The Enigmatic Canal-Associated Neurons Regulate Caenorhabditis elegans Larval Development Through a cAMP Signaling Pathway.

Caenorhabditis elegans larval development requires the function of the two Canal-Associated Neurons (CANs): killing the CANs by laser microsurgery or disrupting their development by mutating the gene ceh-10 results in early larval arrest. How these cells promote larval development, however, remains a mystery. In screens for mutations that bypass CAN function, we identified the gene kin-29, which encodes a member of the Salt-Inducible Kinase (SIK) family and a component of a conserved pathway that regulates various C. elegans phenotypes. Like kin-29 loss, gain-of-function mutations in genes that may act upstream of kin-29 or growth in cyclic-AMP analogs bypassed ceh-10 larval arrest, suggesting that a conserved adenylyl cyclase/PKA pathway inhibits KIN-29 to promote larval development, and that loss of CAN function results in dysregulation of KIN-29 and larval arrest. The adenylyl cyclase ACY-2 mediates CAN-dependent larval development: acy-2 mutant larvae arrested development with a similar phenotype to ceh-10 mutants, and the arrest phenotype was suppressed by mutations in kin-29 ACY-2 is expressed predominantly in the CANs, and we provide evidence that the acy-2 functions in the CANs to promote larval development. By contrast, cell-specific expression experiments suggest that kin-29 acts in both the hypodermis and neurons, but not in the CANs. Based on our findings, we propose two models for how ACY-2 activity in the CANs regulates KIN-29 in target cells

A Serratia marcescens PigP Homolog Controls Prodigiosin Biosynthesis, Swarming Motility and Hemolysis and Is Regulated by cAMP-CRP and HexS

Swarming motility and hemolysis are virulence-associated determinants for a wide array of pathogenic bacteria. The broad host-range opportunistic pathogen Serratia marcescens produces serratamolide, a small cyclic amino-lipid, that promotes swarming motility and hemolysis. Serratamolide is negatively regulated by the transcription factors HexS and CRP. Positive regulators of serratamolide production are unknown. Similar to serratamolide, the antibiotic pigment, prodigiosin, is regulated by temperature, growth phase, HexS, and CRP. Because of this co-regulation, we tested the hypothesis that a homolog of the PigP transcription factor of the atypical Serratia species ATCC 39006, which positively regulates prodigiosin biosynthesis, is also a positive regulator of serratamolide production in S. marcescens. Mutation of pigP in clinical, environmental, and laboratory strains of S. marcescens conferred pleiotropic phenotypes including the loss of swarming motility, hemolysis, and severely reduced prodigiosin and serratamolide synthesis. Transcriptional analysis and electrophoretic mobility shift assays place PigP in a regulatory pathway with upstream regulators CRP and HexS. The data from this study identifies a positive regulator of serratamolide production, describes novel roles for the PigP transcription factor, shows for the first time that PigP directly regulates the pigment biosynthetic operon, and identifies upstream regulators of pigP. This study suggests that PigP is important for the ability of S. marcescens to compete in the environment. © 2013 Shanks et al

Recurrent deletions of ULK4 in schizophrenia : a gene crucial for neuritogenesis and neuronal motility

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THE ROLE OF PHOSPHODIESTERASE 3B IN CAMP-MEDIATED REGULATION OF INSULIN SECRETION

Type 2 diabetes mellitus (T2DM) is characterized by various combinations of ?-cell failure and insulin resistance leading to hyperglycemia and glucose intolerance. In order to maintain glucose tolerance in the insulin resistance state, increased insulin secretion is a requirement and it is because of inadequate islet adaptation that glucose intolerance develops in T2DM. The pathophysiology of T2DM is not fully understood and more knowledge is needed concerning both insulin action and ?-cell physiology and adaptation. The general aim of this thesis was to investigate the role of ?-cell cAMP-degrading phosphodiesterase 3B (PDE3B) in the regulation of insulin secretion and whole body energy homeostasis. The specific aims were (i) to study the physiological importance of well-regulated ?-cell-cAMP levels for insulin release and whole body energy homeostasis during a long-term metabolic challenge, (ii) to evaluate the role of PDE3B in biphasic insulin secretion and its intracellular localization, and (iii) to investigate the mechanisms for regulation of PDE3B activity in ?-cells. It was previously shown that PDE3B attenuates glucose-stimulated insulin secretion and glucagon-like peptide-1 (GLP-1) potentiated-insulin secretion. It is shown here that accurate regulation of ?-cell cAMP is necessary for adequate islet adaptation to a perturbed metabolic environment and protective for the development of glucose intolerance and insulin resistance. This finding is coupled to the novel discovery that PDE3B, shown to localize to the exocytotic machinery, functions as a specific attenuator of cAMP-mediated potentiation of depolarization-induced insulin secretion. Further, we have begun to elucidate the details concerning the regulation of PDE3B activity in ?-cells. Data are presented suggesting that PDE3B activity in ?-cells is intricately regulated by phosphorylation and dephosphorylation, probably by the action of several differentially regulated kinases and phosphatases. In conclusion, this thesis contributes to the knowledge regarding the importance and function of cAMP-mediated regulation of stimulus-secretion coupling in pancreatic ?-cells and demonstrates that dysfunction of cAMP-PDE3B signalling results in a substantially increased sensitivity to the adverse effects of a high-fat diet

Survey of occurrence of bacteria Vibrio cholera in the two provinces Baghdad and Babel

The present study was branched into two lines ,the first line aim to study Tigris River within Baghdad city  and Euphrates  River within Babel city to assess physical ,chemical and biological factors of  rivers water that Affect  the quality of water . the second line is isolating and diagnosis of  Vibrio  cholera bacteria from clinical and environmental sources in the two provinces and study of its  sensitivity against (13) antibiotic also conducting a molecular study to detect  the presence of plasmids and the toxin genes in All  isolated bacteria . the study area included three stations on Tigris river in Baghdad city and three stations on Euphrates river in Babel city , monthly samples were collected from October 2015 to September 2016 in addition to samples were collected from patients in three Hospitals  by using sterile containers . Keywords : Choleragen , Primers , DNA agarose gel electrophoresi

The role of purinergic receptor A1 in neurogenesis modulation from subventricular zone

138 p.La neurogénesis continúa en la edad adulta en regiones específicas del cerebro como la zona subgranular del hipocampo y la zona subventricular (SVZ) de los ventrículos laterales. Resultados previos de nuestro laboratorio demostraron que el ATP liberado tras la deprivación de oxígeno y glucosa inhibe la neurogénesis adulta. Por lo tanto, nuestro objetivo es determinar el papel de la adenosina, uno de los productos de la hidrólisis del ATP, en la modulación de la neurogénesis. Los resultados obtenidos demuestran que altas concentraciones de adenosina (100¿M) inhiben la diferenciación neuronal en cultivos de neuroesferas de la SVZ. Las células multipotentes de la SVZ expresan todos los receptores de adenosina (A1, A2a, A2b y A3); sin embargo el receptor A1 es el involucrado en la inhibición de la diferenciación neuronal como demostramos por PCR cuantitativa, Western Blot y en un ensayo de silenciamiento génico del receptor A1. Además, la activación del receptor A1 indujo una disminución de la expresión de genes relacionados con la neurogénesis como observamos en un análisis de expresión génica. El efecto inhibitorio de la activación del receptor A1 fue también confirmado en un modelo in vivo; de manera que observamos una reducción de la neurogénesis y un aumento de la astrogliogénesis en el bulbo olfatorio de ratas adultas tras la infusión intracerebroventricular del agonista del receptor A1 CPA. A su vez, el estudio de los mecanismos por los que la adenosina inhibe la neurogénesis y sostiene la astrogliogénesis demostraron la implicación de la IL10 y la activación de la ruta STAT3/Bmp2/Smad. Además, dado que la adenosina es liberada de forma masiva durante la isquemia cerebral, estudiamos el efecto del bloqueo del receptor A1 en un modelo de isquemia cerebral (oclusión transitoria de la arteria cerebral media). El antagonismo del receptor A1 produjo un aumento del número de nuevas neuronas (células positivas para DCX/BrdU o NeuN/BrdU) así como una reducción de nuevos astrocitos (células positivas para Thbs4/GFAP/BrdU) en la zona de penumbra isquémica. En definitiva, estos resultados sugieren que la activación del receptor A1 en isquemia puede ser un modulador de neurogénesis y astrogliogénesis

Optogenetic elevation of endogenous glucocorticoid level in larval zebrafish

The stress response is a suite of physiological and behavioral processes that help to maintain or reestablish homeostasis. Central to the stress response is the hypothalamic-pituitary-adrenal (HPA) axis, as it releases crucial hormones in response to stress. Glucocorticoids (GCs) are the final effector hormones of the HPA axis, and exert a variety of actions under both basal and stress conditions. Despite their far-reaching importance for health, specific GC effects have been difficult to pin-down due to a lack of methods for selectively manipulating endogenous GC levels. Hence, in order to study stress-induced GC effects, we developed a novel optogenetic approach to selectively manipulate the rise of GCs triggered by stress. Using this approach, we could induce both transient hypercortisolic states and persistent forms of hypercortisolaemia in freely behaving larval zebrafish. Our results also established that transient hypercortisolism leads to enhanced locomotion shortly after stressor exposure. Altogether, we present a highly specific method for manipulating the gain of the stress axis with high temporal accuracy, altering endocrine and behavioral responses to stress as well as basal GC levels. Our study offers a powerful tool for the analysis of rapid (non-genomic) and delayed (genomic) GC effects on brain function and behavior, feedbacks within the stress axis and developmental programming by GCs

Pharmacological and genetic modulation of adult neurogenesis in animal models relevant to neuropsychiatric disorders

During the past decade, the modulation of adult neurogenesis has been an intensively studied area of neuroscience due to the implications for understanding of physiological mechanisms in the adult brain and the potential clinical applications for neuropsychiatric disorders. This research has resulted in countless discoveries during a relatively short period of time elucidating mechanistic details about where adult neurogenesis takes place, how the process of neurogenesis occurs and how this process can be regulated at several different steps by, not only endogenous mechanisms which normally maintain a homeostasis of adult neurogenesis, but also by exogenous regulation using genetic and pharmacological modulations to manipulate steps of the process. The modulation of adult neurogenesis has been demonstrated to notably occur as a result of chronic antidepressant treatment which affects several stages of this process resulting in increased adult neurogenesis. A consensus of studies examining the importance of this modulation agree that this increase could be an integral and important part of the behavioral effects of antidepressant, indicating that increased neurogenesis is a part of the therapeutic process in the majority of treatment methods. Questions remain though regarding how neurogenesis is involved in modulating mood as a consensus on this matter finds that decreases in adult neurogenesis per se do not induce depression. However, recent studies indicate that adult neurogenesis is important in the regulation of stress, suggesting that a consequence of decreases in adult neurogenesis may play a role in the dysregulation of this endocrine system in combination with severe or chronic stress which may eventually result in depression. These findings highlight the potential significance of treatments which have the potential to increase adult neurogenesis during pathological states to reach stable levels. Current findings indicate that one of the most important and accessible systems in modulating neurogenesis is the serotonergic system, as exemplified by the potent ability of serotonin enhancing drugs such as the antidepressant fluoxetine to increase neurogenesis. A first set of studies present in this thesis investigate the neurogenic potential in the hippocampus of proteins of the S100 family associated with the serotonergic system including p11 and S100B. The first of these studies uses a genetic deletion of p11 in mice. Results from these experiments demonstrate that mice lack a neurogenic and behavioral response to fluoxetine, seen in normal mice. This finding indicates that p11 is involved in the antidepressant mechanism of fluoxetine. Further examination into potential mechanisms revealed that p11 is highly expressed in interneurons which also express low levels of 5-HT1B and 5-HT4 receptors, of which p11 is a known adaptor protein. Interneurons are known to regulate aspects of adult neurogenesis indicating a possible mechanism through which p11 may modulate the neurogenic and furthermore behavioral effects of this antidepressant. A subsequent study identifies other areas of the brain potentially involved in depression which express p11 and 5-HT1B and 5-HT4 receptors. The last of these S100 studies uses a genetic amplification of S100B in mice to investigate its potential role in adult neurogenesis and revealed that S100B mice have an increased baseline level of cell proliferation which however did not translate into an increase in total neurogenesis. Furthermore, these mice display a normal neurogenic and behavioral response to fluoxetine. These results indicate that S100B is involved in cell proliferation though not other aspects of neurogenesis. Furthermore, S100B may be partially involved in aspects of neurogenesis enhancing drugs and highlight the potential benefits of modulation of this protein. Besides the serotonergic system, other neurotransmitter systems have been implicated in the regulation of adult neurogenesis, including the dopaminergic system. Altered dopamine levels are associated with several disorders of the brain with neuropsychiatric complications. Furthermore this system, in similarity to the serotonergic system, is a primary target of pharmacological therapies for neuropsychiatric disorders. A second set of studies therefore further investigated effects of pharmacological and genetic modulation of the dopaminergic system on adult neurogenesis. The first of these studies investigated the neurogenic and behavioral effects of the drug sarizotan which targets both the serotonergic and dopaminergic system. This drug has previously been shown to have potential antidyskinetic beneficial effects against involuntary movements seen in Parkinson’s disease and therefore we investigated effects of this drug in an animal model of Parkinson’s disease in which dopaminergic afferents are lesioned unilaterally. In the lesioned hemisphere, sarizotan increased cell proliferation in two neurogenic regions of the lateral ventricles and the hippocampus. Sarizotan in combination with the anti-Parkinsonian drug L-DOPA, also increased ongoing neurogenesis in the hippocampus. Furthermore, sarizotan had antidepressant-like activity in the forced swim test in lesioned animals. These findings indicate that targeting of both the serotonergic and dopaminergic systems may be an effective modulator of aspects of neurogenesis and behavior in certain pathologies. For example sarizotan may, in addition to antidyskinetic effects, have antidepressant potential in the frequently seen subgroup of Parkinson’s disease patients who also suffer from depression. The numerous studies regarding purely dopaminergic regulation of adult neurogenesis in either the lateral ventricles or hippocampus have resulted in conflicting data suggesting a complex regulation in which several receptors may be involved. Currently available data suggest expression of the D3 receptors in the proliferative zone of the hippocampus indicating a role in adult neurogenesis. The role of the D3 receptor using a genetic deletion of this receptor in mice was therefore investigated. A robust increase was found in baseline levels of cell proliferation and ongoing neurogenesis in these mice, though not in cell survival. Furthermore, pharmacological modulation using the preferential D3 antagonist S33138 had a similar effect on cell proliferation, although less robust. Thus, in the hippocampus, the D3 receptor appears to act inhibitory on cell proliferation. Previous indicating that D3 is expressed in proliferating cells indicates that this may be a direct effect of dopamine whereas expression of D3 and D2 receptors on niche astrocytes may in contrast indirectly stimulate cell proliferation. This study further highlights how modulation of the dopaminergic system affects adult neurogenesis and may ultimately have significance for pathologies in which adult neurogenesis is affected. In summary, these findings exemplify the numerous different ways in which adult neurogenesis can be modulated which is also indicative of the situations in which adult neurogenesis can be defective, potentially contributing to disease. Studies presented in this thesis have via the use of genetic manipulation as well as pharmacological compounds highlighted specific proteins and pharmacological targets which can be used to modulate aspects of neurogenesis, having potential clinical significance for neuropsychiatric disorders in which adult neurogenesis is affected

Functional, metabolic and transcriptional maturation of human pancreatic islets derived from stem cells

Transplantation of pancreatic islet cells derived from human pluripotent stem cells is a promising treatment for diabetes. Despite progress in the generation of stem-cell-derived islets (SC-islets), no detailed characterization of their functional properties has been conducted. Here, we generated functionally mature SC-islets using an optimized protocol and benchmarked them comprehensively against primary adult islets. Biphasic glucose-stimulated insulin secretion developed during in vitro maturation, associated with cytoarchitectural reorganization and the increasing presence of alpha cells. Electrophysiology, signaling and exocytosis of SC-islets were similar to those of adult islets. Glucose-responsive insulin secretion was achieved despite differences in glycolytic and mitochondrial glucose metabolism. Single-cell transcriptomics of SC-islets in vitro and throughout 6 months of engraftment in mice revealed a continuous maturation trajectory culminating in a transcriptional landscape closely resembling that of primary islets. Our thorough evaluation of SC-islet maturation highlights their advanced degree of functionality and supports their use in further efforts to understand and combat diabetes. Pancreatic islets derived from stem cells are benchmarked against primary cells.Peer reviewe