Context and trade-offs characterize real-world threat detection systems: A review and comprehensive framework to improve research practice and resolve the translational crisis.

A better understanding of context in decision-making-that is, the internal and external conditions that modulate decisions-is required to help bridge the gap between natural behaviors that evolved by natural selection and more arbitrary laboratory models of anxiety and fear. Because anxiety and fear are mechanisms evolved to manage threats from predators and other exigencies, the large behavioral, ecological and evolutionary literature on predation risk is useful for re-framing experimental research on human anxiety-related disorders. We review the trade-offs that are commonly made during antipredator decision-making in wild animals along with the context under which the behavior is performed and measured, and highlight their relevance for focused laboratory models of fear and anxiety. We then develop an integrative mechanistic model of decision-making under risk which, when applied to laboratory and field settings, should improve studies of the biological basis of normal and pathological anxiety and may therefore improve translational outcomes.

Biologically meaningful scents: a framework for understanding predator-prey research across disciplines.

Fear of predation is a universal motivator. Because predators hunt using stealth and surprise, there is a widespread ability among prey to assess risk from chemical information - scents - in their environment. Consequently, scents often act as particularly strong modulators of memory and emotions. Recent advances in ecological research and analytical technology are leading to novel ways to use this chemical information to create effective attractants, repellents and anti-anxiolytic compounds for wildlife managers, conservation biologists and health practitioners. However, there is extensive variation in the design, results, and interpretation of studies of olfactory-based risk discrimination. To understand the highly variable literature in this area, we adopt a multi-disciplinary approach and synthesize the latest findings from neurobiology, chemical ecology, and ethology to propose a contemporary framework that accounts for such disparate factors as the time-limited stability of chemicals, highly canalized mechanisms that influence prey responses, and the context within which these scents are detected (e.g. availability of alternative resources, perceived shelter, and ambient physical parameters). This framework helps to account for the wide range of reported responses by prey to predator scents, and explains, paradoxically, how the same individual predator scent can be interpreted as either safe or dangerous to a prey animal depending on how, when and where the cue was deposited. We provide a hypothetical example to illustrate the most common factors that influence how a predator scent (from dingoes, Canis dingo) may both attract and repel the same target organism (kangaroos, Macropus spp.). This framework identifies the catalysts that enable dynamic scents, odours or odorants to be used as attractants as well as deterrents. Because effective scent tools often relate to traumatic memories (fear and/or anxiety) that cause future avoidance, this information may also guide the development of appeasement, enrichment and anti-anxiolytic compounds, and help explain the observed variation in post-traumatic-related behaviours (including post-traumatic stress disorder, PTSD) among diverse terrestrial taxa, including humans.

Are single odorous components of a predator sufficient to elicit defensive behaviors in prey species?

When exposed to the odor of a sympatric predator, prey animals typically display escape or defensive responses. These phenomena have been well-documented, especially in rodents, when exposed to the odor of a cat, ferret, or fox. As a result of these experiments new discussions center on the following questions: (1) is a single volatile compound such as a major or a minor mixture constituent in urine or feces, emitted by the predator sufficient to cause defensive reactions in a potential prey species or (2) is a whole array of odors required to elicit a response and (3) will the relative size or escapability of the prey as compared to the predator influence responsiveness. Most predator-prey studies on this topic have been performed in the laboratory or under semi-natural conditions. Field studies could help to find answers to these questions. Australian mammals are completely naïve toward the introduced placental carnivores. That offers ideal opportunities to analyze in the field the responses of potential prey species to unknown predator odors. During the last decades researchers have accumulated an enormous amount of data exploring the effects of eutherian predator odors on native marsupial mammals. In this review, we will give a survey about the development of olfactory research, chemical signals and their influence on the behavior and-in some cases-physiology of prey species. In addition, we report on the effects of predator odor experiments performed under natural conditions in Australia. When studying all these literature we learned that data gained under controlled laboratory conditions elucidate the role of individual odors on brain structures and ultimately on a comparatively narrow range behaviors. In contrast to single odors odor arrays mimic much more the situation prey animals are confronted to in nature. Therefore, a broad range of methodology-from chemistry to ecology including anatomy, physiology, and behavior-is needed to understand all the different (relevant) stimuli that govern and guide the interactions between a predator and its potential prey.

Behavioral responses of predator-naïve dwarf hamsters (Phodopus campbelli) to odor cues of the European ferret fed with different prey species.

Many mammalian predators are able to identify their prey by odors and, vice versa, numerous prey species recognize predator odors as well. The present paper reports on the behavioral responses of predator-naïve dwarf hamsters (Phodopus campbelli) towards the urine odors of carnivorous ferrets, which were raised on either a chicken, mouse or hamster diet. Chemical composition from ferret urines of the different diet groups was analyzed, while quantitative differences in urinary volatile constituents were observed through capillary gas chromatography­mass spectrometry. In a Y-maze arrangement, hamsters were offered several two-choice odor discrimination tasks and their behavior was quantified. Hamsters were easily able to discriminate the urine odor of ferrets fed with mice against ferrets fed with hamsters. This is probably the first report indicating that a prey species can distinguish urine odors of even an unknown predator species that has been fed different prey species. The analytical data complemented behavioral assays.

Odor detection and odor discrimination in subadult and adult rats for two enantiomeric odorants supported by c-fos data.

The study investigated the olfactory sensitivity and odor discrimination ability of male Wistar rats for the two closely related enantiomic odorants carvone and limonene. Enantiomers have identical chemical and physical properties but many are known to have different odor qualities for humans. Our psychophysical experiments revealed that subadult animals (about 2 months old) demonstrated higher olfactory sensitivities--lower detection thresholds--for D- and L-limonene than the adult conspecifics (about 6 months old). In the discrimination tasks at higher odor concentrations all animals were able to discriminate between the D- and L-odorant but subadults demonstrated lower discrimination thresholds. Within each age group no significant difference could be established. To elucidate neural correlates of our behavioral results, we have used c-fos expression to provide a global map of neuronal activity, with single cell resolution, in the olfactory bulb of D- and L-carvone exposed animals. The two carvone odors elicit different patterns of neuronal activity in the olfactory bulb, however, with partially overlapping activated areas. Our behavioral and immunhistochemical results allow to assume that the D- and L-forms of the two enantiomeric odorants have different odor qualities. Our study further demonstrates that the odor quality of an odorant depends not only on the atoms of the molecule, its chemical structure or on the presence of a functional group. Other parameters like, e.g. interaction profiles with different peptides, optical rotation of the polarized light--as seen in the enantiomers of an odorant--might also contribute to the quality of an odor.

Combined behavioral and c-Fos studies elucidate the vital role of sodium for odor detection.

Salt, known as taste quality, is generally neglected in olfaction, although the olfactory sensory neurons stretch into the salty nasal mucus covering the olfactory epithelium (OE). Using a psychophysical approach, we directly and functionally demonstrate in the awake rat for a variety of structurally diverse odorants that sodium is a critical factor for olfactory perception and sensitivity, both very important components of mammalian communication and sexual behavior. Bathing the olfactory mucus with an iso-osmotic sodium-free buffer solution results in severe deficits in odorant detection. However, sensitivity returns fully within a few hours, indicating continuous mucus production. In the presence of sodium in the mucus covering the OE, all odorants induce odorant-specific c-Fos expression in the olfactory bulb. Yet, if sodium is absent in the mucus, no c-Fos expression is induced as demonstrated for n-octanal. Our noninvasive approach to induce anosmia in mammals here presented--which is fully reversible within hours--opens new possibilities to study the functions of olfactory communication in awake animals.

Behavioral changes induced in rats by exposure to trimethylthiazoline, a component of fox odor.

Trimethylthiazoline (TMT), a component of fox feces, has been used in various studies as a natural predator stimulus to induce autonomic and behavioral signs of fear (e.g., higher levels of stress hormones, freezing, and risk assessment). The present study investigated whether 2 further behavioral signs of fear are induced in rats by TMT exposure: potentiation of the acoustic startle response and inhibition of appetitive behavior. In addition, the authors tested the rats for dose dependency of TMT-induced freezing behavior. The study confirmed that behavioral changes observed during TMT exposure are caused by TMT-induced fear and are dose dependent.

The effects of predator odors in mammalian prey species: a review of field and laboratory studies.

Prey species show specific adaptations that allow recognition, avoidance and defense against predators. For many mammalian species this includes sensitivity towards predator-derived odors. The typical sources of such odors include predator skin and fur, urine, feces and anal gland secretions. Avoidance of predator odors has been observed in many mammalian prey species including rats, mice, voles, deer, rabbits, gophers, hedgehogs, possums and sheep. Field and laboratory studies show that predator odors have distinctive behavioral effects which include (1) inhibition of activity, (2) suppression of non-defensive behaviors such as foraging, feeding and grooming, and (3) shifts to habitats or secure locations where such odors are not present. The repellent effect of predator odors in the field may sometimes be of practical use in the protection of crops and natural resources, although not all attempts at this have been successful. The failure of some studies to obtain repellent effects with predator odors may relate to (1) mismatches between the predator odors and prey species employed, (2) strain and individual differences in sensitivity to predator odors, and (3) the use of predator odors that have low efficacy. In this regard, a small number of recent studies have suggested that skin and fur-derived predator odors may have a more profound lasting effect on prey species than those derived from urine or feces. Predator odors can have powerful effects on the endocrine system including a suppression of testosterone and increased levels of stress hormones such as corticosterone and ACTH. Inhibitory effects of predator odors on reproductive behavior have been demonstrated, and these are particularly prevalent in female rodent species. Pregnant female rodents exposed to predator odors may give birth to smaller litters while exposure to predator odors during early life can hinder normal development. Recent research is starting to uncover the neural circuitry activated by predator odors, leading to hypotheses about how such activation leads to observable effects on reproduction, foraging and feeding.

TMT-induced autonomic and behavioral changes and the neural basis of its processing.

One of the main interests in the field of neuroscience is the investigation of the neural basis of fear. During recent years, an increasing number of studies have used trimethylthiazoline (TMT), a component of red fox feces, as a stimulus to induce fear in predator naive rats, mice, and voles. The aim of the present review is to summarize these studies. We present an overview to the autonomic and behavioral changes that are induced by TMT exposure. Then, we summarize the small number of studies that have examined the neural processing of the TMT stimulus. Finally, we compare these studies with those using a natural predator or predator odor to induce fear and discuss the possible use of TMT exposure in rodents as an animal model of unconditioned fear in humans.

Comparative investigation of the volatile urinary profiles in different Phodopus hamster species.

Stir bar sorptive extraction method was used for investigation of the urinary volatile profiles in male and female Phodopus campbelli and Phodopus sungorus hamsters. Additionally, female Phodopus roborowsky urinary profiles were characterized. A quantitative analytical approach allowed comparisons of 17 selected compounds in urine. Results showed that campbelli and sungorus species show similar urinary volatile profiles for males and females. Differences appeared only in concentrations. Several unique compounds, such as pyrazine derivatives, were found to be gender-and age-specific. P. roborowsky females exhibited a completely different urinary volatile profile from campbelli and sungorus females, featuring a unique set of substituted quinoxalines.

Detecting danger--or just another odorant? Olfactory sensitivity for the fox odor component 2,4,5-trimethylthiazoline in four species of mammals.

2,4,5-trimethylthiazoline (TMT) is a volatile component of the anal gland secretion of the red fox and elicits behavioral and physiological fear responses in the rat. Using instrumental conditioning paradigms, we determined olfactory detection thresholds for TMT in three rats, a natural prey species of the red fox, and compared their performance to that of three squirrel monkeys, three spider monkeys and four pigtail macaques, all non-prey species of the red fox. We found that the rats were able to discriminate concentrations between 0.04 and 0.10 ppt (parts per trillion) of TMT from the odorless solvent which is by far the lowest olfactory detection threshold for an odorant reported in rats so far. In contrast, the spider monkeys needed 0.14-1.38 ppb (parts per billion), the pigtail macaques 0.41-4.07 ppb, and the squirrel monkeys 4.07-13.80 ppb to detect TMT which does not rank among the lowest olfactory thresholds reported for these three primate species. Thus, these results support the assumption that the behavioral relevance of an odorant may be an important determinant of a species' olfactory sensitivity.

Olfactory sensitivity, learning and cognition in young adult and aged male Wistar rats.

Psychophysical experiments with male Wistar rats, ranging from 2 to more than 25 months old, revealed age-related differences in olfactory sensitivity. The highest sensitivities were found in rats 13 months old, and the lowest sensitivity was found in the group aged 25 months and older. Consequently, we considered the hypotheses that young rats will require less time and less trials than aged conspecifics to learn an olfactory discrimination task and that olfactory cognitive abilities will be reduced in older individuals. Rats were initially trained in an olfactometer using operant techniques to discriminate between the odor ethyl acetate (EA) and clean air. Next, young adult and 28-month-old rats were tested on seven different go/no-go odor discrimination tasks. Aged rats performed as well as young adults did on all tasks and we conclude that, for a variety of odor discrimination problems, aged rats show no deterioration in learning ability. This is the first report on olfactory sensitivity, learning ability and cognition in Wistar rats that have passed the normal life span for this strain. Data show that the inability to learn and cognitive deficits do not necessarily develop with age.

Neural correlates of cat odor-induced anxiety in rats: region-specific effects of the benzodiazepine midazolam.

Cat odor elicits a profound defensive reaction in rats that is reduced by benzodiazepine drugs. The neural correlates of this phenomenon were investigated here using Fos immunohistochemistry. Rats received either midazolam (0.75 mg/kg, s.c.) or vehicle and were exposed to pieces of a collar that had been worn by a domestic cat or an unworn (dummy) collar. Cat odor caused midazolam-sensitive defensive behavioral responses, including avoidance of collar contact, inhibition of grooming, and prolonged rearing. Cat odor exposure induced Fos expression in the posterior accessory olfactory bulb (glomerular, mitral, and granule cell layers), with granule cell layer activation attenuated by midazolam. High basal Fos expression, and some cat odor-associated Fos expression, was evident in the main olfactory bulb (glomerular cell layer), and midazolam exerted a strong inhibitory effect in this region. Midazolam inhibited Fos expression in key limbic regions involved in pheromone transduction (medial amygdala and bed nucleus of the stria terminalis) and defensive behavior (prelimbic cortex, lateral septum, lateral and medial preoptic areas, and dorsal premammillary nucleus). However, midazolam failed to affect cat odor-related Fos expression in a range of key defense-related sites, including the ventromedial hypothalamic nucleus, paraventricular nucleus of the hypothalamus, periaqueductal gray, and cuneiform nucleus. These results indicate that midazolam exerts a region-specific effect on the neural substrates activated by predator odor, with effects in the lateral septum and dorsal premammillary nucleus likely to be of major importance. These findings also suggest the intriguing hypothesis that cat odor is processed by rats as a "pheromone-like" stimulus.

Temporary inactivation of the bed nucleus of the stria terminalis but not of the amygdala blocks freezing induced by trimethylthiazoline, a component of fox feces.

Presentation of trimethylthiazoline (TMT, a component of fox feces) to laboratory rats elicits freezing, a prominent behavioral sign of anxiety or fear. The present study investigated the neural basis of this unlearned response. Muscimol, a GABA(A) receptor agonist, was injected (4.4 nmol/0.5 microl) into the bed nucleus of the stria terminalis (BNST) as well as into the amygdala, two brain areas known to be involved in anxiety and fear. Temporary inactivation of the BNST but not of the amygdala significantly blocked TMT-induced freezing. This effect was not caused by an enhancement of motor activity after BNST inactivation. In addition, these results confirm previous studies showing that freezing is possible despite amygdala inactivation. These results, and other findings in the literature, suggest that the BNST is critically involved in unlearned fear, whereas the amygdala is more involved in the acquisition and expression of learned fear.

Concanavalin A application to the olfactory epithelium reveals different sensory neuron populations for the odour pair D- and L-carvone.

Carvone enantiomers (D and L optical isomers) have been shown to be discriminable by humans even though the odor qualities are quite similar. Our experiment is based on a finding (J. Steroid Biochem. Molec. Biol. 1991;39(4B):621) that Concanavalin A (ConA) applied to a frog olfactory epithelium preparation blocks cAMP transduction induced by D- but not by L-carvone. We used standard operant conditioning methods to train animals to discriminate low odor concentrations of D-carvone from clean air, to discriminate L-carvone from clean air; or to discriminate between clean air and the odors of D-carvone, L-carvone, ethyl acetate and methacrylic acid. After perfusion of the nasal cavity with ConA, rats did not respond to D-carvone above or near chance level, while the L-carvone response was not affected at the same or higher ConA doses. However, for rats trained on both enantiomers and the two other unrelated odorants, the D-carvone response remained unaffected by ConA. These results suggest to us that: (1) ConA blocks at least one chiral receptor selective for D-carvone; (2) D-carvone odor quality is modified by ConA so that it is no longer recognized by rats trained on D-carvone only, while rats trained to generalize odors still respond to D-carvone.