PDE-10A inhibitors as insulin secretagogues

Modulation of cAMP levels has been linked to insulin secretion in preclinical animal models and in humans. The high expression of PDE-10A in pancreatic islets suggested that inhibition of this enzyme may provide the necessary modulation to elicit increased insulin secretion. Using an HTS approach, we have identified quinoline-based PDE-10A inhibitors as insulin secretagogues in vitro. Optimized compounds were evaluated in vivo where improvements in glucose tolerance and increases in insulin secretion were measured.     

Central nervous system-specific knockout of steroidogenic factor 1 results in increased anxiety-like behavior

Steroidogenic factor 1 (SF-1) plays key roles in adrenal and gonadal development, expression of pituitary gonadotropins, and development of the ventromedial hypothalamic nucleus (VMH). If kept alive by adrenal transplants, global knockout (KO) mice lacking SF-1 exhibit delayed-onset obesity and decreased locomotor activity. To define specific roles of SF-1 in the VMH, we used the Cre-loxP system to inactivate SF-1 in a central nervous system (CNS)-specific manner. These mice largely recapitulated the VMH structural defect seen in mice lacking SF-1 in all tissues. In multiple behavioral tests, mice with CNS-specific KO of SF-1 had significantly more anxiety-like behavior than wild-type littermates. The CNS-specific SF-1 KO mice had diminished expression or altered distribution in the mediobasal hypothalamus of several genes whose expression has been linked to stress and anxiety-like behavior, including brain-derived neurotrophic factor, the type 2 receptor for CRH (Crhr2), and Ucn 3. Moreover, transfection and EMSAs support a direct role of SF-1 in Crhr2 regulation. These findings reveal important roles of SF-1 in the hypothalamic expression of key regulators of anxiety-like behavior, providing a plausible molecular basis for the behavioral effect of CNS-specific KO of this nuclear receptor.     

Robust Vaginal Colonization of Macaques with a Novel Vaginally Disintegrating Tablet Containing a Live Biotherapeutic Product to Prevent HIV Infection in Women

MucoCept is a biotherapeutic for prevention of HIV-1 infection in women and contains a human, vaginal Lactobacillus jensenii that has been genetically enhanced to express the HIV-1 entry inhibitor, modified cyanovirin-N (mCV-N). The objective of this study was to develop a solid vaginal dosage form that supports sustained vaginal colonization of the MucoCept Lactobacillus at levels previously shown, with freshly prepared cultures, to protect macaques from SHIV infection and to test this formulation in a macaque vaginal colonization model. Vaginally disintegrating tablets were prepared by lyophilizing the formulated bacteria in tablet-shaped molds, then packaging in foil pouches with desiccant. Disintegration time, potency and stability of the tablets were assessed. For colonization, non-synchronized macaques were dosed vaginally with either one tablet or five tablets delivered over five days. Vaginal samples were obtained at three, 14, and 21 days post-dosing and cultured to determine Lactobacillus colonization levels. To confirm identity of the MucoCept Lactobacillus strain, genomic DNA was extracted from samples on days 14 and 21 and a strain-specific PCR was performed. Supernatants from bacteria were tested for the presence of the mCV-N protein by Western blot. The tablets were easy to handle, disintegrated within two minutes, potent (5.7x10¹¹ CFU/g), and stable at 4°C and 25°C. Vaginal administration of the tablets to macaques resulted in colonization of the MucoCept Lactobacillus in 66% of macaques at 14 days post-dosing and 83% after 21 days. There was no significant difference in colonization levels for the one or five tablet dosing regimens (p=0.88 Day 14, p=0.99 Day 21). Strain-specific PCR confirmed the presence of the bacteria even in culture-negative macaques. Finally, the presence of mCV-N protein was confirmed by Western blot analysis using a specific anti-mCV-N antibody.     

Dissociable Influences of Skewness and Valence on Economic Choice and Neural Activity

Asymmetry in distributions of potential outcomes (i.e. skewness), and whether those potential outcomes reflect gains or losses (i.e. their valence), both exert a powerful influence on value-based choice. How valence affects the impact of skewness on choice is unknown. Here by orthogonally manipulating the skewness and valence of economic stimuli we show that both have an influence on choice. We show that the influence of skewness on choice is independent of valence, both across and within subjects. fMRI data revealed skew-related activity in bilateral anterior insula and dorsomedial prefrontal cortex, which shows no interaction with valence. Further, the expression of skew-related activity depends on an individual’s preference for skewness, and this was again independent of valence-related preference. Our findings highlight the importance of skewness in choice and show that its influence, both behaviourally and neurally, is distinct from an influence of valence.     

Non-surgical Intratracheal Instillation of Mice with Analysis of Lungs and Lung Draining Lymph Nodes by Flow Cytometry

Phagocytic cells such as alveolar macrophages and lung dendritic cells (LDCs) continuously sample antigens from the alveolar spaces in the lungs. LDCs, in particular, are known to migrate to the lung draining lymph nodes (LDLNs) where they present inhaled antigens to T cells initiating an appropriate immune response to a variety of immunogens^1,2. To model interactions between the lungs and airborne antigens in mice, antigens can be administered intranasally^1,3,4, intratracheally⁵ or as aerosols⁶. Delivery by each route involves distinct technical skills and limitations that need to be considered before designing an experiment. For example, intranasal and aerosolized exposure delivers antigens to both the lungs and the upper respiratory tract. Hence antigens can access the nasal associated lymphoid tissue (NALT)⁷, potentially complicating interpretation of the results. In addition, swallowing, sneezing and the breathing rate of the mouse may also lead to inconsistencies in the doses delivered. Although the involvement of the upper respiratory tract may be preferred for some studies, it can complicate experiments focusing on events specifically initiated in the lungs. In this setting, the intratracheal (i.t) route is preferable as it delivers test materials directly into the lungs and bypasses the NALT. Many i.t injection protocols involve either blind intubation of the trachea through the oral cavity or surgical exposure of the trachea to access the lungs. Herein, we describe a simple, consistent, non-surgical method for i.t instillation. The opening of the trachea is visualized using a laryngoscope and a bent gavage needle is then inserted directly into the trachea to deliver the innoculum. We also describe procedures for harvesting and processing of LDLNs and lungs for analysis of antigen trafficking by flow cytometry.Download video file.^{(48M, mov)}     

Drosophila follicle cells: morphogenesis in an eggshell

Epithelial morphogenesis is important for organogenesis and pivotal for carcinogenesis, but mechanisms that control it are poorly understood. The Drosophila follicular epithelium is a genetically tractable model to understand these mechanisms in vivo. This epithelium of follicle cells encases germline cells to create an egg. In this review, we summarize progress toward understanding mechanisms that maintain the epithelium or permit migrations essential for oogenesis. Cell-cell communication is important, but the same signals are used repeatedly to control distinct events. Understanding intrinsic mechanisms that alter responses to developmental signals will be important to understand regulation of cell shape and organization.