An efficient and reproducible method for transformation of genetically recalcitrant bifidobacteria

This study describes an efficient transformation system for the introduction of plasmid DNA into Bifidobacterium bifidum PRL2010 and Bifidobacterium asteroides PRL2011, for which to the best of our knowledge no transformation data have been reported previously. The method is based on electroporation of bifidobacterial cells, which were made competent by an optimized methodology based on varying media and growth conditions. Furthermore, the transformation protocol was applied in order to design a PRL2010-derivative, which carries antibiotic resistance against chloramphenicol and which was used to monitor PRL2010 colonization in a murine model

Chronic subordination stress induces hyperphagia and disrupts eating behavior in mice modeling binge-eating-like disorder

Background: Eating disorders are associated with physical morbidity and appear to have causal factors like stressful life events and negative affect. Binge-eating disorder (BED) is characterized by eating in a discrete period of time a larger than normal amount of food, a sense of lack of control over eating, and marked distress. There are still unmet needs for the identification of mechanisms regulating excessive eating, which is in part due to the lack of appropriate animal models. We developed a naturalistic murine model of subordination stress-induced hyperphagia associated with the development of obesity. Here, we tested the hypotheses that the eating responses of subordinate mice recapitulate the BED and that limiting hyperphagia could prevent stress-associated metabolic changes. Methods: Adult male mice were exposed to a model of chronic subordination stress (CSS) associated with the automated acquisition of food intake and we performed a detailed meal pattern analysis. Additionally, using a pair-feeding protocol we tested the hypothesis that the manifestation of obesity and the metabolic syndrome could be prevented by limiting hyperphagia. Results: The architecture of feeding of subordinate mice was disrupted during the stress protocol due to disproportionate amount of food ingested at higher rate and with shorter satiety ratio than control mice. Subordinate mice hyperphagia was further exacerbated in response to either hunger or to the acute application of a social defeat. Notably, the obese phenotype but not the fasting hyperglycemia of subordinate mice was abrogated by preventing hyperphagia in a pair-feeding paradigm. Conclusion: Overall, these results support the validity of our CSS to model BED allowing for the determination of the underlying molecular mechanisms and the generation of testable predictions for innovative therapies, based on the understanding of the regulation and the control of food intake

Aggressive behaviour and physiological responses to pheromones are strongly impaired in mice deficient for the olfactory G-protein -subunit G8

Heterotrimeric G-proteins are critical players in the transduction mechanisms underlying odorant and pheromonal signalling. In the vomeronasal organ (VNO) of the adult mouse, two different G-protein complexes have been identified. Gαoβ2γ8 is preferentially expressed in the basal neurons and coexpresses with type-2 vomeronasal pheromone receptors (V2Rs) whereas Gαi2β2γ2 is found in the apical neurons and coexpresses with type-1 vomeronasal pheromone receptors (V1Rs). V2R-expressing neurons project to the posterior accessory olfactory bulb (AOB) whereas neurons expressing V1Rs send their axon to the anterior AOB. Gγ8 is also expressed in developing olfactory neurons where this protein is probably associated with Go. Here, we generated mice with a targeted deletion of the Gγ8 gene and investigated the behavioural effects and the physiological consequences of this mutation. Gγ8(−/−) mice show a normal development of the main olfactory epithelium; moreover, they do not display major deficits in odour perception. In contrast, the VNO undergoes a slow but remarkable loss of basal neurons starting from the fourth postnatal week, with a 40% reduction of cells at 2 months and 70% at 1 year. This loss is associated with a reduced early-gene expression in the posterior AOB of mice stimulated with pheromones. More interestingly, the Gγ8 deletion specifically leads to a reduced pheromone-mediated aggressiveness in both males and females, all other socio-sexual behaviours remaining unaltered. This study defines a specific role for Gγ8 in maintenance of the neuronal population of the VNO and in the mechanisms of pheromonal signalling that involve the aggressive behaviour towards conspecifics

An efficient and reproducible method for transformation of genetically recalcitrant bifidobacteria

This study describes an efficient transformation system for the introduction of plasmid DNA into Bifidobacterium bifidum PRL2010 and Bifidobacterium asteroides PRL2011, for which to the best of our knowledge no transformation data have been reported previously. The method is based on electroporation of bifidobacterial cells, which were made competent by an optimized methodology based on varying media and growth conditions. Furthermore, the transformation protocol was applied in order to design a PRL2010-derivative, which carries antibiotic resistance against chloramphenicol and which was used to monitor PRL2010 colonization in a murine model

Psychosocial stress induces hyperphagia and exacerbates diet-induced insulin resistance and the manifestations of the Metabolic Syndrome.

Stress and hypercaloric food are recognized risk factors for obesity, Metabolic Syndrome (MetS) and Type 2 Diabetes (T2D). Given the complexity of these metabolic processes and the unavailability of animal models, there is poor understanding of their underlying mechanisms. We established a model of chronic psychosocial stress in which subordinate mice are vulnerable to weight gain while dominant mice are resilient. Subordinate mice fed a standard diet showed marked hyperphagia, high leptin, low adiponectin, and dyslipidemia. Despite these molecular signatures of MetS and T2D, subordinate mice fed a standard diet were still euglycemic. We hypothesized that stress predisposes subordinate mice to develop T2D when synergizing with other risk factors. High fat diet aggravated dyslipidemia and the MetS thus causing a pre-diabetes-like state in subordinate mice. Contrary to subordinates, dominant mice were fully protected from stress-induced metabolic disorders when fed both a standard- and a high fat-diet. Dominant mice showed a hyperphagic response that was similar to subordinate but, unlike subordinates, showed a significant increase in VO2, VCO2, and respiratory exchange ratio when compared to control mice. Overall, we demonstrated a robust stress- and social status-dependent effect on the development of MetS and T2D and provided insights on the physiological mechanisms. Our results are reminiscent of the effect of the individual socioeconomic status on human health and provide an animal model to study the underlying molecular mechanisms

Implication of the VGF-derived peptide TLQP-21 in mouse acute and chronic stress responses.

Abstract The impact of stress is widely recognized in the etiology of multiple disorders. In particular, psychological stress may increase the risk of cardiovascular, metabolic, immune, and mood disorders. Several genes are considered potential candidates to account for the deleterious consequences of stress and recent data point to role of Vgf. VGF mRNA is abundantly expressed in the hypothalamus, where it has been involved in metabolism and energy homeostasis; more recently a link between VGF-derived peptides and mood disorders has been highlighted. The following experiments were performed to address the contribution of the VGF-system to stress induced changes in mice: the distribution of VGF immuno-reactivity in hypothalamic nuclei and its modulation by social stress; the role of VGF-derived peptide TLQP-21 in plasma catecholamine release induced by acute restraint stress (RS); the efficacy of chronic TLQP-21 in a mouse model of chronic subordination stress (CSS). VGF fibers were found in high density in arcuate, dorsomedial, and suprachiasmatic and, at lower density, in lateral, paraventricular, and ventromedial hypothalamic nuclei. Central administration of either 2 or 4 mM TLQP-21 acutely altered the biphasic serum epinephrine release and decreased norepinephrine serum levels in response to RS. Finally, 28-day of 40 μg/day TLQP-21 treatment increased CSS-induced social avoidance of an unfamiliar conspecific. Overall these data support a role for TLQP-21 in stress responses providing a promising starting point to further elucidate its role as a player in stress-related human pathologies