant and subordinate males. As serotonergic activity does not appear to inhibit agonistic behavior during combative social interaction, we investigated the possibility that the negative correlation between serotonergic activity and aggression exists before aggressive behavior begins. To do this, putatively dominant and more aggressive males were determined by their speed overcoming stress (latency to feeding after capture) and their celerity to court females. Serotonergic activities before aggression are differentiated by social rank in a region‐specific manner. Among aggressive males baseline serotonergic activity is lower in the septum, nucleus accumbens, striatum, medial amygdala, anterior hypothalamus, raphe, and locus ceruleus but not in the hippocampus, lateral amygdala, preoptic area, substantia nigra, or ventral tegmental area. However, in regions such as the nucleus accumbens, where low serotonergic activity may help promote aggression, agonistic behavior also stimulates the greatest rise in serotonergic activity among the most aggressive males, most likely as a result of the stress associated with social interaction.

Temporal patterns of limbic monoamine and plasma corticosterone response during social stress

Dominant and subordinate males respond differently to the stress of social interaction. After an hour of social interaction, subordinate male Anolis carolinensis have elevated serotonergic activity in hippocampus, but dominant males do not. In other species, and using other stressors, the activation of hippocampal serotonergic activity is much more rapid than one hour. To elucidate early stress responsiveness, adult male A. carolinensis were divided into four groups: isolated controls, and pairs of males sampled after 10, 20 or 40 minutes of aggressive interaction. Development of dominant-subordinate relationships was determined by behavior and by the celerity of eyespot darkening. Serotonergic activity in the hippocampus, nucleus accumbens and amygdala was elevated rapidly and equally in both dominant and subordinate males, as were plasma corticosterone concentrations. Serotonergic activity remained elevated through 40 minutes in hippocampus and nucleus accumbens. Only subordinate males had elevated corticosterone levels at 40 minutes. Social status does not impede socially induced stress responses. Rather, rapid regulation of serotonergic stress responses appears to be a mediating factor in determining both behavioral output and social status. Temporal expressions of monoaminergic and endocrine stress responses are distinctive between males of dominant and subordinate social status. Such temporal patterns of transmitter and glucocorticoid activity may reflect neurocircuitry adaptations that result in behavior modified to fit social status.

Regional and temporal separation of serotonergic activity mediating social stress

Stressful aggressive interaction stimulates central serotonergic activation in telencephalon as well as brainstem. Social roles can be distinguished by monoamine activity following aggression. Pairs of male lizards, Anolis carolinensis, were allowed to fight and form dominant/subordinate relationships. In micropunched regions of telencephalon, the greatest serotonergic changes occur in subordinate males. In hippocampal cortex and nucleus accumbens, subordinate males have increased 5-hydroxyindoleacetic acid/serotonin at 1 h following the fight. In these areas the ratio gradually decreases over a week of cohabitation, as was previously reported for brainstem. Medial and lateral amygdala develop increased serotonergic activity more slowly, with the greatest increase being evident following a week of interaction. Turnover, serotonin and 5-hydroxyindoleacetic acid levels in amygdala escalate over the first week of interaction in subordinate males, and return to baseline by one month. In dominant males, the pattern is accelerated, with the most extensive serotonin system activity present at 1 h, then decreasing over a month. The patterns of serotonergic activation are so similar in hippocampus, nucleus accumbens and brainstem that a co-ordinated response may be involved in mediating short-term social stress and aggression. Similarly, medial and lateral amygdala exhibit corresponding, but delayed patterns in subordinate males, suggesting a co-ordinated response in these regions mediating longer-term stress responses. These data are consistent with rapid neuroendocrine stress modulation in dominant individuals, and delayed serotonergic activity changes in subordinate males.

Monoaminergic activities of limbic regions are elevated during aggression: Influence of sympathetic social signaling

A visual social signal inhibiting aggression is coincident with limiting serotonergic and noradrenergic activity in subiculum, hippocampus, nucleus accumbens, medial amygdala, but not lateral amygdala, septum, and hypothalamus. Darkening of postorbital skin in the lizard Anolis carolinensis is stimulated by sympathetic activation of beta(2)-adrenergic receptors via adrenal catecholamines, and occurs more rapidly in dominant males during social interaction. Eyespot darkening functions as a social signal limiting aggressive interaction. To assess the effect of this social signal on telencephalic activity of monoamines, males were painted postorbitally with green or black paint, and exposed to a mirror. Serotonergic and noradrenergic turnover, as estimated by ratios of catabolite to transmitter, were elevated in the subiculum, hippocampus, nucleus accumbens, and medial amygdala of animals in which the eyespots were masked by green paint. Conversely, dopaminergic activity in these brain regions was lower in males with hidden eyespots (painted green). Hiding the eyespot evoked significantly increased aggressive activity toward the mirror image. Furthermore, changes in monoaminergic turnover were coincident with altered aggressive behavior, suggesting a relationship between them. Changes of monoaminergic activity were not observed in the septum, lateral amygdala, or hypothalamus, when males with eyespots permanently marked (black) were compared with those with eyespots hidden (painted green). Stimulated (serotonergic and noradrenergic) or inhibited (dopaminergic) activity due to social signal and aggression are confined to regions of the brain similarly activated during social stress, and do not constitute a generalized activation of monoaminergic systems.

Visible sympathetic activity as a social signal in Anolis carolinensis: changes in aggression and plasma catecholamines

Darkening of postorbital skin in Anolis carolinensis occurs during stressful situations and is stimulated by sympathetic activation of beta(2)-adrenergic receptors via adrenal catecholamines. This eyespot forms more rapidly in dominant males during social interaction. Eyespot darkening (green to black) appears to function as a social signal communicating sympathetic activation and limiting aggressive interaction. To assess the value of the eyespot as a social signal, males were painted postorbitally with green, black, or red paint. Each male was exposed to a mirror following acclimation to the cage. The total number of aggressive displays toward the mirror image was greatest when eyespots were masked by green paint. In contrast, black or red artificial eyespots, regardless of size, inhibited biting behavior toward the mirror image. The most aggressive males, those who saw a reflected opponent with no eyespot (hidden with green paint), had significantly higher levels of all plasma catecholamines. These results suggest that A. carolinensis use information from the eyespot to assess their opponents readiness to fight and thereby determine whether to be aggressive. Darkened eyespots are capable of inhibiting aggression, whereas aggressive displays from an opponent in the mirror without darkened eyespots do not. Darkened eyespots reflect rapid changes in plasma NE, DA, and Epi that may signal dominant social status.

Dynamics and mechanics of social rank reversal

Stable social relationships are rearranged over time as resources such as favored territorial positions change. We test the hypotheses that social rank relationships are relatively stable, and although social signals influence aggression and rank, they are not as important as memory of an opponent. In addition, we hypothesize that eyespots, aggression and corticosterone influence serotonin and N-methyl-D-aspartate (NMDA) systems in limbic structures involved in learning and memory. In stable adult dominant-subordinate relationships in the lizard Anolis carolinensis, social rank can be reversed by pharmacological elevation of limbic serotonergic activity. Any pair of specific experiences: behaving aggressively, viewing aggression or perceiving sign stimuli indicative of dominant rank also elevate serotonergic activity. Differences in the extent of serotonergic activation may be a discriminating and consolidating factor in attaining superior rank. For instance, socially aggressive encounters lead to increases in plasma corticosterone that stimulate both serotonergic activity and expression of the NMDA receptor subunit 2B (NR2B) within the CA3 region of the lizard hippocampus. Integration of these systems will regulate opponent recognition and memory, motivation to attack or retreat, and behavioral and physiological reactions to stressful social interactions. Contextually appropriate social responses provide a modifiable basis for coping with the flexibility of social relationships.

Manipulation of visual sympathetic sign stimulus modifies social status and plasma catecholamines.

Darkening of postorbital skin (eyespots) in Anolis carolinensis occurs during stressful situations via adrenal catecholaminergic activation of beta(2)-adrenergic receptors. The eyespots form more rapidly in dominant males during social interaction. It was hypothesized that eyespot darkening from green to black is a social signal that communicates disposition, resulting from sympathetic activation and determines social hierarchy. To assess the value of the eyespot as a signal influencing social dominance, males were paired according to size and weight, and painted postorbitally; one with green and the other with black paint. Painted pairs interacted with aggressive displays, approaches, and bites. Dominant status was determined by continued displacement of an opponent, superior perching position, and lighter color. Males with eyespots darkened by black paint were dominant in 100% of interactions and more aggressive. Males viewing an opponent with eyespots painted black exhibited higher plasma epinephrine (Epi), norepinephrine (NE), and dopamine (DA) levels than males with eyespots hidden with green paint and isolated controls. Results for behavior were similar, but those for plasma catecholamines contrast with those from recent experiments in which interactions occurred vs. a mirrored reflection. The eyespot is a potent stimulus, acting as a social signal that confers dominant status to its bearer, and activates the sympathetic nervous system in opponents that observe it during aggressive interactions.

The role of monoaminergic nuclei during aggression and sympathetic social signaling

A social sign stimulus that is sympathetically induced affects aggressive approaches and influences serotonergic, dopaminergic and noradrenergic activity in the brainstem nuclei of Anolis carolinensis. Darkening of postorbital skin via sympathetic activation of adrenal catecholamines and β2-adrenergic receptors provides a visual signal that forms more rapidly in dominant than subordinate males during social interactions. This signal limits aggressive interactions. Males were painted postorbitally with green or black paint and then exposed to a mirror. Aggressive approaches to the mirror were inhibited in males viewing a reflection with darkened eyespots, and increased in males viewing a reflection without eyespots (hidden). Noradrenergic turnover in the raphe and locus ceruleus were greatest in test subjects that viewed a reflection with eyespots hidden by green paint. Perception of darkened eyespots stimulated greater serotonergic turnover in raphe, locus ceruleus and substantia nigra/ventral tegmental area (SN/VTA). Dopaminergic turnover was higher in the raphe and SN/VTA of Anolis that viewed a reflection with darkened eyespots. However, these animals had lower dopamine turnover in the locus ceruleus than isolated and hidden eyespot groups. Of the possible roles of perikarya on central function and behavior, our results suggest feedback, cross-nuclear regulation, and some independence of function between nuclei and the forebrain terminal fields. Decreased serotonergic activity corresponds with increased aggression only in the raphe, suggesting that the raphe nuclei might be important for this behavioral trait. Increased serotonergic, noradrenergic and dopaminergic activities in SN/VTA in Anolis that view a reflected opponent with dark eyespots suggests that the SN/VTA might be directly involved in recognition of this social sign stimulus and the resulting inhibition of aggression.

Depressive Behavior and Activation of the Orexin/Hypocretin System

The orexin/hypocretin peptide signaling system plays a neuromodulatory role in motivation and stress; two critical components of depression. Although work has been done to identify links between orexin and depression, few specific neuroanatomical associations have been made. These studies have not investigated the relationship between orexin and orexin receptor expression in specific brain regions associated with this disorder. To address this, we examined immobility during the forced swim test (FST) in mice, a commonly used measure of depressive behavior. We analyzed the variation in FST immobility with the distribution of orexin and its receptor mRNA. We found that animals that exhibited more robust depressive behavior had greater or lesser orexin system expression that depended on the limbic brain region analyzed. In the hippocampus there was a negative correlation between orexin expression and FST immobility. Animals that displayed relatively more depressive behavior had lower hippocampal expression of Orexin A (OrxA). In the amygdala, there was a curvilinear relationship between OrxA and FST performance. In addition there was a positive correlation with amygdalar Type I orexin receptor (Orx1) mRNA and depressive behavior. Despite the differences in limbic orexin expression, there was no correlation between immobility and hypothalamic orexin neuron activation as measured by c-Fos. Overall, more severe depressive behavior was associated with reduced hippocampal orexin expression, contrasted with increased orexin plus Orx1 receptor mRNA expression in the amygdala. This divergent pattern between the hippocampus and amygdala mirrors a neurobiological theme seen in depression resulting from reduced hippocampal, but increased amygdalar, size and function.

Female social reproductive roles affect central monoamines

Central monoamines display a variety of activation patterns in different social groups, and among males and females. We addressed three social conditions for female lizards of the species Anolis carolinensis: Isolated, paired with a mate, and in a group of 5 competing for one mate. Among those in a group, only 1 or 2 females exhibited recrudescing ovaries. Individuals paired with a mate (for one month) exhibited ovarian growth, isolated animals (initial controls) had quiescent ovaries. Reproductively dominant females had significantly greater telencephalic 5-HIAA, and serotonergic activation, as indicated by the ratio of 5-HIAA to 5-HT. Telencephalic HVA as well as the HVA/DA ratio were also significantly greater in dominant females compared to all other groups. In contrast, serotonergic activation in brainstem was elevated in subordinate females only. These results suggest that serotonergic activation in telencephalon, found only in dominant females, not in other reproductively active females, is a function of the unique social role of a dominant female, possibly combining submissive behaviors toward a male with dominance over other females and competition for access to that male. Dopaminergic activation in telencephalon, also found only in dominant females, may be related to aggressive interactions with other females. Activation of serotonin in brainstem, found in this study in subordinate females and previously in males [C.H. Summers and N. Greenberg, Activation of central biogenic amines following aggressive interaction in male lizards, Anolis carolinensis, Brain Behav. Evol., 45 (1995) 339-349], may be associated with subordinate social status. Monoamines, involved in social behaviors, appear to be regionally specialized for dominant and subordinate social roles, in males [C.H. Summers and N. Greenberg, Activation of central biogenic amines following aggressive interaction in male lizards, Anolis carolinensis, Brain Behav. Evol., 45 (1995) 339-349][T.R. Summers, E.T. Larson, A.L. Hunter, K.J. Renner, N. Greenberg and C.H. Summers, Amygdalar serotonin mediates long-term social roles following aggressive interaction, Soc. Neurosci. Abs., 22 (1996) 1147] and females. Dominant females exhibit unique social position, behavior and monoamine profile whereas subordinate females and males have a similar serotonergic response in this species.