Marilia Barros

Non-human primate models for investigating fear and anxiety.

Non-human primates exhibit similar physiological and behavioral responses to anxiety-inducing situations as humans and have, in fact, been successfully employed in both conditioned (i.e. conflict paradigms) and ethologically based tests of fear/anxiety (i.e. involuntary isolation, social interaction, human threat, predator confrontation). In the last decade, a renewed and growing interest in non-human primate models has resulted from the use of the small callitrichid species in behavioral pharmacology and neuroscience. This review focuses on the available non-human primate models for investigating fear/anxiety, addressing their advantages, shortcomings, and conceptual framework on which they are based. Lastly, a new ethologically based model to study anxiety and fear-induced avoidance in callitrichids--the marmoset predator confrontation test--is discussed.

Reactions to Potential Predators in Captive-Born Marmosets (Callithrix penicillata)

We describe the behavioral repertoire of captive-born black tufted-eared marmosets (Callithrix penicillata) elicited by brief exposures to three potential mounted taxidermized predators (caracara hawk, Polyborus plancus; rattlesnake, Crotalus durissus; oncilla, Leopardus tigrina), and a stuffed toy. For each of the four stimuli, we submitted the subjects to a 9-min trial divided into three consecutive intervals: a 4-min pre-exposure baseline observation, a 1-min stimulus exposure, and a 4-min postexposure observation period. We positioned stimuli in front of each subjects home cage, and video-taped trials for behavioral analysis. During exposures to the potential taxidermized predators, we heard tsik-tsik vocalization and alarm behavior. After exposures, only the cat induced these reactions. All stimuli elicited observational reaction, albeit only during exposure intervals. Further comparisons between the three trial intervals indicated a decrease in the time spent in proximity to the cat during exposures, while an increase in proximity occurred when subjects were exposed to either the hawk or snake for the same period. Taken together, the behavioral responses during and after exposures to the taxidermized oncilla suggest that this stimulus is capable of inducing strong and persistent emotional reactions in Callithrix penicillata.

Anxiolytic and antidepressant-like effects of Melissa officinalis (lemon balm) extract in rats: Influence of administration and gender

Objective: To analyse the behavioral effects of Melissa officinalis extract in rats following acute or subacute treatment. Materials and Methods: The behavioral effects of an acute or subacute (10-day course) orally administered M. officinalis (MO; 0, 30, 100 or 300 mg/kg) ethanol extract were evaluated in male and female Wistar rats in elevated plus-maze (EPM), forced swimming (FS) and open field (OF) tests. The effects of diazepam (DZP; 1 mg/kg) and fluoxetine (FXT; 10 mg/kg) were also assessed. Results: In the EPM test, the percentage of open arm entries and open arm times of both males and females given the subacute M. officinalis ethanol extract were significantly higher than those of the vehicle-treated animals but were at levels similar to those observed in the DZP group, regardless of the treatment length. In the FS test, immobility duration was significantly lower in both males and females treated with the plant extract when compared to vehicle-treated counterparts. A 10-day treatment with FXT induced the same antidepressant response, regardless of gender, and was more effective than the M. officinalis extract. Male and female rats demonstrated distinct gender profiles, and treatment × gender interactions were observed. Locomotion in the EPM and OF tests was not significantly altered by treatments. Conclusion: The potential psychoactive properties of M. officinalis may provide a unique pharmacological alternative for certain psychiatric disorders; however, the efficacy appears to be dependent on both gender and administration length.

Differences in Aerial and Terrestrial Visual Scanning in Captive Black Tufted-Ear Marmosets (Callithrix penicillata) Exposed to a Novel Environment

Aerial and terrestrial visual scanning were investigated in captive black tufted-ear marmosets, Callithrix penicillata, exposed to a novel environment. Naive adult subjects (n = 24) were individually exposed to a figure-eight maze during seven 30-min trials, 48 h apart. Habituation to the maze was observed, as indicated by the significant decrease in locomotion. The frequency of aerial scanning, however, remained elevated throughout the 7 trials, while its duration rapidly increased to high levels. Frequency and duration of terrestrial scanning persisted at constant low rates, differing significantly from aerial scanning. Males and females did not differ significantly. The different impact of aerial versus terrestrial predators could have a significant influence on vigilant behaviour in this species. Thus, visual scanning is an important and highly organized antipredation strategy in marmosets.

Predatory stress as an experimental strategy to measure fear and anxiety-related behaviors in non-human primates.

Natural defense-inducing stimuli are being increasingly exploited as a means to investigate the neural mechanisms underlying normal and pathological anxiety, as well as for the screening of new compounds with potential therapeutic use in human anxiety disorders. Such an approach, frequently used in rodents, has recently been employed in the Marmoset Predator Confrontation Test (MPCT). In this method, marmoset monkeys are individually confronted with a taxidermized predator (wild oncilla cat) in a previously habituated maze environment, while several easily discernable fear/anxiety-related behaviors are measured. Confrontation with the cat stimulus significantly altered ongoing behaviors, each habituating distinctively during repeated exposures; e.g. complete rapid habituation (alarm call), complete slow habituation (exploration, vigilance) or only partial habituation (proximity avoidance). Pharmacological validating studies with diazepam and buspirone induced a significant dose-dependent reversal of the fear-induced proximic avoidance and scratching/scent-marking behaviors, while exploration (smell/lick the maze, leg stand) was found to increase. The neuropeptide substance P and the selective 5-HT1A receptor antagonist WAY100635 resulted in a similar anxiolytic-like profile. The response pattern observed was not influenced by social isolation, handling/manual restraint, novel environment exposure or habituation to the stimulus or its location. Persistent defensive behavior and response pattern to diazepam was observed when naive versus MPCT-experienced marmosets were tested following a recent predatory stress. Taken together, the results indicate that the MPCT is a valuable experimental procedure to measure fear and anxiety-related behaviors in nonhuman primates.

Decreased methylation of the NK3 receptor coding gene (TACR3) after cocaine-induced place preference in marmoset monkeys

Epigenetic processes have been implicated in neuronal plasticity following repeated cocaine application. Here we measured DNA methylation at promoter CpG sites of the dopamine transporter (DAT1) and serotonin transporter (SERT) and neurokinin3‐receptor (NK3‐R)‐receptor (TACR3) coding genes in marmoset monkeys after repeated cocaine injections in a conditioned place preference paradigm. We found a decrease in DNA methylation at a specific CpG site in TACR3, but not DAT1 or SERT. Thus, TACR3 is a locus for DNA methylation changes in response to repeated cocaine administration and its establishment as a reinforcer, in support of other evidence implicating the NK3‐R in reinforcement‐ and addiction‐related processes.

Superior colliculus lesions impair threat responsiveness in infant capuchin monkeys.

The ability to react fast and efficiently in threatening situations is paramount for the survival of organisms and has been decisive in our evolutionary history. Defense mechanisms in primates rely on the fast recognition of potential predators and facial expressions of conspecifics. The neural circuitry responsible for the detection of threat is generally thought to be centered on the amygdala. Although it is a pivotal structure in the processing of emotional stimuli, the amygdala does not seem necessary for the early stages of this process. Here we show that bilateral neurotoxic lesions of the superior colliculus in infant capuchins monkeys impaired the recognition of a rubber-snake in a threat-reward conflict task. Lesioned monkeys were uninhibited by a snake in a food-reward retrieval task. Lack of inhibition in the task was observed over the course of 15 weeks. The long lasting recognition impairment of a natural predator observed here is similar to the tameness aspects of Kluver-Bucy syndrome, indicating an important role of this structure in threat recognition.

Interaction of the tachykinin NK3 receptor agonist senktide with behavioral effects of cocaine in marmosets (Callithrix penicillata)

Brain neuropeptide transmitters of the tachykinin family are involved in the organization of many behaviors. However, little is known about their contribution to the behavioral effects of drugs of abuse. Recently, antagonism of the tachykinin NK3-receptor (NK3-R), one of the three tachykinin receptors in the brain, was shown to attenuate the acute and chronic behavioral effects of cocaine in rats and the acute effects in non-human primates. In order to expand these findings we investigated the effects of the NK3-R agonist, succinyl-[Asp6, Me-Phe8]SP(6-11) (senktide), on the acute behavioral effects of cocaine in marmoset monkeys (Callithrix penicillata) using a figure-eight maze procedure. Animals were pretreated with senktide (0, 0.1, 0.2, 0.4 mg/kg, s.c.), and received either a treatment with cocaine (10 mg/kg) or saline (i.p.). Cocaine increased locomotor activity and the duration of aerial scanning behavior, but reduced exploratory activity, bodycare activity, the frequency of aerial scanning, and terrestrial glance behavior. Senktide blocked the effects of cocaine on locomotor activity, but enhanced the cocaine effects on exploratory activity, aerial scanning frequency, and terrestrial glance behavior. Senktide alone did not significantly influence monkey behavior in this study. These data expand previous findings suggesting a complex role of the NK3-R in the acute behavioral effects of cocaine in non-human primates.

Rapid habituation of scan behavior in captive marmosets following brief predator encounters

Scan behavior in 10 captive predator-naive adult black tufted-ear marmosets (Callithrix penicillata) was investigated prior, during and following brief predator encounters (taxidermized oncilla cat -- Leopardus tigrinus) versus neutral stimulus exposures (stuffed toy). For each stimulus, three 9 min home-cage trials were conducted > or = 72 h apart. Each trial was divided into three consecutive 3 min intervals: pre-exposure baseline observation, stimulus exposure, and post-exposure observation period. Post-exposure scan duration increased during the first two predator confrontations, while scan frequency increased significantly only after the first. Scan behavior remained constant within the last predator encounter, as it also did within and between the three neutral stimulus exposures. Although marmosets scanned more often and significantly longer after encountering the predator than the neutral stimulus, this response rapidly habituated by the second trial. Therefore, black tufted-ear marmosets in a familiar environment rapidly habituate to brief repeated predator encounters, possibly minimizing anti-predation costs once the degree of a potential threat has been adequately assessed.

Persistent anxiety-like behavior in marmosets following a recent predatory stress condition: Reversal by diazepam

Initial investigations indicated the use of the Marmoset Predator Confrontation Test (MPCT) as an experimental procedure to measure fear/anxiety-related behaviors in non-human primates. However, possible long-term habituation effects and re-use of experimental subjects need to be verified. This study, therefore, compared the behavioral response of experienced versus naïve adult black tufted-ear marmosets (Callithrix penicillata) in the MPCT, with/without diazepam administrations. Subjects were tested in the figure-8 maze and confronted with a taxidermized wild-cat predator stimulus. After four initial 20-min maze habituation sessions, each subject was submitted to two randomly-assigned 20-min predator confrontation sessions: vehicle and 2 mg/kg of diazepam. Confrontation with the predator induced significant behavioral changes; i.e., proximic avoidance and tsik-tsik alarm call. Diazepam administration, concomitant to predator exposure, reversed the behavioral changes observed. In both the experienced and naïve marmosets a similar behavioral profile and response pattern to diazepam was detected, corroborating the important selective pressure that felines seem to have on marmoset behavioral ecology. Therefore, during a more naturalistic-like regimen--i.e., recurring intermittent predator encounters--the general response pattern remains highly consistent, regardless of prior experience. One may consider the re-use of marmoset subjects in the MPCT, particularly under these specific conditions (i.e. repeated 20-min confrontations, 72-h apart).