In animals, emotional memory is classically assessed through pavlovian fear conditioning in which a neutral novel stimulus (conditioned stimulus) is paired with an aversive unconditioned stimulus. After conditioning, the conditioned stimulus elicits a fear response characterized by a wide range of behavioral and physiological responses. Despite the existence of this large repertoire of responses, freezing behavior is often the sole parameter used for quantifying fear response, thus limiting emotional memory appraisal to this unique index. Interestingly, respiratory changes and ultrasonic vocalizations (USV) can occur during fear response, yet very few studies investigated the link between these different parameters and freezing. The aim of the present study was to design an experimental setup allowing the simultaneous recording of respiration, USV, and behavior (RUB cage), and the offline synchronization of the collected data for fine-grain second by second analysis. The setup consisted of a customized plethysmograph for respiration monitoring, equipped with a microphone capturing USV, and with four video cameras for behavior recording. In addition, the bottom of the plethysmograph was equipped with a shock-floor allowing foot-shock delivery, and the top received tubing for odor presentations. Using this experimental setup we first described the characteristics of respiration and USV in different behaviors and emotional states. Then we monitored these parameters during contextual fear conditioning and showed that they bring complementary information about the animal's anxiety state and the strength of aversive memory. The present setup may be valuable in providing a clearer appraisal of the physiological and behavioral changes that occur during acquisition as well as retrieval of emotional memory.

Insulin is involved in multiple regulatory mechanisms, including body weight and food intake, and plays a critical role in metabolic disorders such as obesity and diabetes. An increasing body of evidence indicates that insulin is also involved in the modulation of olfactory function. The olfactory bulb (OB) contains the highest level of insulin and insulin receptors (IRs) in the brain. However, a role for insulin in odor detection and sniffing behavior remains to be elucidated. Using a behavioral paradigm based on conditioned olfactory aversion (COA) to isoamyl-acetate odor, we demonstrated that an intracerebroventricular (ICV) injection of 14 mU insulin acutely decreased olfactory detection of fasted rats to the level observed in satiated animals. In addition, whereas fasted animals demonstrated an increase in respiratory frequency upon food odor detection, this effect was absent in fasted animals receiving a 14 mU insulin ICV injection as well as in satiated animals. In parallel, we showed that the OB and plasma insulin levels were increased in satiated rats compared to fasted rats, and that a 14 mU insulin ICV injection elevated the OB insulin level of fasted rats to that of satiated rats. We further quantified insulin receptors (IRs) distribution and showed that IRs are preferentially expressed in the caudal and lateral parts of the main OB, with the highest labeling found in the mitral cells, the main OB projection neurons. Together, these data suggest that insulin acts on the OB network to modulate olfactory processing and demonstrate that olfactory function is under the control of signals involved in energy homeostasis regulation and feeding behaviors.