Behavioral analysis in laboratory rats: Challenges and usefulness of 50-kHz ultrasonic vocalizations.

Many rodent species emit and detect vocalizations in the ultrasonic range. Rats use three classes of ultrasonic vocalizations depending on developmental stage, experience and the behavioral situation. Calls from one class emitted by juvenile and adult rats, the so-called 50-kHz calls, are typical for appetitive and social situations. This review provides a brief historical account on the introduction of 50-kHz calls in behavioral research followed by a survey of their scientific applications focusing on the last five years, where 50-kHz publications reached a climax. Then, specific methodological challenges will be addressed, like how to measure and report 50-kHz USV, the problem of assignment of acoustic signals to a specific sender in a social situation, and individual variability in call propensity. Finally, the intricacy of interpreting 50-kHz results will be discussed focusing on the most prevalent ones, namely as communicative signals and/or readouts of the sender's emotional status.

Probing the decision-making mechanisms underlying choice between drug and nondrug rewards in rats.

Delineating the decision-making mechanisms underlying choice between drug and nondrug rewards remains a challenge. This study adopts an original approach to probe these mechanisms by comparing response latencies during sampling versus choice trials. While lengthening of latencies during choice is predicted in a deliberative choice model (DCM), the race-like response competition mechanism postulated by the Sequential choice model (SCM) predicts a shortening of latencies during choice compared to sampling. Here, we tested these predictions by conducting a retrospective analysis of cocaine-versus-saccharin choice experiments conducted in our laboratory. We found that rats engage deliberative decision-making mechanisms after limited training, but adopt a SCM-like response selection mechanism after more extended training, while their behavior is presumably habitual. Thus, the DCM and SCM may not be general models of choice, as initially formulated, but could be dynamically engaged to control choice behavior across early and extended training.

The Effect of Personality Traits on Cognitive Behavioral Therapy Outcomes in Student Pharmacists with Rat Phobia: A Randomized Clinical Trial.

Background: Little is known about which personality traits determine the effectiveness of various types of cognitive-behavioral therapy (CBT) on animal phobia. The objective of the present study was to investigate a possible association between personality traits and the outcome of single- and multi-session CBT. Methods: The present randomized clinical trial was conducted from November 2018 to May 2019 in Shiraz, Iran. Forty female students with rat phobia, who met the Diagnostic and Statistical Manual of Mental Disorders (DSM-V) criteria, were systematically allocated into a single- and a multi-session therapy group (odd numbers one-session treatment, even numbers multi-session treatment). In both groups, the students were gradually exposed to rats as part of the treatment. Psychological measures (state-anxiety, rat phobia, and disgust questionnaires) were used to compare pre- and post-intervention outcomes. Multivariate analysis of covariance was used to assess which personality traits influenced the intervention outcome. The statistical analysis was performed using SPSS (version 20.0) and P values<0.05 were considered statistically significant. Results: Rat phobia was positively and significantly affected by conscientiousness (P=0.001) and agreeableness (P=0.003). Of these personality traits, only a higher degree of conscientiousness resulted in a further reduction of state anxiety after the intervention (P=0.005). There were no significant differences between the pre- and post-intervention outcomes. Conclusion: The outcome of single- and multi-session rat phobia therapies was associated with specific personality traits of the participants, namely conscientiousness and agreeableness. Both intervention methods had an equal effect on reducing rat phobia.

Lapses in perceptual decisions reflect exploration.

Perceptual decision-makers often display a constant rate of errors independent of evidence strength. These 'lapses' are treated as a nuisance arising from noise tangential to the decision, e.g. inattention or motor errors. Here, we use a multisensory decision task in rats to demonstrate that these explanations cannot account for lapses' stimulus dependence. We propose a novel explanation: lapses reflect a strategic trade-off between exploiting known rewarding actions and exploring uncertain ones. We tested this model's predictions by selectively manipulating one action's reward magnitude or probability. As uniquely predicted by this model, changes were restricted to lapses associated with that action. Finally, we show that lapses are a powerful tool for assigning decision-related computations to neural structures based on disruption experiments (here, posterior striatum and secondary motor cortex). These results suggest that lapses reflect an integral component of decision-making and are informative about action values in normal and disrupted brain states.

The social life of Norway rats (Rattus norvegicus).

The Norway rat has important impacts on our life. They are amongst the most used research subjects, resulting in ground-breaking advances. At the same time, wild rats live in close association with us, leading to various adverse interactions. In face of this relevance, it is surprising how little is known about their natural behaviour. While recent laboratory studies revealed their complex social skills, little is known about their social behaviour in the wild. An integration of these different scientific approaches is crucial to understand their social life, which will enable us to design more valid research paradigms, develop more effective management strategies, and to provide better welfare standards. Hence, I first summarise the literature on their natural social behaviour. Second, I provide an overview of recent developments concerning their social cognition. Third, I illustrate why an integration of these areas would be beneficial to optimise our interactions with them.

Harm to Others Acts as a Negative Reinforcer in Rats.

Empathy, the ability to share another individual's emotional state and/or experience, has been suggested to be a source of prosocial motivation by attributing negative value to actions that harm others. The neural underpinnings and evolution of such harm aversion remain poorly understood. Here, we characterize an animal model of harm aversion in which a rat can choose between two levers providing equal amounts of food but one additionally delivering a footshock to a neighboring rat. We find that independently of sex and familiarity, rats reduce their usage of the preferred lever when it causes harm to a conspecific, displaying an individually varying degree of harm aversion. Prior experience with pain increases this effect. In additional experiments, we show that rats reduce the usage of the harm-inducing lever when it delivers twice, but not thrice, the number of pellets than the no-harm lever, setting boundaries on the magnitude of harm aversion. Finally, we show that pharmacological deactivation of the anterior cingulate cortex, a region we have shown to be essential for emotional contagion, reduces harm aversion while leaving behavioral flexibility unaffected. This model of harm aversion might help shed light onto the neural basis of psychiatric disorders characterized by reduced harm aversion, including psychopathy and conduct disorders with reduced empathy, and provides an assay for the development of pharmacological treatments of such disorders. VIDEO ABSTRACT.

Freezing Displayed by Others Is a Learned Cue of Danger Resulting from Co-experiencing Own Freezing and Shock.

Social cues of threat are widely reported [1-3], whether actively produced to trigger responses in others such as alarm calls or by-products of an encounter with a predator, like the defensive behaviors themselves such as escape flights [4-14]. Although the recognition of social alarm cues is often innate [15-17], in some instances it requires experience to trigger defensive responses [4, 7]. One mechanism proposed for how learning from self-experience contributes to social behavior is that of auto-conditioning, whereby subjects learn to associate their own behaviors with relevant trigger events. Through this process, the same behaviors, now displayed by others, gain meaning [18, 19] (but see [20]). Although it has been shown that only animals with prior experience with shock display observational freezing [21-25], suggesting that auto-conditioning could mediate this process, evidence for this hypothesis was lacking. Previously we found that, when a rat freezes, the silence that results from immobility triggers observational freezing in its cage-mate, provided the cage-mate had experienced shocks before [24]. Therefore, in our study, auto-conditioning would correspond to rats learning to associate shock with their own response to it-freezing. Using a combination of behavioral and optogenetic manipulations, here, we show that freezing becomes an alarm cue by a direct association with shock. Our work shows that auto-conditioning can indeed modulate social interactions, expanding the repertoire of cues mediating social information exchange, providing a framework to study how the neural circuits involved in the self-experience of defensive behaviors overlap with the ones involved in socially triggered defensive behaviors.

Empathy in stroke rats is modulated by social settings.

Rodents display "empathy" defined as perceived physical pain or psychological stress by cagemates when co-experiencing socially distinct traumatic events. The present study tested the hypothesis that empathy occurs in adult rats subjected to an experimental neurological disorder, by allowing co-experience of stroke with cagemates. Psychological stress was measured by general locomotor activity, Rat Grimace Scale (RGS), and plasma corticosterone. Physiological correlates were measured by Western blot analysis of advanced glycation endproducts (AGE)-related proteins in the thymus. General locomotor activity was impaired in stroke animals and in non-stroke rats housed with stroke rats suggesting transfer of behavioral manifestation of psychological stress from an injured animal to a non-injured animal leading to social inhibition. RGS was higher in stroke rats regardless of social settings. Plasma corticosterone levels at day 3 after stroke were significantly higher in stroke animals housed with stroke rats, but not with non-stroke rats, indicating that empathy upregulated physiological stress level. The expression of five proteins related to AGE in the thymus reflected the observed pattern of general locomotor activity, RGS, and plasma corticosterone levels. These results indicate that stroke-induced psychological stress manifested on both the behavioral and physiological levels and appeared to be affected by empathy-associated social settings.

Perception of predation risk by African giant pouched rats (Cricetomys sp. nov) is higher in forest-edge microhabitats.

How rodents perceive predation risk may alter their seed foraging behaviour and therefore potentially influence the recruitment of tree species. In this study we used two methods to investigate the effect of predation risk on habitat use by the African giant pouched rat (Cricetomys sp. nov) in Ngel Nyaki forest reserve, Nigeria. The first method was 'giving up density' (GUD), an index of perceived risk of predation at an artificial food patch, and the second was the 'spool-and-line' approach, whereby unravelling spools attached to rodent bodies are used to trace their tracks. For our GUD experiment, we chose four major sites in the forest; two representative of core habitat and two at the forest edge. Additionally, three characteristic microsites were used in the GUD experiment: dense understory, open understory and near-burrows. We hypothesised that GUDs would be lower on every succeeding observation day as rats learn to use the food patches, higher GUDs would be observed in the forest edges and open microsites, and rats would show preference for the microhabitats with least exposure to potential predators. In support of our first hypothesis, we found that GUDs were highest on the first experimental nights of every session. We also found that GUDs in the forest edges were higher than GUDs in the forest core. Lower GUDs were observed close to the rat burrows and in dense understory microsites, even though these differences were not statistically significant. Tracking of rat movements using the spool-and-line method overall revealed an even use across microhabitats, with a weak preference for those with logs, dense understory or exposed ground. Overall, our results suggest that vegetation density on a microhabitat scale has little or no effect on the perception of predation risk by African giant pouched rats.

Dissociation between Postrhinal Cortex and Downstream Parahippocampal Regions in the Representation of Egocentric Boundaries.

Navigation requires the integration of many sensory inputs to form a multi-modal cognitive map of the environment, which is believed to be implemented in the hippocampal region by spatially tuned cells [1-10]. These cells encode various aspects of the environment in a world-based (allocentric) reference frame. Although the cognitive map is represented in allocentric coordinates, the environment is sensed through diverse sensory organs, mostly situated in the animal's head, and therefore represented in sensory and parietal cortices in head-centered egocentric coordinates. Yet it is not clear how and where the brain transforms these head-centered egocentric representations to map-like allocentric representations computed in the hippocampal region. Theoretical modeling has predicted a role for both egocentric and head direction (HD) information in performing an egocentric-allocentric transformation [11-15]. Here, we recorded new data and also used data from a previous study [16]. Adapting a generalized linear model (GLM) classification [17]; we show that the postrhinal cortex (POR) contains a population of pure egocentric boundary cells (EBCs), in contrast with the conjunctive EBCs × HD cells, which we found downstream mostly in the parasubiculum (PaS) and in the medial entorhinal cortex (MEC). Our finding corroborates the idea of a brain network performing an egocentric to allocentric transformation by HD cells. This is a fundamental building block in the formation of the brain's internal cognitive map.

"They're always there": resident experiences of living with rats in a disadvantaged urban neighbourhood.

BACKGROUND: The presence of urban rats in the neighbourhood environment may negatively impact the physical and mental health of residents. Our study sought to describe the experiences with, perceptions of, and feelings towards rats and rat control efforts among a group of disadvantaged urban residents in Vancouver, Canada. METHODS: Semi-structured interviews were held with 20 members of the Vancouver Area Network of Drug Users (VANDU) recruited by VANDU staff. Interviews were audio recorded, transcribed, and analyzed using thematic analysis. RESULTS: Participants reported daily sightings of rats and close contact during encounters. Participants generally disliked encountering rats, raising issues of health and safety for themselves and the community due to the belief that rats carry disease. Fear of rats was common, and in some cases resulted in avoidance of rats. Effects of rats on participants were particularly pronounced for those living with rats in the home or for homeless participants who described impacts on sleep due to the sounds made by rats. Although rats were viewed as more problematic in their neighbourhood than elsewhere in Vancouver, participants believed there to be a lack of neighbourhood-level control initiatives that angered and disheartened participants. In combination with other community-level concerns (e.g., housing quality and availability), the presence of rats was viewed by some to align with a general disregard for the community and its residents. CONCLUSIONS: This study suggests that the presence of rats in urban centres may have several consequences on the physical and mental health of residents living in close contact with them. These effects may be exacerbated with continued contact with rats and when residents perceive a lack of initiative to control rats in their neighbourhood. As such, research and policies aimed at mitigating the health risks posed by rats should extend beyond disease-related risk and incorporate diverse health outcomes.

Novel objects elicit greater activation in the basolateral complex of the amygdala of wild rats compared with laboratory rats.

Wild animals tend to avoid novel objects that do not elicit clear avoidance behaviors in domesticated animals. We previously found that the basolateral complex of the amygdala (BLA) and dorsal bed nucleus of the stria terminalis (dBNST) were larger in trapped wild rats compared with laboratory rats. Based on these findings, we hypothesized that the BLA and/or dBNST would be differentially activated when wild and laboratory rats showed different avoidance behaviors towards novel objects. In this study, we placed novel objects at one end of the home cage. We measured the time spent in that half of the cage and expressed the data as a percentage of the time spent in that region with no object placement. We found that this percentage was lower in the wild rats compared with the laboratory rats. These behavioral differences were accompanied by increased Fos expression in the BLA, but not in the dBNST, of the wild rats. These results suggest that wild rats show greater BLA activation compared with laboratory rats in response to novel objects. We also found increased Fos expression in the paraventricular nucleus of the hypothalamus, ventral BNST, and ventromedial hypothalamus, but not in the central amygdala of wild rats. Taken together, our data represent new information regarding differences in behavioral and neural responses towards novel objects in wild vs. laboratory rats.

An Analysis of Decision under Risk in Rats.

In 1979, Daniel Kahneman and Amos Tversky published a ground-breaking paper titled "Prospect Theory: An Analysis of Decision under Risk," which presented a behavioral economic theory that accounted for the ways in which humans deviate from economists' normative workhorse model, Expected Utility Theory [1, 2]. For example, people exhibit probability distortion (they overweight low probabilities), loss aversion (losses loom larger than gains), and reference dependence (outcomes are evaluated as gains or losses relative to an internal reference point). We found that rats exhibited many of these same biases, using a task in which rats chose between guaranteed and probabilistic rewards. However, prospect theory assumes stable preferences in the absence of learning, an assumption at odds with alternative frameworks such as animal learning theory and reinforcement learning [3-7]. Rats also exhibited trial history effects, consistent with ongoing learning. A reinforcement learning model in which state-action values were updated by the subjective value of outcomes according to prospect theory reproduced rats' nonlinear utility and probability weighting functions and also captured trial-by-trial learning dynamics.

Two open source designs for a low-cost operant chamber using Raspberry Pi™.

After almost a century of use and development, operant chambers remain a significant financial investment for scientists. Small powerful single-board computers such as the Raspberry Pi™ offer researchers a low-cost alternative to expensive operant chambers. In this paper, we describe two new operant chambers, one using nose-poke ports as operanda and another using a touchscreen. To validate the chamber designs, rats learned to perform both visual discrimination and delayed alternation tasks in each chamber. Designs and codes are open source and serve as a starting point for researchers to develop behavioral experiments or educational demonstrations.

Rats Generate Vibrissal Sensory Evidence until Boundary Crossing Triggers a Decision.

Behaviors in which primates collect externally generated streams of sensory evidence, such as judgment of random dot motion direction, are explained by a bounded integration decision model. Does this model extend to rodents, and does it account for behavior in which the motor system generates evidence through interactions with the environment? In this study, rats palpated surfaces to identify the texture before them, showing marked trial-to-trial variability in the number of touches prior to expressing their choice. By high-speed video, we tracked whisker kinematic features and characterized how they encoded the contacted texture. Next, we quantified the evidence for each candidate texture transmitted on each touch by the specified whisker kinematic features. The instant of choice was well fit by modeling the brain as an integrator that gives the greatest weight to vibrissal evidence on first touch and exponentially less weight to evidence on successive touches; according to this model, the rat makes a decision when the accumulated quantity of evidence for one texture reaches a boundary. In summary, evidence appears to be accumulated within the brain until sufficient to support a well-grounded choice. These findings extend the framework of bounded sensory integration from primates to rodents and from passively received evidence to evidence that is actively generated by the sensorimotor system.

Texture Identification by Bounded Integration of Sensory Cortical Signals.

Recent work demonstrated that when a rat palpates a surface to identify its texture, signals generated by whisker kinematics are integrated by the brain, one touch at a time, until the accumulated evidence supports a well-grounded choice. The framework of decision making through bounded integration, previously attributed to primates, thus extends to rodents. In the present study, we ask whether vibrissal somatosensory cortex (vS1 and vS2) functions as the integrator of incoming evidence or, alternatively, as a relay of evidence to a downstream integrator. Rats carried out 1-6 touches per trial to discriminate among candidate textures. We calculated the evidence for each texture, per touch, carried by the firing rates of sets of neurons in vS1 and vS2. The quantity of information within vS1 and vS2 did not grow progressively; instead, the decision was accounted for by modeling a downstream integrator that accumulated packets of vS1 and vS2 texture information until the total quantity of evidence for one texture reached a boundary. In this behavioral task, vibrissal somatosensory cortex appears to act as a sensory relay. Bounded integration is likely to take place in regions targeted by somatosensory cortex.

Increasing predation risk with light reduces speed, exploration and visit duration of invasive ship rats (Rattus rattus).

Exploiting predation cues to deter pests remains an untapped management tool for conservationists. We examined foraging and movement patterns of 20 wild ship rats (Rattus rattus) within a large, outdoor 'U maze' that was either illuminated or dark to assess if light (an indirect predation cue) could deter rodents from ecologically vulnerable locations. Light did not alter rats' foraging behaviour (latency to approach seed tray, visits to seed tray, time per visit to seed tray, total foraging duration, foraging rate) within the experimental resource patch but three of seven movement behaviours were significantly impaired (53% fewer visits to the maze, 70% less exploration within the maze, 40% slower movement within the maze). The total time males spent exposed to illumination also declined by 45 minutes per night, unlike females. Individual visits tended to be longer under illumination, but the latency to visit and the latency to cross through the U maze were unaffected by illumination. Elevating predation risk with illumination may be a useful pest management technique for reducing ship rat activity, particularly in island ecosystems where controlling mammalian predators is paramount to preserving biodiversity.

High mutual cooperation rates in rats learning reciprocal altruism: The role of payoff matrix.

Cooperation is one of the most studied paradigms for the understanding of social interactions. Reciprocal altruism -a special type of cooperation that is taught by means of the iterated prisoner dilemma game (iPD)- has been shown to emerge in different species with different success rates. When playing iPD against a reciprocal opponent, the larger theoretical long-term reward is delivered when both players cooperate mutually. In this work, we trained rats in iPD against an opponent playing a Tit for Tat strategy, using a payoff matrix with positive and negative reinforcements, that is food and timeout respectively. We showed for the first time, that experimental rats were able to learn reciprocal altruism with a high average cooperation rate, where the most probable state was mutual cooperation (85%). Although when subjects defected, the most probable behavior was to go back to mutual cooperation. When we modified the matrix by increasing temptation rewards (T) or by increasing cooperation rewards (R), the cooperation rate decreased. In conclusion, we observe that an iPD matrix with large positive reward improves less cooperation than one with small rewards, shown that satisfying the relationship among iPD reinforcement was not enough to achieve high mutual cooperation behavior. Therefore, using positive and negative reinforcements and an appropriate contrast between rewards, rats have cognitive capacity to learn reciprocal altruism. This finding allows to infer that the learning of reciprocal altruism has early appeared in evolution.

Relationship between motor function and ultrasonic vocalizations induced by maternal separation in rat pups.

When rat pups are isolated from their mothers, they emit ultrasonic vocalizations (USVs). Although previous studies have reported that USVs are related to anxiety, others have reported that they are related to simple, nonemotional factors, such as physiological reactions to coldness. In this study, we examined the influence of three maternal separations on rat pups. The number of USVs during 5 min of USV test under maternal separation, latency in the righting reflex as motor function, and body temperature were recorded twice (the first and second tests) before and after the pups were put in various environments for 10 min. The environments were no maternal separation (Control: CON), maternal separation with littermates (LMS), and single maternal separation with a heater (SMS). In the second test, the SMS pups had fewer USVs, a lower body temperature, and a more rapid righting reflex than the CON and LMS pups. In addition, there was no strong correlation between USVs and righting reflex. As a result, pups undergoing 10 min of SMS while being kept warm by the heater showed rapid righting reflex. Thus, by a single maternal separation, the number of USVs decreased but the decrease was unrelated to decrease in motor function.

Rigor and reproducibility in rodent behavioral research.

Behavioral neuroscience research incorporates the identical high level of meticulous methodologies and exacting attention to detail as all other scientific disciplines. To achieve maximal rigor and reproducibility of findings, well-trained investigators employ a variety of established best practices. Here we explicate some of the requirements for rigorous experimental design and accurate data analysis in conducting mouse and rat behavioral tests. Novel object recognition is used as an example of a cognitive assay which has been conducted successfully with a range of methods, all based on common principles of appropriate procedures, controls, and statistics. Directors of Rodent Core facilities within Intellectual and Developmental Disabilities Research Centers contribute key aspects of their own novel object recognition protocols, offering insights into essential similarities and less-critical differences. Literature cited in this review article will lead the interested reader to source papers that provide step-by-step protocols which illustrate optimized methods for many standard rodent behavioral assays. Adhering to best practices in behavioral neuroscience will enhance the value of animal models for the multiple goals of understanding biological mechanisms, evaluating consequences of genetic mutations, and discovering efficacious therapeutics.