Assessment of executive functions in mice with the Fixed-Ratio schedule of reinforcement

The simplest operant schedule of reinforcement is the Fixed Ratio (FR). In this task, animals must provide a determined number of responses (i.e., a ratio of responses) to obtain each food reward. In FR1, a single response is required, 2 responses in a FR2, 5 in a FR5 and so on. The FR is often used only to shape the operant response before to go further with another task, but of course, it can be used with more interesting goals. Classically, measures such as inter-responses time (IRT), latency to collect the delivered food reward, response or reward rate, but also post reinforcement pause are collected. Together, they allow to obtain information about motor ability, learning and motivation. However, there is a lack of interest in the executive aspect potentially implied by the FR3, especially in situations where the ratio is held constant across multiple training sessions. Indeed, in such situations, as training progress the subject can learn that a certain quantity of responses is required before the reward is available. And hence, checking the food tray to see if the reward is available should be more probable as the animal progress in the ratio. With this operant procedure, it seems therefore possible to investigate the animal’s executive function. In this study, we thus focused on the premature head entries into the empty food tray (PHE, i.e., when the reward was not delivered yet) and their evolution across sessions of FR30. The analysis of the distribution of PHE could offer a way to analyze the subject’s ability to optimize its operant behavior. We also compared this aspect of the operant behavior between young and old mice as the age is factor affecting the executive function

How to Improve and Refine the Elevated Plus Maze for Laboratory Mice

peer reviewedThe daily light-dark cycles synchronize behavioural and physiological processes to the external environment. Light is the most important environmental cue that coordinates many aspects such as activity, maintenance behaviours, hormonal regulation or long-term potentiation. Mice are among the main animals used in behavioural neuroscience and preclinical research laboratories. Although nocturnal, they are generally tested during day (i.e. during their resting phase). Even if convenient for the experimenter, a perturbation of the sleep-wake cycle such as manipulations during day can generate some stress to the animal, produce few reliable data, contributing to reproducibility issue, and may lead to negative consequences for health, physiology, behaviour and cognition. Then the testing moment could be an important variable affecting animal behavior. It appears that rodents are less anxious during the dark phase, however, a lack of data regarding the effect the testing moment on behavior has recently been highlighted. Additionally, several studies evaluating the effects of the testing moment have shown conflicting results; these differences can be explained by various methodological aspects such as parameters related to the procedure but also by the way of collecting and interpreting behavioral data. In this study, we focused on an anxiety test. We tested 4 different testing moment to determine if there is a more appropriate testing moment where mice show the less anxiety. Previously, we have shown that the way of analyzing behavior can affect the results. Here, we refined our observation method to improve the data interpretation

Time to switch off lights? Effects of testing moment in the circadian sleep-wake rhythm

peer reviewedThe daily light-dark cycle allows the synchronization of behavioural and physiological processes to the external environment. Light is the most important environmental cue or zeitgeber that coordinate many aspects of physiology and behaviour such as activity, maintenance behaviours, alertness, body temperature, hormonal regulation or long-term potentiation (i.e. a process that plays a key role in memory consolidation). Mice are among the main animals used in behavioural neuroscience and preclinical research laboratories. Although they are nocturnal animals, mice are generally tested during day (i.e. their resting phase). Thus, the testing moment might be a predominant variable that can affect animal behaviour and cognition and from there, all the inferences we make about behavioural and cognitive processes. In this study, several behavioural and cognitive components such as activity, motivation, learning and behavioural flexibility have been evaluated. We assumed that C57/BL6 mice should perform better and show higher activity rate while tested during night. Although we didn’t find any differences between testing conditions in behavioural or cognitive performance, several factors can explain this absence of effects such as nature of the task, presence of zeitgeber, modulation of arousal, light pollution as well as light conditions during the test itself

Effects of testing moment in the circadian sleep-wake rhythm on learning, memory and behavioural flexibility.

The daily light-dark cycles allow the synchronization of behavioural and physiological processes to the external environment. Light is the most important environmental cue or zeitgeber that coordinates many aspects of physiology and behaviour such as body temperature, hormonal regulation, locomotor activity, maintenance behaviours, alertness or long-term potentiation (i.e. a process that plays a key role in memory consolidation). Mice are among the main animals used in behavioural neuroscience and preclinical research laboratories; although nocturnal, they are generally tested during day (i.e. during their resting phase, under the standard laboratory lightning condition). Convenient for the experimenter, manipulations during day can generate some stress to the animal. Moreover, several cognition studies show that mice performed better when tested during their active phase. Thereby, the testing moment might be a predominant variable affecting animal behaviour and therefore all the inferences we make about cognitive processes. Nevertheless, a lack of data related to the effect of testing moment on behavior was recently highlighted. In this study, we focused on a memory test (the Object Recognition Test, ORT) and on a test learning and behavioral flexibility (the operant Attentional Set-Shifting Task, ASST). Due to their ease of use, both tasks are widely used but rarely with the testing moment as an independent variable. Moreover, particularly for the ORT, those studies have led to conflicting results. Theses discrepancies can be due to several methodological differences. According to other published results in the literature and to some common sense, we supposed that mice should perform better when tested during their active phase. 24 female C57/BL6 mice divided in two groups were singly-housed under a standard light-dark cycle (lights on at 08.00 and off at 20.00, N=12) or under a reversed light-dark cycle (lights on at 20.00 and off at 08.00, N=12). Between-group ANOVAs and t-tests were applied to data. Our findings highlight that mice tested during their active phase show a similar performance related to those tested during their inactive phase during the ASST. However, they discriminated better the new object during the ORT. As cognitive indexes are better for the active group or equivalent between groups, testing rodents during their active period should be privileged not only from an ethical perspective but also to improve the data quality. Moreover, further research should be conducted in this area in order to better understand the real effect of the testing moment on cognition

Effets du moment de test et de paramètres méthodologiques sur le comportement et la cognition de la souris de laboratoire

La plupart des espèces terrestres se sont adaptées au cycle nycthéméral en développant des rythmes circadiens physiologiques et comportementaux. La lumière (ou zeitgeber) est l’indice environnemental le plus important qui influence de nombreux aspects tels que l'activité, la vigilance, la température corporelle, la régulation hormonale ou la potentialisation à long terme. Les souris font partie des principaux animaux employés dans les laboratoires de neurosciences comportementales et de recherche préclinique. Bien que nocturnes, ces dernières sont généralement testées pendant leur phase de repos, le jour, et sous des conditions lumineuses classiques. Bien que cette pratique soit plus commode pour l'expérimentateur, toute perturbation du cycle veille-sommeil (comme les manipulations diurnes liées à la maintenance ou à l’expérimentation), pourrait générer un certain stress pour l'animal et affecter son bien-être et aussi mener à la production de données peu fiables. Ainsi, le moment du test pourrait constituer une variable importante affectant le comportement animal et par là certaines conclusions que nous faisons sur les processus cognitifs. Il semblerait que les rongeurs soient moins anxieux durant la phase sombre, cependant, un manque de données relatif à l'effet du moment de test sur le comportement a récemment été mis en avant. En outre, plusieurs études évaluant les effets du moment de test sur diverses capacités cognitives ont montré des résultats contradictoires ; ces écarts peuvent s'expliquer par divers aspects méthodologiques tels que les paramètres liés à la procédure (comme les conditions d'éclairage au moment du test, l’espèce, voire la souche étudiée) mais également par la manière de recueillir et d’interpréter les données comportementales. Dans cette étude, nous nous sommes concentrés sur un test d'anxiété (le labyrinthe en croix surélevé, EPM) et sur un test de mémoire (le test de reconnaissance d’objets, ORT). Nous avons comparé les performances de quatre groupes de sujets testés à 4 moments différents du cycle circadien veille-sommeil (deux dans la phase active et deux dans la phase de repos) pour déterminer s'il y avait un moment de test plus approprié pour l'expérimentation animale où les souris montreraient une anxiété réduite et de meilleures performances. Nous montrons que la manière d'analyser le comportement peut affecter la conclusion concernant l'influence du moment du test sur la performance et suggérons que les paramètres procéduraux et d’analyses soient davantage considérés et justifiés afin d’aboutir à de meilleures conclusions relatives au comportement de l’animal