Leptin, a hormone secreted from adipose tissue, was originally discovered to regulate body weight. The localization of the leptin receptor in limbic structures suggests a potential role for leptin in emotional processes. Here, we show that rats exposed to chronic unpredictable stress and chronic social defeat exhibit low leptin levels in plasma. Systemic leptin treatment reversed the hedoniclike deficit induced by chronic unpredictable stress and improved behavioral despair dose-dependently in the forced swim test (FST), a model widely used for screening potential antidepressant efficacy. The behavioral effects of leptin in the FST were accompanied by increased neuronal activation in limbic structures, particularly in the hippocampus. Intrahippocampal infusion of leptin produced a similar antidepressant-like effect in the FST as its systemic administration. By contrast, infusion of leptin into the hypothalamus decreased body weight but had no effect on FST behavior. These findings suggest that: (i) impaired leptin production and secretion may contribute to chronic stress-induced depression-like phenotypes, (ii) the hippocampus is a brain site mediating leptin's antidepressant-like activity, and (iii) elevating leptin signaling in brain may represent a novel approach for the treatment of depressive disorders.depression ͉ hippocampus ͉ stress ͉ forced swim test ͉ social defeat

INTRODUCTION: Research on the human microbiome has revealed extensive correlations between bacterial populations and host physiology and disease states. However, moving past correlations to understanding causal relationships between the bacteria in our bodies and our health remains a challenge. A well-studied human-bacteria relationship is that of certain gut Escherichia coli strains whose presence correlates with colorectal cancer in humans. These E. coli damage host DNA and cause tumor formation in animal models, and this genotoxic phenotype is thought to derive from a secondary metabolite—known as colibactin—that is synthesized by the bacteria. Because colibactin’s biosynthetic pathway is only partially resolved, the complete structure of colibactin has remained unknown for more than a decade. Similarly, because colibactin is unstable and is produced in vanishingly small quantities, it has yet to be isolated and characterized by means of standard spectroscopic methods. RATIONALE: Determining colibactin’s chemical structure and related biological activity will allow researchers to determine whether the metabolite is the causal agent underlying many colorectal cancers. To that end, we used an interdisciplinary approach to overcome the challenges that have impeded determination of colibactin’s structure. Inspired by an earlier study that showed that colibactin-producing bacteria cross-link DNA, we used DNA as a probe to isolate colibactin from bacterial cultures. Using a combination of isotope labeling and tandem mass spectrometry analysis, we deduced the structure of the colibactin residue when bound to two nucleobases. This information allowed us to then identify and characterize colibactin in bacterial extracts and to identify plausible biosynthetic precolibactin precursors. Last, we developed a method to recreate colibactin in the laboratory and thereby confirm these structure-function relationships. RESULTS: Colibactin is formed through the union of two complex biosynthetic intermediates. This coupling generates a nearly symmetrical structure that contains two electrophilic cyclopropane warheads. We found that each of these residues undergoes ring-opening through nucleotide addition, a determination that is consistent with earlier studies of truncated colibactin derivatives and the observation that colibactin-producing bacteria cross-link DNA. Using genome editing techniques, we were able to show that the production of colibactin’s precursor, precolibactin 1489, requires every biosynthetic gene in the colibactin gene cluster, implicating it as being derived from the long-elusive and now completed biosynthetic pathway. Because natural colibactin remains non-isolable, the chemical synthetic route to colibactin we developed will allow researchers to probe for causal relationships between the metabolite and inflammation-associated colorectal cancer. CONCLUSION: These studies reveal the structure of colibactin, which accounts for the entire gene cluster encoding its biosynthesis, a goal that has remained b...

SUMMARY The hippocampus is an integral brain region for affective disorders. TRIP8b knockout mice lacking functional HCN channels as well as both HCN1 and HCN2 knockout mice have been shown to display antidepressant-like behaviors. The mechanisms or brain regions involved in these alterations in behavior, however, are not clear. We developed a lentiviral shRNA system to examine whether knockdown of HCN1 protein in the dorsal hippocampal CA1 region is sufficient to produce antidepressant-like effects. We found that knockdown of HCN1 channels increased cellular excitability and resulted in physiological changes consistent with a reduction of Ih. Rats infused with lentiviral shRNA-HCN1 in the dorsal hippocampal CA1 region displayed antidepressant- and anxiolytic-like behaviors associated with widespread enhancement of hippocampal activity and upregulation of BDNF-mTOR signaling pathways. Our results suggest that HCN1 protein could be a potential target for treatment of anxiety and depression disorders.

Rationale Our previous studies in rats have shown that the adipocyte-derived hormone leptin induces antidepressant-like effects with a behavioral profile similar to selective serotonin reuptake inhibitor (SSRI) antidepressants. Acute SSRI treatment causes paradoxical anxiogenic responses, although chronic treatment has therapeutic effects on anxiety. However, the role of leptin in anxiety remains to be established. Objectives The scope of this study was to investigate the acute effects of leptin on anxiety-related behaviors in comparison with the SSRI antidepressant fluoxetine. Materials and methods Adult male C57BL/6J mice received intraperitoneal injection of leptin or fluoxetine. Thirty minutes after injection, mice were subjected to the tail suspension test (TST) and forced swim test (FST) for evaluating antidepressant activity. Anxiety-like behavior was assessed in the elevated plus maze (EPM), social interaction, and open field tests 30 min following drug treatment. Results While leptin and fluoxetine showed similar antidepressant-like behavioral effects in the TST and FST, they differed in the behavioral assays for anxiety. Open arm exploration in the EPM was increased by leptin but decreased by fluoxetine. Analysis of social interaction revealed that distinct social behavioral components were modulated by leptin and fluoxetine. The total time of active social behaviors was increased by leptin but reduced by fluoxetine. In addition, self-grooming, a non-social behavior, was suppressed by leptin treatment. Neither leptin nor fluoxetine produced significant effects in the open field test. Conclusions In contrast to anxiogenic-like effects induced by acute fluoxetine, leptin elicits anxiolytic-like effects after acute administration. These results suggest that leptin has both antidepressant-like and anxiolytic-like properties.

The dorsal and ventral hippocampi are functionally and anatomically distinct. Recently, we reported that dorsal Cornu Ammonis area 1 (CA1) neurons have a more hyperpolarized resting membrane potential and a lower input resistance and fire fewer action potentials for a given current injection than ventral CA1 neurons. Differences in the hyperpolarization-activated cyclic nucleotide-gated cation conductance between dorsal and ventral neurons have been reported, but these differences cannot fully account for the different resting properties of these neurons. Here, we show that coupling of A1 adenosine receptors (A1ARs) to G-protein-coupled inwardly rectifying potassium (GIRK) conductance contributes to the intrinsic membrane properties of dorsal CA1 neurons but not ventral CA1 neurons. The block of GIRKs with either barium or the more specific blocker Tertiapin-Q revealed that there is more resting GIRK conductance in dorsal CA1 neurons compared with ventral CA1 neurons. We found that the higher resting GIRK conductance in dorsal CA1 neurons was mediated by tonic A1AR activation. These results demonstrate that the different resting membrane properties between dorsal and ventral CA1 neurons are due, in part, to higher A1AR-mediated GIRK activity in dorsal CA1 neurons.

Escherichia coli is a common inhabitant of the human microbiota and a beacon model organism in biology. However, an understanding of its signaling systems that regulate population-level phenotypes known as quorum sensing remain incomplete. Here, we define the structure and biosynthesis of autoinducer-3 (AI-3), a metabolite of previously unknown structure involved in the pathogenesis of enterohemorrhagic E. coli (EHEC). We demonstrate that novel AI-3 analogs are derived from threonine dehydrogenase (Tdh) products and “abortive” tRNA synthetase reactions, and they are distributed across a variety of Gram-negative and Gram-positive bacterial pathogens. In addition to regulating virulence genes in EHEC, we show that the metabolites exert diverse immunological effects on primary human tissues. The discovery of AI-3 metabolites and their biochemical origins now provides a molecular foundation for investigating the diverse biological roles of these elusive yet widely distributed bacterial signaling molecules.

Certain commensal Escherichia coli contain the clb biosynthetic gene cluster that codes for small molecule prodrugs known as precolibactins. Precolibactins are converted to colibactins by N-deacylation; the latter are postulated to be genotoxic and to contribute to colorectal cancer formation. Though advances toward elucidating (pre)colibactin biosynthesis have been made, the functions and mechanisms of several clb gene products remain poorly understood. Here we report the 2.1 Å X-ray structure and molecular function of ClbS, a gene product that confers resistance to colibactin toxicity in host bacteria and which has been shown to be important for bacterial viability. The structure harbors a potential colibactin binding site and shares similarity to known hydrolases. In vitro studies using a synthetic colibactin analog and ClbS or an active site residue mutant reveal cyclopropane hydrolase activity that converts the electrophilic cyclopropane of the colibactins into an innocuous hydrolysis product. As the cyclopropane has been shown to be essential for genotoxic effects in vitro, this ClbS-catalyzed ring-opening provides a means for the bacteria to circumvent self-induced genotoxicity. Our study provides a molecular-level view of the first reported cyclopropane hydrolase and support for a specific mechanistic role of this enzyme in colibactin resistance.

The clb gene cluster encodes the biosynthesis of metabolites known as precolibactins and colibactins. The clb pathway is found in gut commensal E. coli, and clb metabolites are thought to initiate colorectal cancer via DNA cross-linking. Here we report confirmation of the structural assignment of the complex clb product precolibactin 886 via a biomimetic synthetic pathway. We show that a α-ketoimine linear precursor undergoes spontaneous cyclization to precolibactin 886 upon HPLC purification. Studies of this α-ketoimine and the related α-dicarbonyl revealed that these compounds are unexpectedly susceptible to nucleophilic cleavage under mildly basic conditions. This cleavage pathway forms other known clb metabolites or biosynthetic intermediates and explains the difficulties in isolating fully mature biosynthetic products. This cleavage also accounts for a recently identified colibactin-adenine adduct. The colibactin peptidase ClbP deacylates synthetic precolibactin 886 to form a non-genotoxic pyridone, suggesting precolibactin 886 lies off-path of the major biosynthetic route.