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Sample records for expressed camp phosphodiesterase

  1. Temporal and spatial regulation of cAMP signaling in disease: role of cyclic nucleotide phosphodiesterases.

    PubMed

    Otero, Carolina; Peñaloza, Juan P; Rodas, Paula I; Fernández-Ramires, Ricardo; Velasquez, Luis; Jung, Juan E

    2014-12-01

    Since its discovery, cAMP has been proposed as one of the most versatile second messengers. The remarkable feature of cAMP to tightly control highly diverse physiological processes, including metabolism, homeostasis, secretion, muscle contraction, cell proliferation and migration, immune response, and gene transcription, is reflected by millions of different articles worldwide. Compartmentalization of cAMP in space and time, maintained by mainly phosphodiesterases, contributes to the maintenance of equilibrium inside the cell where one signal can trigger many different events. Novel cAMP sensors seem to carry out certain unexpected signaling properties of cAMP and thereby to permit delicate adaptations of biologic responses. Measuring space and time events with biosensors will increase our current knowledge on the pathophysiology of diseases, such as chronic obstructive pulmonary disease, asthma, cognitive impairment, cancer, and renal and heart failure. Further insights into the cAMP dynamics will help to optimize the pharmacological treatment for these diseases.

  2. Temporal and spatial regulation of cAMP signaling in disease: role of cyclic nucleotide phosphodiesterases.

    PubMed

    Otero, Carolina; Peñaloza, Juan P; Rodas, Paula I; Fernández-Ramires, Ricardo; Velasquez, Luis; Jung, Juan E

    2014-12-01

    Since its discovery, cAMP has been proposed as one of the most versatile second messengers. The remarkable feature of cAMP to tightly control highly diverse physiological processes, including metabolism, homeostasis, secretion, muscle contraction, cell proliferation and migration, immune response, and gene transcription, is reflected by millions of different articles worldwide. Compartmentalization of cAMP in space and time, maintained by mainly phosphodiesterases, contributes to the maintenance of equilibrium inside the cell where one signal can trigger many different events. Novel cAMP sensors seem to carry out certain unexpected signaling properties of cAMP and thereby to permit delicate adaptations of biologic responses. Measuring space and time events with biosensors will increase our current knowledge on the pathophysiology of diseases, such as chronic obstructive pulmonary disease, asthma, cognitive impairment, cancer, and renal and heart failure. Further insights into the cAMP dynamics will help to optimize the pharmacological treatment for these diseases. PMID:24750474

  3. Role of phosphodiesterases in the shaping of sub-plasma-membrane cAMP oscillations and pulsatile insulin secretion.

    PubMed

    Tian, Geng; Sågetorp, Jenny; Xu, Yunjian; Shuai, Hongyan; Degerman, Eva; Tengholm, Anders

    2012-11-01

    Specificity and versatility in cyclic AMP (cAMP) signalling are governed by the spatial localisation and temporal dynamics of the signal. Phosphodiesterases (PDEs) are important for shaping cAMP signals by hydrolyzing the nucleotide. In pancreatic β-cells, glucose triggers sub-plasma-membrane cAMP oscillations, which are important for insulin secretion, but the mechanisms underlying the oscillations are poorly understood. Here, we investigated the role of different PDEs in the generation of cAMP oscillations by monitoring the concentration of cAMP in the sub-plasma-membrane space ([cAMP](pm)) with ratiometric evanescent wave microscopy in MIN6 cells or mouse pancreatic β-cells expressing a fluorescent translocation biosensor. The general PDE inhibitor IBMX increased [cAMP](pm), and whereas oscillations were frequently observed at 50 µM IBMX, 300 µM-1 mM of the inhibitor caused a stable increase in [cAMP](pm). The [cAMP](pm) was nevertheless markedly suppressed by the adenylyl cyclase inhibitor 2',5'-dideoxyadenosine, indicating IBMX-insensitive cAMP degradation. Among IBMX-sensitive PDEs, PDE3 was most important for maintaining a low basal level of [cAMP](pm) in unstimulated cells. After glucose induction of [cAMP](pm) oscillations, inhibitors of PDE1, PDE3 and PDE4 inhibitors the average cAMP level, often without disturbing the [cAMP](pm) rhythmicity. Knockdown of the IBMX-insensitive PDE8B by shRNA in MIN6 cells increased the basal level of [cAMP](pm) and prevented the [cAMP](pm)-lowering effect of 2',5'-dideoxyadenosine after exposure to IBMX. Moreover, PDE8B-knockdown cells showed reduced glucose-induced [cAMP](pm) oscillations and loss of the normal pulsatile pattern of insulin secretion. It is concluded that [cAMP](pm) oscillations in β-cells are caused by periodic variations in cAMP generation, and that several PDEs, including PDE1, PDE3 and the IBMX-insensitive PDE8B, are required for shaping the sub-membrane cAMP signals and pulsatile insulin release.

  4. The cAMP phosphodiesterase Prune localizes to the mitochondrial matrix and promotes mtDNA replication by stabilizing TFAM

    PubMed Central

    Zhang, Fan; Qi, Yun; Zhou, Kiet; Zhang, Guofeng; Linask, Kaari; Xu, Hong

    2015-01-01

    Compartmentalized cAMP signaling regulates mitochondrial dynamics, morphology, and oxidative phosphorylation. However, regulators of the mitochondrial cAMP pathway, and its broad impact on organelle function, remain to be explored. Here, we report that Drosophila Prune is a cyclic nucleotide phosphodiesterase that localizes to the mitochondrial matrix. Knocking down prune in cultured cells reduces mitochondrial transcription factor A (TFAM) and mitochondrial DNA (mtDNA) levels. Our data suggest that Prune stabilizes TFAM and promotes mitochondrial DNA (mtDNA) replication through downregulation of mitochondrial cAMP signaling. In addition, our work demonstrates the prevalence of mitochondrial cAMP signaling in metazoan and its new role in mitochondrial biogenesis. PMID:25648146

  5. Molecular cloning of rat homologues of the Drosophila melanogaster dunce cAMP phosphodiesterase: evidence for a family of genes.

    PubMed Central

    Swinnen, J V; Joseph, D R; Conti, M

    1989-01-01

    To study the structure and function of cyclic nucleotide phosphodiesterases (PDEs) involved in mammalian gametogenesis, a rat testis cDNA library was screened at low stringency with a cDNA clone coding for the Drosophila melanogaster dunce-encoded PDE as a probe. This screening resulted in the isolation of two groups of cDNA clones, differing in their nucleotide sequences (ratPDE1 and ratPDE2). In the rat testis, RNA transcripts corresponding to both groups of clones were expressed predominantly in germ cells. Additional screenings of a Sertoli cell cDNA library with a ratPDE2 clone as a probe led to the isolation of two more groups of clones (rat-PDE3 and ratPDE4). Unlike ratPDE1 and ratPDE2, these clones hybridized to transcripts present predominantly in the Sertoli cell. In the middle of the coding region, all four groups of clones were homologous to each other. The deduced amino acid sequences of part of this region were also homologous to the D. melanogaster dunce PDE and to PDEs from bovine and yeast. These data indicate that a family of genes homologous to the D. melanogaster dunce-encoded PDE is present in the rat and that these genes are differentially expressed in somatic and germ cells of the seminiferous tubule. These findings provide a molecular basis for the observed heterogeneity of cAMP PDEs. Images PMID:2546153

  6. Concerted Regulation of cGMP and cAMP Phosphodiesterases in Early Cardiac Hypertrophy Induced by Angiotensin II

    PubMed Central

    Mokni, Walid; Keravis, Thérèse; Etienne-Selloum, Nelly; Walter, Alison; Kane, Modou O.; Schini-Kerth, Valérie B.; Lugnier, Claire

    2010-01-01

    Left ventricular hypertrophy leads to heart failure and represents a high risk leading to premature death. Cyclic nucleotides (cAMP and cGMP) play a major role in heart contractility and cyclic nucleotide phosphodiesterases (PDEs) are involved in different stages of advanced cardiac diseases. We have investigated their contributions in the very initial stages of left ventricular hypertrophy development. Wistar male rats were treated over two weeks by chronic infusion of angiotensin II using osmotic mini-pumps. Left cardiac ventricles were used as total homogenates for analysis. PDE1 to PDE5 specific activities and protein and mRNA expressions were explored. Rats developed arterial hypertension associated with a slight cardiac hypertrophy (+24%). cAMP-PDE4 activity was specifically increased while cGMP-PDE activities were broadly increased (+130% for PDE1; +76% for PDE2; +113% for PDE5) and associated with increased expressions for PDE1A, PDE1C and PDE5A. The cGMP-PDE1 activation by Ca2+/CaM was reduced. BNP expression was increased by 3.5-fold, while NOX2 expression was reduced by 66% and AMP kinase activation was increased by 64%. In early cardiac hypertrophy induced by angiotensin II, all specific PDE activities in left cardiac ventricles were increased, favoring an increase in cGMP hydrolysis by PDE1, PDE2 and PDE5. Increased cAMP hydrolysis was related to PDE4. We observed the establishment of two cardioprotective mechanisms and we suggest that these mechanisms could lead to increase intracellular cGMP: i) increased expression of BNP could increase “particulate” cGMP pool; ii) increased activation of AMPK, subsequent to increase in PDE4 activity and 5′AMP generation, could elevate “soluble” cGMP pool by enhancing NO bioavailability through NOX2 down-regulation. More studies are needed to support these assumptions. Nevertheless, our results suggest a potential link between PDE4 and AMPK/NOX2 and they point out that cGMP-PDEs, especially PDE1 and PDE2

  7. PDE4 cAMP phosphodiesterases: modular enzymes that orchestrate signalling cross-talk, desensitization and compartmentalization.

    PubMed Central

    Houslay, Miles D; Adams, David R

    2003-01-01

    cAMP is a second messenger that controls many key cellular functions. The only way to inactivate cAMP is to degrade it through the action of cAMP phosphodiesterases (PDEs). PDEs are thus poised to play a key regulatory role. PDE4 cAMP-specific phosphodiesterases appear to have specific functions with selective inhibitors serving as potent anti-inflammatory agents. The recent elucidation of the structure of the PDE4 catalytic unit allows for molecular insight into the mode of catalysis as well as substrate and inhibitor selectivity. The four PDE4 genes encode over 16 isoforms, each of which is characterized by a unique N-terminal region. PDE4 isoforms play a pivotal role in controlling functionally and spatially distinct pools of cAMP by virtue of their unique intracellular targeting. Targeting occurs by association with proteins, such as arrestins, SRC family tyrosyl kinases, A-kinase anchoring proteins ('AKAPs') and receptor for activated C kinase 1 ('RACK1'), and, in the case of isoform PDE4A1, by a specific interaction (TAPAS-1) with phosphatidic acid. PDE4 isoforms are 'designed' to be regulated by extracellular-signal-related protein kinase (ERK), which binds to anchor sites on the PDE4 catalytic domain that it phosphorylates. The upstream conserved region 1 (UCR1) and 2 (UCR2) modules that abut the PDE4 catalytic unit confer regulatory functions by orchestrating the functional outcome of phosphorylation by cAMP-dependent protein kinase ('PKA') and ERK. PDE4 enzymes stand at a crossroads that allows them to integrate various signalling pathways with that of cAMP in spatially distinct compartments. PMID:12444918

  8. Intracellular membrane association of the Aplysia cAMP phosphodiesterase long and short forms via different targeting mechanisms.

    PubMed

    Kim, Kun-Hyung; Jun, Yong-Woo; Park, Yongsoo; Lee, Jin-A; Suh, Byung-Chang; Lim, Chae-Seok; Lee, Yong-Seok; Kaang, Bong-Kiun; Jang, Deok-Jin

    2014-09-12

    Phosphodiesterases (PDEs) play key roles in cAMP compartmentalization, which is required for intracellular signaling processes, through specific subcellular targeting. Previously, we showed that the long and short forms of Aplysia PDE4 (ApPDE4), which are localized to the membranes of distinct subcellular organelles, play key roles in 5-hydroxytryptamine-induced synaptic facilitation in Aplysia sensory and motor synapses. However, the molecular mechanism of the isoform-specific distinct membrane targeting was not clear. In this study, we further investigated the molecular mechanism of the membrane targeting of the ApPDE4 long and short forms. We found that the membrane targeting of the long form was mediated by hydrophobic interactions, mainly via 16 amino acids at the N-terminal region, whereas the short form was targeted solely to the plasma membrane, mainly by nonspecific electrostatic interactions between their N termini and the negatively charged lipids such as the phosphatidylinositol polyphosphates PI4P and PI(4,5)P2, which are embedded in the inner leaflet of the plasma membrane. Moreover, oligomerization of the long or short form by interaction of their respective upstream conserved region domains, UCR1 and UCR2, enhanced their plasma membrane targeting. These results suggest that the long and short forms of ApPDE4 are distinctly targeted to intracellular membranes through their direct association with the membranes via hydrophobic and electrostatic interactions, respectively. PMID:25077971

  9. Intracellular membrane association of the Aplysia cAMP phosphodiesterase long and short forms via different targeting mechanisms.

    PubMed

    Kim, Kun-Hyung; Jun, Yong-Woo; Park, Yongsoo; Lee, Jin-A; Suh, Byung-Chang; Lim, Chae-Seok; Lee, Yong-Seok; Kaang, Bong-Kiun; Jang, Deok-Jin

    2014-09-12

    Phosphodiesterases (PDEs) play key roles in cAMP compartmentalization, which is required for intracellular signaling processes, through specific subcellular targeting. Previously, we showed that the long and short forms of Aplysia PDE4 (ApPDE4), which are localized to the membranes of distinct subcellular organelles, play key roles in 5-hydroxytryptamine-induced synaptic facilitation in Aplysia sensory and motor synapses. However, the molecular mechanism of the isoform-specific distinct membrane targeting was not clear. In this study, we further investigated the molecular mechanism of the membrane targeting of the ApPDE4 long and short forms. We found that the membrane targeting of the long form was mediated by hydrophobic interactions, mainly via 16 amino acids at the N-terminal region, whereas the short form was targeted solely to the plasma membrane, mainly by nonspecific electrostatic interactions between their N termini and the negatively charged lipids such as the phosphatidylinositol polyphosphates PI4P and PI(4,5)P2, which are embedded in the inner leaflet of the plasma membrane. Moreover, oligomerization of the long or short form by interaction of their respective upstream conserved region domains, UCR1 and UCR2, enhanced their plasma membrane targeting. These results suggest that the long and short forms of ApPDE4 are distinctly targeted to intracellular membranes through their direct association with the membranes via hydrophobic and electrostatic interactions, respectively.

  10. PdeH, a High-Affinity cAMP Phosphodiesterase, Is a Key Regulator of Asexual and Pathogenic Differentiation in Magnaporthe oryzae

    PubMed Central

    Ramanujam, Ravikrishna; Naqvi, Naweed I.

    2010-01-01

    Cyclic AMP-dependent pathways mediate the communication between external stimuli and the intracellular signaling machinery, thereby influencing important aspects of cellular growth, morphogenesis and differentiation. Crucial to proper function and robustness of these signaling cascades is the strict regulation and maintenance of intracellular levels of cAMP through a fine balance between biosynthesis (by adenylate cyclases) and hydrolysis (by cAMP phosphodiesterases). We functionally characterized gene-deletion mutants of a high-affinity (PdeH) and a low-affinity (PdeL) cAMP phosphodiesterase in order to gain insights into the spatial and temporal regulation of cAMP signaling in the rice-blast fungus Magnaporthe oryzae. In contrast to the expendable PdeL function, the PdeH activity was found to be a key regulator of asexual and pathogenic development in M. oryzae. Loss of PdeH led to increased accumulation of intracellular cAMP during vegetative and infectious growth. Furthermore, the pdeHΔ showed enhanced conidiation (2–3 fold), precocious appressorial development, loss of surface dependency during pathogenesis, and highly reduced in planta growth and host colonization. A pdeHΔ pdeLΔ mutant showed reduced conidiation, exhibited dramatically increased (∼10 fold) cAMP levels relative to the wild type, and was completely defective in virulence. Exogenous addition of 8-Br-cAMP to the wild type simulated the pdeHΔ defects in conidiation as well as in planta growth and development. While a fully functional GFP-PdeH was cytosolic but associated dynamically with the plasma membrane and vesicular compartments, the GFP-PdeL localized predominantly to the nucleus. Based on data from cAMP measurements and Real-Time RTPCR, we uncover a PdeH-dependent biphasic regulation of cAMP levels during early and late stages of appressorial development in M. oryzae. We propose that PdeH-mediated sustenance and dynamic regulation of cAMP signaling during M. oryzae development is

  11. Traumatic Brain Injury Upregulates Phosphodiesterase Expression in the Hippocampus.

    PubMed

    Wilson, Nicole M; Titus, David J; Oliva, Anthony A; Furones, Concepcion; Atkins, Coleen M

    2016-01-01

    Traumatic brain injury (TBI) results in significant impairments in hippocampal synaptic plasticity. A molecule critically involved in hippocampal synaptic plasticity, 3',5'-cyclic adenosine monophosphate, is downregulated in the hippocampus after TBI, but the mechanism that underlies this decrease is unknown. To address this question, we determined whether phosphodiesterase (PDE) expression in the hippocampus is altered by TBI. Young adult male Sprague Dawley rats received sham surgery or moderate parasagittal fluid-percussion brain injury. Animals were analyzed by western blotting for changes in PDE expression levels in the hippocampus. We found that PDE1A levels were significantly increased at 30 min, 1 h and 6 h after TBI. PDE4B2 and 4D2 were also significantly increased at 1, 6, and 24 h after TBI. Additionally, phosphorylation of PDE4A was significantly increased at 6 and 24 h after TBI. No significant changes were observed in levels of PDE1B, 1C, 3A, 8A, or 8B between 30 min to 7 days after TBI. To determine the spatial profile of these increases, we used immunohistochemistry and flow cytometry at 24 h after TBI. PDE1A and phospho-PDE4A localized to neuronal cell bodies. PDE4B2 was expressed in neuronal dendrites, microglia and infiltrating CD11b(+) immune cells. PDE4D was predominantly found in microglia and infiltrating CD11b(+) immune cells. To determine if inhibition of PDE4 would improve hippocampal synaptic plasticity deficits after TBI, we treated hippocampal slices with rolipram, a pan-PDE4 inhibitor. Rolipram partially rescued the depression in basal synaptic transmission and converted a decaying form of long-term potentiation (LTP) into long-lasting LTP. Overall, these results identify several possible PDE targets for reducing hippocampal synaptic plasticity deficits and improving cognitive function acutely after TBI. PMID:26903822

  12. Expressing Camp, Part 2: Using Key Messages.

    ERIC Educational Resources Information Center

    Schultz, Bob

    1996-01-01

    Camps can play an integral part in raising a child. The American Camping Association (ACA) has developed key messages that correspond to developmental needs of children. To portray a professional image of camp, promotional materials should incorporate these key messages, the benefits of ACA accreditation, and the same language as child development…

  13. Challenge of human Jurkat T-cells with the adenylate cyclase activator forskolin elicits major changes in cAMP phosphodiesterase (PDE) expression by up-regulating PDE3 and inducing PDE4D1 and PDE4D2 splice variants as well as down-regulating a novel PDE4A splice variant.

    PubMed Central

    Erdogan, S; Houslay, M D

    1997-01-01

    The cAMP phosphodiesterase (PDE) 3 and PDE4 isoforms provide the major cAMP-hydrolysing PDE activities in Jurkat T-cells, with additional contributions from the PDE1 and PDE2 isoforms. Challenge of cells with the adenylate cyclase activator forskolin led to a rapid, albeit transient, increase in PDE3 activity occurring over the first 45 min, followed by a sustained increase in PDE3 activity which began after approximately 3 h and continued for at least 24 h. Only this second phase of increase in PDE3 activity was blocked by the transcriptional inhibitor actinomycin D. After approximately 3 h of exposure to forskolin, PDE4 activity had increased, via a process that could be inhibited by actinomycin D, and it remained elevated for at least a 24 h period. Such actions of forskolin were mimicked by cholera toxin and 8-bromo-cAMP. Forskolin increased intracellular cAMP concentrations in a time-dependent fashion and its action was enhanced when PDE induction was blocked with actinomycin D. Reverse transcription (RT)-PCR analysis, using generic primers designed to detect transcripts representing enzymically active products of the four PDE4 genes, identified transcripts for PDE4A and PDE4D but not for PDE4B or PDE4C in untreated Jurkat T-cells. Forskolin treatment did not induce transcripts for either PDE4B or PDE4C; however, it reduced the RT-PCR signal for PDE4A transcripts and markedly enhanced that for PDE4D transcripts. Using RT-PCR primers for PDE4 splice variants, a weak signal for PDE4D1 was evident in control cells whereas, in forskolin-treated cells, clear signals for both PDE4D1 and PDE4D2 were detected. RT-PCR analysis of the PDE4A species indicated that it was not the PDE4A isoform PDE-46 (PDE4A4B). Immunoblotting of control cells for PDE4 forms identified a single PDE4A species of approximately 118 kDa, which migrated distinctly from the PDE4A4B isoform PDE-46, with immunoprecipitation analyses showing that it provided all of the PDE4 activity in control

  14. Iloprost- and isoproterenol-induced increases in cAMP are regulated by different phosphodiesterases in erythrocytes of both rabbits and humans.

    PubMed

    Adderley, Shaquria P; Dufaux, Eileen A; Sridharan, Meera; Bowles, Elizabeth A; Hanson, Madelyn S; Stephenson, Alan H; Ellsworth, Mary L; Sprague, Randy S

    2009-05-01

    Activation of the G protein G(s) results in increases in cAMP, a necessary step in the pathway for ATP release from rabbit and human erythrocytes. In all cells, the level of cAMP is the product of its synthesis by adenylyl cyclase and its hydrolysis by phosphodiesterases (PDEs). Both iloprost (Ilo), a PGI(2) analog, and isoproterenol (Iso), a beta-agonist, stimulate receptor-mediated increases in cAMP in rabbit and human erythrocytes. However, the specific PDEs associated with each of these signaling pathways in the erythrocyte have not been fully characterized. Previously, we reported that PDE3B is present in rabbit and human erythrocyte membranes and that PDE3 inhibitors potentiate Ilo-induced increases in cAMP. Here we report that inhibitors of either PDE2 or PDE4, erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) and rolipram, respectively, potentiate Iso-induced increases in cAMP in rabbit and human erythrocytes. Importantly, these inhibitors had no effect on cAMP increases associated with the incubation of erythrocytes with Ilo. In addition, we establish, for the first time, the presence of PDE2A protein in rabbit and human erythrocyte membranes. Finally, we determined that preincubation of human erythrocytes with EHNA and rolipram together potentiate Iso-induced ATP release, whereas preincubation with cilostazol enhances Ilo-induced release of ATP. These results are consistent with the hypothesis that, in rabbit and human erythrocytes, Ilo-induced increases in cAMP and ATP release are regulated by PDE3, whereas those associated with Iso are regulated by the activities of both PDE2 and PDE4. These studies demonstrate that PDE activity in these cells is localized to specific signaling pathways. PMID:19252089

  15. Identification and tissue-specific expression of PDE7 phosphodiesterase splice variants

    PubMed Central

    Bloom, Timothy J.; Beavo, Joseph A.

    1996-01-01

    Type 7 cyclic nucleotide phosphodiesterases (PDE7s) are a newly described family of enzymes having high affinity and specificity for cAMP. However, little is known about their structure, function, or regulation. We have isolated a mouse skeletal muscle cDNA representing a new alternative splice variant (PDE7A2) of the PDE7 gene. The ORF encodes a 456-amino acid protein having a predicted molecular weight of 52.4 kDa. The 5′ end of the mouse PDE7A2 is divergent from the 5′ end of the human PDE7A1 sequence and is more hydrophobic. A comparison of the 5′ ends of the two cDNA clones with human genomic sequence indicates that they represent alternate splice products rather than species variation. RNase protection analysis of several mouse tissues indicates that PDE7 is expressed widely with highest levels in skeletal muscle. HPLC fractionation and Western blot analysis of two human lymphocyte T-cell lines shows that an unknown PDE activity described by Ichimura and Kase [Ichimura, M. & Kase, H. (1993) Biochem. Biophys. Res. Commun. 193, 985–990] is most likely to be PDE7A1. A single immunoreactive band of ≈55 kDa, which comigrates with PDE7A1, is seen in fractions of the HPLC profile containing this activity suggesting that the original human PDE7A1 clone contains a full-length ORF, and is not truncated at the 5′ end as was originally postulated. In a human lymphocyte B-cell line and also in mouse skeletal muscle, a large amount of PDE7 mRNA but little PDE7 protein or activity is expressed suggesting that the translation or stability of PDE7 protein may be highly regulated in these tissues. PMID:8943082

  16. Phosphodiesterase 10A in the rat pineal gland: localization, daily and seasonal regulation of expression and influence on signal transduction.

    PubMed

    Spiwoks-Becker, Isabella; Wolloscheck, Tanja; Rickes, Oliver; Kelleher, Debra K; Rohleder, Nils; Weyer, Veronika; Spessert, Rainer

    2011-01-01

    The cyclic nucleotide phosphodiesterase 10A (PDE10A) is highly expressed in striatal spiny projection neurons and represents a therapeutic target for the treatment of psychotic symptoms. As reported previously [J Biol Chem 2009; 284:7606-7622], in this study PDE10A was seen to be additionally expressed in the pineal gland where the levels of PDE10A transcript display daily changes. As with the transcript, the amount of PDE10A protein was found to be under daily and seasonal regulation. The observed cyclicity in the amount of PDE10A mRNA persists under constant darkness, is blocked by constant light and is modulated by the lighting regime. It therefore appears to be driven by the master clock in the suprachiasmatic nucleus (SCN). Since adrenergic agonists and dibutyryl-cAMP induce PDE10A mRNA, the in vitro clock-dependent control of Pde10a appears to be mediated via a norepinephrine → β-adrenoceptor → cAMP/protein kinase A signaling pathway. With regard to the physiological role of PDE10A in the pineal gland, the specific PDE10A inhibitor papaverine was seen to enhance the adrenergic stimulation of the second messenger cAMP and cGMP. This indicates that PDE10A downregulates adrenergic cAMP and cGMP signaling by decreasing the half-life of both nucleotides. Consistent with its effect on cAMP, PDE10A inhibition also amplifies adrenergic induction of the cAMP-inducible gene arylalkylamine N-acetyltransferase (Aanat) which codes the rate-limiting enzyme in pineal melatonin formation. The findings of this study suggest that Pde10a expression is under circadian and seasonal regulation and plays a modulatory role in pineal signal transduction and gene expression.

  17. Role of phosphodiesterase 4 expression in the Epac1 signaling-dependent skeletal muscle hypertrophic action of clenbuterol.

    PubMed

    Ohnuki, Yoshiki; Umeki, Daisuke; Mototani, Yasumasa; Shiozawa, Kouichi; Nariyama, Megumi; Ito, Aiko; Kawamura, Naoya; Yagisawa, Yuka; Jin, Huiling; Cai, Wenqian; Suita, Kenji; Saeki, Yasutake; Fujita, Takayuki; Ishikawa, Yoshihiro; Okumura, Satoshi

    2016-05-01

    Clenbuterol (CB), a selective β2-adrenergic receptor (AR) agonist, induces muscle hypertrophy and counteracts muscle atrophy. However, it is paradoxically less effective in slow-twitch muscle than in fast-twitch muscle, though slow-twitch muscle has a greater density of β-AR We recently demonstrated that Epac1 (exchange protein activated by cyclic AMP [cAMP]1) plays a pivotal role in β2-AR-mediated masseter muscle hypertrophy through activation of the Akt and calmodulin kinase II (CaMKII)/histone deacetylase 4 (HDAC4) signaling pathways. Here, we investigated the role of Epac1 in the differential hypertrophic effect of CB using tibialis anterior muscle (TA; typical fast-twitch muscle) and soleus muscle (SOL; typical slow-twitch muscle) of wild-type (WT) and Epac1-null mice (Epac1KO). The TA mass to tibial length (TL) ratio was similar in WT and Epac1KO at baseline and was significantly increased after CB infusion in WT, but not in Epac1KO The SOL mass to TL ratio was also similar in WT and Epac1KO at baseline, but CB-induced hypertrophy was suppressed in both mice. In order to understand the mechanism involved, we measured the protein expression levels of β-AR signaling-related molecules, and found that phosphodiesterase 4 (PDE4) expression was 12-fold greater in SOL than in TA These results are consistent with the idea that increased PDE4-mediated cAMP hydrolysis occurs in SOL compared to TA, resulting in a reduced cAMP concentration that is insufficient to activate Epac1 and its downstream Akt and CaMKII/HDAC4 hypertrophic signaling pathways in SOL of WT This scenario can account for the differential effects of CB on fast- and slow-twitch muscles. PMID:27207782

  18. Bresol inhibits phosphodiesterase 4 gene expression and modulates the levels of select mediators of inflammation in human monocytic cells.

    PubMed

    Sandeep Varma, R; Ashok, G; Vidyashankar, S; Nandakumar, K S; Patki, P S

    2011-01-01

    Bresol-a poly-herbal formulation, has been reported to be effective against bronchial asthma and allergic rhinitis in children. In vivo studies have supported the anti-histaminic and anti-anaphylactic action of bresol. However, the mechanism of action of bresol in modulation of inflammation has not been studied at the cellular and molecular level. The present study was aimed to elucidate the mechanism(s) of action of bresol at the cellular and molecular levels, using human monocyte leukemia cells. The effects of bresol on phosphodiesterase 4B (PDE4B) gene expression were analyzed using human monocytic U937 leukemia cells. The ability of bresol to stimulate cAMP formation in these cells, as well as its effects on mediators of inflammation like tumor necrosis factor-α (TNFα), nitric oxide (NO), and cycloxygenase-2 (COX-2) in lipopolysaccharide (LPS)-stimulated U937 cells, were also studied. The results here indicated that bresol exhibited potential anti-inflammatory properties by inhibiting LPS-induced PDE4B gene expression in the cells. Bresol also dose dependently activated cAMP formation, and inhibited TNFα, NO, as well as COX-2 formation in the LPS-stimulated cells. Based upon the results, we concluded that the reported anti-inflammatory activity of bresol might be attributed to its abilities to inhibit PDE4B and thus elevate cAMP levels in human monocytes. The anti-inflammatory effects of bresol might also be a result of the capacity of bresol to modulate the formation of TNFα, NO, and COX-2 in monocytes.

  19. Expression in bacteria of functional inhibitory subunit of retinal rod cGMP phosphodiesterase.

    PubMed Central

    Brown, R L; Stryer, L

    1989-01-01

    The cGMP phosphodiesterase of vertebrate retinal rod outer segments plays a key role in visual transduction. A functionally active form of the inhibitory gamma subunit of the phosphodiesterase, which keeps the enzyme inactive in the dark, has been obtained in high yield from a synthetic gene expressed in Escherichia coli. A DNA sequence encoding the 87-residue bovine gamma subunit was chemically synthesized and assembled from 10 oligonucleotides. The synthetic gene was cloned into an expression vector that uses the promoter PL of lambda phage. E. coli was transformed with this vector, which encodes a fusion protein consisting of the first 31 residues of the lambda cII protein, a 7-residue joining sequence that is specifically cleaved at its C-terminal end by clotting protease factor Xa, and the 87-residue gamma subunit. The fusion protein was solubilized in 6 M urea and purified by ion-exchange chromatography on a CM-Sephadex column. The typical yield was 1 mg of fusion protein per liter of bacterial culture, which corresponds to the amount of gamma in about 2500 bovine retinas. Proteolytic cleavage of the fusion protein by factor Xa released a synthetic gamma with the same amino acid sequence as that of native gamma. Both fusion protein and synthetic gamma inhibited trypsin-activated phosphodiesterase with high affinity (Kd less than 100 pM). Likewise, both were as effective as native gamma in inhibiting transducin-activated phosphodiesterase in rod outer segment membranes. This inhibition was reversed by the activation of additional transducin. Thus, the N terminus of gamma is not intimately involved in interactions with either the catalytic subunits of the phosphodiesterase or the activated form of transducin. In contrast, a C-terminal deletion mutant terminating at residue 74 of gamma stimulated rather than inhibited the trypsin-activated enzyme. Thus, the C-terminal region of gamma is critical for inhibition of the phosphodiesterase. Images PMID:2544882

  20. Ontogeny of catecholamine and adenosine receptor-mediated cAMP signaling of embryonic red blood cells: role of cGMP-inhibited phosphodiesterase 3 and hemoglobin.

    PubMed

    Baumann, R; Blass, C; Götz, R; Dragon, S

    1999-12-15

    We have previously shown that the cAMP signaling pathway controls major aspects of embryonic red blood cell (RBC) function in avian embryos (Glombitza et al, Am J Physiol 271:R973, 1996; and Dragon et al, Am J Physiol 271:R982, 1996) that are important for adaptation of the RBC gas transport properties to the progressive hypercapnia and hypoxia of later stages of avian embryonic development. Data about the ontogeny of receptor-mediated cAMP signaling are lacking. We have analyzed the response of primitive and definitive chick embryo RBC harvested from day 3 to 18 of development towards forskolin, beta-adrenergic, and A2 receptor agonists. The results show a strong response of immature definitive and primitive RBC to adenosine A2 and beta-adrenergic receptor agonists, which is drastically reduced in the last stage of development, coincident with the appearance of mature, transcriptionally inactive RBC. Modulation of cGMP-inhibited phosphodiesterase 3 (PDE3) has a controlling influence on cAMP accumulation in definitive RBC. Under physiological conditions, PDE3 is inhibited due to activation of soluble guanylyl cyclase (sGC). Inhibition of sGC with the specific inhibitor ODQ decreases receptor-mediated stimulation of cAMP production; this effect is reversed by the PDE3 inhibitor milrinone. sGC is acitivated by nitric oxide (NO), but we found no evidence for production of NO by erythrocyte NO-synthase. However, embryonic hemoglobin releases NO in an oxygen-linked manner that may activate guanylyl cyclase.

  1. Effects of repeated treatment with phosphodiesterase-4 inhibitors on cAMP signaling, hippocampal cell proliferation, and behavior in the forced-swim test.

    PubMed

    Xiao, Lan; O'Callaghan, James P; O'Donnell, James M

    2011-08-01

    The effects of repeated treatment with the phosphodiesterase-4 (PDE4) inhibitors rolipram, piclamilast, and 4-(2-(3-(cyclopentyloxy)-4-methoxyphenyl)-2-phenylethyl)pyridine (CDP840), which differ in their interactions with high- and low-affinity binding conformers of the enzyme, were contrasted to those of acute treatment on cAMP signaling, hippocampal cell proliferation, and immobility in the forced-swim test in rats. Repeated treatment with rolipram (1 and 3 mg/kg), piclamilast (0.3 and 1 mg/kg), or CDP840 (10 and 30 mg/kg) for 16 days increased cAMP and phosphorylation of cAMP response element binding protein (pCREB) in hippocampus and prefrontal cortex. In addition, repeated treatment with the PDE4 inhibitors increased proliferation and survival of newborn cells in the hippocampus and produced antidepressant-like effects on behavior, as evidenced by decreased immobility in the forced-swim test. Acute treatment with rolipram (3 mg/kg), piclamilast (1 mg/kg), or CDP840 (30 mg/kg) induced transient increases in cAMP and pCREB in hippocampus and prefrontal cortex, but the dose and time dependence of these effects did not parallel the behavioral effects. Compared with rolipram and piclamilast, repeated treatment with CDP840 exerted lesser effects on neural and behavioral measures, probably because of its weak interaction with the high-affinity binding conformer of PDE4. This suggests the relative importance of the high-affinity binding conformer in the mediation of the long-term effects of PDE4 inhibition on cAMP/pCREB signaling, hippocampal cell proliferation, and antidepressant-like effects on behavior.

  2. Suppression of mesangial proliferative glomerulonephritis development in rats by inhibitors of cAMP phosphodiesterase isozymes types III and IV.

    PubMed Central

    Tsuboi, Y; Shankland, S J; Grande, J P; Walker, H J; Johnson, R J; Dousa, T P

    1996-01-01

    Excessive mesangial cell (MC) proliferation is a hallmark of many glomerulopathies. In our recent study on cultured rat MC (Matousovic, K., J.P. Grande, C.C.S. Chini, E.N. Chini, and T.P. Dousa. 1995. J. Clin. Invest. 96:401-410) we found that inhibition of isozyme cyclic-3',5'-nucleotide phosphodiesterase (PDE) type III (PDE-III) suppressed MC mitogenesis by activating cAMP-dependent protein kinase (PKA) and by decreasing activity of mitogen-activated protein kinase (MAPK). We also found that inhibition of another PDE isozyme, PDE-IV, suppresses superoxide generation in glomeruli (Chini, C.C.S., E.N. Chini, J.M. Williams, K. Matousovic, and T.P. Dousa. 1994. Kidney Int. 46:28-36). We thus explored whether administration in vivo of the selective PDE-III antagonist, lixazinone (LX), together with the specific PDE-IV antagonist, rolipram (RP), can attenuate development of mesangioproliferative glomerulonephritis (MSGN) induced in rats by anti-rat thymocyte serum (ATS). Unlike the vehicle-treated MSGN rats, rats with MSGN treated with LX and RP did not develop proteinuria and maintained normal renal function when examined 5 d after injection of ATS. In PAS-stained kidneys from PDE-antagonists-treated MSGN-rats the morphology of glomeruli showed a reduction in cellularity compared with control rats with ATS. Compared with MSGN rats receiving vehicle, the MSGN rats receiving PDE-antagonists had less glomerular cell proliferation (PCNA delta -65%), a significantly lesser macrophage infiltration (delta -36% ED-1) and a significant reduction of alpha-smooth muscle actin expression by activated MC; in contrast, immunostaining for platelet antigens and laminin were not different. The beneficial effect of PDE inhibitors was not due to a moderate decrease (approximately -20%) in systolic blood pressure (SBP); as a similar decrease in SBP due to administration of hydralazine, a drug devoid of PDE inhibitory effect, did not reduce severity of MSGN in ATS-injected rats. We

  3. Loss of phosphodiesterase 10A expression is associated with progression and severity in Parkinson's disease.

    PubMed

    Niccolini, Flavia; Foltynie, Thomas; Reis Marques, Tiago; Muhlert, Nils; Tziortzi, Andri C; Searle, Graham E; Natesan, Sridhar; Kapur, Shitij; Rabiner, Eugenii A; Gunn, Roger N; Piccini, Paola; Politis, Marios

    2015-10-01

    The mechanisms underlying neurodegeneration and loss of dopaminergic signalling in Parkinson's disease are still only partially understood. Phosphodiesterase 10A (PDE10A) is a basal ganglia expressed dual substrate enzyme, which regulates cAMP and cGMP signalling cascades, thus having a key role in the regulation of dopaminergic signalling in striatal pathways, and in promoting neuronal survival. This study aimed to assess in vivo the availability of PDE10A in patients with Parkinson's disease using positron emission tomography molecular imaging with (11)C-IMA107, a highly selective PDE10A radioligand. We studied 24 patients with levodopa-treated, moderate to advanced Parkinson's disease. Their positron emission tomography imaging data were compared to those from a group of 12 healthy controls. Parametric images of (11)C-IMA107 binding potential relative to non-displaceable binding (BPND) were generated from the dynamic (11)C-IMA107 scans using the simplified reference tissue model with the cerebellum as the reference tissue. Corresponding region of interest analysis showed lower mean (11)C-IMA107 BPND in the caudate (P < 0.001), putamen (P < 0.001) and globus pallidus (P = 0.025) in patients with Parkinson's disease compared to healthy controls, which was confirmed with voxel-based analysis. Longer Parkinson's duration correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.65; P = 0.005), putamen (r = -0.51; P = 0.025), and globus pallidus (r = -0.47; P = 0.030). Higher Unified Parkinson's Disease Rating Scale part-III motor scores correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.54; P = 0.011), putamen (r = -0.48; P = 0.022), and globus pallidus (r = -0.70; P < 0.001). Higher Unified Dyskinesia Rating Scale scores in those Parkinson's disease with levodopa-induced dyskinesias (n = 12), correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.73; P = 0.031) and putamen (r = -0.74; P = 0.031). Our findings demonstrate striatal and

  4. Novel alternative splice variants of rat phosphodiesterase 7B showing unique tissue-specific expression and phosphorylation.

    PubMed Central

    Sasaki, Takashi; Kotera, Jun; Omori, Kenji

    2002-01-01

    cDNA species coding for novel variants of cyclic-AMP-specific phosphodiesterases (PDEs), namely the PDE7B family, were isolated from rats and characterized. Rat PDE7B1 (RNPDE7B1) was composed of 446 amino acid residues. Rat PDE7B2 (RNPDE7B2) and PDE7B3 (RNPDE7B3), which possessed unique N-terminal sequences, consisted of 359 and 459 residues respectively. Northern hybridization analysis showed that rat PDE7B transcripts were particularly abundant in the striatum and testis. PCR analyses revealed that rat PDE7B2 transcripts were restricted to the testis and that low levels of PDE7B3 transcripts were expressed in the heart, lung and skeletal muscle. In situ hybridization analysis demonstrated that rat PDE7B transcripts were expressed in striatal neurons and spermatocytes. In spermatocytes, rat PDE7B transcripts were expressed in a stage-specific manner during spermatogenesis. The K(m) values of recombinant rat PDE7B1, PDE7B2 and PDE7B3 for cAMP were 0.05, 0.07 and 0.05 microM respectively. Each rat PDE7B variant was the most sensitive to 3-isobutyl-1-methylxanthine (IC(50) 1.5-2.1 microM). Two phosphorylation sites for cAMP-dependent protein kinase (PKA) were found in rat PDE7B1 and PDE7B3, whereas rat PDE7B2 possessed one site. PKA-dependent phosphorylation was observed in C-terminal phosphorylation sites of three rat PDE7B variants, in addition to unique N-terminal regions of rat PDE7B1 and PDE7B3. Unique tissue distribution and PKA-dependent phosphorylation of PDE7B variants suggested that each variant has a specific role for cellular functions via cAMP signalling in various tissues. PMID:11772393

  5. Altered phosphodiesterase 3-mediated cAMP hydrolysis contributes to a hypermotile phenotype in obese JCR:LA-cp rat aortic vascular smooth muscle cells: implications for diabetes-associated cardiovascular disease.

    PubMed

    Netherton, Stuart J; Jimmo, Sandra L; Palmer, Daniel; Tilley, Douglas G; Dunkerley, Heather A; Raymond, Daniel R; Russell, James C; Absher, P Marlene; Sage, E Helene; Vernon, Robert B; Maurice, Donald H

    2002-04-01

    Cardiovascular diseases represent a significant cause of morbidity and mortality in diabetes. Of the many animal models used in the study of non-insulin-dependent (type 2) diabetes, the JCR:LA-cp rat is unique in that it develops insulin resistance in the presence of obesity and manifests both peripheral and coronary vasculopathies. In this animal model, arterial vascular smooth muscle cells (VSMCs) from homozygous obese (cp/cp) rats, but not from age-matched healthy (+/+ or + /cp, collectively defined +/?) littermates, display an " activated" phenotype in vitro and in vivo and have an elevated level of cAMP phosphodiesterase (PDE) activity. In this report, we confirm that cp/cp rat aortic VSMCs have an elevated level of PDE3 activity and show that only particulate PDE3 (PDE3B) activity is elevated. In marked contrast to results obtained in + /? VSMCs, simultaneous activation of adenylyl cyclase and inhibition of PDE3 activity in cp/cp VSMCs synergistically increased cAMP. Although PDE3 inhibition did not potentiate the antimigratory effects of forskolin on +/? VSMCs, PDE3 inhibition did markedly potentiate the forskolin-induced inhibition of migration of cp/cp-derived VSMCs. Although PDE3 activity was elevated in cp/cp rat aortic VSMCs, levels of expression of cytosolic PDE3 (PDE3A) and PDE3B in +/? and cp/cp VSMCs, as well as activation of these enzymes following activation of the cAMP-protein kinase A signaling cascade, were not different. Our data are consistent with an increased role for PDE3 in regulating cAMP-dependent signaling in cp/cp VSMCs and identify PDE3 as a cellular activity potentially responsible for the phenotype of cp/cp VSMCs.

  6. Identification and characterization of the type-IVA cyclic AMP-specific phosphodiesterase RD1 as a membrane-bound protein expressed in cerebellum.

    PubMed

    Shakur, Y; Wilson, M; Pooley, L; Lobban, M; Griffiths, S L; Campbell, A M; Beattie, J; Daly, C; Houslay, M D

    1995-03-15

    An antiserum was generated against a dodecapeptide whose sequence is found at the C-terminus of a cyclic AMP (cAMP)-specific, type-IVA phosphodiesterase encoded by the rat 'dunc-like' cyclic AMP phosphodiesterase (RD1) cDNA. This antiserum identified a single approximately 73 kDa protein species upon immunoblotting of cerebellum homogenates. This species co-migrated upon SDS/PAGE with a single immunoreactive species observed in COS cells transfected with the cDNA for RD1. Native RD1 in cerebellum was found to be predominantly (approximately 93%) membrane-associated and could be found in isolated synaptosome populations, in particular those enriched in post-synaptic densities. Fractionation of lysed synaptosomes on sucrose density gradients identified RD1 as co-migrating with the plasma membrane marker 5'-nucleotidase. Laser scanning confocal and digital deconvolution immunofluorescence studies done on intact COS cells transfected with RD1 cDNA showed RD1 to be predominantly localized to plasma membranes but also associated with the Golgi apparatus and intracellular vesicles. RD1-specific antisera immunoprecipitated phosphodiesterase activity from solubilized cerebellum membranes. This activity had the characteristics expected of the type-IV cAMP phosphodiesterase RD1 in that it was cAMP specific, exhibited a low Km cAMP of 2.3 microM, high sensitivity to inhibition by 4-[3-(cyclopentoxyl)-4-methoxyphenyl]-2-pyrrolidone (rolipram) (Ki approximately 0.7 microM) and was unaffected by Ca2+/calmodulin and low concentrations of cyclic GMP. The phosphodiesterase activities of RD1 solubilized from both cerebellum and transfected COS cell membranes showed identical first-order thermal denaturation kinetics at 50 degrees C. Native RD1 from cerebellum was shown to be an integral protein in that it was solubilized using the non-ionic detergent Triton X-100 but not by either re-homogenization or high NaCl concentrations. The observation that hydroxylamine was unable to cause

  7. Phosphodiesterase-4 Inhibition Alters Gene Expression and Improves Isoniazid – Mediated Clearance of Mycobacterium tuberculosis in Rabbit Lungs

    PubMed Central

    Subbian, Selvakumar; Tsenova, Liana; O'Brien, Paul; Yang, Guibin; Koo, Mi-Sun; Peixoto, Blas; Fallows, Dorothy; Dartois, Veronique; Muller, George; Kaplan, Gilla

    2011-01-01

    Tuberculosis (TB) treatment is hampered by the long duration of antibiotic therapy required to achieve cure. This indolent response has been partly attributed to the ability of subpopulations of less metabolically active Mycobacterium tuberculosis (Mtb) to withstand killing by current anti-TB drugs. We have used immune modulation with a phosphodiesterase-4 (PDE4) inhibitor, CC-3052, that reduces tumor necrosis factor alpha (TNF-α) production by increasing intracellular cAMP in macrophages, to examine the crosstalk between host and pathogen in rabbits with pulmonary TB during treatment with isoniazid (INH). Based on DNA microarray, changes in host gene expression during CC-3052 treatment of Mtb infected rabbits support a link between PDE4 inhibition and specific down-regulation of the innate immune response. The overall pattern of host gene expression in the lungs of infected rabbits treated with CC-3052, compared to untreated rabbits, was similar to that described in vitro in resting Mtb infected macrophages, suggesting suboptimal macrophage activation. These alterations in host immunity were associated with corresponding down-regulation of a number of Mtb genes that have been associated with a metabolic shift towards dormancy. Moreover, treatment with CC-3052 and INH resulted in reduced expression of those genes associated with the bacterial response to INH. Importantly, CC-3052 treatment of infected rabbits was associated with reduced ability of Mtb to withstand INH killing, shown by improved bacillary clearance, from the lungs of co-treated animals compared to rabbits treated with INH alone. The results of our study suggest that changes in Mtb gene expression, in response to changes in the host immune response, can alter the responsiveness of the bacteria to antimicrobial agents. These findings provide a basis for exploring the potential use of adjunctive immune modulation with PDE4 inhibitors to enhance the efficacy of existing anti-TB treatment. PMID:21949656

  8. siRNA-mediated silencing of phosphodiesterase 4B expression affects the production of cytokines in endotoxin-stimulated primary cultured microglia

    PubMed Central

    Cheng, Hao; Wu, Zhifang; He, Xiaoyun; Liu, Qingzhen; Jia, Hongbin; Di, Yan; Ji, Qing

    2016-01-01

    Phosphodiesterase 4 (PDE4) has four subtypes: PDE4A, PDE4B, PDE4C and PDE4D. The expression of PDE4 subtypes in microglial cells and the specific contribution of each subtype to inflammation remain unclear. In this study, the expression of PDE4 subtypes in primary microglial cells was assayed. Primary microglial cells were then transfected with specific small interfering RNA (siRNA) against each PDE4 subtype. PDE4 subtype A-D knockdown was confirmed by quantitative polymerase chain reaction. Secreted cytokines in the supernatant and intracellular cyclic adenosine monophosphate (cAMP) levels of transfected cells were measured. The effect of PDE4B siRNA on the activation of extracellular regulated protein kinase (ERK) induced by lipopolysaccharide (LPS) in microglia was further tested by western blotting. Results showed that the primary microglial cells expressed all four types of PDE4s at the protein level. Transfection with the four siRNAs inhibited PDE4 subtype A-D mRNA expression, respectively. In primary microglial cells, treatment with PDE4B siRNA significantly inhibited the expression of tumor necrosis factor-α and interleukin (IL)-1β, and enhanced the expression of cAMP, while siRNAs to other subtypes had no significant effects. However, none of the four siRNAs had any significant effect on the expression of IL-10. Furthermore, in the PDE4B group, the level of phosphorylated ERK was reduced. Among the four PDE4 subtypes, PDE4B plays an important role in regulating inflammatory responses in microglia, potentially through initially regulating the intracellular cAMP concentration. PMID:27698721

  9. siRNA-mediated silencing of phosphodiesterase 4B expression affects the production of cytokines in endotoxin-stimulated primary cultured microglia

    PubMed Central

    Cheng, Hao; Wu, Zhifang; He, Xiaoyun; Liu, Qingzhen; Jia, Hongbin; Di, Yan; Ji, Qing

    2016-01-01

    Phosphodiesterase 4 (PDE4) has four subtypes: PDE4A, PDE4B, PDE4C and PDE4D. The expression of PDE4 subtypes in microglial cells and the specific contribution of each subtype to inflammation remain unclear. In this study, the expression of PDE4 subtypes in primary microglial cells was assayed. Primary microglial cells were then transfected with specific small interfering RNA (siRNA) against each PDE4 subtype. PDE4 subtype A-D knockdown was confirmed by quantitative polymerase chain reaction. Secreted cytokines in the supernatant and intracellular cyclic adenosine monophosphate (cAMP) levels of transfected cells were measured. The effect of PDE4B siRNA on the activation of extracellular regulated protein kinase (ERK) induced by lipopolysaccharide (LPS) in microglia was further tested by western blotting. Results showed that the primary microglial cells expressed all four types of PDE4s at the protein level. Transfection with the four siRNAs inhibited PDE4 subtype A-D mRNA expression, respectively. In primary microglial cells, treatment with PDE4B siRNA significantly inhibited the expression of tumor necrosis factor-α and interleukin (IL)-1β, and enhanced the expression of cAMP, while siRNAs to other subtypes had no significant effects. However, none of the four siRNAs had any significant effect on the expression of IL-10. Furthermore, in the PDE4B group, the level of phosphorylated ERK was reduced. Among the four PDE4 subtypes, PDE4B plays an important role in regulating inflammatory responses in microglia, potentially through initially regulating the intracellular cAMP concentration.

  10. Angiotensin-converting enzyme inhibition prevents myocardial infarction-induced increase in renal cortical cGMP and cAMP phosphodiesterase activities.

    PubMed

    Clauss, François; Charloux, Anne; Piquard, François; Doutreleau, Stéphane; Talha, Samy; Zoll, Joffrey; Lugnier, Claire; Geny, Bernard

    2015-08-01

    We investigated whether myocardial infarction (MI) enhances renal phosphodiesterases (PDE) activities, investigating particularly the relative contribution of PDE1-5 isozymes in total PDE activity involved in both cGMP and cAMP pathways, and whether angiotensin-converting enzyme inhibition (ACEi) decreases such renal PDE hyperactivities. We also investigated whether ACEi might thereby improve atrial natriuretic peptide (ANP) efficiency. We studied renal cortical PDE1-5 isozyme activities in sham (SH)-operated, MI rats and in MI rats treated with perindopril (ACEi) 1 month after coronary artery ligation. Circulating atrial natriuretic peptide (ANP), its second intracellular messenger cyclic guanosine monophosphate (cGMP) and cGMP/ANP ratio were also determined. Cortical cGMP-PDE2 (80.3 vs. 65.1 pmol/min/mg) and cGMP-PDE1 (50.7 vs. 30.1 pmol/min/mg), and cAMP-PDE2 (161 vs. 104.1 pmol/min/mg) and cAMP-PDE4 (307.5 vs. 197.2 pmol/min/mg) activities were higher in MI than in SH rats. Despite increased ANP plasma level, ANP efficiency tended to be decreased in MI compared to SH rats. Perindopril restored PDE activities and tended to improve ANP efficiency in MI rats. One month after coronary ligation, perindopril treatment of MI rats prevents the increase in renal cortical PDE activities. This may contribute to increase renal ANP efficiency in MI rats.

  11. Angiotensin-converting enzyme inhibition prevents myocardial infarction-induced increase in renal cortical cGMP and cAMP phosphodiesterase activities.

    PubMed

    Clauss, François; Charloux, Anne; Piquard, François; Doutreleau, Stéphane; Talha, Samy; Zoll, Joffrey; Lugnier, Claire; Geny, Bernard

    2015-08-01

    We investigated whether myocardial infarction (MI) enhances renal phosphodiesterases (PDE) activities, investigating particularly the relative contribution of PDE1-5 isozymes in total PDE activity involved in both cGMP and cAMP pathways, and whether angiotensin-converting enzyme inhibition (ACEi) decreases such renal PDE hyperactivities. We also investigated whether ACEi might thereby improve atrial natriuretic peptide (ANP) efficiency. We studied renal cortical PDE1-5 isozyme activities in sham (SH)-operated, MI rats and in MI rats treated with perindopril (ACEi) 1 month after coronary artery ligation. Circulating atrial natriuretic peptide (ANP), its second intracellular messenger cyclic guanosine monophosphate (cGMP) and cGMP/ANP ratio were also determined. Cortical cGMP-PDE2 (80.3 vs. 65.1 pmol/min/mg) and cGMP-PDE1 (50.7 vs. 30.1 pmol/min/mg), and cAMP-PDE2 (161 vs. 104.1 pmol/min/mg) and cAMP-PDE4 (307.5 vs. 197.2 pmol/min/mg) activities were higher in MI than in SH rats. Despite increased ANP plasma level, ANP efficiency tended to be decreased in MI compared to SH rats. Perindopril restored PDE activities and tended to improve ANP efficiency in MI rats. One month after coronary ligation, perindopril treatment of MI rats prevents the increase in renal cortical PDE activities. This may contribute to increase renal ANP efficiency in MI rats. PMID:25939307

  12. Ca(2+)/calmodulin-activated phosphodiesterase 1A is highly expressed in rabbit cardiac sinoatrial nodal cells and regulates pacemaker function.

    PubMed

    Lukyanenko, Yevgeniya O; Younes, Antoine; Lyashkov, Alexey E; Tarasov, Kirill V; Riordon, Daniel R; Lee, Joonho; Sirenko, Syevda G; Kobrinsky, Evgeny; Ziman, Bruce; Tarasova, Yelena S; Juhaszova, Magdalena; Sollott, Steven J; Graham, David R; Lakatta, Edward G

    2016-09-01

    Constitutive Ca(2+)/calmodulin (CaM)-activation of adenylyl cyclases (ACs) types 1 and 8 in sinoatrial nodal cells (SANC) generates cAMP within lipid-raft-rich microdomains to initiate cAMP-protein kinase A (PKA) signaling, that regulates basal state rhythmic action potential firing of these cells. Mounting evidence in other cell types points to a balance between Ca(2+)-activated counteracting enzymes, ACs and phosphodiesterases (PDEs) within these cells. We hypothesized that the expression and activity of Ca(2+)/CaM-activated PDE Type 1A is higher in SANC than in other cardiac cell types. We found that PDE1A protein expression was 5-fold higher in sinoatrial nodal tissue than in left ventricle, and its mRNA expression was 12-fold greater in the corresponding isolated cells. PDE1 activity (nimodipine-sensitive) accounted for 39% of the total PDE activity in SANC lysates, compared to only 4% in left ventricular cardiomyocytes (LVC). Additionally, total PDE activity in SANC lysates was lowest (10%) in lipid-raft-rich and highest (76%) in lipid-raft-poor fractions (equilibrium sedimentation on a sucrose density gradient). In intact cells PDE1A immunolabeling was not localized to the cell surface membrane (structured illumination microscopy imaging), but located approximately within about 150nm inside of immunolabeling of hyperpolarization-activated cyclic nucleotide-gated potassium channels (HCN4), which reside within lipid-raft-rich microenvironments. In permeabilized SANC, in which surface membrane ion channels are not functional, nimodipine increased spontaneous SR Ca(2+) cycling. PDE1A mRNA silencing in HL-1 cells increased the spontaneous beating rate, reduced the cAMP, and increased cGMP levels in response to IBMX, a broad spectrum PDE inhibitor (detected via fluorescence resonance energy transfer microscopy). We conclude that signaling via cAMP generated by Ca(2+)/CaM-activated AC in SANC lipid raft domains is limited by cAMP degradation by Ca(2+)/Ca

  13. Defining the role of a FYVE domain in the localization and activity of a cAMP phosphodiesterase implicated in osmoregulation in Trypanosoma cruzi

    PubMed Central

    Schoijet, Alejandra C.; Miranda, Kildare; Medeiros, Lia Carolina Soares; de Souza, Wanderley; Flawiá, Mirtha M.; Torres, Héctor N.; Pignataro, Omar P.; Docampo, Roberto; Alonso, Guillermo D.

    2010-01-01

    Summary Intracellular levels of cyclic nucleotide second messengers are regulated predominantly by a large superfamily of phosphodiesterases. Trypanosoma cruzi, the causative agent of Chagas disease, encodes four different phosphodiesterase (PDE) families. One of these PDEs, T. cruzi phosphodiesterase C2 (TcrPDEC2) has been characterized as a FYVE-domain containing protein. Here, we report a novel role for TcrPDEC2 in osmoregulation in T. cruzi and reveal the relevance of its FYVE domain. Our data show that treatment of epimastigotes with TcrPDEC2 inhibitors improves their regulatory volume decrease, whereas cells overexpressing this enzyme are unaffected by the same inhibitors. Consistent with these results, TcrPDEC2 localizes to the contractile vacuole complex, showing strong labeling in the region corresponding to the spongiome. Furthermore, transgenic parasites overexpressing a truncated version of TcrPDEC2 without the FYVE domain show a failure in its targeting to the contractile vacuole complex and a marked decrease in phosphodiesterase activity, supporting the importance of this domain to the localization and activity of TcrPDEC2. Taking together, the results here presented are consistent with the importance of the cyclic AMP signaling pathway in regulatory volume decrease and implicate TcrPDEC2 as a specifically localized phosphodiesterase involved in osmoregulation in T. cruzi. PMID:21166893

  14. Dysregulation of hepatic cAMP levels via altered Pde4b expression plays a critical role in alcohol-induced steatosis.

    PubMed

    Avila, Diana V; Barker, David F; Zhang, JingWen; McClain, Craig J; Barve, Shirish; Gobejishvili, Leila

    2016-09-01

    Alcohol-induced hepatic steatosis is a significant risk factor for progressive liver disease. Cyclic adenosine monophosphate (cAMP) signalling has been shown to significantly regulate lipid metabolism; however, the role of altered cAMP homeostasis in alcohol-mediated hepatic steatosis has never been studied. Our previous work demonstrated that increased expression of hepatic phosphodiesterase 4 (Pde4), which specifically hydrolyses and decreases cAMP levels, plays a pathogenic role in the development of liver inflammation/injury. The aim of this study was to examine the role of PDE4 in alcohol-induced hepatic steatosis. C57BL/6 wild-type and Pde4b knockout (Pde4b(-/-) ) mice were pair-fed control or ethanol liquid diets. One group of wild-type mice received rolipram, a PDE4-specific inhibitor, during alcohol feeding. We demonstrate for the first time that an early increase in PDE4 enzyme expression and a resultant decrease in hepatic cAMP levels are associated with the significant reduction in carnitine palmitoyltransferase 1A (Cpt1a) expression. Notably, alcohol-fed (AF) Pde4b(-/-) mice and AF wild-type mice treated with rolipram had significantly lower hepatic free fatty acid content compared with AF wild-type mice. Importantly, PDE4 inhibition in alcohol-fed mice prevented the decrease in hepatic Cpt1a expression via the Pparα/Sirt1/Pgc1α pathway. These results demonstrate that the alcohol- induced increase in hepatic Pde4, specifically Pde4b expression, and compromised cAMP signalling predispose the liver to impaired fatty acid oxidation and the development of steatosis. Moreover, these data also suggest that hepatic PDE4 may be a clinically relevant therapeutic target for the treatment of alcohol-induced hepatic steatosis. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. PMID:27287961

  15. Evidence that protein kinase C may not be involved in the insulin action on cAMP phosphodiesterase: studies with electroporated rat adipocytes that were highly responsive to insulin.

    PubMed

    Shibata, H; Robinson, F W; Benzing, C F; Kono, T

    1991-02-15

    Partially permeabilized rat adipocytes with a high responsiveness to insulin were prepared by electroporation and used to study the effect of 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7) on insulin actions in adipocytes. H-7 is a well-documented inhibitor of several protein kinases, including protein kinase C; however, it does not rapidly enter adipocytes protected with the intact plasma membrane. The cells were suspended in Buffer X [4.74 mM NaCl, 118.0 mM KCl, 0.38 mM CaCl2, 1.00 mM EGTA, 1.19 mM Mg2SO4, 1.19 mM KH2PO4, 25.0 mM Hepes/K, 20 mg/ml bovine serum albumin, and 3 mM pyruvate/Na, pH 7.4] and electroporated six times with a Gene-Pulser (from Bio-Rad) set at 25 microF and 2 kV/cm. In cells electroporated as above, insulin stimulated (a) membrane-bound, cAMP phosphodiesterase approximately 2.6-fold when the hormone concentration was 10 nM and (b) glucose transport activity approximately 4.5-fold when the hormone concentration was raised to 100 nM. H-7 strongly inhibited the actions of insulin on both glucose transport (apparent Ki = 0.3 mM) and cAMP phosphodiesterase (apparent Ki = 1.2 mM) in electroporated adipocytes. H-7 also inhibited lipolysis in adipocytes; the apparent Ki value for the reaction in intact cells was 0.45 mM, and that in electroporated cells was 0.075 mM. It is suggested that a certain protein kinase or kinases that are significantly sensitive to H-7 may be involved in the insulin-dependent stimulation of glucose transport and that of phosphodiesterase. However, protein kinase C (or Ca2+/phospholipid-dependent protein kinase) may not be involved, at least, in the hormonal action on phosphodiesterase since the apparent Ki value of H-7 for the reaction is too high. PMID:1846737

  16. Altered gene expression in rat mesenteric tissue following in vivo exposure to a phosphodiesterase 4 inhibitor

    SciTech Connect

    Dagues, Nicolas . E-mail: nicolas.dagues@pfizer.com; Pawlowski, Valerie; Guigon, Ghislaine; Ledieu, David; Sobry, Cecile; Hanton, Gilles; Freslon, Jean-Louis; Chevalier, Stephan

    2007-01-01

    Vascular injury is a relatively common finding during the pre-clinical toxicity testing of drugs. The mechanisms of the injury are poorly understood and in turn, sensitive and specific biomarkers for pre-clinical and clinical monitoring do not exist. The present study was undertaken to investigate the molecular mechanisms of drug-induced vascular injury in mesenteric tissue of rats treated with the selective phosphodiesterase 4 (PDE4) inhibitor CI-1044. In a time-course study, male Sprague Dawley rats were given daily doses of 40 or 80 mg/kg for 1, 2 or 3 successive days and were euthanized the following day. Gene expression profiles in mesenteric tissue were determined using Affymetrix RG{sub U}34A microarrays and fibrinogen and cytokine measurements were performed in blood samples. Hierarchical clustering analysis produced a clear pattern separation of the animals with inflammation, animal with inflammation and necrosis and animals without any lesion. Genes associated with inflammation, procoagulation, extracellular matrix remodeling were up-regulated. An altered expression of genes involved in vascular tone regulation, lipid and glucose metabolism was also observed. Selected genes expression changes were confirmed by TaqMan real-time RT-PCR. The inflammatory process was also detected in the bloodstream at the protein level since fibrinogen, IL6 and IL1{beta} concentrations were increased in treated animals. Overall, the present study reveals several molecular changes supporting the hypothesis by which PDE4 inhibitor-induced vascular lesions in rats are triggered by an inflammatory mechanism and/or a vascular tone dysregulation.

  17. Cloning and expression of a cohemolysin, the CAMP factor of Actinobacillus pleuropneumoniae.

    PubMed Central

    Frey, J; Perrin, J; Nicolet, J

    1989-01-01

    The genetic determinant of the cohemolysin which is responsible for the CAMP phenomenon, a cohemolysis, of Actinobacillus pleuropneumoniae was cloned in Escherichia coli. Total DNA from the A. pleuropneumoniae serotype 1 type strain 4074 was used to construct a gene library in plasmid pUC18 in E. coli JM83. A total of 10,500 clones containing recombinant plasmids have been screened for hemolysis on blood plates. Fifty-five clones which showed a weak hemolytic response after 24 to 48 h of incubation were screened for the CAMP reaction with Staphylococcus aureus. This led to the identification of one clone which showed a positive CAMP reaction. Immunoblot analysis revealed that the recombinant strain expressed a protein with a molecular mass of 27,000 daltons, similar in size to the CAMP protein of the group B streptococci. Rabbit antibodies against the CAMP+ clone neutralized the CAMP reaction mediated by the E. coli strain containing the cloned CAMP gene as well as that of A. pleuropneumoniae. Antibodies raised against the cloned CAMP cohemolysin cross-reacted with Streptococcus agalactiae protein B. We designate the 27,000-dalton molecule CAMP factor protein and name its corresponding gene cfp. Images PMID:2659534

  18. Loss of extra-striatal phosphodiesterase 10A expression in early premanifest Huntington's disease gene carriers.

    PubMed

    Wilson, Heather; Niccolini, Flavia; Haider, Salman; Marques, Tiago Reis; Pagano, Gennaro; Coello, Christopher; Natesan, Sridhar; Kapur, Shitij; Rabiner, Eugenii A; Gunn, Roger N; Tabrizi, Sarah J; Politis, Marios

    2016-09-15

    Huntington's disease (HD) is a monogenic neurodegenerative disorder with an underlying pathology involving the toxic effect of mutant huntingtin protein primarily in striatal and cortical neurons. Phosphodiesterase 10A (PDE10A) regulates intracellular signalling cascades, thus having a key role in promoting neuronal survival. Using positron emission tomography (PET) with [(11)C]IMA107, we investigated the in vivo extra-striatal expression of PDE10A in 12 early premanifest HD gene carriers. Image processing and kinetic modelling was performed using MIAKAT™. Parametric images of [(11)C]IMA107 non-displaceable binding potential (BPND) were generated from the dynamic [(11)C]IMA107 scans using the simplified reference tissue model with the cerebellum as the reference tissue for nonspecific binding. We set a threshold criterion for meaningful quantification of [(11)C]IMA107 BPND at 0.30 in healthy control data; regions meeting this criterion were designated as regions of interest (ROIs). MRI-based volumetric analysis showed no atrophy in ROIs. We found significant differences in mean ROIs [(11)C]IMA107 BPND between HD gene carriers and healthy controls. HD gene carriers had significant loss of PDE10A within the insular cortex and occipital fusiform gyrus compared to healthy controls. Insula and occipital fusiform gyrus are important brain areas for the regulation of cognitive and limbic function that is impaired in HD. Our findings suggest that dysregulation of PDE10A-mediated intracellular signalling could be an early phenomenon in the course of HD with relevance also for extra-striatal brain areas.

  19. Loss of extra-striatal phosphodiesterase 10A expression in early premanifest Huntington's disease gene carriers.

    PubMed

    Wilson, Heather; Niccolini, Flavia; Haider, Salman; Marques, Tiago Reis; Pagano, Gennaro; Coello, Christopher; Natesan, Sridhar; Kapur, Shitij; Rabiner, Eugenii A; Gunn, Roger N; Tabrizi, Sarah J; Politis, Marios

    2016-09-15

    Huntington's disease (HD) is a monogenic neurodegenerative disorder with an underlying pathology involving the toxic effect of mutant huntingtin protein primarily in striatal and cortical neurons. Phosphodiesterase 10A (PDE10A) regulates intracellular signalling cascades, thus having a key role in promoting neuronal survival. Using positron emission tomography (PET) with [(11)C]IMA107, we investigated the in vivo extra-striatal expression of PDE10A in 12 early premanifest HD gene carriers. Image processing and kinetic modelling was performed using MIAKAT™. Parametric images of [(11)C]IMA107 non-displaceable binding potential (BPND) were generated from the dynamic [(11)C]IMA107 scans using the simplified reference tissue model with the cerebellum as the reference tissue for nonspecific binding. We set a threshold criterion for meaningful quantification of [(11)C]IMA107 BPND at 0.30 in healthy control data; regions meeting this criterion were designated as regions of interest (ROIs). MRI-based volumetric analysis showed no atrophy in ROIs. We found significant differences in mean ROIs [(11)C]IMA107 BPND between HD gene carriers and healthy controls. HD gene carriers had significant loss of PDE10A within the insular cortex and occipital fusiform gyrus compared to healthy controls. Insula and occipital fusiform gyrus are important brain areas for the regulation of cognitive and limbic function that is impaired in HD. Our findings suggest that dysregulation of PDE10A-mediated intracellular signalling could be an early phenomenon in the course of HD with relevance also for extra-striatal brain areas. PMID:27538642

  20. Reduced expression of cytochrome oxidases largely explains cAMP inhibition of aerobic growth in Shewanella oneidensis

    PubMed Central

    Yin, Jianhua; Meng, Qiu; Fu, Huihui; Gao, Haichun

    2016-01-01

    Inhibition of bacterial growth under aerobic conditions by elevated levels of cyclic adenosine 3′,5′-monophosphate (cAMP), first revealed more than 50 years ago, was attributed to accumulation of toxic methylglyoxal (MG). Here, we report a Crp-dependent mechanism rather than MG accumulation that accounts for the phenotype in Shewanella oneidensis, an emerging research model for the bacterial physiology. We show that a similar phenotype can be obtained by removing CpdA, a cAMP phosphodiesterase that appears more effective than its Escherichia coli counterpart. Although production of heme c and cytochromes c is correlated well with cAMP levels, neither is sufficient for the retarded growth. Quantities of overall cytochromes c increased substantially in the presence of elevated cAMP, a phenomenon resembling cells respiring on non-oxygen electron acceptors. In contrast, transcription of Crp-dependent genes encoding both cytochromes bd and cbb3 oxidases is substantially repressed under the same condition. Overall, our results suggest that cAMP of elevated levels drives cells into a low-energetic status, under which aerobic respiration is inhibited. PMID:27076065

  1. Reduced expression of cytochrome oxidases largely explains cAMP inhibition of aerobic growth in Shewanella oneidensis.

    PubMed

    Yin, Jianhua; Meng, Qiu; Fu, Huihui; Gao, Haichun

    2016-01-01

    Inhibition of bacterial growth under aerobic conditions by elevated levels of cyclic adenosine 3',5'-monophosphate (cAMP), first revealed more than 50 years ago, was attributed to accumulation of toxic methylglyoxal (MG). Here, we report a Crp-dependent mechanism rather than MG accumulation that accounts for the phenotype in Shewanella oneidensis, an emerging research model for the bacterial physiology. We show that a similar phenotype can be obtained by removing CpdA, a cAMP phosphodiesterase that appears more effective than its Escherichia coli counterpart. Although production of heme c and cytochromes c is correlated well with cAMP levels, neither is sufficient for the retarded growth. Quantities of overall cytochromes c increased substantially in the presence of elevated cAMP, a phenomenon resembling cells respiring on non-oxygen electron acceptors. In contrast, transcription of Crp-dependent genes encoding both cytochromes bd and cbb3 oxidases is substantially repressed under the same condition. Overall, our results suggest that cAMP of elevated levels drives cells into a low-energetic status, under which aerobic respiration is inhibited. PMID:27076065

  2. Depletion of tyrosyl-DNA phosphodiesterase 1α (MtTdp1α) affects transposon expression in Medicago truncatula.

    PubMed

    Sabatini, Maria Elisa; Donà, Mattia; Leonetti, Paola; Minio, Andrea; Delledonne, Massimo; Carboneral, Daniela; Confalonieri, Massimo; Giraffa, Giorgio; Balestrazzi, Alma

    2016-07-01

    The role of plant tyrosyl-DNA phosphodiesterase 1α in genome stability is studied using a Medicago truncatula MtTdp1α-depleted line. Lack of MtTdp1α results in a 39% reduction of methylated cytosines as compared to control. RNA-Seq analyses revealed that 11 DNA transposons and 22 retrotransposons were differentially expressed in the Tdp1α-2a line. Among them all, DNA transposons (MuDR, hAT, DNA3-11_Mad) and seven retrotransposons (LTR (Long Terminal Repeat)/Gipsy, LTR/Copia, LTR and NonLTR/L1) were down-regulated, while the 15 retrotransposons were up-regulated. Results suggest that the occurrence of stress-responsive cis-elements as well as changes in the methylation pattern at the LTR promoters might be responsible for the enhanced retrotransposon transcription. PMID:26699667

  3. Diminished phosphodiesterase-8B potentiates biphasic insulin response to glucose.

    PubMed

    Dov, Avital; Abramovitch, Eva; Warwar, Nasim; Nesher, Rafael

    2008-02-01

    cAMP activates multiple signal pathways, crucial for the pancreatic beta-cells function and survival and is a major potentiator of insulin release. A family of phosphodiesterases (PDEs) terminate the cAMP signals. We examined the expression of PDEs in rat beta-cells and their role in the regulation of insulin response. Using RT-PCR and Western blot analyses, we identified PDE3A, PDE3B, PDE4B, PDE4D, and PDE8B in rat islets and in INS-1E cells and several possible splice variants of these PDEs. Specific depletion of PDE3A with small interfering (si) RNA (siPDE3A) led to a small (67%) increase in the insulin response to glucose in INS-1E cells but not rat islets. siPDE3A had no effect on the glucagon-like peptide-1 (10 nmol/liter) potentiated insulin response in rat islets. Depletion in PDE8B levels in rat islets using similar technology (siPDE8B) increased insulin response to glucose by 70%, the potentiation being of similar magnitude during the first and second phase insulin release. The siPDE8B-potentiated insulin response was further increased by 23% when glucagon-like peptide-1 was included during the glucose stimulus. In conclusion, PDE8B is expressed in a small number of tissues unrelated to glucose or fat metabolism. We propose that PDE8B, an 3-isobutyl-1-methylxanthine-insensitive cAMP-specific phosphodiesterase, could prove a novel target for enhanced insulin response, affecting a specific pool of cAMP involved in the control of insulin granule trafficking and exocytosis. Finally, we discuss evidence for functional compartmentation of cAMP in pancreatic beta-cells.

  4. Acidosis is a key regulator of osteoblast ecto-nucleotidase pyrophosphatase/phosphodiesterase 1 (NPP1) expression and activity.

    PubMed

    Orriss, Isabel R; Key, Michelle L; Hajjawi, Mark O R; Millán, José L; Arnett, Timothy R

    2015-12-01

    Previous work has shown that acidosis prevents bone nodule formation by osteoblasts in vitro by inhibiting mineralisation of the collagenous matrix. The ratio of phosphate (Pi ) to pyrophosphate (PPi ) in the bone microenvironment is a fundamental regulator of bone mineralisation. Both Pi and PPi , a potent inhibitor of mineralisation, are generated from extracellular nucleotides by the actions of ecto-nucleotidases. This study investigated the expression and activity of ecto-nucleotidases by osteoblasts under normal and acid conditions. We found that osteoblasts express mRNA for a number of ecto-nucleotidases including NTPdase 1-6 (ecto-nucleoside triphosphate diphosphohydrolase) and NPP1-3 (ecto-nucleotide pyrophosphatase/phosphodiesterase). The rank order of mRNA expression in differentiating rat osteoblasts (day 7) was Enpp1 > NTPdase 4 > NTPdase 6 > NTPdase 5 >  alkaline phosphatase > ecto-5-nucleotidase > Enpp3 > NTPdase 1 > NTPdase 3 > Enpp2 > NTPdase 2. Acidosis (pH 6.9) upregulated NPP1 mRNA (2.8-fold) and protein expression at all stages of osteoblast differentiation compared to physiological pH (pH 7.4); expression of other ecto-nucleotidases was unaffected. Furthermore, total NPP activity was increased up to 53% in osteoblasts cultured in acid conditions (P < 0.001). Release of ATP, one of the key substrates for NPP1, from osteoblasts, was unaffected by acidosis. Further studies showed that mineralised bone formation by osteoblasts cultured from NPP1 knockout mice was increased compared with wildtypes (2.5-fold, P < 0.001) and was partially resistant to the inhibitory effect of acidosis. These results indicate that increased NPP1 expression and activity might contribute to the decreased mineralisation observed when osteoblasts are exposed to acid conditions.

  5. Alterations of Phosphodiesterases in Adrenocortical Tumors

    PubMed Central

    Hannah-Shmouni, Fady; Faucz, Fabio R.; Stratakis, Constantine A.

    2016-01-01

    Alterations in the cyclic (c)AMP-dependent signaling pathway have been implicated in the majority of benign adrenocortical tumors (ACTs) causing Cushing syndrome (CS). Phosphodiesterases (PDEs) are enzymes that regulate cyclic nucleotide levels, including cyclic adenosine monophosphate (cAMP). Inactivating mutations and other functional variants in PDE11A and PDE8B, two cAMP-binding PDEs, predispose to ACTs. The involvement of these two genes in ACTs was initially revealed by a genome-wide association study in patients with micronodular bilateral adrenocortical hyperplasia. Thereafter, PDE11A or PDE8B genetic variants have been found in other ACTs, including macronodular adrenocortical hyperplasias and cortisol-producing adenomas. In addition, downregulation of PDE11A expression and inactivating variants of the gene have been found in hereditary and sporadic testicular germ cell tumors, as well as in prostatic cancer. PDEs confer an increased risk of ACT formation probably through, primarily, their action on cAMP levels, but other actions might be possible. In this report, we review what is known to date about PDE11A and PDE8B and their involvement in the predisposition to ACTs. PMID:27625633

  6. Alterations of Phosphodiesterases in Adrenocortical Tumors

    PubMed Central

    Hannah-Shmouni, Fady; Faucz, Fabio R.; Stratakis, Constantine A.

    2016-01-01

    Alterations in the cyclic (c)AMP-dependent signaling pathway have been implicated in the majority of benign adrenocortical tumors (ACTs) causing Cushing syndrome (CS). Phosphodiesterases (PDEs) are enzymes that regulate cyclic nucleotide levels, including cyclic adenosine monophosphate (cAMP). Inactivating mutations and other functional variants in PDE11A and PDE8B, two cAMP-binding PDEs, predispose to ACTs. The involvement of these two genes in ACTs was initially revealed by a genome-wide association study in patients with micronodular bilateral adrenocortical hyperplasia. Thereafter, PDE11A or PDE8B genetic variants have been found in other ACTs, including macronodular adrenocortical hyperplasias and cortisol-producing adenomas. In addition, downregulation of PDE11A expression and inactivating variants of the gene have been found in hereditary and sporadic testicular germ cell tumors, as well as in prostatic cancer. PDEs confer an increased risk of ACT formation probably through, primarily, their action on cAMP levels, but other actions might be possible. In this report, we review what is known to date about PDE11A and PDE8B and their involvement in the predisposition to ACTs.

  7. Alterations of Phosphodiesterases in Adrenocortical Tumors.

    PubMed

    Hannah-Shmouni, Fady; Faucz, Fabio R; Stratakis, Constantine A

    2016-01-01

    Alterations in the cyclic (c)AMP-dependent signaling pathway have been implicated in the majority of benign adrenocortical tumors (ACTs) causing Cushing syndrome (CS). Phosphodiesterases (PDEs) are enzymes that regulate cyclic nucleotide levels, including cyclic adenosine monophosphate (cAMP). Inactivating mutations and other functional variants in PDE11A and PDE8B, two cAMP-binding PDEs, predispose to ACTs. The involvement of these two genes in ACTs was initially revealed by a genome-wide association study in patients with micronodular bilateral adrenocortical hyperplasia. Thereafter, PDE11A or PDE8B genetic variants have been found in other ACTs, including macronodular adrenocortical hyperplasias and cortisol-producing adenomas. In addition, downregulation of PDE11A expression and inactivating variants of the gene have been found in hereditary and sporadic testicular germ cell tumors, as well as in prostatic cancer. PDEs confer an increased risk of ACT formation probably through, primarily, their action on cAMP levels, but other actions might be possible. In this report, we review what is known to date about PDE11A and PDE8B and their involvement in the predisposition to ACTs. PMID:27625633

  8. Expression and Characterization of a Novel Glycerophosphodiester Phosphodiesterase from Pyrococcus furiosus DSM 3638 That Possesses Lysophospholipase D Activity.

    PubMed

    Wang, Fanghua; Lai, Linhui; Liu, Yanhua; Yang, Bo; Wang, Yonghua

    2016-01-01

    Glycerophosphodiester phosphodiesterases (GDPD) are enzymes which degrade various glycerophosphodiesters to produce glycerol-3-phosphate and the corresponding alcohol moiety. Apart from this, a very interesting finding is that this enzyme could be used in the degradation of toxic organophosphorus esters, which has resulted in much attention on the biochemical and application research of GDPDs. In the present study, a novel GDPD from Pyrococcus furiosus DSM 3638 (pfGDPD) was successfully expressed in Escherichia coli and biochemically characterized. This enzyme hydrolyzed bis(p-nitrophenyl) phosphate, one substrate analogue of organophosphorus diester, with an optimal reaction temperature 55 °C and pH 8.5. The activity of pfGDPD was strongly dependent on existing of bivalent cations. It was strongly stimulated by Mn(2+) ions, next was Co(2+) and Ni(2+) ions. Further investigations were conducted on its substrate selectivity towards different phospholipids. The results indicated that except of glycerophosphorylcholine (GPC), this enzyme also possessed lysophospholipase D activity toward both sn1-lysophosphatidylcholine (1-LPC) and sn2-lysophosphatidylcholine (2-LPC). Higher activity was found for 1-LPC than 2-LPC; however, no hydrolytic activity was found for phosphatidylcholine (PC). Molecular docking based on the 3D-modeled structure of pfGDPD was conducted in order to provide a structural foundation for the substrate selectivity. PMID:27248999

  9. Expression and Characterization of a Novel Glycerophosphodiester Phosphodiesterase from Pyrococcus furiosus DSM 3638 That Possesses Lysophospholipase D Activity

    PubMed Central

    Wang, Fanghua; Lai, Linhui; Liu, Yanhua; Yang, Bo; Wang, Yonghua

    2016-01-01

    Glycerophosphodiester phosphodiesterases (GDPD) are enzymes which degrade various glycerophosphodiesters to produce glycerol-3-phosphate and the corresponding alcohol moiety. Apart from this, a very interesting finding is that this enzyme could be used in the degradation of toxic organophosphorus esters, which has resulted in much attention on the biochemical and application research of GDPDs. In the present study, a novel GDPD from Pyrococcus furiosus DSM 3638 (pfGDPD) was successfully expressed in Escherichia coli and biochemically characterized. This enzyme hydrolyzed bis(p-nitrophenyl) phosphate, one substrate analogue of organophosphorus diester, with an optimal reaction temperature 55 °C and pH 8.5. The activity of pfGDPD was strongly dependent on existing of bivalent cations. It was strongly stimulated by Mn2+ ions, next was Co2+ and Ni2+ ions. Further investigations were conducted on its substrate selectivity towards different phospholipids. The results indicated that except of glycerophosphorylcholine (GPC), this enzyme also possessed lysophospholipase D activity toward both sn1-lysophosphatidylcholine (1-LPC) and sn2-lysophosphatidylcholine (2-LPC). Higher activity was found for 1-LPC than 2-LPC; however, no hydrolytic activity was found for phosphatidylcholine (PC). Molecular docking based on the 3D-modeled structure of pfGDPD was conducted in order to provide a structural foundation for the substrate selectivity. PMID:27248999

  10. Diatom acclimation to elevated CO2 via cAMP signalling and coordinated gene expression

    NASA Astrophysics Data System (ADS)

    Hennon, Gwenn M. M.; Ashworth, Justin; Groussman, Ryan D.; Berthiaume, Chris; Morales, Rhonda L.; Baliga, Nitin S.; Orellana, Mónica V.; Armbrust, E. V.

    2015-08-01

    Diatoms are responsible for ~40% of marine primary productivity, fuelling the oceanic carbon cycle and contributing to natural carbon sequestration in the deep ocean. Diatoms rely on energetically expensive carbon concentrating mechanisms (CCMs) to fix carbon efficiently at modern levels of CO2 (refs , , ). How diatoms may respond over the short and long term to rising atmospheric CO2 remains an open question. Here we use nitrate-limited chemostats to show that the model diatom Thalassiosira pseudonana rapidly responds to increasing CO2 by differentially expressing gene clusters that regulate transcription and chromosome folding, and subsequently reduces transcription of photosynthesis and respiration gene clusters under steady-state elevated CO2. These results suggest that exposure to elevated CO2 first causes a shift in regulation, and then a metabolic rearrangement. Genes in one CO2-responsive cluster included CCM and photorespiration genes that share a putative cAMP-responsive cis-regulatory sequence, implying these genes are co-regulated in response to CO2, with cAMP as an intermediate messenger. We verified cAMP-induced downregulation of CCM gene δ-CA3 in nutrient-replete diatom cultures by inhibiting the hydrolysis of cAMP. These results indicate an important role for cAMP in downregulating CCM and photorespiration genes under elevated CO2 and provide insights into mechanisms of diatom acclimation in response to climate change.

  11. Selective Phosphodiesterase 4B Inhibitors: A Review

    PubMed Central

    Azam, Mohammed Afzal; Tripuraneni, Naga Srinivas

    2014-01-01

    Abstract Phosphodiesterase 4B (PDE4B) is a member of the phosphodiesterase family of proteins that plays a critical role in regulating intracellular levels of cyclic adenosine monophosphate (cAMP) by controlling its rate of degradation. It has been demonstrated that this isoform is involved in the orchestra of events which includes inflammation, schizophrenia, cancers, chronic obstructive pulmonary disease, contractility of the myocardium, and psoriatic arthritis. Phosphodiesterase 4B has constituted an interesting target for drug development. In recent years, a number of PDE4B inhibitors have been developed for their use as therapeutic agents. In this review, an up-to-date status of the inhibitors investigated for the inhibition of PDE4B has been given so that this rich source of structural information of presently known PDE4B inhibitors could be helpful in generating a selective and potent inhibitor of PDE4B. PMID:25853062

  12. Host cell contact induces expression of virulence factors and VieA, a cyclic di-GMP phosphodiesterase, in Vibrio cholerae.

    PubMed

    Dey, Amit K; Bhagat, Abha; Chowdhury, Rukhsana

    2013-05-01

    Vibrio cholerae, a noninvasive bacterium, colonizes the intestinal epithelium and secretes cholera toxin (CT), a potent enterotoxin that causes the severe fluid loss characteristic of the disease cholera. In this study, we demonstrate that adherence of V. cholerae to the intestinal epithelial cell line INT 407 strongly induces the expression of the major virulence genes ctxAB and tcpA and the virulence regulatory gene toxT. No induction of toxR and tcpP, which encode transcriptional activators of toxT, was observed in adhered bacteria, and the adherence-dependent upregulation of toxT expression was independent of ToxR and TcpP. A sharp increase in the expression of the vieA gene, which encodes a cyclic di-GMP (c-di-GMP) phosphodiesterase, was observed in INT 407-adhered V. cholerae immediately after infection. Induction of toxT, ctxAB, and tcpA in INT 407-adhered vieA mutant strain O395 ΔvieA was consistently lower than in the parent strain, although no effect was observed in unadhered bacteria, suggesting that VieA has a role in the upregulation of toxT expression specifically in host cell-adhered V. cholerae. Furthermore, though VieA has both a DNA binding helix-turn-helix domain and an EAL domain conferring c-di-GMP phosphodiesterase activity, the c-di-GMP phosphodiesterase activity of VieA is necessary and sufficient for the upregulation of toxT expression.

  13. Repeated immobilization stress disturbed steroidogenic machinery and stimulated the expression of cAMP signaling elements and adrenergic receptors in Leydig cells.

    PubMed

    Stojkov, Natasa J; Janjic, Marija M; Bjelic, Maja M; Mihajlovic, Aleksandar I; Kostic, Tatjana S; Andric, Silvana A

    2012-05-01

    This study was designed to evaluate the effect of acute (2 h daily) and repeated (2 h daily for 2 or 10 consecutive days) immobilization stress (IMO) on: 1) the steroidogenic machinery homeostasis; 2) cAMP signaling; and the expression of receptors for main markers of 3) adrenergic and 4) glucocorticoid signaling in Leydig cells of adult rats. The results showed that acute IMO inhibited steroidogenic machinery in Leydig cells by downregulation of Scarb1 (scavenger receptor class B), Cyp11a1 (cholesterol side-chain cleavage enzyme), Cyp17a1 (17α-hydroxylase/17,20 lyase), and Hsd17b3 (17β-hydroxysteroid dehydrogenase) expression. In addition to acute IMO effects, repeated IMO increased transcription of Star (steroidogenic acute regulatory protein) and Arr19 (androgen receptor corepressor 19 kDa) in Leydig cells. In the same cells, the transcription of adenylyl cyclases (Adcy7, Adcy9, Adcy10) and cAMP-specific phosphodiesterases (Pde4a, Pde4b, Pde4d, Pde7a, Pde8a) was stimulated, whereas the expression of the genes encoding protein kinase A subunits were unaffected. Ten times repeated IMO increased the levels of all adrenergic receptors and β-adrenergic receptor kinase (Adrbk1) in Leydig cells. The transcription analysis was supported by cAMP/testosterone production. In this signaling scenario, partial recovery of testosterone production in medium/content was detected. The physiological significance of the present results was proven by ex vivo application of epinephrine, which increased cAMP/testosterone production by Leydig cells from control rats in greater fashion than from stressed. IMO did not affect the expression of transcripts for Crhr1/Crhr2 (corticotropin releasing hormone receptors), Acthr (adrenocorticotropin releasing hormone receptor), Gr (glucocorticoid receptor), and Hsd11b1 [hydroxysteroid (11-β) dehydrogenase 1], while all types of IMO stimulated the expression of Hsd11b2, the unidirectional oxidase with high affinity to inactivate

  14. Exchange factors directly activated by cAMP mediate melanocortin 4 receptor-induced gene expression

    PubMed Central

    Glas, Evi; Mückter, Harald; Gudermann, Thomas; Breit, Andreas

    2016-01-01

    Gs protein-coupled receptors regulate many vital body functions by activation of cAMP response elements (CRE) via cAMP-dependent kinase A (PKA)-mediated phosphorylation of the CRE binding protein (CREB). Melanocortin 4 receptors (MC4R) are prototypical Gs-coupled receptors that orchestrate the hypothalamic control of food-intake and metabolism. Remarkably, the significance of PKA for MC4R-induced CRE-dependent transcription in hypothalamic cells has not been rigorously interrogated yet. In two hypothalamic cell lines, we observed that blocking PKA activity had only weak or no effects on reporter gene expression. In contrast, inhibitors of exchange factors directly activated by cAMP-1/2 (EPAC-1/2) mitigated MC4R-induced CRE reporter activation and mRNA induction of the CREB-dependent genes c-fos and thyrotropin-releasing hormone. Furthermore, we provide first evidence that extracellular-regulated kinases-1/2 (ERK-1/2) activated by EPACs and not PKA are the elusive CREB kinases responsible for MC4R-induced CREB/CRE activation in hypothalamic cells. Overall, these data emphasize the pivotal role of EPACs rather than PKA in hypothalamic gene expression elicited by a prototypical Gs-coupled receptor. PMID:27612207

  15. Exchange factors directly activated by cAMP mediate melanocortin 4 receptor-induced gene expression.

    PubMed

    Glas, Evi; Mückter, Harald; Gudermann, Thomas; Breit, Andreas

    2016-01-01

    Gs protein-coupled receptors regulate many vital body functions by activation of cAMP response elements (CRE) via cAMP-dependent kinase A (PKA)-mediated phosphorylation of the CRE binding protein (CREB). Melanocortin 4 receptors (MC4R) are prototypical Gs-coupled receptors that orchestrate the hypothalamic control of food-intake and metabolism. Remarkably, the significance of PKA for MC4R-induced CRE-dependent transcription in hypothalamic cells has not been rigorously interrogated yet. In two hypothalamic cell lines, we observed that blocking PKA activity had only weak or no effects on reporter gene expression. In contrast, inhibitors of exchange factors directly activated by cAMP-1/2 (EPAC-1/2) mitigated MC4R-induced CRE reporter activation and mRNA induction of the CREB-dependent genes c-fos and thyrotropin-releasing hormone. Furthermore, we provide first evidence that extracellular-regulated kinases-1/2 (ERK-1/2) activated by EPACs and not PKA are the elusive CREB kinases responsible for MC4R-induced CREB/CRE activation in hypothalamic cells. Overall, these data emphasize the pivotal role of EPACs rather than PKA in hypothalamic gene expression elicited by a prototypical Gs-coupled receptor. PMID:27612207

  16. Involvement of Type 4 cAMP-Phosphodiesterase in the Myogenic Differentiation of L6 Cells

    PubMed Central

    Naro, Fabio; Sette, Claudio; Vicini, Elena; De Arcangelis, Vania; Grange, Muriel; Conti, Marco; Lagarde, Michel; Molinaro, Mario; Adamo, Sergio; Némoz, Georges

    1999-01-01

    Myogenic cell differentiation is induced by Arg8-vasopressin, whereas high cAMP levels and protein kinase A (PKA) activity inhibit myogenesis. We investigated the role of type 4 phosphodiesterase (PDE4) during L6-C5 myoblast differentiation. Selective PDE4 inhibition resulted in suppression of differentiation induced by vasopressin. PDE4 inhibition prevented vasopressin-induced nuclear translocation of the muscle-specific transcription factor myogenin without affecting its overall expression level. The effects of PDE4 inhibition could be attributed to an increase of cAMP levels and PKA activity. RNase protection, reverse transcriptase PCR, immunoprecipitation, Western blot, and enzyme activity assays demonstrated that the PDE4D3 isoform is the major PDE4 expressed in L6-C5 myoblasts and myotubes, accounting for 75% of total cAMP-hydrolyzing activity. Vasopressin cell stimulation caused a biphasic increase of PDE4 activity, which peaked at 2 and 15 min and remained elevated for 48 h. In the continuous presence of vasopressin, cAMP levels and PKA activity were lowered. PDE4D3 overexpression increased spontaneous and vasopressin-dependent differentiation of L6-C5 cells. These results show that PDE4D3 plays a key role in the control of cAMP levels and differentiation of L6-C5 cells. Through the modulation of PDE4 activity, vasopressin inhibits the cAMP signal transduction pathway, which regulates myogenesis possibly by controlling the subcellular localization of myogenin. PMID:10588663

  17. Leptin receptor expressing neurons express phosphodiesterase-3B (PDE3B) and leptin induces STAT3 activation in PDE3B neurons in the mouse hypothalamus.

    PubMed

    Sahu, Maitrayee; Sahu, Abhiram

    2015-11-01

    Leptin signaling in the hypothalamus is critical for normal food intake and body weight regulation. Cumulative evidence suggests that besides the signal transducer and activator of transcription-3 (STAT3) pathway, several non-STAT3 pathways including the phosphodiesterase-3B (PDE3B) pathway mediate leptin signaling in the hypothalamus. We have shown that PDE3B is localized in various hypothalamic sites implicated in the regulation of energy homeostasis and that the anorectic and body weight reducing effects of leptin are mediated by the activation of PDE3B. It is still unknown if PDE3B is expressed in the long form of the leptin-receptor (ObRb)-expressing neurons in the hypothalamus and whether leptin induces STAT3 activation in PDE3B-expressing neurons. In this study, we examined co-localization of PDE3B with ObRb neurons in various hypothalamic nuclei in ObRb-GFP mice that were treated with leptin (5mg/kg, ip) for 2h. Results showed that most of the ObRb neurons in the arcuate nucleus (ARC, 93%), ventromedial nucleus (VMN, 94%), dorsomedial nucleus (DMN, 95%), ventral premammillary nucleus (PMv, 97%) and lateral hypothalamus (LH, 97%) co-expressed PDE3B. We next examined co-localization of p-STAT3 and PDE3B in the hypothalamus in C57BL6 mice that were treated with leptin (5mg/kg, ip) for 1h. The results showed that almost all p-STAT3 positive neurons in different hypothalamic nuclei including ARC, VMN, DMN, LH and PMv areas expressed PDE3B. These results suggest the possibility for a direct role for the PDE3B pathway in mediating leptin action in the hypothalamus.

  18. Atrazine Acts as an Endocrine Disrupter by Inhibiting cAMP-specific Phosphodiesterase-4

    PubMed Central

    Kucka, Marek; Pogrmic-Majkic, Kristina; Fa, Svetlana; Stojilkovic, Stanko S.; Kovacevic, Radmila

    2014-01-01

    Atrazine, one of the most commonly used herbicides worldwide, acts as an endocrine disruptor, but the mechanism of its action has not been characterized. In this study, we show that atrazine rapidly increases cAMP levels in cultured rat pituitary and testicular Leydig cells in a concentration-dependent manner, but less effectively than 3-isobutyl-1-methylxanthine, a competitive non-specific inhibitor of phosphodiesterases (PDEs). In forskolin (an activator of adenylyl cyclase)- and probenecid (an inhibitor of cyclic nucleotide transporters)-treated cells, but not in 3-isobutyl-1-methylxanthine-treated cells, atrazine further increased cAMP levels, indicating that inhibition of PDEs accounts for accumulation of cAMP. In contrast to cAMP, atrazine did not alter cGMP levels, further indicating that it inhibits cAMP-specific PDEs. Atrazine-induced changes in cAMP levels were sufficient to stimulate prolactin release in pituitary cells and androgen production in Leydig cells, indicating that it acts as an endocrine disrupter both in cells that secrete by exocytosis of prestored hormones and in cells that secrete by de novo hormone synthesis. Rolipram abolished the stimulatory effect of atrazine on cAMP release in both cell types, suggesting that it acts as an inhibitor of PDE4s, isoforms whose mRNA transcripts dominate in pituitary and Leydig cells together with mRNA for PDE8A. In contrast, immortalized lacto-somatotrophs showed low expression of these mRNA transcripts and several fold higher cAMP levels compared to normal pituitary cells, and atrazine was unable to further increase cAMP levels. These results indicate that atrazine acts as a general endocrine disrupter by inhibiting cAMP-specific PDE4s. PMID:23022511

  19. The cyclic AMP phosphodiesterase RegA critically regulates encystation in social and pathogenic amoebas.

    PubMed

    Du, Qingyou; Schilde, Christina; Birgersson, Elin; Chen, Zhi-hui; McElroy, Stuart; Schaap, Pauline

    2014-02-01

    Amoebas survive environmental stress by differentiating into encapsulated cysts. As cysts, pathogenic amoebas resist antibiotics, which particularly counteracts treatment of vision-destroying Acanthamoeba keratitis. Limited genetic tractability of amoeba pathogens has left their encystation mechanisms unexplored. The social amoeba Dictyostelium discoideum forms spores in multicellular fruiting bodies to survive starvation, while other dictyostelids, such as Polysphondylium pallidum can additionally encyst as single cells. Sporulation is induced by cAMP acting on PKA, with the cAMP phosphodiesterase RegA critically regulating cAMP levels. We show here that RegA is deeply conserved in social and pathogenic amoebas and that deletion of the RegA gene in P. pallidum causes precocious encystation and prevents cyst germination. We heterologously expressed and characterized Acanthamoeba RegA and performed a compound screen to identify RegA inhibitors. Two effective inhibitors increased cAMP levels and triggered Acanthamoeba encystation. Our results show that RegA critically regulates Amoebozoan encystation and that components of the cAMP signalling pathway could be effective targets for therapeutic intervention with encystation.

  20. cAMP increases surface expression of NKCC2 in rat thick ascending limbs: role of VAMP.

    PubMed

    Ortiz, Pablo A

    2006-03-01

    NaCl absorption by the thick ascending limb of Henle's loop (TAL) is mediated by the apical Na-K-2Cl cotransporter NKCC2. cAMP increases NaCl absorption in the TAL by stimulating NKCC2. In oocytes, cAMP increases NKCC2 activity by regulating its trafficking. However, the mechanism by which cAMP stimulates NKCC2 in TALs is not clear. We hypothesized that cAMP increases surface expression of NKCC2 and NaCl absorption in TALs and that vesicle-associated membrane protein (VAMP) is involved in this mechanism. We used surface biotinylation of rat medullary TALs (mTAL) to examine surface and total NKCC2 levels. When mTAL suspensions were treated with dibutyryl cAMP (db-cAMP) or forskolin plus IBMX for 20 min, surface NKCC2 expression increased by 126 +/- 23 and 92 +/- 17% above basal, respectively (P < 0.03). No changes in total NKCC2 expression were observed, suggesting that cAMP increased translocation of NKCC2. We studied the role of VAMP in NKCC2 translocation and found that incubating mTALs with tetanus toxin (30 nM), which inhibits vesicle trafficking by inactivating VAMP-2 and -3, completely blocked the stimulatory effect of db-cAMP on surface NKCC2 expression (tetanus toxin = 100% vs. tetanus toxin + db-cAMP = 102 +/- 21% of control; not significant). We studied VAMP-2 and -3 expression and localization in isolated perfused TALs by confocal microscopy and found that both of them were located in the subapical space of the TAL. Finally, in isolated perfused mTALs, db-cAMP increased net Cl absorption by 95.0 +/- 34.8% (P < 0.03), and pretreatment of TALs with tetanus toxin blocked the stimulation of Cl absorption (from 110.9 +/- 15.9 to 109.7 +/- 15.6 pmol.min(-1).mm(-1); not significant). We concluded that cAMP increases NKCC2 surface expression by a mechanism involving VAMP and that NKCC2 trafficking to the apical membrane is involved in the stimulation of TAL NaCl absorption by cAMP.

  1. Gene Expression Patterns Define Key Transcriptional Events InCell-Cycle Regulation By cAMP And Protein Kinase A

    SciTech Connect

    Zambon, Alexander C.; Zhang, Lingzhi; Minovitsky, Simon; Kanter, Joan R.; Prabhakar, Shyam; Salomonis, Nathan; Vranizan, Karen; Dubchak Inna,; Conklin, Bruce R.; Insel, Paul A.

    2005-06-01

    Although a substantial number of hormones and drugs increase cellular cAMP levels, the global impact of cAMP and its major effector mechanism, protein kinase A (PKA), on gene expression is not known. Here we show that treatment of murine wild-type S49 lymphoma cells for 24 h with 8-(4-chlorophenylthio)-cAMP (8-CPTcAMP), a PKA-selective cAMP analog, alters the expression of approx equal to 4,500 of approx. equal to 13,600 unique genes. By contrast, gene expression was unaltered in Kin- S49 cells (that lack PKA) incubated with 8-CPTcAMP. Changes in mRNA and protein expression of several cell cycle regulators accompanied cAMP-induced G1-phase cell-cycle arrest of wild-type S49 cells. Within 2h, 8-CPT-cAMP altered expression of 152 genes that contain evolutionarily conserved cAMP-response elements within 5 kb of transcriptional start sites, including the circadian clock gene Per1. Thus, cAMP through its activation of PKA produces extensive transcriptional regulation in eukaryotic cells. These transcriptional networks include a primary group of cAMP-response element-containing genes and secondary networks that include the circadian clock.

  2. Phosphodiesterases in endocrine physiology and disease.

    PubMed

    Vezzosi, Delphine; Bertherat, Jérôme

    2011-08-01

    The cAMP-protein kinase A pathway plays a central role in the development and physiology of endocrine tissues. cAMP mediates the intracellular effects of numerous peptide hormones. Various cellular and molecular alterations of the cAMP-signaling pathway have been observed in endocrine diseases. Phosphodiesterases (PDEs) are key regulatory enzymes of intracellular cAMP levels. Indeed, PDEs are the only known mechanism for inactivation of cAMP by catalysis to 5'-AMP. It has been suggested that disruption of PDEs could also have a role in the pathogenesis of many endocrine diseases. This review summarizes the most recent advances concerning the role of the PDEs in the physiopathology of endocrine diseases. The potential significance of this knowledge can be easily envisaged by the development of drugs targeting specific PDEs.

  3. Expression, intracellular distribution and basis for lack of catalytic activity of the PDE4A7 isoform encoded by the human PDE4A cAMP-specific phosphodiesterase gene.

    PubMed Central

    Johnston, Lee Ann; Erdogan, Suat; Cheung, York Fong; Sullivan, Michael; Barber, Rachael; Lynch, Martin J; Baillie, George S; Van Heeke, Gino; Adams, David R; Huston, Elaine; Houslay, Miles D

    2004-01-01

    PDE4A7 is an isoform encoded by the human PDE4A cAMP-specific phosphodiesterase gene that fails to hydrolyse cAMP and whose transcripts are widely expressed. Removal of either the N- or C-terminal unique portions of PDE4A7 did not reconstitute catalytic activity, showing that they did not exert a chronic inhibitory effect. A chimera (Hyb2), formed by swapping the unique N-terminal portion of PDE4A7 with that of the active PDE4A4C form, was not catalytically active. However, one formed (Hyb1) by swapping the unique C-terminal portion of PDE4A7 with that common to all active PDE4 isoforms was catalytically active. Compared with the active PDE4A4B isoform, Hyb1 exhibited a similar K(m) value for cAMP and IC50 value for rolipram inhibition, but was less sensitive to inhibition by Ro-20-1724 and denbufylline, and considerably more sensitive to thermal denaturation. The unique C-terminal region of PDE4A7 was unable to support an active catalytic unit, whereas its unique N-terminal region can. The N-terminal portion of the PDE4 catalytic unit is essential for catalytic activity and can be supplied by either highly conserved sequence found in active PDE4 isoforms from all four PDE4 subfamilies or the unique N-terminal portion of PDE4A7. A discrete portion of the conserved C-terminal region in active PDE4A isoforms underpins their aberrant migration on SDS/PAGE. Unlike active PDE4A isoforms, PDE4A7 is exclusively localized to the P1 particulate fraction in cells. A region located within the C-terminal portion of active PDE4 isoforms prevents such exclusive targeting. Three functional regions in PDE4A isoforms are identified, which influence catalytic activity, subcellular targeting and conformational status. PMID:15025561

  4. Levodopa-induced dyskinesias are associated with transient down-regulation of cAMP and cGMP in the caudate-putamen of hemiparkinsonian rats: reduced synthesis or increased catabolism?

    PubMed

    Sancesario, Giuseppe; Morrone, Luigi Antonio; D'Angelo, Vincenza; Castelli, Valentina; Ferrazzoli, Davide; Sica, Francesco; Martorana, Alessandro; Sorge, Roberto; Cavaliere, Federica; Bernardi, Giorgio; Giorgi, Mauro

    2014-12-01

    Second messenger cAMP and cGMP represent a key step in the action of dopamine that modulates directly or indirectly their synthesis. We aimed to verify whether levodopa-induced dyskinesias are associated with changes of the time course of levodopa/dopamine stimulated cAMP and cGMP levels, and/or with changes of their catabolism by phosphodiesterase activity in rats with experimental hemiparkinsonism. Microdialysis and tissue homogenates of the striatal tissues demonstrated that extracellular and intracellular cAMP/cGMP levels were lower in dyskinetic animals during the increasing phase of dyskinesias compared to eukinetic animals, but cAMP/cGMP levels increased in dyskinetic animals during the phase of decreasing and extinction of dyskinesias. Dyskinesias and the abnormal lowering of striatal cGMP and cAMP after levodopa were prevented by pretreatment with the multipotent drug amantadine, outlining the inverse relationship of cAMP/cGMP to dyskinesias. Moreover, dyskinetic animals showed higher striatal hydrolyzing cGMP-phosphodiesterase but not hydrolyzing cAMP-phosphodiesterase activity, suggesting that low cGMP but not cAMP levels could be due to increased catabolism. However, expressions of isozyme phosphodiesterase-1B and -10A highly and specifically located in the basal ganglia were not changed after levodopa in dyskinetic and eukinetic animals: accordingly, selective inhibitors of phosphodiesterase-1B and -10A were ineffective on levodopa dyskinesias. Therefore, the isozyme(s) expressing higher cGMP-phosphodiesterase activity in the striatum of dyskinetic animal should be determined. These observations suggest that dopamine-mediated processes of synthesis and/or degradation of cAMP/cGMP could be acutely impaired in levodopa dyskinesias, opening new ways to understanding physiopathology and treatment.

  5. Cytokine-Induced S-Nitrosylation of Soluble Guanylyl Cyclase and Expression of Phosphodiesterase 1A Contribute to Dysfunction of Longitudinal Smooth Muscle Relaxation.

    PubMed

    Rajagopal, Senthilkumar; Nalli, Ancy D; Kumar, Divya P; Bhattacharya, Sayak; Hu, Wenhui; Mahavadi, Sunila; Grider, John R; Murthy, Karnam S

    2014-09-01

    The following manuscript was published as a Fast Forward article on September 9, 2014: Rajagopal S, Nalli AD, Kumar DP, Bhattacharya S, Hu W, Mahavadi S, Grider JR, and Murthy KS, Cytokine-Induced S-Nitrosylation of Soluble Guanylyl Cyclase and Expression of Phosphodiesterase 1A Contribute to Dysfunction of Longitudinal Smooth Muscle Relaxation. J Pharmacol Exp Ther jpet.114.218156; doi:10.1124/jpet.114.218156. It was later found that the chemical identity of IC86340 was not sufficiently disclosed. The authors are unable, at this time, to provide this information in accordance with the editorial policies of The Journal of Pharmacology and Experimental Therapeutics. As a result, the manuscript has been withdrawn from publication. We apologize for any inconvenience this may cause JPET's readers. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

  6. Coupling gene expression to cAMP signalling: role of CREB and CREM.

    PubMed

    Sassone-Corsi, P

    1998-01-01

    Several endocrine and neuronal functions are governed by the cAMP-dependent pathway. Transcriptional regulation upon stimulation of this pathway is mediated by a family of cAMP-responsive nuclear factors. This family consists of a large number of members, which may act as activators or repressors. These factors contain the basic domain/leucine zipper motifs and bind as dimers to cAMP-response elements (CRE). CRE-binding protein (CREBs) function is modulated by phosphorylation by several kinases. Direct activation of gene expression by CREB requires phosphorylation by the cAMP-dependent PKA to serine 133. Among the repressors, ICER (Inducible cAMP Early Repressor) deserves special mention. ICER is generated from an alternative CREM promoter and is the only inducible CRE-binding protein. ICER negatively autoregulates the alternative promoter, generating a feedback loop. ICER expression is tissue specific and developmentally regulated. The kinetics of ICER expression are characteristic of an early response gene. CREM plays a key physiological and developmental role within the hypothalamic-pituitary-gonadal axis. The transcriptional activator CREM is highly expressed in postmeiotic cells. The role of CREM in spermiogenesis was addressed using CREM knock-out mice. Spermatogenesis stops at the first step of spermiogenesis in the mutants and there is a significant increase in apoptotic germ cells. This phenotype is reminiscent of cases of human infertility. ICER is regulated in a circadian manner in the pineal gland, the site of the hormone melatonin production. This night-day oscillation is driven by the endogenous clock (located in the suprachiasmatic nucleus). The synthesis of melatonin is regulated by a rate-limiting enzyme, serotonin N-acetyltransferase (NAT). Analysis of the CREM-null mice and of the promoter of the NAT gene revealed that ICER controls the amplitude and rhythmicity of NAT, and thus the oscillation in the hormonal synthesis of melatonin.

  7. Cyclic Nucleotide Compartmentalization: Contributions of Phosphodiesterases and ATP-Binding Cassette Transporters

    PubMed Central

    Cheepala, Satish; Hulot, Jean-Sebastien; Morgan, Jessica A.; Sassi, Yassine; Zhang, Weiqiang; Naren, Anjaparavanda P.; Schuetz, John D.

    2015-01-01

    Cyclic nucleotides [e.g., cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP)] are ubiquitous second messengers that affect multiple cell functions from maturation of the egg to cell division, growth, differentiation, and death. The concentration of cAMP can be regulated by processes within membrane domains (local regulation) as well as throughout a cell (global regulation). The phosphodiesterases (PDEs) that degrade cAMP have well-known roles in both these processes. It has recently been discovered that ATP-binding cassette (ABC) transporters contribute to both local and global regulation of cAMP. This regulation may require the formation of macromolecular complexes. Some of these transporters are ubiquitously expressed, whereas others are more tissue restricted. Because some PDE inhibitors are also ABC transporter inhibitors, it is conceivable that the therapeutic benefits of their use result from the combined inhibition of both PDEs and ABC transporters. Deciphering the individual contributions of PDEs and ABC transporters to such drug effects may lead to improved therapeutic benefits. PMID:23072381

  8. PPARgamma-dependent regulation of adenylate cyclase 6 amplifies the stimulatory effect of cAMP on renin gene expression.

    PubMed

    Desch, Michael; Schubert, Thomas; Schreiber, Andrea; Mayer, Sandra; Friedrich, Björn; Artunc, Ferruh; Todorov, Vladimir T

    2010-11-01

    The second messenger cAMP plays an important role in the regulation of renin gene expression. Nuclear receptor peroxisome proliferator-activated receptor-γ (PPARγ) is known to stimulate renin gene transcription acting through PPARγ-binding sequences in renin promoter. We show now that activation of PPARγ by unsaturated fatty acids or thiazolidinediones drastically augments the cAMP-dependent increase of renin mRNA in the human renin-producing cell line Calu-6. The underlying mechanism involves potentiation of agonist-induced cAMP increase and up-regulation of adenylate cyclase 6 (AC6) gene expression. We identified a palindromic element with a 3-bp spacer (Pal3) in AC6 intron 1 (AC6Pal3). AC6Pal3 bound PPARγ and mediated trans-activation by PPARγ agonist. AC6 knockdown decreased basal renin mRNA level and attenuated the maximal PPARγ-dependent stimulation of the cAMP-induced renin gene expression. AC6Pal3 decoy oligonucleotide abrogated the PPARγ-dependent potentiation of cAMP-induced renin gene expression. Treatment of mice with PPARγ agonist increased AC6 mRNA kidney levels. Our data suggest that in addition to its direct effect on renin gene transcription, PPARγ "sensitizes" renin gene to cAMP via trans-activation of AC6 gene. AC6 has been identified as PPARγ target gene with a functional Pal3 sequence.

  9. Lesbian camp: An unearthing.

    PubMed

    Nielsen, Elly-Jean

    2016-01-01

    Camp-a sensibility, a style, and a form of artistic self-expression-is an elusive concept said to be in the eye of the beholder. To refute Susan Sontag's ( 1966 ) claims that camp is apolitical and not especially homosexual, a number of recent scholarly works have been geared toward revealing camp's fundamental gayness. With the odd footnote aside, lesbian camp has been collapsed into the category of gay male camp, if not eclipsed entirely. Despite the negligible efforts made to legitimize lesbian camp, there are numerous salient cultural examples one might draw on to illustrate, typify, and substantiate a lesbian camp sensibility. I lay the ground work for this scholarly exercise by outlining various definitions and critiques of camp, and by discussing its history and application to queer theory. Then, to unveil lesbian camp, three non-mutually exclusive categories are discussed: classic, erotic, and radical. By gathering various strands of inquiry, and various textual examples (e.g., photography, artistic performances, and literary tropes), this article attempts to reach a more inclusive and nuanced understanding of lesbian camp. PMID:26701773

  10. Misoprostol modulates cytokine expression through a cAMP pathway: Potential therapeutic implication for liver disease.

    PubMed

    Gobejishvili, Leila; Ghare, Smita; Khan, Rehan; Cambon, Alexander; Barker, David F; Barve, Shirish; McClain, Craig; Hill, Daniell

    2015-12-01

    Dysregulated cytokine metabolism plays a critical role in the pathogenesis of many forms of liver disease, including alcoholic and non-alcoholic liver disease. In this study we examined the efficacy of Misoprostol in modulating LPS-inducible TNFα and IL-10 expression in healthy human subjects and evaluated molecular mechanisms for Misoprostol modulation of cytokines in vitro. Healthy subjects were given 14day courses of Misoprostol at doses of 100, 200, and 300μg four times a day, in random order. Baseline and LPS-inducible cytokine levels were examined ex vivo in whole blood at the beginning and the end of the study. Additionally, in vitro studies were performed using primary human PBMCs and the murine macrophage cell line, RAW 264.7, to investigate underlying mechanisms of misoprostol on cytokine production. Administration of Misoprostol reduced LPS inducible TNF production by 29%, while increasing IL-10 production by 79% in human subjects with no significant dose effect on ex vivo cytokine activity; In vitro, the effect of Misoprostol was largely mediated by increased cAMP levels and consequent changes in CRE and NFκB activity, which are critical for regulating IL-10 and TNF expression. Additionally, chromatin immunoprecipitation (ChIP) studies demonstrated that Misoprostol treatment led to changes in transcription factor and RNA Polymerase II binding, resulting in changes in mRNA levels. In summary, Misoprostol was effective at beneficially modulating TNF and IL-10 levels both in vivo and in vitro; these studies suggest a potential rationale for Misoprostol use in ALD, NASH and other liver diseases where inflammation plays an etiologic role.

  11. The human beta-subunit of rod photoreceptor cGMP phosphodiesterase: complete retinal cDNA sequence and evidence for expression in brain.

    PubMed

    Collins, C; Hutchinson, G; Kowbel, D; Riess, O; Weber, B; Hayden, M R

    1992-07-01

    We have identified and sequenced cDNA clones that encode for the human beta-subunit of rod cGMP phosphodiesterase (PDEB). A single 2565-bp open reading frame that codes for an 854-amino-acid protein was identified. The human beta-subunit protein is 90% identical to the bovine beta-subunit and 91% identical to the mouse protein. Northern blot analysis indicates that the gene is expressed as an abundant 3.5-kb transcript in retina and as a rare 2.9-kb transcript in brain. The isolation of cDNAs from human brain cDNA libraries confirms the brain as a site of expression for this gene. The molecular defect underlying retinal degeneration in the rd mouse has been found to be a nonsense mutation in the beta-subunit of the mouse cGMP PDE, resulting in a truncated protein (Pittler et al., 1991b, Proc. Natl. Acad. Sci. USA. 88: 8322-8326). The molecular cloning of the cDNA encoding for the PDEB represents the first step in establishing whether this gene plays a causative role in any one of the several human hereditary retinopathies or, based on its localization to chromosome 4p 16.3, in the pathogenesis of Huntington disease.

  12. Atrazine acts as an endocrine disrupter by inhibiting cAMP-specific phosphodiesterase-4

    SciTech Connect

    Kucka, Marek; Pogrmic-Majkic, Kristina; Fa, Svetlana; Stojilkovic, Stanko S.; Kovacevic, Radmila

    2012-11-15

    Atrazine, one of the most commonly used herbicides worldwide, acts as an endocrine disruptor, but the mechanism of its action has not been characterized. In this study, we show that atrazine rapidly increases cAMP levels in cultured rat pituitary and testicular Leydig cells in a concentration-dependent manner, but less effectively than 3-isobutyl-1-methylxanthine, a competitive non-specific inhibitor of phosphodiesterases (PDEs). In forskolin (an activator of adenylyl cyclase)- and probenecid (an inhibitor of cyclic nucleotide transporters)-treated cells, but not in 3-isobutyl-1-methylxanthine-treated cells, atrazine further increased cAMP levels, indicating that inhibition of PDEs accounts for accumulation of cAMP. In contrast to cAMP, atrazine did not alter cGMP levels, further indicating that it inhibits cAMP-specific PDEs. Atrazine-induced changes in cAMP levels were sufficient to stimulate prolactin release in pituitary cells and androgen production in Leydig cells, indicating that it acts as an endocrine disrupter both in cells that secrete by exocytosis of prestored hormones and in cells that secrete by de novo hormone synthesis. Rolipram abolished the stimulatory effect of atrazine on cAMP release in both cell types, suggesting that it acts as an inhibitor of PDE4s, isoforms whose mRNA transcripts dominate in pituitary and Leydig cells together with mRNA for PDE8A. In contrast, immortalized lacto-somatotrophs showed low expression of these mRNA transcripts and several fold higher cAMP levels compared to normal pituitary cells, and atrazine was unable to further increase cAMP levels. These results indicate that atrazine acts as a general endocrine disrupter by inhibiting cAMP-specific PDE4s. -- Highlights: ► Atrazine stimulates cAMP accumulation in pituitary and Leydig cells. ► Atrazine also stimulates PRL and androgens secretion. ► Stimulatory effects of atrazine were abolished in cells with IBMX-inhibited PDEs. ► Atrazine specificity toward c

  13. Identification and Characterization of a Novel Phosphodiesterase from the Metagenome of an Indian Coalbed

    PubMed Central

    Singh, Durgesh Narain; Gupta, Ankush; Singh, Vijay Shankar; Mishra, Rajeev; Kateriya, Suneel; Tripathi, Anil Kumar

    2015-01-01

    Phosphoesterases are involved in the degradation of organophosphorus compounds. Although phosphomonoesterases and phosphotriesterases have been studied in detail, studies on phosphodiesterases are rather limited. In our search to find novel phosphodiesterases using metagenomic approach, we cloned a gene encoding a putative phosphodiesterase (PdeM) from the metagenome of the formation water collected from an Indian coal bed. Bioinformatic analysis showed that PdeM sequence possessed the characteristic signature motifs of the class III phosphodiesterases and phylogenetic study of PdeM enabled us to identify three distinct subclasses (A, B, and C) within class III phosphodiesterases, PdeM clustering in new subclass IIIB. Bioinformatic, biochemical and biophysical characterization of PdeM further revealed some of the characteristic features of the phosphodiesterases belonging to newly described subclass IIIB. PdeM is a monomer of 29.3 kDa, which exhibits optimum activity at 25°C and pH 8.5, but low affinity for bis(pNPP) as well as pNPPP. The recombinant PdeM possessed phosphodiesterase, phosphonate-ester hydrolase and nuclease activity. It lacked phosphomonoesterase, phosphotriesterase, and RNAse activities. Overexpression of PdeM in E.coli neither affected catabolite respression nor did the recombinant protein hydrolyzed cAMP in vitro, indicating its inability to hydrolyze cAMP. Although Mn2+ was required for the activity of PdeM, but addition of metals (Mn2+ or Fe3+) did not induce oligomerization. Further increase in concentration of Mn2+ upto 3 mM, increased α-helical content as well as the phosphodiesterase activity. Structural comparison of PdeM with its homologs showed that it lacked critical residues required for dimerization, cAMP hydrolysis, and for the high affinity binding of bis(pNPP). PdeM, thus, is a novel representative of new subclass of class III phosphodiesterases. PMID:25658120

  14. Vasopressin V2 receptor mRNA expression and cAMP accumulation in aging rat kidney.

    PubMed

    Klingler, C; Preisser, L; Barrault, M B; Lluel, P; Horgen, L; Teillet, L; Ancellin, N; Corman, B

    1997-06-01

    The ability of the kidney to regulate water balance is impaired with age, although the secretion of vasopressin is maintained in senescent animals. This suggests that the cellular response to antidiuretic hormone is reduced in aging kidney. To test this hypothesis, the relationship between the expression of the vasopressin. V2 receptor mRNA and adenosine 3',5'-cyclic monophosphate (cAMP) accumulation was investigated in the medullary thick ascending limb of Henle's loop (MTAL) of adult and aging rats. Tubular suspensions of MTAL were prepared from 10- and 30-mo-old female WAG/Rij rats. The accumulation of cAMP for maximal concentration of vasopressin was 34% larger in adult than in old animals (9.5 +/- 0.5 pmol/4 min, n = 16, and 7.1 +/- 0.6 pmol/4 min, n = 12, respectively). The concentration of vasopressin corresponding to half-maximal stimulation was similar in the two groups (0.66 +/- 0.20 and 0.52 +/- 0.09 nmol, n = 5, in adult and old animals), indicating comparable sensitivity of the renal cells with age. The age-related impaired response to vasopressin of the V2 receptor was specific for females and was not observed in males. Direct stimulation of adenylyl cyclase by forskolin induced a comparable accumulation of cAMP in adult and senescent rats. The V2 receptor mRNA level in the MTAL was constant between 10 and 30 mo whether the animals were normally hydrated or dehydrated for 2 days. These data indicate that, in MTAL, the age-related impaired cAMP accumulation by vasopressin would be linked to a change either in the translation of V2 mRNA or in posttranslational processing mechanisms or in the coupling between the V2 receptor and adenylyl cyclase. PMID:9227590

  15. Changes in the expression of extracellular regulated kinase (ERK 1/2) in the R6/2 mouse model of Huntington's disease after phosphodiesterase IV inhibition.

    PubMed

    Fusco, Francesca R; Anzilotti, Serenella; Giampà, Carmela; Dato, Clemente; Laurenti, Daunia; Leuti, Alessandro; Colucci D'Amato, Luca; Perrone, Lorena; Bernardi, Giorgio; Melone, Mariarosa A B

    2012-04-01

    The mitogen-activated protein kinases (MAPKs) superfamily comprises three major signaling pathways: the extracellular signal-regulated protein kinases (ERKs), the c-Jun N-terminal kinases or stress-activated protein kinases (JNKs/SAPKs) and the p38 family of kinases. ERK 1/2 signaling has been implicated in a number of neurodegenerative disorders, including Huntington's disease (HD). Phosphorylation patterns of ERK 1/2 and JNK are altered in cell models of HD. In this study, we aimed at studying the correlations between ERK 1/2 and the neuronal vulnerability to HD degeneration in the R6/2 transgenic mouse model of HD. Single and double-label immunofluorescence for phospho-ERK (pERK, the activated form of ERK) and for each of the striatal neuronal markers were employed on perfusion-fixed brain sections from R6/2 and wild-type mice. Moreover, Phosphodiesterase 4 inhibition through rolipram was used to study the effects on pERK expression in the different types of striatal neurons. We completed our study with western blot analysis. Our study shows that pERK levels increase with age in the medium spiny striatal neurons and in the parvalbumin interneurons, and that rolipram counteracts such increase in pERK. Conversely, cholinergic and somatostatinergic interneurons of the striatum contain higher levels of pERK in the R6/2 mice compared to the controls. Rolipram induces an increase in pERK expression in these interneurons. Thus, our study confirms and extends the concept that the expression of phosphorylated ERK 1/2 is related to neuronal vulnerability and is implicated in the pathophysiology of cell death in HD.

  16. Evidence suggesting phosphodiesterase-3B regulation of NPY/AgRP gene expression in mHypoE-46 hypothalamic neurons.

    PubMed

    Anamthathmakula, Prashanth; Sahu, Maitrayee; Sahu, Abhiram

    2015-09-14

    Hypothalamic neurons expressing neuropeptide Y (NPY) and agouti related-protein (AgRP) are critical regulators of feeding behavior and body weight, and transduce the action of many peripheral signals including leptin and insulin. However, intracellular signaling molecules involved in regulating NPY/AgRP neuronal activity are incompletely understood. Since phosphodiesterase-3B (PDE3B) mediates the hypothalamic action of leptin and insulin on feeding, and is expressed in NPY/AgRP neurons, PDE3B could play a significant role in regulating NPY/AgRP neuronal activity. To investigate the direct regulation of NPY/AgRP neuronal activity by PDE3B, we examined the effects of gain-of-function or reduced function of PDE3B on NPY/AgRP gene expression in a clonal hypothalamic neuronal cell line, mHypoE-46, which endogenously express NPY, AgRP and PDE3B. Overexpression of PDE3B in mHypoE-46 cells with transfection of pcDNA-3.1-PDE3B expression plasmid significantly decreased NPY and AgRP mRNA levels and p-CREB levels as compared to the control plasmid. For the PDE3B knockdown study, mHypoE-46 cells transfected with lentiviral PDE3BshRNAmir plasmid or non-silencing lentiviral shRNAmir control plasmid were selected with puromycin, and stably transfected cells were grown in culture for 48h. Results showed that PDE3BshRNAmir mediated knockdown of PDE3B mRNA and protein levels (∼60-70%) caused an increase in both NPY and AgRP gene expression and in p-CREB levels. Together, these results demonstrate a reciprocal change in NPY and AgRP gene expression following overexpression and knockdown of PDE3B, and suggest a significant role for PDE3B in the regulation of NPY/AgRP gene expression in mHypoE-46 hypothalamic neurons.

  17. Crosstalk between Phosphodiesterase 7 and Glycogen Synthase Kinase-3: Two Relevant Therapeutic Targets for Neurological Disorders

    PubMed Central

    2014-01-01

    Chronic neuroinflammation has been increasingly recognized as a primary mechanism underlying acute brain injury and neurodegenerative diseases. Enhanced expression of diverse pro-inflammatory agents in glial cells has been shown to contribute to the cell death that takes place in these disorders. Previous data from our group have shown that different inhibitors of the cyclic adenosine monophosphate (cAMP) specific phosphodiesterase 7 (PDE7) and glycogen synthase kinase-3 (GSK-3) enzymes are potent anti-inflammatory agents in different models of brain injury. In this study, we investigated cross-talk between PDE7 and GSK-3, two relevant therapeutic targets for neurological disorders, using a chemical approach. To this end, we compared specific inhibitors of GSK-3 and PDE7 with dual inhibitors of both enzymes with regard to anti-inflammatory effects in primary cultures of glial cells treated with lipopolysaccharide. Our results show that the GSK-3 inhibitors act exclusively by inhibition of this enzyme. By contrast, PDE7 inhibitors exert their effects via inhibition of PDE7 to increase intracellular cAMP levels but also through indirect inhibition of GSK-3. Activation of protein kinase A by cAMP results in phosphorylation of Ser9 of GSK-3 and subsequent inhibition. Our results indicate that the indirect inhibition of GSK-3 by PDE7 inhibitors is an important mechanism that should be considered in the future development of pharmacological treatments. PMID:24437940

  18. Expression, immunolocalization, and serological reactivity of a novel sphingomyelin phosphodiesterase-like protein, an excretory/secretory antigen from Clonorchis sinensis.

    PubMed

    Huang, Yanwei; Zheng, Youwei; Li, Yuzhe; Yang, Mei; Li, Ting; Zeng, Suxiang; Yu, Xinbing; Huang, Huaiqiu; Hu, Xuchu

    2013-06-01

    Clonorchiasis, caused by Clonorchis sinensis infection, is a zoonotic parasitic disease of hepatobiliary system in which the proteins released by adult are major pathogenetic factors. In this study, we first characterized a putative sphingomyelin phosphodiesterase (CsSMPase) A-like secretory protein, which was highly expressed in the adult worm. The full-length gene was cloned. The putative protein is of relatively low homology comparing with SMPase from other species, and of rich T cell and B cell epitopes, suggesting that it is an antigen of strong antigenicity. The complete coding sequence of the gene was expressed in the Escherichia coli. The recombinant CsSMPase (rCsSMPase) can be recognized by C. sinensis-infected serum, and the protein immunoserum can recognize a specific band in excretory/secretory products (ESPs) of C. sinensis adult by western blotting. Immunolocalization revealed that CsSMPase was not only localized on tegument, ventral sucker of metacercaria, and the intestine of adult but also on the nearby epithelium of bile duct of the infected Sprague-Dawley rats, implying that CsSMPase was mainly secreted and excreted through adult intestine and directly interacted with bile duct epithelium. Although immunized rats evoked high level antibody response, the antigen level was low in clonorchiasis patients. And the sensitivity and specificity of rCsSMPase were 50.0 % (12/24) and 88.4 % (61/69), in sera IgG-ELISA, respectively. It is likely due to the fact that CsSMPase binding to the plasma membrane of biliary epithelium decreases the antigen immune stimulation.

  19. UCR1C is a novel activator of phosphodiesterase 4 (PDE4) long isoforms and attenuates cardiomyocyte hypertrophy

    PubMed Central

    Wang, Li; Burmeister, Brian T.; Johnson, Keven R.; Baillie, George S.; Karginov, Andrei V.; Skidgel, Randal A.; O’Bryan, John P.; Carnegie, Graeme K.

    2015-01-01

    Hypertrophy increases the risk of heart failure and arrhythmia. Prevention or reversal of the maladaptive hypertrophic phenotype has thus been proposed to treat heart failure. Chronic β-adrenergic receptor (β-AR) stimulation induces cardiomyocyte hypertrophy by elevating 3′, 5′-cyclic adenosine monophosphate (cAMP) levels and activating downstream effectors such protein kinase A (PKA). Conversely, hydrolysis of cAMP by phosphodiesterases (PDEs) spatiotemporally restricts cAMP signaling. Here, we demonstrate that PDE4, but not PDE3, is critical in regulating cardiomyocyte hypertrophy, and may represent a potential target for preventing maladaptive hypertrophy. We identify a sequence within the upstream conserved region 1 of PDE4D, termed UCR1C, as a novel activator of PDE4 long isoforms. UCR1C activates PDE4 in complex with A-Kinase anchoring protein (AKAP)-Lbc resulting in decreased PKA signaling facilitated by AKAP-Lbc. Expression of UCR1C in cardiomyocytes inhibits hypertrophy in response to chronic β-AR stimulation. This effect is partially due to inhibition of nuclear PKA activity, which decreases phosphorylation of the transcription factor cAMP response element-binding protein (CREB). In conclusion, PDE4 activation by UCR1C attenuates cardiomyocyte hypertrophy by specifically inhibiting nuclear PKA activity. PMID:25683917

  20. Selective inhibition of two soluble adenosine cyclic 3',5'-phosphate phosphodiesterases partially purified from calf liver.

    PubMed

    Yamamoto, T; Lieberman, F; Osborne, J C; Manganiello, V C; Vaughan, M; Hidaka, H

    1984-02-14

    "Low Km" cAMP phosphodiesterase and cGMP-stimulated cyclic nucleotide phosphodiesterase activities were partially purified from calf liver supernatant by chromatography on DEAE-cellulose and DEAE-Sepharose and ammonium sulfate precipitation. The low Km phosphodiesterase was not retained on N6-H2N(CH2)2-cAMP-agarose and could be separated from the cGMP-stimulated phosphodiesterase which was absorbed by this matrix. From the proteins that did not bind, two distinct low Km cAMP phosphodiesterases were separated on Ultrogel AcA 34. One form (fraction C) hydrolyzed cAMP with an apparent Km of approximately 0.5 microM and was very sensitive to inhibition by cGMP. Lineweaver-Burk plots of cAMP hydrolysis by a second form (fraction B) were nonlinear, with an apparent low Km component of approximately 2 microM. This form was rather insensitive to inhibition by cGMP. With both fractions, hydrolysis of cAMP relative to cGMP was much greater at low (approximately 1 microM) than at high (approximately 100 microM) substrate concentrations. Maximal velocities for cAMP and cGMP were similar. From sedimentation equilibrium, the apparent weight-average molecular weight of fraction B was estimated as 174000, and that of fraction C was 85000. Another fraction (A) of cAMP phosphodiesterase eluted at the void volume of the AcA 34 column. On the basis of the relative affinities for cAMP and cGMP and inhibition by cGMP, fraction A is most likely an aggregated form of fraction B. No apparent interconversion of fractions A, B, or C was observed on high-performance liquid chromatography.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:6324851

  1. Mechanically induced c-fos expression is mediated by cAMP in MC3T3-E1 osteoblasts

    NASA Technical Reports Server (NTRS)

    Fitzgerald, J.; Hughes-Fulford, M.

    1999-01-01

    In serum-deprived MC3T3-E1 osteoblasts, mechanical stimulation caused by mild (287 x g) centrifugation induced a 10-fold increase in mRNA levels of the proto-oncogene, c-fos. Induction of c-fos was abolished by the cAMP-dependent protein kinase inhibitor H-89, suggesting that the transient c-fos mRNA increase is mediated by cAMP. Down-regulation of protein kinase C (PKC) activity by chronic TPA treatment failed to significantly reduce c-fos induction, suggesting that TPA-sensitive isoforms of PKC are not responsible for c-fos up-regulation. In addition, 287 x g centrifugation increased intracellular prostaglandin E2 (PGE2) levels 2.8-fold (P<0. 005). Since we have previously shown that prostaglandin E2 (PGE2) can induce c-fos expression via a cAMP-mediated mechanism, we asked whether the increase in c-fos mRNA was due to centrifugation-induced PGE2 release. Pretreatment with the cyclooxygenase inhibitors indomethacin and flurbiprofen did not hinder the early induction of c-fos by mechanical stimulation. We conclude that c-fos expression induced by mild mechanical loading is dependent primarily on cAMP, not PKC, and initial induction of c-fos is not necessarily dependent on the action of newly synthesized PGE2.

  2. Phosphodiesterase 8B and cyclic AMP signaling in the adrenal cortex.

    PubMed

    Leal, Leticia Ferro; Szarek, Eva; Faucz, Fabio; Stratakis, Constantine A

    2015-09-01

    Bilateral adrenocortical hyperplasia (BAH) in humans and mice has been recently linked to phosphodiesterase (PDE) 8B (PDE8B) and 11 (PDE11A) defects. These findings have followed the discovery that defects of primary genes of the cyclic monophosphatase (cAMP) signaling pathway, such as guanine nucleotide binding alpha subunit and PRKAR1A, are involved in the pathogenesis of BAH in humans; complete absence of Prkar1a in the adrenal cortex of mice also led to pathology that mimicked the human disease. Here, we review the most recent findings in human and mouse studies on PDE8B, a cAMP-specific PDE that appears to be highly expressed in the adrenal cortex and whose deficiency may underlie predisposition to BAH and possibly other human diseases. PMID:25971952

  3. Cyclic AMP Control Measured in Two Compartments in HEK293 Cells: Phosphodiesterase KM Is More Important than Phosphodiesterase Localization

    PubMed Central

    Matthiesen, Karina; Nielsen, Jacob

    2011-01-01

    The intracellular second messenger cyclic AMP (cAMP) is degraded by phosphodiesterases (PDE). The knowledge of individual families and subtypes of PDEs is considerable, but how the different PDEs collaborate in the cell to control a cAMP signal is still not fully understood. In order to investigate compartmentalized cAMP signaling, we have generated a membrane-targeted variant of the cAMP Bioluminiscence Resonance Energy Transfer (BRET) sensor CAMYEL and have compared intracellular cAMP measurements with it to measurements with the cytosolic BRET sensor CAMYEL in HEK293 cells. With these sensors we observed a slightly higher cAMP response to adenylyl cyclase activation at the plasma membrane compared to the cytosol, which is in accordance with earlier results from Fluorescence Resonance Energy Transfer (FRET) sensors. We have analyzed PDE activity in fractionated lysates from HEK293 cells using selective PDE inhibitors and have identified PDE3 and PDE10A as the major membrane-bound PDEs and PDE4 as the major cytosolic PDE. Inhibition of membrane-bound or cytosolic PDEs can potentiate the cAMP response to adenylyl cyclase activation, but we see no significant difference between the potentiation of the cAMP response at the plasma membrane and in cytosol when membrane-bound and cytosolic PDEs are inhibited. When different levels of stimulation were tested, we found that PDEs 3 and 10 are mainly responsible for cAMP degradation at low intracellular cAMP concentrations, whereas PDE4 is more important for control of cAMP at higher concentrations. PMID:21931705

  4. Identification and characterization of DdPDE3, a cGMP-selective phosphodiesterase from Dictyostelium.

    PubMed Central

    Kuwayama, H; Snippe, H; Derks, M; Roelofs, J; Van Haastert, P J

    2001-01-01

    In Dictyostelium cAMP and cGMP have important functions as first and second messengers in chemotaxis and development. Two cyclic-nucleotide phosphodiesterases (DdPDE 1 and 2) have been identified previously, an extracellular dual-specificity enzyme and an intracellular cAMP-specific enzyme (encoded by the psdA and regA genes respectively). Biochemical data suggest the presence of at least one cGMP-specific phosphodiesterase (PDE) that is activated by cGMP. Using bioinformatics we identified a partial sequence in the Dictyostelium expressed sequence tag database that shows a high degree of amino acid sequence identity with mammalian PDE catalytic domains (DdPDE3). The deduced amino acid sequence of a full-length DdPDE3 cDNA isolated in this study predicts a 60 kDa protein with a 300-residue C-terminal PDE catalytic domain, which is preceded by approx. 200 residues rich in asparagine and glutamine residues. Expression of the DdPDE3 catalytic domain in Escherichia coli shows that the enzyme has Michaelis-Menten kinetics and a higher affinity for cGMP (K(m)=0.22 microM) than for cAMP (K(m)=145 microM); cGMP does not stimulate enzyme activity. The enzyme requires bivalent cations for activity; Mn(2+) is preferred to Mg(2+), whereas Ca(2+) yields no activity. DdPDE3 is inhibited by 3-isobutyl-1-methylxanthine with an IC(50) of approx. 60 microM. Overexpression of the DdPDE3 catalytic domain in Dictyostelium confirms these kinetic properties without indications of its activation by cGMP. The properties of DdPDE3 resemble those of mammalian PDE9, which also shows the highest sequence similarity within the catalytic domains. DdPDE3 is the first cGMP-selective PDE identified in lower eukaryotes. PMID:11171061

  5. Phosphodiesterase sequence variants may predispose to prostate cancer

    PubMed Central

    de Alexandre, Rodrigo Bertollo; Horvath, Anelia; Szarek, Eva; Manning, Allison D.; Leal, Leticia Ferro; Kardauke, Fabio; Epstein, Jonathan A.; Carraro, Dirce Maria; Soares, Fernando Augusto; Apanasovich, Tatiyana; Stratakis, Constantine A.; Faucz, Fabio Rueda

    2015-01-01

    We hypothesized that mutations that inactivate phosphodiesterase (PDE) activity and lead to increased cyclic AMP (cAMP) and cyclic GMP (cGMP) levels may be associated with prostate cancer (PCa). We sequenced the entire PDE coding sequences in the DNA of 16 biopsy samples from PCa patients. Novel mutations were confirmed in the somatic or germline state by Sanger sequencing. Data were then compared to the 1000 Genome Project. PDE, CREB and pCREB protein expression was also studied in all samples, in both normal and abnormal tissue, by immunofluorescence. We identified 3 previously described PDE sequence variants that were significantly higher in PCa. Four novel sequence variations, one each in the PDE4B, PDE6C, PDE7B and PDE10A genes, respectively, were also found in the PCa samples. Interestingly, PDE10A and PDE4B novel variants that were present in 19% and 6% of the patients, respectively, were found in the tumor tissue only. In patients carrying PDE defects, there was pCREB accumulation (p<0.001), and an increase of the pCREB/CREB ratio (patients 0.97± 0.03; controls 0.52± 0.03; p-value < 0.001) by immunohistochemical analysis. We conclude that PDE sequence variants may play a role in the predisposition and/or progression to PCa at the germline and/or somatic state, respectively. Larger such studies are needed to confirm these findings. PMID:25979379

  6. Cyclic Nucleotide Phosphodiesterases: important signaling modulators and therapeutic targets

    PubMed Central

    Ahmad, Faiyaz; Murata, Taku; Simizu, Kasumi; Degerman, Eva; Maurice, Donald; Manganiello, Vincent

    2014-01-01

    By catalyzing hydrolysis of cAMP and cGMP, cyclic nucleotide phosphodiesterases are critical regulators of their intracellular concentrations and their biological effects. Since these intracellular second messengers control many cellular homeostatic processes, dysregulation of their signals and signaling pathways initiate or modulate pathophysiological pathways related to various disease states, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication, chronic obstructive pulmonary disease, and psoriasis. Alterations in expression of PDEs and PDE-gene mutations (especially mutations in PDE6, PDE8B, PDE11A and PDE4) have been implicated in various diseases and cancer pathologies. PDEs also play important role in formation and function of multi-molecular signaling/regulatory complexes called signalosomes. At specific intracellular locations, individual PDEs, together with pathway-specific signaling molecules, regulators, and effectors, are incorporated into specific signalosomes, where they facilitate and regulate compartmentalization of cyclic nucleotide signaling pathways and specific cellular functions. Currently, only a limited number of PDE inhibitors (PDE3, PDE4, PDE5 inhibitors) are used in clinical practice. Future paths to novel drug discovery include the crystal structure-based design approach, which has resulted in generation of more effective family-selective inhibitors, as well as burgeoning development of strategies to alter compartmentalized cyclic nucleotide signaling pathways by selectively targeting individual PDEs and their signalosome partners. PMID:25056711

  7. Camp Chinkapin.

    ERIC Educational Resources Information Center

    Crook County School District, Prineville, OR.

    The Camp Chinkapin program, begun in 1957-58 as a pilot program for the State of Oregon, provides all sixth grade students in Crook County (Oregon) with a 5-day session in a resident camp setting in the early summer. The book serves as an introduction to and workbook for students attending the Crook County Outdoor Classroom at Suttle Lake. The…

  8. Identification and Localization of the Cyclic Nucleotide Phosphodiesterase 10A in Bovine Testis and Mature Spermatozoa.

    PubMed

    Goupil, Serge; Maréchal, Loïze; El Hajj, Hassan; Tremblay, Marie-Ève; Richard, François J; Leclerc, Pierre

    2016-01-01

    In mammals, adenosine 3', 5'-cyclic monophosphate (cAMP) is known to play highly important roles in sperm motility and acrosomal exocytosis. It is known to act through protein phosphorylation via PRKA and through the activation of guanine nucleotide exchange factors like EPAC. Sperm intracellular cAMP levels depend on the activity of adenylyl cyclases, mostly SACY, though transmembrane-containing adenylyl cyclases are also present, and on the activity of cyclic nucleotide phosphodiesterases (PDE) whose role is to degrade cAMP into 5'-AMP. The PDE superfamily is subdivided into 11 families (PDE1 to 11), which act on either cAMP or cGMP, or on both cAMP and cGMP although with different enzymatic properties. PDE10, which is more effective on cAMP than cGMP, has been known for almost 15 years and is mostly studied in the brain where it is associated with neurological disorders. Although a high level of PDE10A gene expression is observed in the testis, information on the identity of the isoforms or on the cell type that express the PDE10 protein is lacking. The objective of this study was to identify the PDE10A isoforms expressed in the testis and germ cells, and to determine the presence and localization of PDE10A in mature spermatozoa. As a sub-objective, since PDE10A transcript variants were reported strictly through analyses of bovine genomic sequence, we also wanted to determine the nucleotide and amino acid sequences by experimental evidence. Using RT-PCR, 5'- and 3'-RACE approaches we clearly show that PDE10A transcript variants X3 and X5 are expressed in bovine testis as well as in primary spermatocytes and spermatids. We also reveal using a combination of immunological techniques and proteomics analytical tools that the PDE10A isoform X4 is present in the area of the developing acrosome of spermatids and of the acrosome of mature spermatozoa. PMID:27548062

  9. Identification and Localization of the Cyclic Nucleotide Phosphodiesterase 10A in Bovine Testis and Mature Spermatozoa

    PubMed Central

    Goupil, Serge; Maréchal, Loïze; El Hajj, Hassan; Tremblay, Marie-Ève; Richard, François J.

    2016-01-01

    In mammals, adenosine 3’, 5’-cyclic monophosphate (cAMP) is known to play highly important roles in sperm motility and acrosomal exocytosis. It is known to act through protein phosphorylation via PRKA and through the activation of guanine nucleotide exchange factors like EPAC. Sperm intracellular cAMP levels depend on the activity of adenylyl cyclases, mostly SACY, though transmembrane-containing adenylyl cyclases are also present, and on the activity of cyclic nucleotide phosphodiesterases (PDE) whose role is to degrade cAMP into 5’-AMP. The PDE superfamily is subdivided into 11 families (PDE1 to 11), which act on either cAMP or cGMP, or on both cAMP and cGMP although with different enzymatic properties. PDE10, which is more effective on cAMP than cGMP, has been known for almost 15 years and is mostly studied in the brain where it is associated with neurological disorders. Although a high level of PDE10A gene expression is observed in the testis, information on the identity of the isoforms or on the cell type that express the PDE10 protein is lacking. The objective of this study was to identify the PDE10A isoforms expressed in the testis and germ cells, and to determine the presence and localization of PDE10A in mature spermatozoa. As a sub-objective, since PDE10A transcript variants were reported strictly through analyses of bovine genomic sequence, we also wanted to determine the nucleotide and amino acid sequences by experimental evidence. Using RT-PCR, 5’- and 3’-RACE approaches we clearly show that PDE10A transcript variants X3 and X5 are expressed in bovine testis as well as in primary spermatocytes and spermatids. We also reveal using a combination of immunological techniques and proteomics analytical tools that the PDE10A isoform X4 is present in the area of the developing acrosome of spermatids and of the acrosome of mature spermatozoa. PMID:27548062

  10. Constellation of HCN channels and cAMP regulating proteins in dendritic spines of the primate prefrontal cortex: potential substrate for working memory deficits in schizophrenia.

    PubMed

    Paspalas, Constantinos D; Wang, Min; Arnsten, Amy F T

    2013-07-01

    Schizophrenia associates with impaired prefrontal cortical (PFC) function and alterations in cyclic AMP (cAMP) signaling pathways. These include genetic insults to disrupted-in-schizophrenia (DISC1) and phosphodiesterases (PDE4s) regulating cAMP hydrolysis, and increased dopamine D1 receptor (D1R) expression that elevates cAMP. We used immunoelectron microscopy to localize DISC1, PDE4A, PDE4B, and D1R in monkey PFC and to view spatial interactions with hyperpolarization-activated cyclic nucleotide-gated (HCN) channels that gate network inputs when opened by cAMP. Physiological interactions between PDE4s and HCN channels were tested in recordings of PFC neurons in monkeys performing a spatial working memory task. The study reveals a constellation of cAMP-related proteins (DISC1, PDE4A, and D1R) and HCN channels next to excitatory synapses and the spine neck in thin spines of superficial PFC, where working memory microcircuits interconnect and spine loss is most evident in schizophrenia. In contrast, channels in dendrites were distant from synapses and cAMP-related proteins, and were associated with endosomal trafficking. The data suggest that a cAMP signalplex is selectively positioned in the spines to gate PFC pyramidal cell microcircuits. Single-unit recordings confirmed physiological interactions between cAMP and HCN channels, consistent with gating actions. These data may explain why PFC networks are especially vulnerable to genetic insults that dysregulate cAMP signaling.

  11. Phosphodiesterase 7 Inhibition Preserves Dopaminergic Neurons in Cellular and Rodent Models of Parkinson Disease

    PubMed Central

    Morales-Garcia, Jose A.; Redondo, Miriam; Alonso-Gil, Sandra; Gil, Carmen; Perez, Concepción; Martinez, Ana; Santos, Angel; Perez-Castillo, Ana

    2011-01-01

    Background Phosphodiesterase 7 plays a major role in down-regulation of protein kinase A activity by hydrolyzing cAMP in many cell types. This cyclic nucleotide plays a key role in signal transduction in a wide variety of cellular responses. In the brain, cAMP has been implicated in learning, memory processes and other brain functions. Methodology/Principal Findings Here we show a novel function of phosphodiesterase 7 inhibition on nigrostriatal dopaminergic neuronal death. We found that S14, a heterocyclic small molecule inhibitor of phosphodiesterase 7, conferred significant neuronal protection against different insults both in the human dopaminergic cell line SH-SY5Y and in primary rat mesencephalic cultures. S14 treatment also reduced microglial activation, protected dopaminergic neurons and improved motor function in the lipopolysaccharide rat model of Parkinson disease. Finally, S14 neuroprotective effects were reversed by blocking the cAMP signaling pathways that operate through cAMP-dependent protein kinase A. Conclusions/Significance Our findings demonstrate that phosphodiesterase 7 inhibition can protect dopaminergic neurons against different insults, and they provide support for the therapeutic potential of phosphodiesterase 7 inhibitors in the treatment of neurodegenerative disorders, particularly Parkinson disease. PMID:21390306

  12. cAMP stimulation of StAR expression and cholesterol metabolism is modulated by co-expression of labile suppressors of transcription and mRNA turnover.

    PubMed

    Jefcoate, Colin R; Lee, Jinwoo; Cherradi, Nadia; Takemori, Hiroshi; Duan, Haichuan

    2011-04-10

    The steroidogenic acute regulatory (StAR) protein is generated in rodents from 1.6 kb and 3.5 kb mRNA formed by alternative polyadenylation. The zinc finger protein, TIS11B (also Znf36L1), is elevated by cAMP in adrenal cells in parallel with StAR mRNA. TIS11b selectively destabilizes the 3.5 kb mRNA through AU-rich sequences at the end of the 3'UTR. siRNA suppression shows that TIS11b surprisingly increases StAR protein and cholesterol metabolism. StAR transcription is directly activated by PKA phosphorylation. cAMP responsive element binding (CREB) protein 1 phosphorylation is a key step leading to recruitment of the co-activator, CREB binding protein (CBP). A second protein, CREB regulated transcription coactivator (TORC/CRTC), enhances this recruitment, but is inhibited by salt inducible kinase (SIK). Basal StAR transcription is constrained through this phosphorylation of TORC. PKA provides an alternative stimulation by phosphorylating SIK, which prevents TORC inactivation. PKA stimulation of StAR nuclear transcripts substantially precedes TORC recruitment to the StAR promoter, which may, therefore, mediate a later step in mRNA production. Inhibition of SIK by staurosporine elevates StAR transcription and TORC recruitment to maximum levels, but without CREB phosphorylation. TORC suppression by SIK evidently limits basal StAR transcription. Staurosporine and cAMP stimulate synergistically. SIK targets the phosphatase, PP2a (activation), and Type 2 histone de-acetylases (inhibition), which may each contribute to suppression. Staurosporine stimulation through SIK inhibition is repeated in cAMP stimulation of many steroidogenic genes regulated by steroidogenic factor 1 (SF-1) and CREB. TIS11b and SIK may combine to attenuate StAR expression when hormonal stimuli decline.

  13. Inhibition of hormone-sensitive lipase gene expression by cAMP and phorbol esters in 3T3-F442A and BFC-1 adipocytes.

    PubMed

    Plée-Gautier, E; Grober, J; Duplus, E; Langin, D; Forest, C

    1996-09-15

    Hormone-sensitive lipase (HSL) catalyses the rate-limiting step in adipocyte lipolysis. Short-term hormonal regulation of HSL activity is well characterized, whereas little is known about the control of HSL gene expression. We have measured HSL mRNA content of 3T3-F442A and BFC-1 adipocytes in response to the cAMP analogue 8-(4-chlorophenylthio)-cAMP (8-CPT-cAMP) and to the phorbol ester phorbol 12-myristate 13-acetate (PMA) by Northern blot, using a specific mouse cDNA fragment. Treatment of the cells for 12 or 6 h with, respectively, 0.5 mM 8-CPT-cAMP or 1 microM PMA produced a maximal decrease of about 60% in HSL mRNA. These effects were unaffected by the protein-synthesis inhibitor anisomycin, suggesting that cAMP and PMA actions were direct. The reduction in HSL mRNA was accompanied by a reduction in HSL total activity. The intracellular routes that cAMP and PMA follow for inducing such an effect seemed clearly independent. (i) After desensitization of the protein kinase C regulation pathway by a 24 h treatment of the cells with 1 microM PMA, PMA action was abolished whereas cAMP was still fully active. (ii) Treatment with saturating concentrations of both agents produced an additive effect. (iii) The synthetic glucocorticoid dexamethasone had no proper effect on HSL gene expression but potentiated cAMP action without affecting PMA action. cAMP inhibitory action on HSL is unexpected. Indeed, the second messenger of catecholamines is the main activator of HSL by phosphorylation. We envision that a long-term cAMP treatment of adipocytes induces a counter-regulatory process that reduces HSL content and, ultimately, limits fatty acid depletion from stored triacylglycerols.

  14. Low aquaporin-2 levels in polyuric DI +/+ severe mice with constitutively high cAMP-phosphodiesterase activity.

    PubMed

    Frøkiaer, J; Marples, D; Valtin, H; Morris, J F; Knepper, M A; Nielsen, S

    1999-02-01

    In the renal collecting duct, vasopressin acutely activates cAMP production, resulting in trafficking of aquaporin-2 water channels (AQP2) to the apical plasma membrane, thereby increasing water permeability. This acute response is modulated by long-term changes in AQP2 expression. Recently, a cAMP-responsive element has been identified in the AQP2 gene, raising the possibility that changes in cAMP levels may control AQP2 expression. To investigate this possibility, we determined AQP2 protein levels in a strain of mice, DI +/+ severe (DI), which have genetically high levels of cAMP-phosphodiesterase activity, and hence low cellular cAMP levels, and severe polyuria. Semiquantitative immunoblotting of membrane fractions prepared from whole kidneys revealed that AQP2 levels in DI mice were only 26 +/- 7% (+/-SE) of those in control mice (n = 10, P < 0.01). In addition, semiquantitative Northern blotting revealed a significantly lower AQP2 mRNA expression in kidneys from DI mice compared with control mice (43 +/- 6% vs. 100 +/- 10%; n = 6 in each group, P < 0.05). AQP3 levels were also reduced. The mice were polyuric and urine osmolalities were accordingly substantially lower in the DI mice than in controls (496 +/- 53 vs. 1,696 +/- 105 mosmol/kgH2O, respectively). Moreover, there was a linear correlation between urine osmolalities and AQP2 levels (P < 0.05). Immunoelectron microscopy confirmed the markedly lower expression of AQP2 in collecting duct principal cells in kidneys of DI mice and, furthermore, demonstrated that AQP2 was almost completely absent from the apical plasma membrane. Thus expression of AQP2 and AQP2 trafficking were severely impaired in DI mice. These results are consistent with the view that in vivo regulation of AQP2 expression by vasopressin is mediated by cAMP. PMID:9950948

  15. Steroid hormone modulation of cAMP production in response to beta adrenergic receptor stimulation in genital tract myocytes.

    PubMed

    DiGiovanni, L; Austin, R; Phillippe, M

    1992-01-01

    beta-Adrenergic receptor stimulation results in smooth muscle relaxation through activation of adenylyl cyclase and subsequent cyclic AMP (cAMP) production. The present study was performed to evaluate the effects of steroid hormones (i.e. testosterone and hydrocortisone) on beta 2-adrenergic receptors and their signal transduction in the DDT1 MF-2 genital tract myocyte. Radioligand binding studies demonstrated that these two steroid hormones produced a 70 to 80% increase in the density of beta 2-adrenergic receptors in these myocytes. Stimulation of the beta 2-adrenergic receptors with isoproterenol resulted in a significant increase of cAMP in control myocytes; cells treated with testosterone for 24 h demonstrated a comparable response to isoproterenol, whereas hydrocortisone for 24 h resulted in a 50% greater cAMP response. In contrast to the response at 24 h, stimulation of myocytes after testosterone treatment for 48 h resulted in a cAMP response comparable to that seen in response to hydrocortisone at 24 h. Studies performed using theophylline demonstrated similar cAMP responses at 24 h between the control and testosterone-treated myocytes, thereby ruling out the possibility that the delayed increase of the cAMP response after testosterone was caused by stimulation of phosphodiesterase. Direct stimulation with forskolin resulted in greater cAMP production in the testosterone-treated myocytes compared to controls, thereby refuting the possibility that testosterone directly suppresses adenylyl cyclase activity at 24 h. These findings suggest that although both testosterone and hydrocortisone produce a twofold increase in beta 2-adrenergic receptor density in the DDT1 myocytes, beta 2-adrenergic receptors expressed in response to hydrocortisone appear functional at 24 h resulting in increased cAMP production, whereas those expressed in response to testosterone require 48 h to demonstrate increased functional activity.

  16. The ever unfolding story of cAMP signaling in trypanosomatids: vive la difference!

    PubMed Central

    Tagoe, Daniel N. A.; Kalejaiye, Titilola D.; de Koning, Harry P.

    2015-01-01

    Kinetoplastids are unicellular, eukaryotic, flagellated protozoans containing the eponymous kinetoplast. Within this order, the family of trypanosomatids are responsible for some of the most serious human diseases, including Chagas disease (Trypanosoma cruzi), sleeping sickness (Trypanosoma brucei spp.), and leishmaniasis (Leishmania spp). Although cAMP is produced during the life cycle stages of these parasites, its signaling pathways are very different from those of mammals. The absence of G-protein-coupled receptors, the presence of structurally different adenylyl cyclases, the paucity of known cAMP effector proteins and the stringent need for regulation of cAMP in the small kinetoplastid cells all suggest a significantly different biochemical pathway and likely cell biology. However, each of the main kinetoplastid parasites express four class 1-type cyclic nucleotide-specific phosphodiesterases (PDEA-D), which have highly similar catalytic domains to that of human PDEs. To date, only TbrPDEB, expressed as two slightly different isoforms TbrPDEB1 and B2, has been found to be essential when ablated. Although the genomes contain reasonably well conserved genes for catalytic and regulatory domains of protein kinase A, these have been shown to have varied structural and functional roles in the different species. Recent discovery of a role of cAMP/AMP metabolism in a quorum-sensing signaling pathway in T. brucei, and the identification of downstream cAMP Response Proteins (CARPs) whose expression levels correlate with sensitivity to PDE inhibitors, suggests a complex signaling cascade. The interplay between the roles of these novel CARPs and the quorum-sensing signaling pathway on cell division and differentiation makes for intriguing cell biology and a new paradigm in cAMP signal transduction, as well as potential targets for trypanosomatid-specific cAMP pathway-based therapeutics. PMID:26441645

  17. Effect of cAMP signaling on expression of glucocorticoid receptor, Bim and Bad in glucocorticoid-sensitive and resistant leukemic and multiple myeloma cells.

    PubMed

    Dong, Hongli; Carlton, Michael E; Lerner, Adam; Epstein, Paul M

    2015-01-01

    Stimulation of cAMP signaling induces apoptosis in glucocorticoid-sensitive and resistant CEM leukemic and MM.1 multiple myeloma cell lines, and this effect is enhanced by dexamethasone in both glucocorticoid-sensitive cell types and in glucocorticoid-resistant CEM cells. Expression of the mRNA for the glucocorticoid receptor alpha (GR) promoters 1A3, 1B and 1C, expression of mRNA and protein for GR, and the BH3-only proapoptotic proteins, Bim and Bad, and the phosphorylation state of Bad were examined following stimulation of the cAMP and glucocorticoid signaling pathways. Expression levels of GR promoters were increased by cAMP and glucocorticoid signaling, but GR protein expression was little changed in CEM and decreased in MM.1 cells. Stimulation of these two signaling pathways induced Bim in CEM cells, induced Bad in MM.1 cells, and activated Bad, as indicated by its dephosphorylation on ser112, in both cell types. This study shows that leukemic and multiple myeloma cells, including those resistant to glucocorticoids, can be induced to undergo apoptosis by stimulating the cAMP signaling pathway, with enhancement by glucocorticoids, and the mechanism by which this occurs may be related to changes in Bim and Bad expression, and in all cases, to activation of Bad.

  18. Relationship between Platelet PPARs, cAMP Levels, and P-Selectin Expression: Antiplatelet Activity of Natural Products.

    PubMed

    Fuentes, Eduardo; Palomo, Iván

    2013-01-01

    Platelets are no longer considered simply as cells participating in thrombosis. In atherosclerosis, platelets are regulators of multiple processes, with the recruitment of inflammatory cells towards the lesion sites, inflammatory mediators release, and regulation of endothelial function. The antiplatelet therapy has been used for a long time in an effort to prevent and treat cardiovascular diseases. However, limited efficacy in some patients, drug resistance, and side effects are limitations of current antiplatelet therapy. In this context, a large number of natural products (polyphenols, terpenoids, alkaloids, and fatty acids) have been reported with antiplatelet activity. In this sense, the present paper describes mechanisms of antiplatelet action of natural products on platelet P-selectin expression through cAMP levels and its role as peroxisome proliferator-activated receptors agonists.

  19. Relationship between Platelet PPARs, cAMP Levels, and P-Selectin Expression: Antiplatelet Activity of Natural Products

    PubMed Central

    Fuentes, Eduardo; Palomo, Iván

    2013-01-01

    Platelets are no longer considered simply as cells participating in thrombosis. In atherosclerosis, platelets are regulators of multiple processes, with the recruitment of inflammatory cells towards the lesion sites, inflammatory mediators release, and regulation of endothelial function. The antiplatelet therapy has been used for a long time in an effort to prevent and treat cardiovascular diseases. However, limited efficacy in some patients, drug resistance, and side effects are limitations of current antiplatelet therapy. In this context, a large number of natural products (polyphenols, terpenoids, alkaloids, and fatty acids) have been reported with antiplatelet activity. In this sense, the present paper describes mechanisms of antiplatelet action of natural products on platelet P-selectin expression through cAMP levels and its role as peroxisome proliferator-activated receptors agonists. PMID:24324520

  20. Characterization of CHO-K1 cells stably expressing PDE-IV enzymes. Whole-cell cAMP determinations vs broken-cell enzymatic assays.

    PubMed

    Pon, D J; Plant, M; Tkach, J; Boulet, L; Muise, E; Allen, R A; Rodger, I W

    1998-01-01

    A CHO-K1 cell line stably expressing a recombinant full-length human PDE-IVa (HSPDE4A4B) enzyme was established under hygromycin B selection. Full-length expression of the protein was determined by Western blot analysis, which revealed the presence of a 125-kDa immunoreactive band using rabbit anti-PDE-IVa antibodies. The potency of inhibitor compounds was examined by their ability to increase cAMP in the whole-cell, and by their ability to inhibit cAMP hydrolysis in a 100,000 g supernatant (soluble enzyme preparation) obtained from the same cell line. Inhibition of the expressed PDE-IVa activity by selective PDE-IV inhibitors--(R) and (S)-rolipram, RS 14203, and CDP 840--at 100 nM substrate demonstrated that RS 14203 and CDP 840 were the most potent with IC50 = 9 nM, followed by (R)-rolipram (IC50 = 110 nM) and (S)-rolipram (IC50 = 420 nM). The rank order of potencies of the inhibitors in elevating cAMP in the whole-cell assay was quite different from that on the soluble enzyme. RS 14203 was still the most potent compound in elevating cAMP. Moreover, the relative rank order of potencies between CDP 840 and (R)-rolipram changed dramatically, such that (R)-rolipram was more potent than CDP 840 = (S)-rolipram. An apparent 30-fold stereoselectivity between (R)- and (S)-rolipram was also noted. The whole-cell rank order of potencies was also maintained when PKA activity ratios were measured in place of cAMP levels. The ability of the compounds to elevate cAMP in the stable CHO-K1 cells appeared to track better with the potency of the compounds against the high-affinity (Sr) conformer of the enzyme rather than the low-affinity catalytic state.

  1. Astro Camp

    NASA Technical Reports Server (NTRS)

    1995-01-01

    Children who attend NASA's summer Astro Camp at Stennis Space Center enjoy a week of fun-filled activities. Campers learn what it is like to be a couple of inches taller in space and go through an astronaut obstacle course. They also learn how to build their own model rockets, which are launched on the last day of each camp. Campers also attend field trips to places such as the Challenger Learning Center at the Louisiana Arts and Science Center in Baton Rouge. Four weeks of Astro Camp are held during the summer each year-two camps for 8- to 10-year-olds and two for 11- to 13-year olds.

  2. Astro Camp

    NASA Technical Reports Server (NTRS)

    1996-01-01

    Children who attend NASA's summer Astro Camp at Stennis Space Center enjoy a week of fun-filled activities. Campers learn what it feels like to be a couple of inches taller in space and treck through an astronaut obstacle course. They also have the opportunity to build their own model rockets, which are then launched on the last day of each camp. Campers also travel on field trips to places such as the Challenger Learning Center at the Louisiana Arts and Science Center in Baton Rouge. Four weeks of Astro Camp are held each year during the summer-two camps for 8- to 10-year-olds and two for 11- to 13-year olds.

  3. Is zucchini a phosphodiesterase or a ribonuclease?

    PubMed

    Nureki, Osamu

    2014-01-01

    Zucchini (Zuc), a member of the phospholipase D (PLD) superfamily, is essential for the primary PIWI-interacting RNA (piRNA) biogenesis and the suppression of transposon expression, which are crucial for the genome integrity of germline cells. However, it has been ambiguous whether Zuc acts as a phosphodiesterase to produce phosphatidic acid (PA), the lipid signaling molecule, or as a nuclease. The recent three papers describing the crystal structures and functional analyses of fly and mouse Zuc proteins have elucidated that Zuc is a PLD family single-strand ribonuclease, not a phosphodiesterase, and functions in the maturation of primary piRNAs. This review will discuss in detail how the crystal structures clearly predict the function of Zuc, which is subsequently demonstrated by biochemical analysis to conclude the previous controversial discussion on the real function of Zuc.

  4. The Distribution of Phosphodiesterase 2a in the Rat Brain

    PubMed Central

    Stephenson, D. T.; Coskran, T. M.; Kelly, M. P.; Kleiman, R. J.; Morton, D.; O'neill, S. M.; Schmidt, C. J.; Weinberg, R. J.; Menniti, F. S.

    2015-01-01

    The phosphodiesterases (PDEs) are a superfamily of enzymes that regulate spatio-temporal signaling by the intracellular second messengers cAMP and cGMP. PDE2A is expressed at high levels in the mammalian brain. To advance our understanding of the role of this enzyme in regulation of neuronal signaling, we here describe the distribution of PDE2A in the rat brain. PDE2A mRNA was prominently expressed in glutamatergic pyramidal cells in cortex, and in pyramidal and dentate granule cells in the hippocampus. Protein concentrated in the axons and nerve terminals of these neurons; staining was markedly weaker in the cell bodies and proximal dendrites. In addition, in both hippocampus and cortex, small populations of non-pyramidal cells, presumed to be interneurons, were strongly immunoreactive. PDE2A mRNA was expressed in medium spiny neurons in neostriatum. Little immunoreactivity was observed in cell bodies, whereas dense immunoreactivity was found in the axon tracts of these neurons and their terminal regions in globus pallidus and substantia nigra pars reticulata. Immunostaining was dense in the medial habenula, but weak in other diencephalic regions. In midbrain and hindbrain, immunostaining was restricted to discrete regions of the neuropil or clusters of cell bodies. These results suggest that PDE2A may modulate cortical, hippocampal and striatal networks at several levels. Preferential distribution of PDE2A into axons and terminals of the principal neurons suggests roles in regulation of axonal excitability or transmitter release. The enzyme is also in forebrain interneurons, and in mid- and hindbrain neurons that may modulate forebrain networks and circuits. PMID:23000621

  5. Novel reciprocal regulation of cAMP signaling and apoptosis by orphan G-protein-coupled receptor GPRC5A gene expression

    SciTech Connect

    Hirano, Minoru; Zang, Liqing; Oka, Takehiko; Ito, Yoshiyuki; Shimada, Yasuhito; Nishimura, Yuhei; Tanaka, Toshio . E-mail: tanaka@doc.medic.mie-u.ac.jp

    2006-12-08

    GPRC5A is a member of G-protein-coupled receptors, which was originally identified as an all-trans-retinoic acid-induced gene. Although recent studies reported that this gene was highly expressed in the cancer cell lines and that GPRC5A might positively regulate cell proliferation, its mechanism remains unknown. We investigated the upstream and downstream signaling of GPRC5A and its biological function, and found that cAMP signaling is the novel GPRC5A induction pathway. When GPRC5A gene was overexpressed, intracellular cAMP concentration was decreased, and Gs{alpha} gene expression was downregulated. On the other hand, RNA interference of GPRC5A increased mRNA levels of Gs{alpha} and intracellular cAMP, reduced cell number, and induced apoptosis. Conversely, cell number was increased by GPRC5A overexpression. We first report the novel negative feedback model of cAMP signaling through GPRC5A gene expression. This evidence explains one of the mechanisms of the GPRC5A-regulated cell growth in some cancer cell lines.

  6. Characterization of phosphodiesterase 2A in human malignant melanoma PMP cells.

    PubMed

    Morita, Hiroshi; Murata, Taku; Shimizu, Kasumi; Okumura, Kenya; Inui, Madoka; Tagawa, Toshiro

    2013-04-01

    The prognosis for malignant melanoma is poor; therefore, new diagnostic methods and treatment strategies are urgently needed. Phosphodiesterase 2 (PDE2) is one of 21 phosphodiesterases, which are divided into 11 families (PDE1-PDE11). PDE2 hydrolyzes cyclic AMP (cAMP) and cyclic GMP (cGMP), and its binding to cGMP enhances the hydrolysis of cAMP. We previously reported the expression of PDE1, PDE3 and PDE5 in human malignant melanoma cells. However, the expression of PDE2 in these cells has not been investigated. Herein, we examined the expression of PDE2A and its role in human oral malignant melanoma PMP cells. Sequencing of RT-PCR products revealed that PDE2A2 was the only variant expressed in PMP cells. Four point mutations were detected; one missense mutation at nucleotide position 734 (from C to T) resulted in the substitution of threonine with isoleucine at amino acid position 214. The other three were silent mutations. An in vitro migration assay and a terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay revealed that suppressing PDE2 activity with its specific inhibitor, erythro-9-(2-hydroxy-3-nonyl)-adenine (EHNA), had no impact on cell motility or apoptosis. Furthermore, the cytotoxicity of EHNA, assessed using a trypan blue exclusion assay, was negligible. On the other hand, assessment of cell proliferation by BrdU incorporation and cell cycle analysis by flow cytometry revealed that EHNA treatment inhibited DNA synthesis and increased the percentage of G2/M-arrested cells. Furthermore, cyclin A mRNA expression was downregulated, while cyclin E mRNA expression was upregulated in EHNA-treated cells. Our results demonstrated that the PDE2A2 variant carrying point mutations is expressed in PMP cells and may affect cell cycle progression by modulating cyclin A expression. Thus, PDE2A2 is a possible new molecular target for the treatment of malignant melanoma.

  7. Exposure to an Extremely-Low-Frequency Magnetic Field Stimulates Adrenal Steroidogenesis via Inhibition of Phosphodiesterase Activity in a Mouse Adrenal Cell Line

    PubMed Central

    Kitaoka, Kazuyoshi; Kawata, Shiyori; Yoshida, Tomohiro; Kadoriku, Fumiya; Kitamura, Mitsuo

    2016-01-01

    Extremely low-frequency magnetic fields (ELF-MFs) are generated by power lines and household electrical devices. In the last several decades, some evidence has shown an association between ELF-MF exposure and depression and/or anxiety in epidemiological and animal studies. The mechanism underlying ELF-MF-induced depression is considered to involve adrenal steroidogenesis, which is triggered by ELF-MF exposure. However, how ELF-MFs stimulate adrenal steroidogenesis is controversial. In the current study, we investigated the effect of ELF-MF exposure on the mouse adrenal cortex-derived Y-1 cell line and the human adrenal cortex-derived H295R cell line to clarify whether the ELF-MF stimulates adrenal steroidogenesis directly. ELF-MF exposure was found to significantly stimulate adrenal steroidogenesis (p < 0.01–0.05) and the expression of adrenal steroid synthetic enzymes (p < 0.05) in Y-1 cells, but the effect was weak in H295R cells. Y-1 cells exposed to an ELF-MF showed significant decreases in phosphodiesterase activity (p < 0.05) and intracellular Ca2+ concentration (p < 0.01) and significant increases in intracellular cyclic adenosine monophosphate (cAMP) concentration (p < 0.001–0.05) and cAMP response element-binding protein phosphorylation (p < 0.05). The increase in cAMP was not inhibited by treatment with NF449, an inhibitor of the Gs alpha subunit of G protein. Our results suggest that ELF-MF exposure stimulates adrenal steroidogenesis via an increase in intracellular cAMP caused by the inhibition of phosphodiesterase activity in Y-1 cells. The same mechanism may trigger the increase in adrenal steroid secretion in mice observed in our previous study. PMID:27100201

  8. Long-term niacin treatment induces insulin resistance and adrenergic responsiveness in adipocytes by adaptive downregulation of phosphodiesterase 3B.

    PubMed

    Heemskerk, Mattijs M; van den Berg, Sjoerd A A; Pronk, Amanda C M; van Klinken, Jan-Bert; Boon, Mariëtte R; Havekes, Louis M; Rensen, Patrick C N; van Dijk, Ko Willems; van Harmelen, Vanessa

    2014-04-01

    The lipid-lowering effect of niacin has been attributed to the inhibition of cAMP production in adipocytes, thereby inhibiting intracellular lipolysis and release of nonesterified fatty acids (NEFA) to the circulation. However, long-term niacin treatment leads to a normalization of plasma NEFA levels and induces insulin resistance, for which the underlying mechanisms are poorly understood. The current study addressed the effects of long-term niacin treatment on insulin-mediated inhibition of adipocyte lipolysis and focused on the regulation of cAMP levels. APOE*3-Leiden.CETP transgenic mice treated with niacin for 15 wk were subjected to an insulin tolerance test and showed whole body insulin resistance. Similarly, adipocytes isolated from niacin-treated mice were insulin resistant and, interestingly, exhibited an increased response to cAMP stimulation by 8Br-cAMP, β1- and β2-adrenergic stimulation. Gene expression analysis of the insulin and β-adrenergic pathways in adipose tissue indicated that all genes were downregulated, including the gene encoding the cAMP-degrading enzyme phosphodiesterase 3B (PDE3B). In line with this, we showed that insulin induced a lower PDE3B response in adipocytes isolated from niacin-treated mice. Inhibiting PDE3B with cilostazol increased lipolytic responsiveness to cAMP stimulation in adipocytes. These data show that long-term niacin treatment leads to a downregulation of PDE3B in adipocytes, which could explain part of the observed insulin resistance and the increased responsiveness to cAMP stimulation.

  9. Does phosphodiesterase 11A (PDE11A) hold promise as a future therapeutic target?

    PubMed

    Kelly, Michy P

    2015-01-01

    Phosphodiesterase 11A (PDE11A) is the most recently discovered 3', 5'-cyclic nucleotide phosphodiesterase. By breaking down both cAMP and cGMP, PDE11A is a critical regulator of intracellular signaling. To date, PDE11A has been implicated to play a role in tumorigenesis, brain function, and inflammation. Here, we consolidate and, where necessary, reconcile the PDE11A literature to evaluate this enzyme as a potential therapeutic target. We compare the results and methodologies of numerous studies that report conflicting tissue expression profiles for PDE11A. We conclude that PDE11A expression is relatively restricted in the body, with reliable expression reported in tissues such as the brain (particularly the hippocampus), the prostate, and the adrenal gland. Each of the four PDE11A splice variants (PDE11A1-4) appears to exhibit a distinct tissue expression profile and has a unique N-terminal regulatory region, suggesting that each isoform could be individually targeted with a small molecule or biologic. Progress has been made in identifying a tool PDE11A inhibitor as well as an activator; however, the functional effects of these pharmacological tools remain to be determined. Importantly, PDE11A knockout mice do exist and appear healthy into late age, suggesting a potential safety window for targeting this enzyme. Considering the implication of PDE11A in disease-relevant biology, the potential to selectively target specific PDE11A variants, and the possibility of either activating or inhibiting the enzyme, we believe PDE11A holds promise as a potential future therapeutic target.

  10. Null mutations of the Dictyostelium cyclic nucleotide phosphodiesterase gene block chemotactic cell movement in developing aggregates.

    PubMed

    Sucgang, R; Weijer, C J; Siegert, F; Franke, J; Kessin, R H

    1997-12-01

    Extracellular cAMP is a critical messenger in the multicellular development of the cellular slime mold Dictyostelium discoideum. The levels of cAMP are controlled by a cyclic nucleotide phosphodiesterase (PDE) that is secreted by the cells. The PDE gene (pdsA) is controlled by three promoters that permit expression during vegetative growth, during aggregation, and in prestalk cells of the older structures. Targeted disruption of the gene aborts development, and complementation with a modified pdsA restores development. Two distinct promoters must be used for full complementation, and an inhibitory domain of the PDE must be removed. We took advantage of newly isolated PDE-null cells and the natural chimerism of the organism to ask whether the absence of PDE affected individual cell behavior. PDE-null cells aggregated with isogenic wild-type cells in chimeric mixtures, but could not move in a coordinated manner in mounds. The wild-type cells move inward toward the center of the mound, leaving many of the PDE-null cells at the periphery of the aggregate. During the later stages of development, PDE-null cells in the chimera segregate to regions which correspond to the prestalk region and the rear of the slug. Participation in the prespore/spore population returns with the restoration of a modified pdsA to the null cells. PMID:9405107

  11. Intrinsic sex-specific differences in microvascular endothelial cell phosphodiesterases

    PubMed Central

    Bingaman, Susan; Huxley, Virginia H.

    2010-01-01

    The importance of gonadal hormones in the regulation of vascular function has been documented. An alternate and essential contribution of the sex chromosomes to sex differences in vascular function is poorly understood. We reported previously sex differences in microvessel permeability (Ps) responses to adenosine that were mediated by the cAMP signaling pathway (Wang J, PhD thesis, 2005; Wang J and Huxley V, Proceedings of the VIII World Congress of Microcirculation, 2007; Wang J and Huxley VH, Am J Physiol Heart Circ Physiol 291: H3094–H3105, 2006). The two cyclic nucleotides, cAMP and cGMP, central to the regulation of vascular barrier integrity, are hydrolyzed by phosphodiesterases (PDE). We hypothesized that microvascular endothelial cells (EC) would retain intrinsic and inheritable sexually dimorphic genes with respect to the PDEs modulating EC barrier function. Primary cultured microvascular EC from skeletal muscles isolated from male and female rats, respectively, were used. SRY (a sex-determining region Y gene) mRNA expression was observed exclusively in male, not female, cells. The predominant isoform among PDE1–5, present in both XY and XX EC, was PDE4. Expression mRNA levels of PDE1A (male > female) and PDE3B (male < female) were sex dependent; PDE2A, PDE4D, and PDE5A were sex independent. Barrier function, Ps, was determined from measures of albumin flux across confluent primary cultured microvessel XY and XX EC monolayers. Consistent with intact in situ microvessels, basal monolayer Ps did not differ between XY (1.7 ± 0.2 × 10−6 cm/s; n = 8) and XX (1.8 ± 0.1 × 10−6 cm/s; n = 10) EC. Cilostazol, a PDE3 inhibitor, reduced (11%, P < 0.05) Ps in XX, not XY, cells. These findings demonstrate the presence and maintenance of intrinsic sex-related differences in gene expression and cellular phenotype by microvascular EC in a gonadal-hormone-free environment. Furthermore, intrinsic cell-sex likely contributes significantly to sexual dimorphism in

  12. Phosphodiesterase-5 inhibitors.

    PubMed

    Cockrill, Barbara A; Waxman, Aaron B

    2013-01-01

    Nitric oxide (NO) signaling plays a key role in modulating vascular tone and remodeling in the pulmonary circulation. The guanylate cyclase/cyclic guanylate monophosphate-signaling pathway primarily mediates nitric oxide signaling. This pathway is critical in normal regulation of the pulmonary vasculature, and is an important target for therapy in patients with pulmonary hypertension. In the pulmonary vasculature, degradation of cGMP is primarily regulated by PDE-5, and inhibition of this enzyme has important effects on pulmonary vasculature smooth muscle tone. Large randomized placebo-controlled trials of PDE-5 inhibitors demonstrated improved exercise capacity, hemodynamics and quality of life in adult patients with PAH. This chapter will discuss the mechanisms of NO signaling in the vasculature, characteristics of the PDE5-inhibitors approved for treatment of PH, and review available data on the use of phosphodiesterase inhibitors in PH. PMID:24092343

  13. [Hemophilia camps.

    PubMed

    Juárez-Sierra, Julieta; Del Pilar Torres-Arreola, Laura; Marín-Palomares, Teresa; Dueñas-González, María Teresa; Monteros-Rincón, Martha Patricia; Osorio-Guzmán, Maricela

    2013-01-01

    We reported the experience of hemophilia camps which was accomplished with patients from hospitals of the Instituto Mexicano del Seguro Social. The aim was to prepare the families and patients regarding the disease treatment, in order to promote the self sufficiency and to know the impact of the program on the course of the disease. Surveys were applied about treatment items and personal opinions were collected. The results of the national hemophilia camp were: group of 56 patients, average 14 years, 2 % women, 51 % severe hemophilia and 43 % had hemophilic brothers. Benefits: patients increased their knowledge about earlier bleeding identification and the self-infusion method; they became aware on their responsibility in self care, timely treatment and duties at home. Hemophilia camps with patients are an option for attitude change before disease complications. Social network creation and the increase in self-sufficiency are other benefits.

  14. [Hemophilia camps.

    PubMed

    Juárez-Sierra, Julieta; Del Pilar Torres-Arreola, Laura; Marín-Palomares, Teresa; Dueñas-González, María Teresa; Monteros-Rincón, Martha Patricia; Osorio-Guzmán, Maricela

    2013-01-01

    We reported the experience of hemophilia camps which was accomplished with patients from hospitals of the Instituto Mexicano del Seguro Social. The aim was to prepare the families and patients regarding the disease treatment, in order to promote the self sufficiency and to know the impact of the program on the course of the disease. Surveys were applied about treatment items and personal opinions were collected. The results of the national hemophilia camp were: group of 56 patients, average 14 years, 2 % women, 51 % severe hemophilia and 43 % had hemophilic brothers. Benefits: patients increased their knowledge about earlier bleeding identification and the self-infusion method; they became aware on their responsibility in self care, timely treatment and duties at home. Hemophilia camps with patients are an option for attitude change before disease complications. Social network creation and the increase in self-sufficiency are other benefits. PMID:24290020

  15. Dual specificity and novel structural folding of yeast phosphodiesterase-1 for hydrolysis of second messengers cyclic adenosine and guanosine 3',5'-Monophosphate

    DOE PAGES

    Tian, Yuanyuan; Cui, Wenjun; Huang, Manna; Robinson, Howard; Wan, Yiqian; Wang, Yousheng; Ke, Hengming

    2014-08-05

    Cyclic nucleotide phosphodiesterases (PDEs) decompose second messengers cAMP and cGMP that play critical roles in many physiological processes. PDE1 of Saccharomyces cerevisiae has been subcloned and expressed in Escherichia coli. Recombinant yPDE1 has a KM of 110 μM and a kcat of 16.9 s⁻¹ for cAMP and a KM of 105 μM and a kcat of 11.8 s₅⁻¹ for cGMP. Thus, the specificity constant (kcat/KMcAMP)/(kcat/KMcGMP) of 1.4 indicates a dual specificity of yPDE1 for hydrolysis of both cAMP and cGMP. The crystal structures of unliganded yPDE1 and its complex with GMP at 1.31 Å resolution reveal a new structural foldingmore » that is different from those of human PDEs but is partially similar to that of some other metalloenzymes such as metallo-β-lactamase. In spite of their different structures and divalent metals, yPDE1 and human PDEs may share a common mechanism for hydrolysis of cAMP and cGMP.« less

  16. Schwann Cells Metabolize Extracellular 2',3'-cAMP to 2'-AMP.

    PubMed

    Verrier, Jonathan D; Kochanek, Patrick M; Jackson, Edwin K

    2015-08-01

    The 3',5'-cAMP-adenosine pathway (3',5'-cAMP→5'-AMP→adenosine) and the 2',3'-cAMP-adenosine pathway (2',3'-cAMP→2'-AMP/3'-AMP→adenosine) are active in the brain. Oligodendrocytes participate in the brain 2',3'-cAMP-adenosine pathway via their robust expression of 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase; converts 2',3'-cAMP to 2'-AMP). Because Schwann cells also express CNPase, it is conceivable that the 2',3'-cAMP-adenosine pathway exists in the peripheral nervous system. To test this and to compare the 2',3'-cAMP-adenosine pathway to the 3',5'-cAMP-adenosine pathway in Schwann cells, we examined the metabolism of 2',3'-cAMP, 2'-AMP, 3'-AMP, 3',5'-cAMP, and 5'-AMP in primary rat Schwann cells in culture. Addition of 2',3'-cAMP (3, 10, and 30 µM) to Schwann cells increased levels of 2'-AMP in the medium from 0.006 ± 0.002 to 21 ± 2, 70 ± 3, and 187 ± 10 nM/µg protein, respectively; in contrast, Schwann cells had little ability to convert 2',3'-cAMP to 3'-AMP or 3',5'-cAMP to either 3'-AMP or 5'-AMP. Although Schwann cells slightly converted 2',3'-cAMP and 2'-AMP to adenosine, they did so at very modest rates (e.g., 5- and 3-fold, respectively, more slowly compared with our previously reported studies in oligodendrocytes). Using transected myelinated rat sciatic nerves in culture medium, we observed a time-related increase in endogenous intracellular 2',3'-cAMP and extracellular 2'-AMP. These findings indicate that Schwann cells do not have a robust 3',5'-cAMP-adenosine pathway but do have a 2',3'-cAMP-adenosine pathway; however, because the pathway mostly involves 2'-AMP formation rather than 3'-AMP, and because the conversion of 2'-AMP to adenosine is slow, metabolism of 2',3'-cAMP mostly results in the accumulation of 2'-AMP. Accumulation of 2'-AMP in peripheral nerves postinjury could have pathophysiological consequences. PMID:25998049

  17. The effect of resveratrol on beta amyloid-induced memory impairment involves inhibition of phosphodiesterase-4 related signaling

    PubMed Central

    Wang, Gang; Chen, Ling; Pan, Xiaoyu; Chen, Jiechun; Wang, Liqun; Wang, Weijie; Cheng, Ruochuan; Wu, Fan; Feng, Xiaoqing; Yu, Yingcong; Zhang, Han-Ting; O'Donnell, James M.; Xu, Ying

    2016-01-01

    Resveratrol, a natural polyphenol found in red wine, has wide spectrum of pharmacological properties including antioxidative and antiaging activities. Beta amyloid peptides (Aβ) are known to involve cognitive impairment, neuroinflammatory and apoptotic processes in Alzheimer's disease (AD). Activation of cAMP and/or cGMP activities can improve memory performance and decrease the neuroinflammation and apoptosis. However, it remains unknown whether the memory enhancing effect of resveratrol on AD associated cognitive disorders is related to the inhibition of phosphodiesterase 4 (PDE4) subtypes and subsequent increases in intracellular cAMP and/or cGMP activities. This study investigated the effect of resveratrol on Aβ1-42-induced cognitive impairment and the participation of PDE4 subtypes related cAMP or cGMP signaling. Mice microinfused with Aβ1-42 into bilateral CA1 subregions displayed learning and memory impairment, as evidenced by reduced memory acquisition and retrieval in the water maze and retention in the passive avoidance tasks; it was also significant that neuroinflammatory and pro-apoptotic factors were increased in Aβ1-42-treated mice. Aβ1-42-treated mice also increased in PDE4A, 4B and 4D expression, and decreased in PKA level. However, PKA inhibitor H89, but not PKG inhibitor KT5823, prevented resveratrol's effects on these parameters. Resveratrol also reversed Aβ1-42-induced decreases in phosphorylated cAMP response-element binding protein (pCREB), brain derived neurotrophic factor (BDNF) and anti-apoptotic factor BCl-2 expression, which were reversed by H89. These findings suggest that resveratrol reversing Aβ-induced learning and memory disorder may involve the regulation of neuronal inflammation and apoptosis via PDE4 subtypes related cAMP-CREB-BDNF signaling. PMID:26980711

  18. The effect of resveratrol on beta amyloid-induced memory impairment involves inhibition of phosphodiesterase-4 related signaling.

    PubMed

    Wang, Gang; Chen, Ling; Pan, Xiaoyu; Chen, Jiechun; Wang, Liqun; Wang, Weijie; Cheng, Ruochuan; Wu, Fan; Feng, Xiaoqing; Yu, Yingcong; Zhang, Han-Ting; O'Donnell, James M; Xu, Ying

    2016-04-01

    Resveratrol, a natural polyphenol found in red wine, has wide spectrum of pharmacological properties including antioxidative and antiaging activities. Beta amyloid peptides (Aβ) are known to involve cognitive impairment, neuroinflammatory and apoptotic processes in Alzheimer's disease (AD). Activation of cAMP and/or cGMP activities can improve memory performance and decrease the neuroinflammation and apoptosis. However, it remains unknown whether the memory enhancing effect of resveratrol on AD associated cognitive disorders is related to the inhibition of phosphodiesterase 4 (PDE4) subtypes and subsequent increases in intracellular cAMP and/or cGMP activities. This study investigated the effect of resveratrol on Aβ1-42-induced cognitive impairment and the participation of PDE4 subtypes related cAMP or cGMP signaling. Mice microinfused with Aβ1-42 into bilateral CA1 subregions displayed learning and memory impairment, as evidenced by reduced memory acquisition and retrieval in the water maze and retention in the passive avoidance tasks; it was also significant that neuroinflammatory and pro-apoptotic factors were increased in Aβ1-42-treated mice. Aβ1-42-treated mice also increased in PDE4A, 4B and 4D expression, and decreased in PKA level. However, PKA inhibitor H89, but not PKG inhibitor KT5823, prevented resveratrol's effects on these parameters. Resveratrol also reversed Aβ1-42-induced decreases in phosphorylated cAMP response-element binding protein (pCREB), brain derived neurotrophic factor (BDNF) and anti-apoptotic factor BCl-2 expression, which were reversed by H89. These findings suggest that resveratrol reversing Aβ-induced learning and memory disorder may involve the regulation of neuronal inflammation and apoptosis via PDE4 subtypes related cAMP-CREB-BDNF signaling.

  19. The effect of resveratrol on beta amyloid-induced memory impairment involves inhibition of phosphodiesterase-4 related signaling.

    PubMed

    Wang, Gang; Chen, Ling; Pan, Xiaoyu; Chen, Jiechun; Wang, Liqun; Wang, Weijie; Cheng, Ruochuan; Wu, Fan; Feng, Xiaoqing; Yu, Yingcong; Zhang, Han-Ting; O'Donnell, James M; Xu, Ying

    2016-04-01

    Resveratrol, a natural polyphenol found in red wine, has wide spectrum of pharmacological properties including antioxidative and antiaging activities. Beta amyloid peptides (Aβ) are known to involve cognitive impairment, neuroinflammatory and apoptotic processes in Alzheimer's disease (AD). Activation of cAMP and/or cGMP activities can improve memory performance and decrease the neuroinflammation and apoptosis. However, it remains unknown whether the memory enhancing effect of resveratrol on AD associated cognitive disorders is related to the inhibition of phosphodiesterase 4 (PDE4) subtypes and subsequent increases in intracellular cAMP and/or cGMP activities. This study investigated the effect of resveratrol on Aβ1-42-induced cognitive impairment and the participation of PDE4 subtypes related cAMP or cGMP signaling. Mice microinfused with Aβ1-42 into bilateral CA1 subregions displayed learning and memory impairment, as evidenced by reduced memory acquisition and retrieval in the water maze and retention in the passive avoidance tasks; it was also significant that neuroinflammatory and pro-apoptotic factors were increased in Aβ1-42-treated mice. Aβ1-42-treated mice also increased in PDE4A, 4B and 4D expression, and decreased in PKA level. However, PKA inhibitor H89, but not PKG inhibitor KT5823, prevented resveratrol's effects on these parameters. Resveratrol also reversed Aβ1-42-induced decreases in phosphorylated cAMP response-element binding protein (pCREB), brain derived neurotrophic factor (BDNF) and anti-apoptotic factor BCl-2 expression, which were reversed by H89. These findings suggest that resveratrol reversing Aβ-induced learning and memory disorder may involve the regulation of neuronal inflammation and apoptosis via PDE4 subtypes related cAMP-CREB-BDNF signaling. PMID:26980711

  20. Expression of orphan G-protein coupled receptor GPR174 in CHO cells induced morphological changes and proliferation delay via increasing intracellular cAMP

    SciTech Connect

    Sugita, Kazuya; Yamamura, Chiaki; Tabata, Ken-ichi; Fujita, Norihisa

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer Expression of GPR174 in CHO cells induces morphological changes and proliferation delay. Black-Right-Pointing-Pointer These are due to increase in intracellular cAMP concentration. Black-Right-Pointing-Pointer Lysophosphatidylserine was identified to stimulate GPR174 leading to activate ACase. Black-Right-Pointing-Pointer The potencies of fatty acid moiety on LysoPS were oleoyl Greater-Than-Or-Slanted-Equal-To stearoyl > palmitoyl. Black-Right-Pointing-Pointer We propose that GPR174 is a lysophosphatidylserine receptor. -- Abstract: We established cell lines that stably express orphan GPCR GPR174 using CHO cells, and studied physiological and pharmacological features of the receptor. GPR174-expressing cells showed cell-cell adhesion with localization of actin filaments to cell membrane, and revealed significant delay of cell proliferation. Since the morphological changes of GPR174-cells were very similar to mock CHO cells treated with cholera toxin, we measured the concentration of intracellular cAMP. The results showed the concentration was significantly elevated in GPR174-cells. By measuring intracellular cAMP concentration in GPR174-cells, we screened lipids and nucleotides to identify ligands for GPR174. We found that lysophosphatidylserine (LysoPS) stimulated increase in intracellular cAMP in a dose-dependent manner. Moreover, phosphorylation of Erk was elevated by LysoPS in GPR174 cells. These LysoPS responses were inhibited by NF449, an inhibitor of G{alpha}{sub s} protein. These results suggested that GPR174 was a putative LysoPS receptor conjugating with G{alpha}{sub s}, and its expression induced morphological changes in CHO cells by constitutively activating adenylyl cycles accompanied with cell conjunctions and delay of proliferation.

  1. Phosphodiesterase 4D Inhibitors Limit Prostate Cancer Growth Potential

    PubMed Central

    Powers, Ginny L.; Hammer, Kimberly D.P.; Domenech, Maribella; Frantskevich, Katsiaryna; Malinowski, Rita L.; Bushman, Wade; Beebe, David J.; Marker, Paul C.

    2014-01-01

    Phosphodiesterase 4D (PDE4D) has recently been implicated as a proliferation-promoting factor in prostate cancer and is over-expressed in human prostate carcinoma. However, the effects of PDE4D inhibition using pharmacological inhibitors have not been examined in prostate cancer. These studies examined the effects of selective PDE4D inhibitors, NVP-ABE171 and cilomilast, as anti-prostate cancer therapies in both in vitro and in vivo models. The effects of PDE4D inhibitors on pathways that are critical in prostate cancer and/or downstream of cyclic AMP (cAMP) were examined. Both NVP-ABE171 and cilomilast decreased cell growth. In vitro, PDE4D inhibitors lead to decreased signaling of the sonic hedgehog (SHH), Androgen Receptor (AR), and MAPK pathways, but growth inhibition was best correlated to the sonic hedgehog pathway. PDE4D inhibition also reduced proliferation of epithelial cells induced by paracrine signaling from co-cultured stromal cells that had activated hedgehog signaling. In addition, PDE4D inhibitors decreased the weight of the prostate in wild-type mice. Prostate cancer xenografts grown in nude mice that were treated with cilomilast or NVP-ABE171 had decreased wet weight and increased apoptosis compared to vehicle treated controls. These studies suggest the pharmacological inhibition of PDE4D using small molecule inhibitors is an effective option for prostate cancer therapy. Implications PDE4D inhibitors decrease the growth of prostate cancer cells in vivo and in vitro, and PDE4D inhibition has therapeutic potential in prostate cancer. PMID:25149359

  2. The Molecular Basis for Different Recognition of Substrates by Phosphodiesterase Families 4 and 10

    SciTech Connect

    Wang,H.; Robinson, H.; Ke, H.

    2007-01-01

    Phosphodiesterases (PDEs) are key enzymes that control the cellular concentrations of the second messengers cAMP and cGMP. The mechanism for selective recognition of substrates cAMP and cGMP by individual PDE families remains a puzzle. To understand the mechanism for substrate recognition by PDE enzymes, the crystal structure of the catalytic domain of an inactive D201N mutant of PDE4D2 in complex with substrate cAMP has been determined at 1.56 Angstroms resolution. The structure shows that Gln369 forms only one hydrogen bond with the adenine of cAMP. This finding provides experimental evidence against the hypothesis of two hydrogen bonds between the invariant glutamine and the substrate cAMP in PDE4, and thus suggests that the widely circulated 'glutamine switch' model is unlikely the mechanism for substrate recognition by PDEs. A structure comparison between PDE4D2-cAMP and PDE10A2-cAMP reveals an anti configuration of cAMP in PDE4D2 but syn in PDE10A2, in addition to different contact patterns of cAMP in these two structures. These observations imply that individual PDE families have their characteristic mechanisms for substrate recognition.

  3. cAMP target sequences enhCRE and CNRE sense low-salt intake to increase human renin gene expression in vivo.

    PubMed

    Desch, Michael; Harlander, Sabine; Neubauer, Björn; Gerl, Melanie; Germain, Stephane; Castrop, Hayo; Todorov, Vladimir T

    2011-05-01

    This study aimed to assess the role of cAMP target sequences enhancer cAMP response element (enhCRE) and cAMP and overlapping negative response element (CNRE) in the control of human renin gene (REN) in vivo. enhCRE and CNRE were silenced by mutations in a 12.2-kb human renin promoter fused to LacZ reporter gene. This construct was used to generate transgenic mice (RENMut-LacZ). The expression of the transgene was correctly targeted to the juxtaglomerular portions of renal afferent arterioles which express endogenous mouse renin. Therefore, enhCRE and CNRE do not seem to be relevant for the control of the cell-specific expression of the human renin gene. The β-adrenoreceptor agonist isoproterenol (10 mg/kg/day, for 2 days) stimulated the endogenous renin, but not the LacZ mRNA expression. Treatment of RENMut-LacZ mice with the angiotensin converting enzyme inhibitor (enalapril 10 mg/kg/day, for 7 days) or their crossing to angiotensin receptor type 1a knockout mice led to increased renin and LacZ mRNA levels. Renin expression was upregulated by low-salt diet (0.03% NaCl, for 10 days) and downregulated by high-salt diet (4% NaCl, for 10 days). In contrast, low-salt diet did not influence, while high-salt diet inhibited the expression of LacZ. In summary, enhCRE and CNRE appear to be necessary for the transactivation of the human renin gene through β-adrenoreceptors and by low-salt diet. Our data also suggest that different intracellular mechanisms mediate the effect of low- and high-salt intake on renin expression in vivo.

  4. Cyclic AMP control measured in two compartments in HEK293 cells: phosphodiesterase K(M) is more important than phosphodiesterase localization.

    PubMed

    Matthiesen, Karina; Nielsen, Jacob

    2011-01-01

    The intracellular second messenger cyclic AMP (cAMP) is degraded by phosphodiesterases (PDE). The knowledge of individual families and subtypes of PDEs is considerable, but how the different PDEs collaborate in the cell to control a cAMP signal is still not fully understood. In order to investigate compartmentalized cAMP signaling, we have generated a membrane-targeted variant of the cAMP Bioluminiscence Resonance Energy Transfer (BRET) sensor CAMYEL and have compared intracellular cAMP measurements with it to measurements with the cytosolic BRET sensor CAMYEL in HEK293 cells. With these sensors we observed a slightly higher cAMP response to adenylyl cyclase activation at the plasma membrane compared to the cytosol, which is in accordance with earlier results from Fluorescence Resonance Energy Transfer (FRET) sensors. We have analyzed PDE activity in fractionated lysates from HEK293 cells using selective PDE inhibitors and have identified PDE3 and PDE10A as the major membrane-bound PDEs and PDE4 as the major cytosolic PDE. Inhibition of membrane-bound or cytosolic PDEs can potentiate the cAMP response to adenylyl cyclase activation, but we see no significant difference between the potentiation of the cAMP response at the plasma membrane and in cytosol when membrane-bound and cytosolic PDEs are inhibited. When different levels of stimulation were tested, we found that PDEs 3 and 10 are mainly responsible for cAMP degradation at low intracellular cAMP concentrations, whereas PDE4 is more important for control of cAMP at higher concentrations.

  5. Structural Insight into Substrate Specificity of Phosphodiesterase 10

    SciTech Connect

    Wang,H.; Liu, Y.; Hou, J.; Zheng, M.; Robinson, H.; Ke, H.

    2007-01-01

    Phosphodiesterases (PDEs) hydrolyze the second messengers cAMP and cGMP. It remains unknown how individual PDE families selectively recognize cAMP and cGMP. This work reports structural studies on substrate specificity. The crystal structures of the catalytic domains of the D674A and D564N mutants of PDE10A2 in complex with cAMP and cGMP reveal that two substrates bind to the active site with the same syn configuration but different orientations and interactions. The products AMP and GMP bind PDE10A2 with the anti configuration and interact with both divalent metals, in contrast to no direct contact of the substrates. The structures suggest that the syn configurations of cAMP and cGMP are the genuine substrates for PDE10 and the specificity is achieved through the different interactions and conformations of the substrates. The PDE10A2 structures also show that the conformation of the invariant glutamine is locked by two hydrogen bonds and is unlikely to switch for substrate recognition. Sequence alignment shows a potential pocket, in which variation of amino acids across PDE families defines the size and shape of the pocket and thus determines the substrate specificity.

  6. Increased expression of adenylylcyclase type VI proportionately increases β-adrenergic receptor-stimulated production of cAMP in neonatal rat cardiac myocytes

    PubMed Central

    Gao, Meihua; Ping, Peipei; Post, Steven; Insel, Paul A.; Tang, Ruoying; Hammond, H. Kirk

    1998-01-01

    Cellular content of cAMP generated by activation of adenylylcyclase (AC; EC 4.6.1.1) is a key determinant of functional responsiveness in the heart and other tissues. We have tested two hypotheses regarding the relationship between AC content and β-adrenergic receptor (βAR)-mediated signal transduction in cardiac myocytes. First, that AC content limits adrenergic signal transduction, and, second, that increased AC, independent of (βAR) number and G-protein content, yields a proportional increase in βAR-mediated transmembrane signaling. We used recombinant adenovirus to increase AC isoform VI (ACVI) expression in neonatal cardiac myocytes. Cells that overexpressed ACVI responded to agonist stimulation with marked increases in cAMP production in proportion to protein expressed. In parallel experiments performed on cells transfected with lacZ (control) or ACVI, [3H]forskolin binding, used to assess AC protein expression, was amplified 6-fold, while βAR-stimulated cAMP production from these cells was increased 7-fold. No changes in βAR number, or in the heterotrimeric GTP-binding proteins, Gαs or Gαi2, were observed. Previous studies indicate that increased cardiac expression of βAR or Gαs does not yield proportional increases in transmembrane adrenergic signaling. In contrast, the current data demonstrate that increased ACVI expression provides a proportional increase in β-adrenergic signal transduction. Our results show that the amount of AC sets a limit on transmembrane β-adrenergic signaling. We speculate that similar functional responses are possible in other cell types in which AC plays an important physiological role. PMID:9448281

  7. Heterozygous mutations in cyclic AMP phosphodiesterase-4D (PDE4D) and protein kinase A (PKA) provide new insights into the molecular pathology of acrodysostosis.

    PubMed

    Kaname, Tadashi; Ki, Chang-Seok; Niikawa, Norio; Baillie, George S; Day, Jonathan P; Yamamura, Ken-Ichi; Ohta, Tohru; Nishimura, Gen; Mastuura, Nobuo; Kim, Ok-Hwa; Sohn, Young Bae; Kim, Hyun Woo; Cho, Sung Yoon; Ko, Ah-Ra; Lee, Jin Young; Kim, Hyun Wook; Ryu, Sung Ho; Rhee, Hwanseok; Yang, Kap-Seok; Joo, Keehyoung; Lee, Jooyoung; Kim, Chi Hwa; Cho, Kwang-Hyun; Kim, Dongsan; Yanagi, Kumiko; Naritomi, Kenji; Yoshiura, Ko-Ichiro; Kondoh, Tatsuro; Nii, Eiji; Tonoki, Hidefumi; Houslay, Miles D; Jin, Dong-Kyu

    2014-11-01

    Acrodysostosis without hormone resistance is a rare skeletal disorder characterized by brachydactyly, nasal hypoplasia, mental retardation and occasionally developmental delay. Recently, loss-of-function mutations in the gene encoding cAMP-hydrolyzing phosphodiesterase-4D (PDE4D) have been reported to cause this rare condition but the pathomechanism has not been fully elucidated. To understand the pathogenetic mechanism of PDE4D mutations, we conducted 3D modeling studies to predict changes in the binding efficacy of cAMP to the catalytic pocket in PDE4D mutants. Our results indicated diminished enzyme activity in the two mutants we analyzed (Gly673Asp and Ile678Thr; based on PDE4D4 residue numbering). Ectopic expression of PDE4D mutants in HEK293 cells demonstrated this reduction in activity, which was identified by increased cAMP levels. However, the cells from an acrodysostosis patient showed low cAMP accumulation, which resulted in a decrease in the phosphorylated cAMP Response Element-Binding Protein (pCREB)/CREB ratio. The reason for this discrepancy was due to a compensatory increase in expression levels of PDE4A and PDE4B isoforms, which accounted for the paradoxical decrease in cAMP levels in the patient cells expressing mutant isoforms with a lowered PDE4D activity. Skeletal radiographs of 10-week-old knockout (KO) rats showed that the distal part of the forelimb was shorter than in wild-type (WT) rats and that all the metacarpals and phalanges were also shorter in KO, as the name acrodysostosis implies. Like the G-protein α-stimulatory subunit and PRKAR1A, PDE4D critically regulates the cAMP signal transduction pathway and influences bone formation in a way that activity-compromising PDE4D mutations can result in skeletal dysplasia. We propose that specific inhibitory PDE4D mutations can lead to the molecular pathology of acrodysostosis without hormone resistance but that the pathological phenotype may well be dependent on an over-compensatory induction

  8. Parallel Allostery by cAMP and PDE Coordinates Activation and Termination Phases in cAMP Signaling.

    PubMed

    Krishnamurthy, Srinath; Tulsian, Nikhil Kumar; Chandramohan, Arun; Anand, Ganesh S

    2015-09-15

    The second messenger molecule cAMP regulates the activation phase of the cAMP signaling pathway through high-affinity interactions with the cytosolic cAMP receptor, the protein kinase A regulatory subunit (PKAR). Phosphodiesterases (PDEs) are enzymes responsible for catalyzing hydrolysis of cAMP to 5' AMP. It was recently shown that PDEs interact with PKAR to initiate the termination phase of the cAMP signaling pathway. While the steps in the activation phase are well understood, steps in the termination pathway are unknown. Specifically, the binding and allosteric networks that regulate the dynamic interplay between PKAR, PDE, and cAMP are unclear. In this study, PKAR and PDE from Dictyostelium discoideum (RD and RegA, respectively) were used as a model system to monitor complex formation in the presence and absence of cAMP. Amide hydrogen/deuterium exchange mass spectrometry was used to monitor slow conformational transitions in RD, using disordered regions as conformational probes. Our results reveal that RD regulates its interactions with cAMP and RegA at distinct loci by undergoing slow conformational transitions between two metastable states. In the presence of cAMP, RD and RegA form a stable ternary complex, while in the absence of cAMP they maintain transient interactions. RegA and cAMP each bind at orthogonal sites on RD with resultant contrasting effects on its dynamics through parallel allosteric relays at multiple important loci. RD thus serves as an integrative node in cAMP termination by coordinating multiple allosteric relays and governing the output signal response.

  9. Clinical and Molecular Genetics of the Phosphodiesterases (PDEs)

    PubMed Central

    Azevedo, Monalisa F.; Faucz, Fabio R.; Bimpaki, Eirini; Horvath, Anelia; Levy, Isaac; de Alexandre, Rodrigo B.; Ahmad, Faiyaz; Manganiello, Vincent

    2014-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) are enzymes that have the unique function of terminating cyclic nucleotide signaling by catalyzing the hydrolysis of cAMP and GMP. They are critical regulators of the intracellular concentrations of cAMP and cGMP as well as of their signaling pathways and downstream biological effects. PDEs have been exploited pharmacologically for more than half a century, and some of the most successful drugs worldwide today affect PDE function. Recently, mutations in PDE genes have been identified as causative of certain human genetic diseases; even more recently, functional variants of PDE genes have been suggested to play a potential role in predisposition to tumors and/or cancer, especially in cAMP-sensitive tissues. Mouse models have been developed that point to wide developmental effects of PDEs from heart function to reproduction, to tumors, and beyond. This review brings together knowledge from a variety of disciplines (biochemistry and pharmacology, oncology, endocrinology, and reproductive sciences) with emphasis on recent research on PDEs, how PDEs affect cAMP and cGMP signaling in health and disease, and what pharmacological exploitations of PDEs may be useful in modulating cyclic nucleotide signaling in a way that prevents or treats certain human diseases. PMID:24311737

  10. Interprofessional Flight Camp.

    PubMed

    Alfes, Celeste M; Rowe, Amanda S

    2016-01-01

    The Dorothy Ebersbach Academic Center for Flight Nursing in Cleveland, OH, holds an annual flight camp designed for master's degree nursing students in the acute care nurse practitioner program, subspecializing in flight nursing at the Frances Payne Bolton School of Nursing at Case Western Reserve University. The weeklong interprofessional training is also open to any health care provider working in an acute care setting and focuses on critical care updates, trauma, and emergency care within the critical care transport environment. This year, 29 graduate nursing students enrolled in a master's degree program from Puerto Rico attended. Although the emergency department in Puerto Rico sees and cares for trauma patients, there is no formal trauma training program. Furthermore, the country only has 1 rotor wing air medical transport service located at the Puerto Rico Medical Center in San Juan. Flight faculty and graduate teaching assistants spent approximately 9 months planning for their participation in our 13th annual flight camp. Students from Puerto Rico were extremely pleased with the learning experiences at camp and expressed particular interest in having more training time within the helicopter flight simulator.

  11. Astro Camp Plus

    NASA Technical Reports Server (NTRS)

    2006-01-01

    Stennis Space Center's new Astro Camp Plus camp kicked off June 19 for teens ages 13-15. The new camp delves more deeply into the science, math and technology concepts introduced in the center's popular Astro Camp series. Campers including Jasmyne White (left) and Dana Yingst, both of Slidell, La., learn how NASA uses 'podcasting' to broadcast video, and made their own podcasts.

  12. Victory Junction Gang Camp

    ERIC Educational Resources Information Center

    Shell, Ryan

    2007-01-01

    This article describes the Victory Junction Gang Camp, a not-for-profit, NASCAR-themed camp for children with chronic medical conditions that serves 24 different disease groups. The mission of the camp is to give children life-changing camping experiences that are exciting, fun, and empowering in a safe and medically sound environment. While doing…

  13. Camp through the Decades.

    ERIC Educational Resources Information Center

    Nicodemus, Teresa

    2003-01-01

    Camp pioneers relate how camping has grown to become more diverse, environmentally aware, safe, and conscious of its responsibility to promote healthy development of children. Changing trends in clothing, transportation, and food preparation at camp are described. The joys, discoveries, and teachable moments that camp offers children have endured.…

  14. A Computational Modeling and Simulation Approach to Investigate Mechanisms of Subcellular cAMP Compartmentation.

    PubMed

    Yang, Pei-Chi; Boras, Britton W; Jeng, Mao-Tsuen; Docken, Steffen S; Lewis, Timothy J; McCulloch, Andrew D; Harvey, Robert D; Clancy, Colleen E

    2016-07-01

    Subcellular compartmentation of the ubiquitous second messenger cAMP has been widely proposed as a mechanism to explain unique receptor-dependent functional responses. How exactly compartmentation is achieved, however, has remained a mystery for more than 40 years. In this study, we developed computational and mathematical models to represent a subcellular sarcomeric space in a cardiac myocyte with varying detail. We then used these models to predict the contributions of various mechanisms that establish subcellular cAMP microdomains. We used the models to test the hypothesis that phosphodiesterases act as functional barriers to diffusion, creating discrete cAMP signaling domains. We also used the models to predict the effect of a range of experimentally measured diffusion rates on cAMP compartmentation. Finally, we modeled the anatomical structures in a cardiac myocyte diad, to predict the effects of anatomical diffusion barriers on cAMP compartmentation. When we incorporated experimentally informed model parameters to reconstruct an in silico subcellular sarcomeric space with spatially distinct cAMP production sites linked to caveloar domains, the models predict that under realistic conditions phosphodiesterases alone were insufficient to generate significant cAMP gradients. This prediction persisted even when combined with slow cAMP diffusion. When we additionally considered the effects of anatomic barriers to diffusion that are expected in the cardiac myocyte dyadic space, cAMP compartmentation did occur, but only when diffusion was slow. Our model simulations suggest that additional mechanisms likely contribute to cAMP gradients occurring in submicroscopic domains. The difference between the physiological and pathological effects resulting from the production of cAMP may be a function of appropriate compartmentation of cAMP signaling. Therefore, understanding the contribution of factors that are responsible for coordinating the spatial and temporal

  15. A Computational Modeling and Simulation Approach to Investigate Mechanisms of Subcellular cAMP Compartmentation

    PubMed Central

    Yang, Pei-Chi; Boras, Britton W.; Jeng, Mao-Tsuen; Lewis, Timothy J.; McCulloch, Andrew D.; Harvey, Robert D.; Clancy, Colleen E.

    2016-01-01

    Subcellular compartmentation of the ubiquitous second messenger cAMP has been widely proposed as a mechanism to explain unique receptor-dependent functional responses. How exactly compartmentation is achieved, however, has remained a mystery for more than 40 years. In this study, we developed computational and mathematical models to represent a subcellular sarcomeric space in a cardiac myocyte with varying detail. We then used these models to predict the contributions of various mechanisms that establish subcellular cAMP microdomains. We used the models to test the hypothesis that phosphodiesterases act as functional barriers to diffusion, creating discrete cAMP signaling domains. We also used the models to predict the effect of a range of experimentally measured diffusion rates on cAMP compartmentation. Finally, we modeled the anatomical structures in a cardiac myocyte diad, to predict the effects of anatomical diffusion barriers on cAMP compartmentation. When we incorporated experimentally informed model parameters to reconstruct an in silico subcellular sarcomeric space with spatially distinct cAMP production sites linked to caveloar domains, the models predict that under realistic conditions phosphodiesterases alone were insufficient to generate significant cAMP gradients. This prediction persisted even when combined with slow cAMP diffusion. When we additionally considered the effects of anatomic barriers to diffusion that are expected in the cardiac myocyte dyadic space, cAMP compartmentation did occur, but only when diffusion was slow. Our model simulations suggest that additional mechanisms likely contribute to cAMP gradients occurring in submicroscopic domains. The difference between the physiological and pathological effects resulting from the production of cAMP may be a function of appropriate compartmentation of cAMP signaling. Therefore, understanding the contribution of factors that are responsible for coordinating the spatial and temporal

  16. Effects of carvedilol treatment on cardiac cAMP response element binding protein expression and phosphorylation in acute coxsackievirus B3-induced myocarditis

    PubMed Central

    2013-01-01

    Background The role of β-adrenergic stimulation on viral myocarditis has been investigated in animal models of viral myocarditis. Excess stimulation of β-adrenergic receptors by catecholamines causes phosphorylation/activation of cAMP response element binding protein (CREB) by the cAMP signaling pathway. CREB as an important regulator of gene expression mediates the cardiovascular remodeling process and promotes anti-inflammatory immune responses. However, the CREB expression and phosphorylation have not been studied, and the effects of carvedilol (a nonselective β-adrenoceptor antagonist) on the CREB has not been investigated in the setting of acute viral myocarditis. Methods This study was therefore designed to examine the effects of carvedilol on the transcriptional factor CREB in a murine model of acute viral myocarditis. In a coxsackievirus B3 murine myocarditis model (Balb/c), effects of carvedilol on plasma noradrenaline, heart rate and blood pressure, myocardial histopathological changes and fibrosis, cardiomyocyte apoptosis, cardiac CREB and phosphorylated CREB, cytokine levels, and viral RNA were studied. Results The expression and phosphorylation of CREB were decreased with concomitant increase of IL-6 and TNF-α in murine coxsackievirus-induced acute viral myocarditis. The levels of IL-6 and TNF-α were correlated with the expression of CREB or phosphorylated CREB. Carvedilol increased the cardiac CREB expression and phosphorylation and decreased the plasma catecholamine levels and the production of IL-6 and TNF-α with amelioration of acute viral myocarditis. Conclusion These results show that CREB may be involved in the pathophysiology of viral myocarditis and carvedilol exerts some of its beneficial effects by increasing the CREB expression and phosphorylation. PMID:24225056

  17. Bacterial Cyclic AMP-Phosphodiesterase Activity Coordinates Biofilm Formation

    PubMed Central

    Kalivoda, Eric J.; Brothers, Kimberly M.; Stella, Nicholas A.; Schmitt, Matthew J.; Shanks, Robert M. Q.

    2013-01-01

    Biofilm-related infections are a major contributor to human disease, and the capacity for surface attachment and biofilm formation are key attributes for the pathogenesis of microbes. Serratia marcescens type I fimbriae-dependent biofilms are coordinated by the adenylate cyclase, CyaA, and the cyclic 3′,5′-adenosine monophosphate (cAMP)-cAMP receptor protein (CRP) complex. This study uses S. marcescens as a model system to test the role of cAMP-phosphodiesterase activity in controlling biofilm formation. Herein we describe the characterization of a putative S. marcescens cAMP-phosphodiesterase gene (SMA3506), designated as cpdS, and demonstrated to be a functional cAMP-phosphodiesterase both in vitro and in vivo. Deletion of cpdS resulted in defective biofilm formation and reduced type I fimbriae production, whereas multicopy expression of cpdS conferred a type I fimbriae-dependent hyper-biofilm. Together, these results support a model in which bacterial cAMP-phosphodiesterase activity modulates biofilm formation. PMID:23923059

  18. Phosphodiesterase4D (PDE4D)--A risk factor for atrial fibrillation and stroke?

    PubMed

    Jørgensen, Carina; Yasmeen, Saiqa; Iversen, Helle K; Kruuse, Christina

    2015-12-15

    Mutations in the gene encoding phosphodiesterase 4D (PDE4D) enzyme are associated with ischemic stroke; however the functional implications of such mutations are not well understood. PDE4D is part of a complex protein family modulating intracellular signalling by cyclic nucleotides. The PDE4 family includes subtypes A-D, all of which show unique intracellular, cellular and tissue distribution. PDE4D is the major subtype expressed in human atrial myocytes and involved in the pathophysiology of arrhythmias, such as atrial fibrillation. The PDE4D enzyme hydrolyses cyclic adenosine monophosphate (cAMP). Though diverging results are reported, several population based studies describe association of various PDE4D single nucleotide polymorphisms (SNP) with cardio-embolic stroke in particular. Functionally, a down regulation of PDE4D variants has been reported in stroke patients. The anti-inflammatory and vasodilator properties of PDE4 inhibitors make them suitable for treatment of stroke and cardiovascular disease. PDE4D has recently been suggested as factor in atrial fibrillation. This review summarizes the possible function of PDE4D in the brain, heart, and vasculature. Further, association of the described SNPs, in particular, with cardioembolic stroke, is reviewed. Current findings on the PDE4D mutations suggest functionality involves an increased cardiac risk factor as well as augmented risk of atrial fibrillation. PMID:26671126

  19. Distribution and function of 3',5'-Cyclic-AMP phosphodiesterases in the human ovary.

    PubMed

    Petersen, T S; Kristensen, S G; Jeppesen, J V; Grøndahl, M L; Wissing, M L; Macklon, K T; Andersen, C Y

    2015-03-01

    The concentration of the important second messenger cAMP is regulated by phosphodiesterases (PDEs) and hence an attractive drug target. However, limited human data are available about the PDEs in the ovary. The aim of the present study was to describe and characterise the PDEs in the human ovary. Results were obtained by analysis of mRNA microarray data from follicles and granulosa cells (GCs), combined RT-PCR and enzymatic activity analysis in GCs, immunohistochemical analysis of ovarian sections and by studying the effect of PDE inhibitors on progesterone production from cultured GCs. We found that PDE3, PDE4, PDE7 and PDE8 are the major families present while PDE11A was not detected. PDE8B was differentially expressed during folliculogenesis. In cultured GCs, inhibition of PDE7 and PDE8 increased basal progesterone secretion while PDE4 inhibition increased forskolin-stimulated progesterone secretion. In conclusion, we identified PDE3, PDE4, PDE7 and PDE8 as the major PDEs in the human ovary.

  20. Regulation of the pro-inflammatory cytokine osteopontin by GIP in adipocytes - A role for the transcription factor NFAT and phosphodiesterase 3B

    SciTech Connect

    Omar, Bilal; Banke, Elin; Guirguis, Emilia; Aakesson, Lina; Manganiello, Vincent; Lyssenko, Valeriya; Groop, Leif; Gomez, Maria F.; Degerman, Eva

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer GIP stimulates lipogenesis and osteopontin expression in primary adipocytes. Black-Right-Pointing-Pointer GIP-induced osteopontin expression is NFAT-dependent. Black-Right-Pointing-Pointer Osteopontin expression is PDE3-dependent. Black-Right-Pointing-Pointer Osteopontin expression is increased in PDE3B KO mice. -- Abstract: The incretin - glucose-dependent insulinotropic polypeptide (GIP) - and the pro-inflammatory cytokine osteopontin are known to have important roles in the regulation of adipose tissue functions. In this work we show that GIP stimulates lipogenesis and osteopontin expression in primary adipocytes. The GIP-induced increase in osteopontin expression was inhibited by the NFAT (the transcription factor nuclear factor of activated T-cells) inhibitor A-285222. Also, the NFAT kinase glycogen synthase kinase (GSK) 3 was upregulated by GIP. To test whether cAMP might be involved in GIP-mediated effects on osteopontin a number of strategies were used. Thus, the {beta}3-adrenergic receptor agonist CL316,243 stimulated osteopontin expression, an effects which was mimicked by OPC3911, a specific inhibitor of phosphodiesterase 3. Furthermore, treatment of phosphodiesterase 3B knock-out mice with CL316,243 resulted in a dramatic upregulation of osteopontin in adipose tissue which was not the case in wild-type mice. In summary, we delineate mechanisms by which GIP stimulates osteopontin in adipocytes. Given the established link between osteopontin and insulin resistance, our data suggest that GIP by stimulating osteopontin expression, also could promote insulin resistance in adipocytes.

  1. Bidirectional promoters link cAMP signaling with irreversible differentiation through promoter-associated non-coding RNA (pancRNA) expression in PC12 cells.

    PubMed

    Yamamoto, Naoki; Agata, Kiyokazu; Nakashima, Kinichi; Imamura, Takuya

    2016-06-20

    Bidirectional promoters are the major source of gene activation-associated noncoding RNA (ncRNA). PC12 cells offer an interesting model for understanding the mechanism underlying bidirectional promoter-mediated cell cycle control. Nerve growth factor (NGF)-stimulated PC12 cells elongate neurites, and are in a reversible cell-cycle-arrested state. In contrast, these cells irreversibly differentiate and cannot re-enter the normal cell cycle after NGF plus cAMP treatment. In this study, using directional RNA-seq, we found that bidirectional promoters for protein-coding genes with promoter-associated ncRNA (pancRNA) were enriched for cAMP response element consensus sequences, and were preferred targets for transcriptional regulation by the transcription factors in the cAMP-dependent pathway. A spindle-formation-associated gene, Nusap1 and pancNusap1 were among the most strictly co-transcribed pancRNA-mRNA pairs. This pancRNA-mRNA pair was specifically repressed in irreversibly differentiated PC12 cells. Knockdown (KD) and overexpression experiments showed that pancNusap1 positively regulated the Nusap1 expression in a sequence-specific manner, which was accompanied by histone acetylation at the Nusap1 promoter. Furthermore, pancNusap1 KD recapitulated the effects of cAMP on cell cycle arrest. Thus, we conclude that pancRNA-mediated histone acetylation contributes to the establishment of the cAMP-induced transcription state of the Nusap1 locus and contributes to the irreversible cell cycle exit for terminal differentiation of PC12 cells. PMID:26945044

  2. Role of phosphodiesterase 5 in synaptic plasticity and memory

    PubMed Central

    Puzzo, Daniela; Sapienza, Salvatore; Arancio, Ottavio; Palmeri, Agostino

    2008-01-01

    Phosphodiesterases (PDEs) are enzymes that break down the phosphodiesteric bond of the cyclic nucleotides, cAMP and cGMP, second messengers that regulate many biological processes. PDEs participate in the regulation of signal transduction by means of a fine regulation of cyclic nucleotides so that the response to cell stimuli is both specific and activates the correct third messengers. Several PDE inhibitors have been developed and used as therapeutic agents because they increase cyclic nucleotide levels by blocking the PDE function. In particular, sildenafil, an inhibitor of PDE5, has been mainly used in the treatment of erectile dysfunction but is now also utilized against pulmonary hypertension. This review examines the physiological role of PDE5 in synaptic plasticity and memory and the use of PDE5 inhibitors as possible therapeutic agents against disorders of the central nervous system (CNS). PMID:18728748

  3. Intracellular tortuosity underlies slow cAMP diffusion in adult ventricular myocytes

    PubMed Central

    Richards, Mark; Lomas, Oliver; Jalink, Kees; Ford, Kerrie L.; Vaughan-Jones, Richard D.; Lefkimmiatis, Konstantinos; Swietach, Pawel

    2016-01-01

    Aims 3′,5′-Cyclic adenosine monophosphate (cAMP) signals in the heart are often confined to concentration microdomains shaped by cAMP diffusion and enzymatic degradation. While the importance of phosphodiesterases (degradative enzymes) in sculpting cAMP microdomains is well established in cardiomyocytes, less is known about cAMP diffusivity (DcAMP) and factors affecting it. Many earlier studies have reported fast diffusivity, which argues against sharply defined microdomains. Methods and results [cAMP] dynamics in the cytoplasm of adult rat ventricular myocytes were imaged using a fourth generation genetically encoded FRET-based sensor. The [cAMP]-response to the addition and removal of isoproterenol (β-adrenoceptor agonist) quantified the rates of cAMP synthesis and degradation. To obtain a read out of DcAMP, a stable [cAMP] gradient was generated using a microfluidic device which delivered agonist to one half of the myocyte only. After accounting for phosphodiesterase activity, DcAMP was calculated to be 32 µm2/s; an order of magnitude lower than in water. Diffusivity was independent of the amount of cAMP produced. Saturating cAMP-binding sites with the analogue 6-Bnz-cAMP did not accelerate DcAMP, arguing against a role of buffering in restricting cAMP mobility. cAMP diffused at a comparable rate to chemically unrelated but similar sized molecules, arguing for a common physical cause of restricted diffusivity. Lower mitochondrial density and order in neonatal cardiac myocytes allowed for faster diffusion, demonstrating the importance of mitochondria as physical barriers to cAMP mobility. Conclusion In adult cardiac myocytes, tortuosity due to physical barriers, notably mitochondria, restricts cAMP diffusion to levels that are more compatible with microdomain signalling. PMID:27089919

  4. Role of phosphodiesterase-4 on ethanol elicited locomotion and narcosis.

    PubMed

    Baliño, Pablo; Ledesma, Juan Carlos; Aragon, Carlos M G

    2016-02-01

    The cAMP signaling pathway has emerged as an important modulator of the pharmacological effects of ethanol. In this respect, the cAMP-dependent protein kinase has been shown to play an important role in the modulation of several ethanol-induced behavioral actions. Cellular levels of cAMP are maintained by the activity of adenylyl cyclases and phosphodiesterases. In the present work we have focused on ascertaining the role of PDE4 in mediating the neurobehavioral effects of ethanol. For this purpose, we have used the selective PDE4 inhibitor Ro 20-1724. This compound has been proven to enhance cellular cAMP response by PDE4 blockade and can be administered systemically. Swiss mice were injected intraperitoneally (i.p.) with Ro 20-1724 (0-5 mg/kg; i.p.) at different time intervals before ethanol (0-4 g/kg; i.p.) administration. Immediately after the ethanol injection, locomotor activity, loss of righting reflex, PKA footprint and enzymatic activity were assessed. Pretreatment with Ro 20-1724 increased ethanol-induced locomotor stimulation in a dose-dependent manner. Doses that increased locomotor stimulation did not modify basal locomotion or the suppression of motor activity produced by high doses of this alcohol. Ro 20-1724 did not alter the locomotor activation produced by amphetamine or cocaine. The time of loss of righting reflex evoked by ethanol was increased after pretreatment with Ro 20-1724. This effect was selective for the narcotic effects of ethanol since Ro 20-1724 did not affect pentobarbital-induced narcotic effects. Moreover, Ro 20-1724 administration increased the PKA footprint and enzymatic activity response elicited by ethanol. These data provide further evidence of the key role of the cAMP signaling pathway in the central effects of ethanol.

  5. Inhibitors of phosphodiesterases PDE2, PDE3, and PDE4 do not increase the sinoatrial tachycardia of noradrenaline and prostaglandin PGE₁ in mice.

    PubMed

    Galindo-Tovar, Alejandro; Vargas, María Luisa; Kaumann, Alberto J

    2016-02-01

    Phosphodiesterases PDE2, PDE3, and PDE4 are expressed in murine sinoatrial cells. PDE3 and/or PDE4 reduce heart rate but apparently do not influence the tachycardia mediated through sinoatrial β1- and β2-adrenoceptors despite the high content of sinoatrial cAMP. The function of PDE2 is, however, uncertain. Prostaglandin PGE1 elicits sinoatrial tachycardia through EP receptors, but the control by phosphodiesterases is unknown. We investigated on spontaneously beating right atria of mice the effects of the PDE2 inhibitors Bay 60-7550 and EHNA on basal beating and the tachycardia produced by noradrenaline (3 nM) and PGE1 (1 μM). Bay 60-7550 (1 μM), but not EHNA (10 μM), increased basal sinoatrial beating. EHNA also failed to produce tachycardia in the presence of the adenosine deaminase inhibitor 2'-deoxycoformycin (10 μM), remaining inconclusive whether PDE2 reduces basal sinoatrial beating. Rolipram (10 μM) and cilostamide (300 nM) caused moderate tachycardia. The tachycardia evoked by Bay 60-7550 was similar in the absence and presence of rolipram. Noradrenaline elicited stable tachycardia that was not increased by Bay 60-7550. A stable tachycardia caused by PGE1 was not increased by the inhibitors of PDE2, PDE3, and PDE4. Unlike PDE3 and PDE4 which reduce murine basal sinoatrial beating, a possible effect of PDE2 needs further research. The stable tachycardia produced by noradrenaline and PGE1, together with the lack potentiation by the inhibitors of PDE2, PDE3, and PDE4, suggests that cAMP generated at the receptor compartments is hardly hydrolyzed by these phophodiesterases. Evidence from human volunteers is consistent with this proposal.

  6. Phosphodiesterase inhibitors block the acceleration of skin permeability barrier repair by red light.

    PubMed

    Goto, Makiko; Ikeyama, Kazuyuki; Tsutsumi, Moe; Denda, Sumiko; Denda, Mitsuhiro

    2011-07-01

    We previously demonstrated that exposure to red light (550-670 nm) accelerates epidermal permeability barrier recovery after barrier disruption. Furthermore, we showed that photosensitive proteins, originally found in retina, are also expressed in epidermis. In retina, transducin and phosphodiesterase 6 play key roles in signal transmission. In this study, we evaluate the role of phosphodiesterese 6 in the acceleration by red light of epidermal permeability barrier recovery. Immunohistochemical study and reverse transcription-PCR assays confirmed the expression of both transducin and phosphodiesterase 6 in epidermal keratinocytes. Topical application of 3-isobutyl-1-methylxanthine, a non-specific phosphodiesterase inhibitor, blocked the acceleration of the barrier recovery by red light. Topical application of zaprinast, a specific inhibitor of phosphodiesterases 5 and 6, also blocked the acceleration, whereas T0156, a specific inhibitor of phosphodiesterase 5, had no effect. Red light exposure reduced the epidermal hyperplasia induced by barrier disruption under low humidity, and the effect was blocked by pretreatment with zaprinast. Our results indicate phosphodiesterase 6 is involved in the recovery-accelerating effect of red light on the disrupted epidermal permeability barrier.

  7. Phosphodiesterase inhibitor-dependent inverse agonism of agouti-related protein on melanocortin 4 receptor in sea bass (Dicentrarchus labrax)

    PubMed Central

    Sánchez, Elisa; Rubio, Vera Cruz; Thompson, Darren; Metz, Juriaan; Flik, Gert; Millhauser, Glenn L.; Cerdá-Reverter, José Miguel

    2009-01-01

    The melanocortin 4 receptor (MC4R) is a G protein-coupled receptor mainly expressed in the central nervous system of vertebrates. Activation of the MC4R leads to a decrease in food intake, whereas inactivating mutations are a genetic cause of obesity. The binding of agouti-related protein (AGRP) reduces not only agonist-stimulated cAMP production (competitive antagonist) but also the basal activity of the receptor, as an inverse agonist. Transgenic zebrafish overexpressing AGRP display increased food intake and linear growth, indicative of a physiological role for the melanocortin system in the control of the energy balance in fish. We report on the cloning, pharmacological characterization, tissue distribution, and detailed brain mapping of a sea bass (Dicentrarchus labrax) MC4R ortholog. Sea bass MC4R is profusely expressed within food intake-controlling pathways of the fish brain. However, the activity of the melanocortin system during progressive fasting does not depend on the hypothalamic/pituitary proopiomelanocortin (POMC) and MC4R expression, which suggests that sea bass MC4R is constitutively activated and regulated by AGRP binding. We demonstrate that AGRP acts as competitive antagonist and reduces MTII-induced cAMP production. AGRP also decreases the basal activity of the receptor as an inverse agonist. This observation suggests that MC4R is constitutively active and supports the evolutionary conservation of the AGRP/MC4R interactions. The inverse agonism, but not the competitive antagonism, depends on the presence of a phosphodiesterase inhibitor (IBMX). This suggests that inverse agonism and competitive antagonism operate through different intracellular signaling pathways, a view that opens up new targets for the treatment of melanocortin-induced metabolic syndrome. PMID:19225141

  8. Michelle: Growing Through Camping

    ERIC Educational Resources Information Center

    Ouellet, Annette M.

    1977-01-01

    A mother, whose physically handicapped 7-year-old daughter has prosthetic feet, describes how her child adjusted well first to a summer day camp and then to a week long camping program run by the Girl Scouts. (GW)

  9. 1,3-Dichloro-2-propanol inhibits progesterone production through the expression of steroidogenic enzymes and cAMP concentration in Leydig cells.

    PubMed

    Sun, Jianxia; Bai, Shun; Bai, Weibin; Zou, Feiyan; Zhang, Lei; Li, Guoqiang; Hu, Yunfeng; Li, Mingwei; Yan, Rian; Su, Zhijian; Huang, Yadong

    2014-07-01

    1,3-Dichloro-2-propanol (1,3-DCP) is a well-known food processing contaminant that has been shown to impede male reproductive function. However, its mechanism of action remains elusive. In this study, the effects of 1,3-DCP on progesterone production were investigated using the R2C Leydig cell model. 1,3-DCP significantly reduced cell viability from 7.48% to 97.4% at doses comprised between 0.5 and 6mM. Single cell gel/comet assays and atomic force microscopy assays showed that 1,3-DCP induced early phase cell apoptosis. In addition, 1,3-DCP significantly reduced progesterone production detected by radioimmunoassay (RIA). The results from quantitative polymerase chain reaction and western blotting demonstrated that the mRNA expression levels of steroidogenic acute regulatory protein (StAR), cytochrome P450 side-chain cleavage enzyme and 3β-hydroxysteroid dehydrogenase were significantly down-regulated in R2C cells. Particularly, the change rhythm of Star expression was highly consistent with progesterone production. Furthermore, the cyclic adenosine monophosphate (cAMP) and the mitochondrial membrane potential mediated by ROS, which are involved in regulating progesterone synthesis were also decreased in response to the 1,3-DCP treatment. Overall, the data presented here suggested that 1,3-DCP interferes with the male steroidogenic capacity mainly by down-regulating the level of cAMP and the key enzymes involved in the androgen synthesis pathway. PMID:24518350

  10. Spilanthol from Acmella Oleracea Lowers the Intracellular Levels of cAMP Impairing NKCC2 Phosphorylation and Water Channel AQP2 Membrane Expression in Mouse Kidney

    PubMed Central

    Gerbino, Andrea; Schena, Giorgia; Milano, Serena; Milella, Luigi; Barbosa, Alan Franco; Armentano, Francesca; Procino, Giuseppe; Svelto, Maria; Carmosino, Monica

    2016-01-01

    Acmella oleracea is well recognized in Brazilian traditional medicine as diuretic, although few scientific data have been published to support this effect. Aim of this study was to determine the molecular effect of Acmella oleracea extract and its main alkylamide spilanthol on two major processes involved in the urine concentrating mechanism: Na-K-2Cl symporter (NKCC2) activity in the thick ascending limb and water channel aquaporin 2 accumulation at the apical plasma membrane of collecting duct cells. Phosphorylation of NKCC2 was evaluated as index of its activation by Western blotting. Rate of aquaporin 2 apical expression was analyzed by confocal laser microscopy. Spilanthol-induced intracellular signalling events were dissected by video-imaging experiments. Exposure to spilanthol reduced the basal phosphorylation level of NKCC2 both in freshly isolated mouse kidney slices and in NKCC2-expresing HEK293 cells. In addition, exposure to spilanthol strongly reduced both desmopressin and low Cl−-dependent increase in NKCC2 phosphorylation in mouse kidney slices and NKCC2-expressing HEK293 cells, respectively. Similarly, spilanthol reduced both desmopressin- and forskolin-stimulated aquaporin 2 accumulation at the apical plasma membrane of collecting duct in mouse kidney slice and MCD4 cells, respectively. Of note, when orally administered, spilanthol induced a significant increase in both urine output and salt urinary excretion associated with a markedly reduced urine osmolality compared with control mice. Finally, at cellular level, spilanthol rapidly reduced or reversed basal and agonist-increased cAMP levels through a mechanism involving increases in intracellular [Ca2+]. In conclusion, spilanthol-induced inhibition of cAMP production negatively modulates urine-concentrating mechanisms thus holding great promise for its use as diuretic. PMID:27213818

  11. The mechanisms of action of cAMP. A quantum chemical study.

    PubMed

    van Ool, P J; Buck, H M

    1982-01-01

    Quantum chemical calculations were performed on the formation of intermediates with trigonal bipyramidal (TBP) configurations in the hydrolysis of adenosine 3',5'-monophosphate (cAMP) with phosphodiesterases and the activation of protein kinases by cAMP. The results show that in the reaction sequence concerning the hydrolysis of cAMP with phosphodiesterase the TBP intermediate must possess an equatorial-apical cyclic phosphate ring with the 3'-oxygen atom in the apical position. This could be an additional reason for the sensitivity of the 3' position in cAMP towards modifications in comparison with the 5' position. According to the calculations, a mechanistic model is presented for the enzymatic hydrolysis of cAMP with the involvement of a covalently bonded enzyme-nucleotide intermediate. Also a model is offered for the activation of protein kinase by cAMP. The activation of protein kinase is assumed to proceed via diequatorial-ring-positioned TBP intermediates resulting in the formation of a covalent bond between cAMP and the protein kinase with retention of the cyclic phosphate ring. It seems likely that the enzyme-nucleotide intermediate enforces a conformational change in the enzyme, which causes the dissociation of the regulatory and catalytic subunit of the protein kinase, necessary for a physiological response.

  12. Camp Market Research.

    ERIC Educational Resources Information Center

    Chenery, Mary Faeth; Akers, Ward

    1987-01-01

    By interviewing parents and reviewing letters of complaint, Camp Hy-Lake (Tennessee) identified six conditions/outcomes parents expected of a good camp experience. By providing those conditions the camp improved its camper return rate from 60 to 80% in approximately two years and increased camper returns from two to three and a half years. (JHZ)

  13. Phosphodiesterase Inhibition to Target the Synaptic Dysfunction in Alzheimer's Disease

    NASA Astrophysics Data System (ADS)

    Bales, Kelly R.; Plath, Niels; Svenstrup, Niels; Menniti, Frank S.

    Alzheimer's Disease (AD) is a disease of synaptic dysfunction that ultimately proceeds to neuronal death. There is a wealth of evidence that indicates the final common mediator of this neurotoxic process is the formation and actions on synaptotoxic b-amyloid (Aβ). The premise in this review is that synaptic dysfunction may also be an initiating factor in for AD and promote synaptotoxic Aβ formation. This latter hypothesis is consistent with the fact that the most common risk factors for AD, apolipoprotein E (ApoE) allele status, age, education, and fitness, encompass suboptimal synaptic function. Thus, the synaptic dysfunction in AD may be both cause and effect, and remediating synaptic dysfunction in AD may have acute effects on the symptoms present at the initiation of therapy and also slow disease progression. The cyclic nucleotide (cAMP and cGMP) signaling systems are intimately involved in the regulation of synaptic homeostasis. The phosphodiesterases (PDEs) are a superfamily of enzymes that critically regulate spatial and temporal aspects of cyclic nucleotide signaling through metabolic inactivation of cAMP and cGMP. Thus, targeting the PDEs to promote improved synaptic function, or 'synaptic resilience', may be an effective and facile approach to new symptomatic and disease modifying therapies for AD. There continues to be a significant drug discovery effort aimed at discovering PDE inhibitors to treat a variety of neuropsychiatric disorders. Here we review the current status of those efforts as they relate to potential new therapies for AD.

  14. Cadmium-induced decrement of the LH receptor expression and cAMP levels in the testis of rats.

    PubMed

    Gunnarsson, David; Nordberg, Gunnar; Lundgren, Per; Selstam, Gunnar

    2003-02-01

    Cadmium (Cd) is a widespread environmental pollutant, characterized by its ability to affect various organs. Adverse effect of Cd on the testis including decreased testosterone production are well-known phenomena, but the cellular events explaining these effects have not yet been established. In the present study the initial steps of gonadotropin mediated testosterone biosynthesis were examined in vivo in rats, in relation to Cd dose and time after injection. In the dose-response experiment Male Sprague-Dawley rats received a single subcutaneous (s.c.) injection of CdCl(2) (1, 5 or 10 micromol/kg body weight) and were sacrificed 48 h after injection. A statistically significant decrease in luteinizing hormone (LH) receptor mRNA level in the testicular tissue was demonstrated at the highest dose (10 micromol/kg). In the temporal-response experiment rats were given 10 micromol/kg of CdCl(2) s.c. and sacrificed 0.48, 4.8, 48 or 144 h after injection. LH receptor mRNA levels as well as cyclic adenosine monophosphate (cAMP) levels were found to be significantly lowered at 48 and 144 h. These observations of the mechanisms whereby Cd exerts its effect on the initial steps of testosterone biosynthesis are the first from in vivo experiments. PMID:12504342

  15. Creating healthy camp experiences.

    PubMed

    Walton, Edward A; Tothy, Alison S

    2011-04-01

    The American Academy of Pediatrics has created recommendations for health appraisal and preparation of young people before participation in day or resident camps and to guide health and safety practices for children at camp. These recommendations are intended for parents, primary health care providers, and camp administration and health center staff. Although camps have diverse environments, there are general guidelines that apply to all situations and specific recommendations that are appropriate under special conditions. This policy statement has been reviewed and is supported by the American Camp Association. PMID:21444589

  16. Engineering of a red-light–activated human cAMP/cGMP-specific phosphodiesterase

    PubMed Central

    Gasser, Carlos; Taiber, Sandra; Yeh, Chen-Min; Wittig, Charlotte Helene; Hegemann, Peter; Ryu, Soojin; Wunder, Frank; Möglich, Andreas

    2014-01-01

    Sensory photoreceptors elicit vital physiological adaptations in response to incident light. As light-regulated actuators, photoreceptors underpin optogenetics, which denotes the noninvasive, reversible, and spatiotemporally precise perturbation by light of living cells and organisms. Of particular versatility, naturally occurring photoactivated adenylate cyclases promote the synthesis of the second messenger cAMP under blue light. Here, we have engineered a light-activated phosphodiesterase (LAPD) with complementary light sensitivity and catalytic activity by recombining the photosensor module of Deinococcus radiodurans bacterial phytochrome with the effector module of Homo sapiens phosphodiesterase 2A. Upon red-light absorption, LAPD up-regulates hydrolysis of cAMP and cGMP by up to sixfold, whereas far-red light can be used to down-regulate activity. LAPD also mediates light-activated cAMP and cGMP hydrolysis in eukaryotic cell cultures and in zebrafish embryos; crucially, the biliverdin chromophore of LAPD is available endogenously and does not need to be provided exogenously. LAPD thus establishes a new optogenetic modality that permits light control over diverse cAMP/cGMP-mediated physiological processes. Because red light penetrates tissue more deeply than light of shorter wavelengths, LAPD appears particularly attractive for studies in living organisms. PMID:24889611

  17. Ustilago maydis phosphodiesterases play a role in the dimorphic switch and in pathogenicity.

    PubMed

    Agarwal, Charu; Aulakh, Kavita B; Edelen, Kaly; Cooper, Michael; Wallen, R Margaret; Adams, Seth; Schultz, David J; Perlin, Michael H

    2013-05-01

    Components of the cAMP (cyclic AMP) signalling cascades are conserved from fungi to humans, and are particularly important for fungal dimorphism and pathogenicity. Previous work has described two phosphodiesterases, UmPde1 and UmPde2, in Ustilago maydis which show strong phosphodiesterase activity. We further characterized the biological function(s) of these phosphodiesterases in U. maydis. Specifically, we examined their possible role(s) in regulation of the cAMP-dependent protein kinase A (PKA) pathway and their roles in filamentous growth and pathogenicity. We found that UmPde1, which shares 35 % similarity with Cryptococcus neoformans Pde1, also displays functional homology with this enzyme. UmPde1 complements the capsule-formation defect of C. neoformans strains deleted for Pde1. In U. maydis, the cell morphology of the umpde1 deletion mutant resembled the multiple budding phenotypes seen with the ubc1 mutant, which lacks the regulatory subunit of PKA. Interestingly, on low-ammonium medium, umpde2 deletion strains showed a reduction in filamentation that was comparable to that of ubc1 deletion strains; however, umpde1 deletion strains showed normal filamentation on low-ammonium medium. Furthermore, both the ubc1 deletion strain in which the PKA pathway was constitutively active and the umpde1 deletion strains were significantly reduced in pathogenicity, while the umpde2 deletion strains showed a trend for reduced pathogenicity compared with wild-type strains. These data support a role for the phosphodiesterases UmPde1 and UmPde2 in regulating the U. maydis cAMP-dependent PKA pathway through modulation of cAMP levels, thus affecting dimorphic growth and pathogenicity.

  18. Stimulation of StAR expression by cAMP is controlled by inhibition of highly inducible SIK1 via CRTC2, a co-activator of CREB.

    PubMed

    Lee, Jinwoo; Tong, Tiegang; Takemori, Hiroshi; Jefcoate, Colin

    2015-06-15

    In mouse steroidogenic cells the activation of cholesterol metabolism is mediated by steroidogenic acute regulatory protein (StAR). Here, we visualized a coordinated regulation of StAR transcription, splicing and post-transcriptional processing, which are synchronized by salt inducible kinase (SIK1) and CREB-regulated transcription coactivator (CRTC2). To detect primary RNA (pRNA), spliced primary RNA (Sp-RNA) and mRNA in single cells, we generated probe sets by using fluorescence in situ hybridization (FISH). These methods allowed us to address the nature of StAR gene expression and to visualize protein-nucleic acid interactions through direct detection. We show that SIK1 represses StAR expression in Y1 adrenal and MA10 testis cells through inhibition of processing mediated by CRTC2. Digital image analysis matches qPCR analyses of the total cell culture. Evidence is presented for spatially separate accumulation of StAR pRNA and Sp-RNA at the gene loci in the nucleus. These findings establish that cAMP, SIK and CRTC mediate StAR expression through activation of individual StAR gene loci.

  19. Phosphodiesterase 11A (PDE11A), Enriched in Ventral Hippocampus Neurons, is Required for Consolidation of Social but not Nonsocial Memories in Mice

    PubMed Central

    Hegde, Shweta; Capell, Will R; Ibrahim, Baher A; Klett, Jennifer; Patel, Neema S; Sougiannis, Alexander T; Kelly, Michy P

    2016-01-01

    The capacity to form long-lasting social memories is critical to our health and survival. cAMP signaling in the ventral hippocampal formation (VHIPP) appears to be required for social memory formation, but the phosphodiesterase (PDE) involved remains unknown. Previously, we showed that PDE11A, which degrades cAMP and cGMP, is preferentially expressed in CA1 and subiculum of the VHIPP. Here, we determine whether PDE11A is expressed in neurons where it could directly influence synaptic plasticity and whether expression is required for the consolidation and/or retrieval of social memories. In CA1, and possibly CA2, PDE11A4 is expressed throughout neuronal cell bodies, dendrites (stratum radiatum), and axons (fimbria), but not astrocytes. Unlike PDE2A, PDE9A, or PDE10A, PDE11A4 expression begins very low at postnatal day 7 (P7) and dramatically increases until P28, at which time it stabilizes to young adult levels. This expression pattern is consistent with the fact that PDE11A is required for social long-term memory (LTM) formation during adolescence and adulthood. Male and female PDE11 knockout (KO) mice show normal short-term memory (STM) for social odor recognition (SOR) and social transmission of food preference (STFP), but no LTM 24 h post training. Importantly, PDE11A KO mice show normal LTM for nonsocial odor recognition. Deletion of PDE11A may impair memory consolidation by impairing requisite protein translation in the VHIPP. Relative to WT littermates, PDE11A KO mice show reduced expression of RSK2 and lowered phosphorylation of S6 (pS6–235/236). Together, these data suggest PDE11A is selectively required for the proper consolidation of recognition and associative social memories. PMID:27339393

  20. Cyclic nucleotide phosphodiesterases in heart and vessels: A therapeutic perspective.

    PubMed

    Bobin, Pierre; Belacel-Ouari, Milia; Bedioune, Ibrahim; Zhang, Liang; Leroy, Jérôme; Leblais, Véronique; Fischmeister, Rodolphe; Vandecasteele, Grégoire

    2016-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) degrade the second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), thereby regulating multiple aspects of cardiac and vascular muscle functions. This highly diverse class of enzymes encoded by 21 genes encompasses 11 families that are not only responsible for the termination of cyclic nucleotide signalling, but are also involved in the generation of dynamic microdomains of cAMP and cGMP, controlling specific cell functions in response to various neurohormonal stimuli. In the myocardium and vascular smooth muscle, the PDE3 and PDE4 families predominate, degrading cAMP and thereby regulating cardiac excitation-contraction coupling and smooth muscle contractile tone. PDE3 inhibitors are positive inotropes and vasodilators in humans, but their use is limited to acute heart failure and intermittent claudication. PDE5 is particularly important for the degradation of cGMP in vascular smooth muscle, and PDE5 inhibitors are used to treat erectile dysfunction and pulmonary hypertension. There is experimental evidence that these PDEs, as well as other PDE families, including PDE1, PDE2 and PDE9, may play important roles in cardiac diseases, such as hypertrophy and heart failure, as well as several vascular diseases. After a brief presentation of the cyclic nucleotide pathways in cardiac and vascular cells, and the major characteristics of the PDE superfamily, this review will focus on the current use of PDE inhibitors in cardiovascular diseases, and the recent research developments that could lead to better exploitation of the therapeutic potential of these enzymes in the future. PMID:27184830

  1. A CaMKII/PDE4D negative feedback regulates cAMP signaling

    PubMed Central

    Mika, Delphine; Richter, Wito; Conti, Marco

    2015-01-01

    cAMP production and protein kinase A (PKA) are the most widely studied steps in β-adrenergic receptor (βAR) signaling in the heart; however, the multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII) is also activated in response to βAR stimulation and is involved in the regulation of cardiac excitation-contraction coupling. Its activity and expression are increased during cardiac hypertrophy, in heart failure, and under conditions that promote arrhythmias both in animal models and in the human heart, underscoring the clinical relevance of CaMKII in cardiac pathophysiology. Both CaMKII and PKA phosphorylate a number of protein targets critical for Ca2+ handling and contraction with similar, but not always identical, functional consequences. How these two pathways communicate with each other remains incompletely understood, however. To maintain homeostasis, cyclic nucleotide levels are regulated by phosphodiesterases (PDEs), with PDE4s predominantly responsible for cAMP degradation in the rodent heart. Here we have reassessed the interaction between cAMP/PKA and Ca2+/CaMKII signaling. We demonstrate that CaMKII activity constrains basal and βAR-activated cAMP levels. Moreover, we show that these effects are mediated, at least in part, by CaMKII regulation of PDE4D. This regulation establishes a negative feedback loop necessary to maintain cAMP/CaMKII homeostasis, revealing a previously unidentified function for PDE4D as a critical integrator of cAMP/PKA and Ca2+/CaMKII signaling. PMID:25646485

  2. The activity of cAMP-phosphodiesterase 4D7 (PDE4D7) is regulated by protein kinase A-dependent phosphorylation within its unique N-terminus

    PubMed Central

    Byrne, Ashleigh M.; Elliott, Christina; Hoffmann, Ralf; Baillie, George S.

    2015-01-01

    The cyclic AMP phosphodiesterases type 4 (PDE4s) are expressed in a cell specific manner, with intracellular targeting directed by unique N-terminal anchor domains. All long form PDE4s are phosphorylated and activated by PKA phosphorylation within their upstream conserved region 1 (UCR1). Here, we identify and characterise a novel PKA site (serine 42) within the N-terminal region of PDE4D7, an isoform whose activity is known to be important in prostate cancer progression and ischemic stroke. In contrast to the UCR1 site, PKA phosphorylation of the PDE4D7 N-terminus appears to occur constitutively and inhibits PDE4 activity to allow cAMP signalling under basal conditions. PMID:25680530

  3. Regulation of cyclooxygenase-2 expression by cAMP response element and mRNA stability in a human airway epithelial cell line exposed to zinc

    SciTech Connect

    Wu Weidong Silbajoris, Robert A.; Cao Dongsun; Bromberg, Philip A.; Zhang Qiao; Peden, David B.; Samet, James M.

    2008-09-01

    Exposure to zinc-laden particulate matter in ambient and occupational settings has been associated with proinflammatory responses in the lung. Cyclooxygenase 2-derived eicosanoids are important modulators of airway inflammation. In this study, we characterized the transcriptional and posttranscriptional events that regulate COX-2 expression in a human bronchial epithelial cell line BEAS-2B exposed to Zn{sup 2+}. Zn{sup 2+} exposure resulted in pronounced increases in COX-2 mRNA and protein expression, which were prevented by pretreatment with the transcription inhibitor actinomycin D, implying the involvement of transcriptional regulation. This was supported by the observation of increased COX-2 promoter activity in Zn{sup 2+}-treated BEAS-2B cells. Mutation of the cAMP response element (CRE), but not the {kappa}B-binding sites in the COX-2 promoter markedly reduced COX-2 promoter activity induced by Zn{sup 2+}. Inhibition of NF{kappa}B activation did not block Zn{sup 2+}-induced COX-2 expression. Measurement of mRNA stability demonstrated that Zn{sup 2+} exposure impaired the degradation of COX-2 mRNA in BEAS-2B cells. This message stabilization effect of Zn{sup 2+} exposure was shown to be dependent on the integrity of the 3'-untranslated region found in the COX-2 transcript. Taken together, these data demonstrate that the CRE and mRNA stability regulates COX-2 expression induced in BEAS-2B cells exposed to extracellular Zn{sup 2+}.

  4. Molecular cloning and subcellular distribution of the novel PDE4B4 cAMP-specific phosphodiesterase isoform.

    PubMed Central

    Shepherd, Malcolm; McSorley, Theresa; Olsen, Aileen E; Johnston, Lee Ann; Thomson, Neil C; Baillie, George S; Houslay, Miles D; Bolger, Graeme B

    2003-01-01

    We have isolated cDNAs encoding PDE4B4, a new cAMP-specific phosphodiesterase (PDE4) isoform with novel properties. The amino acid sequence of PDE4B4 demonstrates that it is encoded by the PDE4B gene, but that it differs from the previously isolated PDE4B1, PDE4B2 and PDE4B3 isoforms by the presence of a novel N-terminal region of 17 amino acids. PDE4B4 contains both of the upstream conserved region 1 (UCR1) and UCR2 regulatory units that are characteristic of 'long' PDE4 isoforms. RNase protection demonstrated that PDE4B4 mRNA is expressed preferentially in liver, skeletal muscle and various regions of the brain, which differs from the pattern of tissue distribution of the other known PDE4B long forms, PDE4B1 and PDE4B3. Expression of PDE4B4 cDNA in COS7 cells produced a protein of 85 kDa under denaturing conditions. Subcellular fractionation of recombinant, COS7-cell expressed PDE4B4 showed that the protein was localized within the cytosol, which was confirmed by confocal microscopic analysis of living COS7 cells transfected with a green fluorescent protein-PDE4B4 chimaera. PDE4B4 exhibited a K(m) for cAMP of 5.4 microM and a V(max), relative to that of the long PDE4B1 isoform, of 2.1. PDE4B4 was inhibited by the prototypical PDE4 inhibitor rolipram [4-[3-(cyclopentoxyl)-4-methoxyphenyl]-2-pyrrolidinone] with an IC(50) of 83 nM. Treatment of COS7 cells with forskolin, to elevate cAMP levels, produced activation of PDE4B4, which was associated with the phosphorylation of PDE4B4 on Ser-56 within UCR1. The unique tissue distribution and intracellular targeting of PDE4B4 suggests that this isoform may have a distinct functional role in regulating cAMP levels in specific cell types. PMID:12441002

  5. Development of a New Radiofluorinated Quinoline Analog for PET Imaging of Phosphodiesterase 5 (PDE5) in Brain

    PubMed Central

    Liu, Jianrong; Wenzel, Barbara; Dukic-Stefanovic, Sladjana; Teodoro, Rodrigo; Ludwig, Friedrich-Alexander; Deuther-Conrad, Winnie; Schröder, Susann; Chezal, Jean-Michel; Moreau, Emmanuel; Brust, Peter; Maisonial-Besset, Aurélie

    2016-01-01

    Phosphodiesterases (PDEs) are enzymes that play a major role in cell signalling by hydrolysing the secondary messengers cyclic adenosine monophosphate (cAMP) and/or cyclic guanosine monophosphate (cGMP) throughout the body and brain. Altered cyclic nucleotide-mediated signalling has been associated with a wide array of disorders, including neurodegenerative disorders. Recently, PDE5 has been shown to be involved in neurodegenerative disorders such as Alzheimer’s disease, but its precise role has not been elucidated yet. To visualize and quantify the expression of this enzyme in brain, we developed a radiotracer for specific PET imaging of PDE5. A quinoline-based lead compound has been structurally modified resulting in the fluoroethoxymethyl derivative ICF24027 with high inhibitory activity towards PDE5 (IC50 = 1.86 nM). Radiolabelling with fluorine-18 was performed by a one-step nucleophilic substitution reaction using a tosylate precursor (RCY(EOB) = 12.9% ± 1.8%; RCP > 99%; SA(EOS) = 70–126 GBq/μmol). In vitro autoradiographic studies of [18F]ICF24027 on different mouse tissue as well as on porcine brain slices demonstrated a moderate specific binding to PDE5. In vivo studies in mice revealed that [18F]ICF24027 was metabolized under formation of brain penetrable radiometabolites making the radiotracer unsuitable for PET imaging of PDE5 in brain. PMID:27110797

  6. BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition and P2Y12 antagonism.

    PubMed

    Zhang, Si; Hu, Liang; Du, Hongguang; Guo, Yan; Zhang, Yan; Niu, Haixia; Jin, Jianguo; Zhang, Jian; Liu, Junling; Zhang, Xiaohui; Kunapuli, Satya P; Ding, Zhongren

    2010-10-01

    Though antiplatelet drugs are proven beneficial to patients with coronary heart disease and stroke, more effective and safer antiplatelet drugs are still needed. In this study we report the antiplatelet effects and mechanism of BF0801, a novel adenine derivative. BF0801 dramatically inhibited platelet aggregation and ATP release induced by ADP, 2MeSADP, AYPGKF, SFLLRN or convulxin without affecting shape change in vitro . It also potentiated the inhibitory effects of adenosine-based P2Y12 antagonist AR-C69931MX or phosphodiesterase (PDE) inhibitor IBMX on platelet aggregation. The cAMP levels in both resting and forskolin-stimulated platelets were increased by BF0801 suggesting its PDE inhibitor activity, which is further confirmed by the concentration-dependent suppression of BF0801 on the native and recombinant PDE. Similar to AR-C69931MX, BF0801 drastically inhibited 2MeSADP- induced adenylyl cyclase inhibition in platelets indicating its P2Y12 antagonism activity, which is substantiated by the inhibition of BF0801 on the interaction between ADP and P2Y12 receptor expressed in CHO-K1 cells measured by atomic force microscopy. Moreover, we confirmed the antiplatelet effects of BF0801 using platelets from rats intravenously given BF0801. In summary, for the first time we developed a novel adenine derivative bearing dual activities of PDE inhibition and P2Y12 antagonism, which may have therapeutic advantage as a potential antithrombotic drug. PMID:20806121

  7. RNA interference-mediated knockdown of long-form phosphodiesterase-4D (PDE4D) enzyme reverses amyloid-β42-induced memory deficits in mice.

    PubMed

    Zhang, Cong; Cheng, Yufang; Wang, Haitao; Wang, Chuang; Wilson, Steven P; Xu, Jiangping; Zhang, Han-Ting

    2014-01-01

    Phosphodiesterase-4 (PDE4) inhibitors enhance memory, increase hippocampal neurogenesis, and reverse amyloid-β (Aβ)-induced memory deficits. Here, we examined whether long-form PDE4D knockdown by lentiviral RNA construct containing a specific microRNA/miRNA-mir hairpin structure (4DmiRNA) reversed memory impairment caused by amyloid-β1-42 (Aβ42) in mice using the Morris water maze (MWM) and novelty object recognition tests. Western blotting analysis was used to assess protein levels of cAMP response element-binding protein (CREB, unphosphorylated and phosphorylated [pCREB]), brain-derived neurotrophic factor (BDNF), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and nuclear factor-κB (NF-κB) to explore the neurochemical mechanisms. Aggregated Aβ42 (0.5 μg/side) bilaterally infused in dentate gyrus decreased cAMP levels (p < 0.01) and produced memory deficits in the MWM (p < 0.01) and object recognition tests (p < 0.01). Microinfusions of lentiviruses resulted in downregulated expression of PDE4D4 and 4D5 proteins and reversed Aβ42-induced cAMP decline (p < 0.05) and memory deficits. Treatment also concomitantly increased pCREB (p < 0.05) and BDNF (p < 0.01) and reduced IL-1β (p < 0.05), TNF-α (p < 0.01), and NF-κB (p65) (p < 0.05) in the hippocampus of Aβ42-challenged mice. These results suggest that long-form PDE4D knockdown may offer a promising treatment for memory loss associated with Alzheimer's disease.

  8. Auphen and dibutyryl cAMP suppress growth of hepatocellular carcinoma by regulating expression of aquaporins 3 and 9 in vivo

    PubMed Central

    Peng, Rui; Zhao, Guang-Xi; Li, Jing; Zhang, Yu; Shen, Xi-Zhong; Wang, Ji-Yao; Sun, Jian-Yong

    2016-01-01

    AIM: To investigate whether the regulation of aquaporin 3 (AQP3) and AQP9 induced by Auphen and dibutyryl cAMP (dbcAMP) inhibits hepatic tumorigenesis. METHODS: Expression of AQP3 and AQP9 was detected by Western blot, immunohistochemistry (IHC), and RT-PCR in HCC samples and paired non-cancerous liver tissue samples from 30 hepatocellular carcinoma (HCC) patients. A xenograft tumor model was used in vivo. Nine nude mice were divided into control, Auphen-treated, and dbcAMP-treated groups (n = 3 for each group). AQP3 and AQP9 protein expression after induction of xenograft tumors was detected by IHC and mRNA by RT-PCR analysis. The terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay and histological evaluation were used to detect apoptosis of tumor cells, and the concentration of serum α-fetoprotein (AFP) was measured using RT-PCR and an ELISA kit. RESULTS: The volumes and weights of tumors decreased significantly in the Auphen- and dbcAMP-treated mice compared with the control mice (P < 0.01). The levels of AQP3 were significantly lower in the Auphen treatment group, and levels of AQP9 were significantly higher in thedbcAMP treatment mice than in the control mice (P < 0.01). The reduction of AQP3 by Auphen and increase of AQP9 by dbcAMP in nude mice suppressed tumor growth of HCC, which resulted in reduced AFP levels in serum and tissues, and apoptosis of tumor cells in the Auphen- and dbcAMP-treated mice, when compared with control mice (P < 0.01). Compared with para-carcinoma tissues, AQP3 expression increased in tumor tissues whereas the expression of AQP9 decreased. By correlating clinicopathological and expression levels, we demonstrated that the expression of AQP3 and AQP9 was correlated with clinical progression of HCC and disease outcomes. CONCLUSION: AQP3 increases in HCC while AQP9 decreases. Regulation of AQP3 and AQP9 expression by Auphen and dbcAMP inhibits the development and growth of HCC. PMID:27022216

  9. Cyclooxygenase-2 expression in lipopolysaccharide-stimulated human monocytes is modulated by cyclic AMP, prostaglandin E(2), and nonsteroidal anti-inflammatory drugs.

    PubMed

    Hinz, B; Brune, K; Pahl, A

    2000-11-30

    Using human blood monocytes (for determination of cyclooxygenase-2 (COX-2) mRNA by RT-PCR) and human whole blood (for prostanoid determination), the present study investigates the influence of the second messenger cAMP on lipopolysaccharide (LPS)-induced COX-2 expression with particular emphasis on the role of prostaglandin E(2) (PGE(2)) in this process. Elevation of intracellular cAMP with a cell-permeable cAMP analogue (dibutyryl cAMP), an adenylyl cyclase activator (cholera toxin), or a phosphodiesterase inhibitor (3-isobutyl-1-methylxanthine) substantially enhanced LPS-induced PGE(2) formation and COX-2 mRNA expression, but did not modify COX-2 enzyme activity. Moreover, up-regulation of LPS-induced COX-2 expression was caused by PGE(2), butaprost (selective agonist of the adenylyl cyclase-coupled EP(2) receptor) and 11-deoxy PGE(1) (EP(2)/EP(4) agonist), whereas sulprostone (EP(3)/EP(1) agonist) left COX-2 expression unaltered. Abrogation of LPS-induced PGE(2) synthesis with the selective COX-2 inhibitor NS-398 caused a decrease in COX-2 mRNA levels that was restored by exogenous PGE(2) and mimicked by S(+)-flurbiprofen and ketoprofen. Overall, these results indicate a modulatory role of cAMP in the regulation of COX-2 expression. PGE(2), a cAMP-elevating final product of the COX-2 pathway, may autoregulate COX-2 expression in human monocytes via a positive feedback mechanism.

  10. Inhibition of Phosphodiesterase 10A Increases the Responsiveness of Striatal Projection Neurons to Cortical Stimulation.

    PubMed

    Threlfell, Sarah; Sammut, Stephen; Menniti, Frank S; Schmidt, Christopher J; West, Anthony R

    2009-03-01

    The cyclic nucleotide phosphodiesterase 10A (PDE10A) is highly expressed in striatal medium-sized spiny projection neurons (MSNs), apparently playing a critical role in the regulation of both cGMP and cAMP signaling cascades. Genetic disruption or pharmacological inhibition of PDE10A reverses behavioral abnormalities associated with subcortical hyperdopaminergia. Here, we investigate the effect of PDE10A inhibition on the activity of MSNs using single-unit extracellular recordings performed in the dorsal striatum of anesthetized rats. Antidromic stimulation of the substantia nigra pars reticulata was used to identify striatonigral (SNr+) MSNs. Intrastriatal infusion of the selective PDE10A inhibitors papaverine or TP-10 [2-{4-[-pyridin-4-yl-1-(2,2,2-trifluoroethyl)-1H-pyrazol-3-yl]-phenoxymethyl}-quinoline succinic acid] by reverse microdialysis did not affect spontaneous firing but robustly increased measures of cortically evoked spike activity in a stimulus intensity-dependent manner. Systemic administration of TP-10 also increased cortically evoked spike activity in a stimulus intensity- and dose-dependent manner. A robust increase in cortically evoked activity was apparent in SNr- MSNs (primarily striatopallidal). It is interesting that TP-10 administration did not affect cortically evoked activity in SNr+ MSNs. However, TP-10 administration increased the incidence of antidromically activated (i.e., SNr+) MSNs. These findings indicate that inhibition of striatal PDE10A activity increases the responsiveness of MSNs to depolarizing stimuli. Furthermore, given the lack of effect of TP-10 on SNr+ MSNs, we speculate that PDE10A inhibition may have a greater facilitatory effect on corticostriatal synaptic activity in striatopallidal MSNs. These data support further investigation of selective targeting of PDE signaling pathways in MSN subpopulations because this may represent a promising novel approach for treating brain disorders involving dysfunctional glutamatergic

  11. Erythroid pyrimidine 5'-nucleotidase: cloning, developmental expression, and regulation by cAMP and in vivo hypoxia.

    PubMed

    Mass, Markus; Simo, Erika; Dragon, Stefanie

    2003-12-01

    A characteristic process of terminal erythroid differentiation is the degradation of ribosomal RNA into mononucleotides. The pyrimidine mononucleotides can be dephosphorylated by pyrimidine 5'-nucleotidase (P5N-I). In humans, a lack of this enzyme causes hemolytic anemia with ribosomal structures and trinucleotides retained in the red blood cells (RBCs). Although the protein/nucleotide sequence of P5N-I is known in mammals, the onset and regulation of P5N-I during erythroid maturation is unknown. However, in circulating chicken embryonic RBCs, the enzyme is induced together with carbonic anhydrase (CAII) and 2,3-bisphosphoglycerate (2,3-BPG) by norepinephrine (NE) and adenosine, which are released by the embryo under hypoxic conditions. Here, we present the chicken P5N-I sequence and the gene expression of P5N-I during RBC maturation; the profile of gene expression follows the enzyme activity with a rise between days 13 and 16 of embryonic development. The p5n-I expression is induced (1) in definitive but not primitive RBCs by stimulation of beta-adrenergic/adenosine receptors, and (2) in definitive RBCs by hypoxic incubation of the chicken embryo. Since embryonic RBCs increase their hemoglobin-oxygen affinity by degradation of nucleotides such as uridine triphosphate (UTP) and cytidine triphosphate (CTP), the induction of p5n-I expression can be seen as an adaptive response to hypoxia.

  12. Marketing for Camp Trends.

    ERIC Educational Resources Information Center

    Biddle, Alicia

    1998-01-01

    To effectively market a camp, current trends and issues must be considered: specialty programming, the Americans With Disabilities Act, competing recreational programs, changes in the school year, programming for seniors, and accountability. Camps should have a marketing strategy that includes public relations, a marketing plan, a pricing…

  13. Friends' Discovery Camp

    ERIC Educational Resources Information Center

    Seymour, Seth

    2008-01-01

    This article features Friends' Discovery Camp, a program that allows children with and without autism spectrum disorder to learn and play together. In Friends' Discovery Camp, campers take part in sensory-rich experiences, ranging from hands-on activities and performing arts to science experiments and stories teaching social skills. Now in its 7th…

  14. Camp's "Disneyland" Effect.

    ERIC Educational Resources Information Center

    Renville, Gary

    1999-01-01

    Describes the positive mental, physical, and social growth impacts that the camping experience had on the author, and urges camp program evaluation to plan and implement such changes. Sidebar lists steps of effective evaluation: program goals and objectives, goals of evaluation, implementation of evaluation, data analysis, and findings and…

  15. p54nrb/NONO Regulates Cyclic AMP-Dependent Glucocorticoid Production by Modulating Phosphodiesterase mRNA Splicing and Degradation

    PubMed Central

    Lu, Jia Yang

    2015-01-01

    Glucocorticoid production in the adrenal cortex is activated in response to an increase in cyclic AMP (cAMP) signaling. The nuclear protein p54nrb/NONO belongs to the Drosophila behavior/human splicing (DBHS) family and has been implicated in several nuclear processes, including transcription, splicing, and RNA export. We previously identified p54nrb/NONO as a component of a protein complex that regulates the transcription of CYP17A1, a gene required for glucocorticoid production. Based on the multiple mechanisms by which p54nrb/NONO has been shown to control gene expression and the ability of the protein to be recruited to the CYP17A1 promoter, we sought to further define the molecular mechanism by which p54nrb/NONO confers optimal cortisol production. We show here that silencing p54nrb/NONO expression in H295R human adrenocortical cells decreases the ability of the cells to increase intracellular cAMP production and subsequent cortisol biosynthesis in response to adrenocorticotropin hormone (ACTH) stimulation. Interestingly, the expression of multiple phosphodiesterase (PDE) isoforms, including PDE2A, PDE3A, PDE3B, PDE4A, PDE4D, and PDE11A, was induced in p54nrb/NONO knockdown cells. Investigation of the mechanism by which silencing of p54nrb/NONO led to increased expression of select PDE isoforms revealed that p54nrb/NONO regulates the splicing of a subset of PDE isoforms. Importantly, we also identify a role for p54nrb/NONO in regulating the stability of PDE transcripts by facilitating the interaction between the exoribonuclease XRN2 and select PDE transcripts. In summary, we report that p54nrb/NONO modulates cAMP-dependent signaling, and ultimately cAMP-stimulated glucocorticoid biosynthesis by regulating the splicing and degradation of PDE transcripts. PMID:25605330

  16. cAMP signalling in trypanosomatids: role in pathogenesis and as a drug target

    PubMed Central

    Makin, Laura; Gluenz, Eva

    2015-01-01

    Despite recent research linking cAMP signalling to virulence in trypanosomatids and detailed studies of trypanosomatid adenylyl cyclases (ACs) and phosphodiesterases (PDEs) since their discoveries 40 years ago, downstream components of the pathway and their biological functions have remained remarkably elusive. However, in recent years, significant discoveries have been made: a role for parasite ACs has been proposed in cytokinesis, evasion of the host immune response, and social motility. cAMP phosphodiesterases PDEB1 and PDEB2 were found to be essential for survival and virulence of Trypanosoma brucei and, in Trypanosoma cruzi, PDEC2 was shown to be required for normal osmoregulation. As we discuss here, these breakthroughs have led to an ongoing surge in the development of PDE inhibitors as lead compounds for trypanocidal drugs. PMID:26004537

  17. cAMP signalling in trypanosomatids: role in pathogenesis and as a drug target.

    PubMed

    Makin, Laura; Gluenz, Eva

    2015-08-01

    Despite recent research linking cAMP signalling to virulence in trypanosomatids and detailed studies of trypanosomatid adenylyl cyclases (ACs) and phosphodiesterases (PDEs) since their discoveries 40 years ago, downstream components of the pathway and their biological functions have remained remarkably elusive. However, in recent years, significant discoveries have been made: a role for parasite ACs has been proposed in cytokinesis, evasion of the host immune response, and social motility. cAMP phosphodiesterases PDEB1 and PDEB2 were found to be essential for survival and virulence of Trypanosoma brucei and, in Trypanosoma cruzi, PDEC2 was shown to be required for normal osmoregulation. As we discuss here, these breakthroughs have led to an ongoing surge in the development of PDE inhibitors as lead compounds for trypanocidal drugs. PMID:26004537

  18. Phosphodiesterase 11A (PDE11A) Gene Defects in Patients with ACTH-Independent Macronodular Adrenal Hyperplasia (AIMAH): Functional Variants May Contribute to Genetic Susceptibility of Bilateral Adrenal Tumors

    PubMed Central

    Vezzosi, Delphine; Libé, Rossella; Baudry, Camille; Rizk-Rabin, Marthe; Horvath, Anelia; Levy, Isaac; René-Corail, Fernande; Ragazzon, Bruno; Stratakis, Constantine A.; Vandecasteele, Grégoire

    2012-01-01

    Context: Phosphodiesterases (PDEs) are key regulatory enzymes of intracellular cAMP levels. PDE11A function has been linked to predisposition to adrenocortical tumors. Objective: The aim of the study was to study the PDE11A gene in a large cohort of patients with ACTH-independent macronodular adrenal hyperplasia (AIMAH) and in control subjects. Design: The PDE11A entire coding region was sequenced in 46 patients with AIMAH and 192 controls. Two variants found in AIMAH patients were transiently expressed in HEK 293 and adrenocortical H295R cells for further functional studies. Results: The frequency of all PDE11A variants was significantly higher among patients with AIMAH (28%) compared to controls (7.2%) (P = 5 × 10−5). Transfection of the two PDE11A variants found in AIMAH patients only (D609N or M878V) showed that cAMP levels were higher, after forskolin stimulation, in cells transfected with the PDE11A mutants, compared to cells transfected with the wild-type PDE11A in HEK 293 cells (P < 0.05). Moreover, transfection with mutants PDE11A increased transcriptional activity of a cAMP-response element reporter construct compared to wild-type PDE11A in HEK 293 cells (P < 0.0004 for D609N and P < 0.003 for M878V) and in the adrenocortical H295R cells (P < 0.05 for D609N and M878V). In addition, analysis of cAMP levels in intact living culture cells by fluorescence resonance energy transfer probes showed increased cAMP in forskolin-treated cells transfected with PDE11A variants compared with wild-type PDE11A (P < 0.05). Conclusion: We conclude that PDE11A genetic variants may increase predisposition to AIMAH. PMID:22996146

  19. Phosphodiesterase MoPdeH targets MoMck1 of the conserved mitogen-activated protein (MAP) kinase signalling pathway to regulate cell wall integrity in rice blast fungus Magnaporthe oryzae.

    PubMed

    Yin, Ziyi; Tang, Wei; Wang, Jingzhen; Liu, Xinyu; Yang, Lina; Gao, Chuyun; Zhang, Jinlong; Zhang, Haifeng; Zheng, Xiaobo; Wang, Ping; Zhang, Zhengguang

    2016-06-01

    In the rice blast fungus Magnaporthe oryzae, the high-affinity cyclic adenosine monophosphate (cAMP) phosphodiesterase MoPdeH is important not only for cAMP signalling and pathogenicity, but also for cell wall integrity (CWI) maintenance through an unknown mechanism. By utilizing affinity purification, we found that MoPdeH interacts with MoMck1, one of the components of the mitogen-activated protein (MAP) kinase cascade that regulates CWI. Overexpression of MoMCK1 suppressed defects in autolysis and pathogenicity of the ΔMopdeH mutant, although partially, suggesting that MoPdeH plays a critical role in CWI maintenance mediated by the MAP kinase pathway. We found that MoMck1 and two other MAP kinase cascade components, MoMkk1 and MoMps1, modulate intracellular cAMP levels by regulating the expression of MoPDEH through a feedback loop. In addition, disruption of MoMKK1 resulted in less aerial hyphal formation, defective asexual development and attenuated pathogenicity. Moreover, MoMkk1 plays a role in the response to osmotic stress via regulation of MoOsm1 phosphorylation levels, whereas endoplasmic reticulum (ER) stress enhances MoMps1 phosphorylation and loss of the MAP kinase cascade component affects the unfolded protein response (UPR) pathway. Taken together, our findings demonstrate that MoPdeH functions upstream of the MoMck1-MoMkk1-MoMps1 MAP kinase pathway to regulate CWI, and that MoPdeH also mediates crosstalk between the cAMP signalling pathway, the osmotic sensing high osmolarity glycerol (HOG) pathway and the dithiothreitol (DTT)-induced UPR pathway in M. oryzae.

  20. Phosphodiesterase MoPdeH targets MoMck1 of the conserved mitogen-activated protein (MAP) kinase signalling pathway to regulate cell wall integrity in rice blast fungus Magnaporthe oryzae.

    PubMed

    Yin, Ziyi; Tang, Wei; Wang, Jingzhen; Liu, Xinyu; Yang, Lina; Gao, Chuyun; Zhang, Jinlong; Zhang, Haifeng; Zheng, Xiaobo; Wang, Ping; Zhang, Zhengguang

    2016-06-01

    In the rice blast fungus Magnaporthe oryzae, the high-affinity cyclic adenosine monophosphate (cAMP) phosphodiesterase MoPdeH is important not only for cAMP signalling and pathogenicity, but also for cell wall integrity (CWI) maintenance through an unknown mechanism. By utilizing affinity purification, we found that MoPdeH interacts with MoMck1, one of the components of the mitogen-activated protein (MAP) kinase cascade that regulates CWI. Overexpression of MoMCK1 suppressed defects in autolysis and pathogenicity of the ΔMopdeH mutant, although partially, suggesting that MoPdeH plays a critical role in CWI maintenance mediated by the MAP kinase pathway. We found that MoMck1 and two other MAP kinase cascade components, MoMkk1 and MoMps1, modulate intracellular cAMP levels by regulating the expression of MoPDEH through a feedback loop. In addition, disruption of MoMKK1 resulted in less aerial hyphal formation, defective asexual development and attenuated pathogenicity. Moreover, MoMkk1 plays a role in the response to osmotic stress via regulation of MoOsm1 phosphorylation levels, whereas endoplasmic reticulum (ER) stress enhances MoMps1 phosphorylation and loss of the MAP kinase cascade component affects the unfolded protein response (UPR) pathway. Taken together, our findings demonstrate that MoPdeH functions upstream of the MoMck1-MoMkk1-MoMps1 MAP kinase pathway to regulate CWI, and that MoPdeH also mediates crosstalk between the cAMP signalling pathway, the osmotic sensing high osmolarity glycerol (HOG) pathway and the dithiothreitol (DTT)-induced UPR pathway in M. oryzae. PMID:27193947

  1. Eukaryotic-Type Ser/Thr Protein Kinase Mediated Phosphorylation of Mycobacterial Phosphodiesterase Affects its Localization to the Cell Wall.

    PubMed

    Malhotra, Neha; Chakraborti, Pradip K

    2016-01-01

    Phosphodiesterase enzymes, involved in cAMP hydrolysis reaction, are present throughout phylogeny and their phosphorylation mediated regulation remains elusive in prokaryotes. In this context, we focused on this enzyme from Mycobacterium tuberculosis. The gene encoded by Rv0805 was PCR amplified and expressed as a histidine-tagged protein (mPDE) utilizing Escherichia coli based expression system. In kinase assays, upon incubation with mycobacterial Clade I eukaryotic-type Ser/Thr kinases (PknA, PknB, and PknL), Ni-NTA purified mPDE protein exhibited transphosphorylation ability albeit with varying degree. When mPDE was co-expressed one at a time with these kinases in E. coli, it was also recognized by an anti-phosphothreonine antibody, which further indicates its phosphorylating ability. Mass spectrometric analysis identified Thr-309 of mPDE as a phosphosite. In concordance with this observation, anti-phosphothreonine antibody marginally recognized mPDE-T309A mutant protein; however, such alteration did not affect the enzymatic activity. Interestingly, mPDE expressed in Mycobacterium smegmatis yielded a phosphorylated protein that preferentially localized to cell wall. In contrast, mPDE-T309A, the phosphoablative variant of mPDE, did not show such behavior. On the other hand, phosphomimics of mPDE (T309D or T309E), exhibited similar cell wall anchorage as was observed with the wild-type. Thus, our results provide credence to the fact that eukaryotic-type Ser/Thr kinase mediated phosphorylation of mPDE renders negative charge to the protein, promoting its localization on cell wall. Furthermore, multiple sequence alignment revealed that Thr-309 is conserved among mPDE orthologs of M. tuberculosis complex, which presumably emphasizes evolutionary significance of phosphorylation at this residue.

  2. Eukaryotic-Type Ser/Thr Protein Kinase Mediated Phosphorylation of Mycobacterial Phosphodiesterase Affects its Localization to the Cell Wall

    PubMed Central

    Malhotra, Neha; Chakraborti, Pradip K.

    2016-01-01

    Phosphodiesterase enzymes, involved in cAMP hydrolysis reaction, are present throughout phylogeny and their phosphorylation mediated regulation remains elusive in prokaryotes. In this context, we focused on this enzyme from Mycobacterium tuberculosis. The gene encoded by Rv0805 was PCR amplified and expressed as a histidine-tagged protein (mPDE) utilizing Escherichia coli based expression system. In kinase assays, upon incubation with mycobacterial Clade I eukaryotic-type Ser/Thr kinases (PknA, PknB, and PknL), Ni-NTA purified mPDE protein exhibited transphosphorylation ability albeit with varying degree. When mPDE was co-expressed one at a time with these kinases in E. coli, it was also recognized by an anti-phosphothreonine antibody, which further indicates its phosphorylating ability. Mass spectrometric analysis identified Thr-309 of mPDE as a phosphosite. In concordance with this observation, anti-phosphothreonine antibody marginally recognized mPDE-T309A mutant protein; however, such alteration did not affect the enzymatic activity. Interestingly, mPDE expressed in Mycobacterium smegmatis yielded a phosphorylated protein that preferentially localized to cell wall. In contrast, mPDE-T309A, the phosphoablative variant of mPDE, did not show such behavior. On the other hand, phosphomimics of mPDE (T309D or T309E), exhibited similar cell wall anchorage as was observed with the wild-type. Thus, our results provide credence to the fact that eukaryotic-type Ser/Thr kinase mediated phosphorylation of mPDE renders negative charge to the protein, promoting its localization on cell wall. Furthermore, multiple sequence alignment revealed that Thr-309 is conserved among mPDE orthologs of M. tuberculosis complex, which presumably emphasizes evolutionary significance of phosphorylation at this residue. PMID:26904001

  3. cAMP enhances BMP2-signaling through PKA and MKP1-dependent mechanisms

    SciTech Connect

    Ghayor, Chafik; Ehrbar, Martin; Miguel, Blanca San; Graetz, Klaus W.; Weber, Franz E.

    2009-04-03

    Recent studies suggest that the elevation of intracellular cyclic adenosine monophosphate (cAMP) and the activation of the protein kinase A regulate BMP-induced osteogenesis. However, the precise mechanisms underlying the enhancing effect of cAMP on BMP2 signaling were not completely revealed. In this study we investigated the effect of elevated cAMP level and PKA activation on the BMP2-induced osteoblastic differentiation in pluripotent C2C12 cells. Alkaline phosphatase activity and its mRNA were consistently induced by BMP2 treatment. The pretreatment of C2C12 cells with Forskolin, a cAMP generating agent, dbcAMP, an analogue of cAMP, or IBMX (3-isobutyl 1-methyl xanthine), and a nonspecific inhibitor of phosphodiesterases elicited further activation of alkaline phosphatase. Furthermore, elevated intracellular cAMP level increased BMP2-induced MKP1. On the other hand, BMP2-induced Erk phosphorylation (p44/p42) and cell proliferation were suppressed in the presence of cAMP. Thus, cAMP might enhance BMP2-induced osteoblastic differentiation by a MKP1-Erk-dependent mechanism.

  4. Are diabetes camps effective?

    PubMed

    Barone, Mark Thomaz Ugliara; Vivolo, Marco Antonio; Madden, Paul B

    2016-04-01

    In the present article data about Diabetes Camps (DC) from all continents were reviewed in order to answer the title question "are diabetes camps effective?". Articles from peer reviewed journals and abstracts published in international conferences proceedings were raised. The effectiveness was considered in terms of knowledge acquisition, and psychosocial and physiological changes. Even though expected improvements were not found in all studies, in a deeper and wider analysis the aspects that influence the most toward gains are identified. Among them are: number of participations in a DC, post-camp educational opportunities, staff training, and program oriented toward campers' autonomy. To conclude, practical recommendations are addressed intending to amplify DC's potential.

  5. Camp Invention Connects to Classrooms.

    ERIC Educational Resources Information Center

    Krebs, Danute V.; Clark, Barbara D.

    2000-01-01

    This article describes Camp Invention, a national creativity day camp that integrates science, math, social studies, and the arts. The one week camp for children entering grades 2-6 attracts many academically gifted children because of its hands-on curriculum. The camp's curriculum and activities are discussed. (Contains two references.) (CR)

  6. Pre-Camp Checklists.

    ERIC Educational Resources Information Center

    Camping Magazine, 1995

    1995-01-01

    Provides checklists to prepare for camp opening. Covers such areas as health histories of all campers and staff, staff credentials, condition of facilities and equipment, staff training, horse stables and equipment, and safety of swimming areas. (LP)

  7. Hitler's Death Camps.

    ERIC Educational Resources Information Center

    Wieser, Paul

    1995-01-01

    Presents a high school lesson on Hitler's death camps and the widespread policy of brutality and oppression against European Jews. Includes student objectives, instructional procedures, and a chart listing the value of used clothing taken from the Jews. (CFR)

  8. Camping in the Snow.

    ERIC Educational Resources Information Center

    Brown, Constance

    1979-01-01

    Describes the experience of winter snow camping. Provides suggestions for shelter, snow kitchens, fires and stoves, cooking, latrines, sleeping warm, dehydration prevention, and clothing. Illustrated with full color photographs. (MA)

  9. A role for phosphodiesterase 3B in acquisition of brown fat characteristics by white adipose tissue in male mice.

    PubMed

    Guirguis, Emilia; Hockman, Steven; Chung, Youn Wook; Ahmad, Faiyaz; Gavrilova, Oksana; Raghavachari, Nalini; Yang, Yanqin; Niu, Gang; Chen, Xiaoyuan; Yu, Zu Xi; Liu, Shiwei; Degerman, Eva; Manganiello, Vincent

    2013-09-01

    Obesity is linked to various diseases, including insulin resistance, diabetes, and cardiovascular disorders. The idea of inducing white adipose tissue (WAT) to assume characteristics of brown adipose tissue (BAT), and thus gearing it to fat burning instead of storage, is receiving serious consideration as potential treatment for obesity and related disorders. Phosphodiesterase 3B (PDE3B) links insulin- and cAMP-signaling networks in tissues associated with energy metabolism, including WAT. We used C57BL/6 PDE3B knockout (KO) mice to elucidate mechanisms involved in the formation of BAT in epididymal WAT (EWAT) depots. Examination of gene expression profiles in PDE3B KO EWAT revealed increased expression of several genes that block white and promote brown adipogenesis, such as C-terminal binding protein, bone morphogenetic protein 7, and PR domain containing 16, but a clear BAT-like phenotype was not completely induced. However, acute treatment of PDE3B KO mice with the β3-adrenergic agonist, CL316243, markedly increased the expression of cyclooxygenase-2, which catalyzes prostaglandin synthesis and is thought to be important in the formation of BAT in WAT and the elongation of very long-chain fatty acids 3, which is linked to BAT recruitment upon cold exposure, causing a clear shift toward fat burning and the induction of BAT in KO EWAT. These data provide insight into the mechanisms of BAT formation in mouse EWAT, suggesting that, in a C57BL/6 background, an increase in cAMP, caused by ablation of PDE3B and administration of CL316243, may promote differentiation of prostaglandin-responsive progenitor cells in the EWAT stromal vascular fraction into functional brown adipocytes.

  10. Theophylline Represses IL-8 Secretion from Airway Smooth Muscle Cells Independently of Phosphodiesterase Inhibition. Novel Role as a Protein Phosphatase 2A Activator.

    PubMed

    Patel, Brijeshkumar S; Rahman, Md Mostafizur; Rumzhum, Nowshin N; Oliver, Brian G; Verrills, Nicole M; Ammit, Alaina J

    2016-06-01

    Theophylline is an old drug experiencing a renaissance owing to its beneficial antiinflammatory effects in chronic respiratory diseases, such as asthma and chronic obstructive pulmonary disease. Multiple modes of antiinflammatory action have been reported, including inhibition of the enzymes that degrade cAMP-phosphodiesterase (PDE). Using primary cultures of airway smooth muscle (ASM) cells, we recently revealed that PDE4 inhibitors can potentiate the antiinflammatory action of β2-agonists by augmenting cAMP-dependent expression of the phosphatase that deactivates mitogen-activated protein kinase (MAPK)-MAPK phosphatase (MKP)-1. Therefore, the aim of this study was to address whether theophylline repressed cytokine production in a similar, PDE-dependent, MKP-1-mediated manner. Notably, theophylline did not potentiate cAMP release from ASM cells treated with the long-acting β2-agonist formoterol. Moreover, theophylline (0.1-10 μM) did not increase formoterol-induced MKP-1 messenger RNA expression nor protein up-regulation, consistent with the lack of cAMP generation. However, theophylline (at 10 μM) was antiinflammatory and repressed secretion of the neutrophil chemoattractant cytokine IL-8, which is produced in response to TNF-α. Because theophylline's effects were independent of PDE4 inhibition or antiinflammatory MKP-1, we then wished to elucidate the novel mechanisms responsible. We investigated the impact of theophylline on protein phosphatase (PP) 2A, a master controller of multiple inflammatory signaling pathways, and show that theophylline increases TNF-α-induced PP2A activity in ASM cells. Confirmatory results were obtained in A549 lung epithelial cells. PP2A activators have beneficial effects in ex vivo and in vivo models of respiratory disease. Thus, our study is the first to link theophylline with PP2A activation as a novel mechanism to control respiratory inflammation.

  11. Inactivation of oncogenic cAMP-specific phosphodiesterase 4D by miR-139-5p in response to p53 activation.

    PubMed

    Cao, Bo; Wang, Kebing; Liao, Jun-Ming; Zhou, Xiang; Liao, Peng; Zeng, Shelya X; He, Meifang; Chen, Lianzhou; He, Yulong; Li, Wen; Lu, Hua

    2016-01-01

    Increasing evidence highlights the important roles of microRNAs in mediating p53's tumor suppression functions. Here, we report miR-139-5p as another new p53 microRNA target. p53 induced the transcription of miR-139-5p, which in turn suppressed the protein levels of phosphodiesterase 4D (PDE4D), an oncogenic protein involved in multiple tumor promoting processes. Knockdown of p53 reversed these effects. Also, overexpression of miR-139-5p decreased PDE4D levels and increased cellular cAMP levels, leading to BIM-mediated cell growth arrest. Furthermore, our analysis of human colorectal tumor specimens revealed significant inverse correlation between the expression of miR-139-5p and that of PDE4D. Finally, overexpression of miR-139-5p suppressed the growth of xenograft tumors, accompanied by decrease in PDE4D and increase in BIM. These results demonstrate that p53 inactivates oncogenic PDE4D by inducing the expression of miR-139-5p. PMID:27383270

  12. Methoxychlor and its metabolite HPTE inhibit cAMP production and expression of estrogen receptors α and β in the rat granulosa cell in vitro.

    PubMed

    Harvey, Craig N; Chen, Joseph C; Bagnell, Carol A; Uzumcu, Mehmet

    2015-01-01

    The major metabolite of the estrogenic pesticide methoxychlor (MXC) HPTE is a stronger ESR1 agonist than MXC and acts also as an ESR2 antagonist. In granulosa cells (GCs), FSH stimulates estradiol via the second messenger cAMP. HPTE inhibits estradiol biosynthesis, and this effect is greater in FSH-treated GCs than in cAMP-treated GCs. Therefore; we examined the effect of MXC/HPTE on FSH-stimulated cAMP production in cultured GCs. To test involvement of ESR-signaling, we used the ESR1 and ESR2 antagonist ICI 182,780, ESR2 selective antagonist PHTPP, and ESR2 selective agonist DPN. ESR1 and ESR2 mRNA and protein levels were quantified. Both HPTE and MXC inhibited the FSH-induced cAMP production. ICI 182,780 and PHTPP mimicked the inhibitory action of HPTE. MXC/HPTE reduced FSH-stimulated Esr2 mRNA and protein to basal levels. MXC/HPTE also inhibited FSH-stimulated Esr1. The greater inhibition on FSH-stimulated GCs is likely due to reduced cAMP level that involves ESR-signaling, through ESR2.

  13. Methoxychlor and Its Metabolite HPTE Inhibit cAMP Production and Expression of Estrogen Receptors α and β in the Rat Granulosa Cell In Vitro

    PubMed Central

    Harvey, Craig N.; Chen, Joseph C.; Bagnell, Carol A.; Uzumcu, Mehmet

    2015-01-01

    The major metabolite of the estrogenic pesticide methoxychlor (MXC) HPTE is a stronger ESR1 agonist than MXC and acts also as an ESR2 antagonist. In granulosa cells (GCs), FSH stimulates estradiol via the second messenger cAMP. HPTE inhibits estradiol biosynthesis, and this effect is greater in FSH-treated GCs than in cAMP-treated GCs. Therefore; we examined the effect of MXC/HPTE on FSH-stimulated cAMP production in cultured GCs. To test involvement of ESR-signaling, we used the ESR1 and ESR2 antagonist ICI 182,780, ESR2 selective antagonist PHTPP, and ESR2 selective agonist DPN. ESR1 and ESR2 mRNA and protein levels were quantified. Both HPTE and MXC inhibited the FSH-induced cAMP production. ICI 182,780 and PHTPP mimicked the inhibitory action of HPTE. MXC/HPTE reduced FSH-stimulated Esr2 mRNA and protein to basal levels. MXC/HPTE also inhibited FSH-stimulated Esr1. The greater inhibition on FSH-stimulated GCs is likely due to reduced cAMP level that involves ESR-signaling, through ESR2. PMID:25549949

  14. Turning Neighbors into Friends: The Los Angeles Camp Experience.

    ERIC Educational Resources Information Center

    Jenya, Judith

    2002-01-01

    A camp for Los Angeles (California) children traumatized by war, community violence, and hatred gives them time to heal and feel safe. The camp's theme of reconciliation and nonviolence is expressed by an all-volunteer staff through community, teamwork, and cooperation. Nature's diversity is used to show how diverse groups can interact, thrive,…

  15. Investigation of the relationship between protein-protein interaction and catalytic activity of a heme-regulated phosphodiesterase from Escherichia coli (Ec DOS) by protein microarray.

    PubMed

    Sasakura, Yukie; Kanda, Katsuhiro; Yoshimura-Suzuki, Tokiko; Matsui, Takuya; Fukuzono, Shinichi; Shimizu, Toru

    2005-07-19

    Ec DOS, a heme-regulated phosphodiesterase from Escherichia coli, is composed of an N-terminal heme-bound PAS domain and a C-terminal phosphodiesterase domain. The heme redox state in the PAS domain regulates Ec DOS phosphodiesterase activity. Interestingly, the isolated heme-bound PAS fragment enhances phosphodiesterase activity of full-length Ec DOS. The enhancement is also regulated by the heme redox state of the isolated PAS domain. In the present study, we used a newly developed protein microarray system to examine the relationship between catalytic activity and the interaction of full-length Ec DOS and the isolated PAS fragment. Adenosine 3',5'-cyclic monophosphate (cAMP), a substrate of the Ec DOS phosphodiesterase, was found to be indispensable for the interaction between Ec DOS and the PAS fragment, and two phosphodiesterase inhibitors, 3-isobutyl-methyl-xanthine and etazolate hydrochloride, hindered the interaction. In addition, an enzyme with a mutation in the putative cAMP-binding sites (H590 and H594) was unable to interact with Ec DOS and lacked enzymatic activity. These results strongly suggest a close relationship between Ec DOS phosphodiesterase activity and interaction with the isolated PAS fragment. Therefore, this study provides insights into the mechanism of how the isolated PAS domain activates Ec DOS, which has important implications for the general role of the isolated PAS domain in cells. Moreover, we found that multiple microscale analyses using the protein microarray system had several advantages over conventional affinity column methods, including the quantity of protein needed, the sensitivity, the variability of immobilized protein, and the time required for the experiment. PMID:16008345

  16. Downregulation of steroidogenic acute regulatory protein (StAR) gene expression by cyclic AMP in cultured Schwann cells.

    PubMed

    Benmessahel, Yasmina; Troadec, Jean-Denis; Cadepond, Françoise; Guennoun, Rachida; Hales, Dale Buchanan; Schumacher, Michael; Groyer, Ghislaine

    2004-02-01

    Steroidogenic acute regulatory protein (StAR) plays a key role in the availability of cholesterol to the inner mitochondrial membrane, where the first step of steroidogenesis, its conversion to pregnenolone, takes place. Here, we demonstrate for the first time that the StAR gene is also expressed in the rat sciatic nerve and in cultured Schwann cells. The addition to the culture medium of the cAMP-elevating agent forskolin or of the cAMP analogue 8Br-cAMP produced a time-course extinction of StAR gene expression. An inverse relationship was demonstrated between StAR gene expression and the intracellular cAMP content. Accordingly, pharmacological inhibition of the activities of Schwann cell adenylyl cyclase or of phosphodiesterase IV resulted in modifications of StAR gene expression. Since StAR gene expression is stimulated by cAMP in classical steroidogenic cells, our work is the first demonstration of a negative regulation of StAR gene by cAMP.

  17. In vivo blockade of α1-adrenergic receptors mitigates stress-disturbed cAMP and cGMP signaling in Leydig cells.

    PubMed

    Stojkov, Natasa J; Baburski, Aleksandar Z; Bjelic, Maja M; Sokanovic, Srdjan J; Mihajlovic, Aleksandar I; Drljaca, Dragana M; Janjic, Marija M; Kostic, Tatjana S; Andric, Silvana A

    2014-01-01

    The molecular mechanism of stress-associated reproductive dysfunction is complex and largely unknown. This study was designed to systematically analyze molecular effects of systemic in vivo blockade of α1-adrenergic receptors (α1-ADRs) on stress-induced disturbance of cAMP/cGMP signaling in testosterone-producing Leydig cells using the following parameters (i) level of circulating stress hormones, LH and testosterone; (ii) level of main molecular markers of Leydig cell functionality (testosterone, Insl3, cAMP); (iii) expression of cAMP signaling (cAMP 'producers'/'effectors'/'removers') and (iv) expression of NO-cGMP signaling (NO-cGMP 'producers'/'effectors'/'removers'). The results showed that oral administration of α1-ADR blocker before stress increased cGMP and diminished stress-reduced cAMP production in Leydig cells. In the same cells, stress-induced effects on cAMP/cGMP signaling pathways elements were changed. Sustained in vivo α1-ADR blockade completely abolished stress-increased transcription of most abundantly expressed phosphodiesterase that remove cAMP (Pde4b) and potentiated stress-increased expression of PRKA, the main stimulator of Leydig cell steroidogenesis. In the same Leydig cells, stress-decreased NOS3 expression was abolished, while stress-increased GUCY1 (cGMP 'producer') and PRKG1 (cGMP 'effector') were potentiated. It is possible that all molecules mentioned could contribute, at least in part, in recovery of Leydig cell testosterone production. Presented data provide new role of α1-ADRs in stress-triggered disturbance of cAMP/cGMP signaling, and new molecular insights into the relationship between stress and mammalian reproduction. Regardless of whether the effects of α1-blocker + stress are direct or indirect, the results are important in terms of human reproductive health and the wide use of α1-ADR antagonists, alone or in combination, to treat post-traumatic stress disorders, hypertension, benign prostatic hyperplasia symptoms and

  18. Modulation of cellular interactions between C3H/10T1/2 cells and their transformed counterparts by phosphodiesterase inhibitors.

    PubMed

    Bertram, J S

    1979-09-01

    It has been demonstrated previously that nontransformed C3H/10T1/2CL8 mouse embryo fibroblasts (10T1/2) can induce a state of reversible growth inhibition in cocultured malignantly transformed mouse fibroblasts and that this inhibition is modulated by serum concentration. The present study suggests that cyclic nucleotides may be implicated in this intercellular communication. The phosphodiesterase inhibitors theophylline, caffeine, and 3-isobutyl-1-methylxanthine (IBX) at concentrations of 10(-3) M, maintained continuously, were all found to inhibit the expression of 3-methylcholanthrene-induced malignant transformation when added 7 days after removal of carcinogen. IBX was the most potent, causing 100% inhibition at 10(-4) M and 70% inhibition at 10(-5) M. This inhibition was partially reversible in the former case and completely reversible in the latter case by removal of drug. Complete inhibition by 10(-4) M IBX was still observed when treatment was delayed 21 days postcarcinogen. In reconstruction experiments, utilizing confluent monolayers of 10T1/2 cells overlaid with transformed cells, IBX caused a dose-dependent inhibition of colony size of the transformed cells. Adenosine cyclic 2':3'-monophosphoric acid (cAMP) and N6,O2'-dibutyryladenosine cyclic 3':5'-monophophoric acid potentiated this response. The presence of non-transformed 10T1/2 cells was required for this effect, since a concentration of IBX (10(-4) M) inhibitory for the growth of transformed cells in mixed cultures was without effect on the growth rate, plating efficiency, or saturation density of pure cultures of 10T1/2 cells or of their transformed counterparts. Conditioned medium removed from IBX-treated 10T1/2 cells was not growth inhibitory for transformed cells, indicating a requirement for cell-cell contact. IBX caused a dose-dependent increase in intracellular cAMP in confluent 10T1/2 cells and a more pronounced increase in cAMP concentration in the culture medium of these cells. The dose

  19. Are diabetes camps effective?

    PubMed

    Barone, Mark Thomaz Ugliara; Vivolo, Marco Antonio; Madden, Paul B

    2016-04-01

    In the present article data about Diabetes Camps (DC) from all continents were reviewed in order to answer the title question "are diabetes camps effective?". Articles from peer reviewed journals and abstracts published in international conferences proceedings were raised. The effectiveness was considered in terms of knowledge acquisition, and psychosocial and physiological changes. Even though expected improvements were not found in all studies, in a deeper and wider analysis the aspects that influence the most toward gains are identified. Among them are: number of participations in a DC, post-camp educational opportunities, staff training, and program oriented toward campers' autonomy. To conclude, practical recommendations are addressed intending to amplify DC's potential. PMID:27103364

  20. A selective phosphodiesterase 3 inhibitor rescues low Po2-induced ATP release from erythrocytes of humans with type 2 diabetes: implication for vascular control

    PubMed Central

    Bowles, Elizabeth A.; Achilleus, David; Stephenson, Alan H.; Ellis, Christopher G.; Ellsworth, Mary L.

    2011-01-01

    Erythrocytes, via release of ATP in areas of low oxygen (O2) tension, are components of a regulatory system for the distribution of perfusion in skeletal muscle ensuring optimal O2 delivery to meet tissue needs. In type 2 diabetes (DM2), there are defects in O2 supply to muscle as well as a failure of erythrocytes to release ATP. The goal of this study was to ascertain if a phosphodiesterase 3 (PDE3) inhibitor, cilostazol, would rescue low O2-induced ATP release from DM2 erythrocytes and, thereby, enable these cells to dilate isolated erythrocyte-perfused skeletal muscle arterioles exposed to decreased extraluminal O2. Erythrocytes were obtained from healthy humans (HH; n = 12) and humans with DM2 (n = 17). We determined that 1) PDE3B is similarly expressed in both groups, 2) mastoparan 7 (Gi activation) stimulates increases in cAMP in HH but not in DM2 erythrocytes, and 3) pretreatment of DM2 erythrocytes with cilostazol resulted in mastoparan 7-induced increases in cAMP not different from those in HH cells. Most importantly, cilostazol restored the ability of DM2 erythrocytes to release ATP in response to low O2. In contrast with perfusion with HH erythrocytes, isolated hamster retractor muscle arterioles perfused with DM2 erythrocytes constricted in response to low extraluminal PO2. However, in the presence of cilostazol (100 μM), DM2 erythrocytes induced vessel dilation not different from that seen with HH erythrocytes. Thus rescue of low O2-induced ATP release from DM2 erythrocytes by cilostazol restored the ability of erythrocytes to participate in the regulation of perfusion distribution in skeletal muscle. PMID:21963837

  1. A Flying Summer Camp

    ERIC Educational Resources Information Center

    Mercurio, Frank X.

    1975-01-01

    Describes a five-day summer camp which provided 12 children, ages 9-14, with a complete flying experience. The training consisted of ground school and one hour actual flying time, including the basics of aircraft control and a flight prepared and executed by the students. (MLH)

  2. Camping and Outdoor Education.

    ERIC Educational Resources Information Center

    Bucher, Charles A.

    Outdoor education has become an integral part of the curriculum in a number of schools across the nation. Outdoor education activities can be readily integrated into physical education, recreation, and adult education programs, as well as science, mathematics, and related fields. Camping and outdoor education should become a part of each child's…

  3. Summer Fish Camp.

    ERIC Educational Resources Information Center

    Remick, Dennis; Pulu, Tupou L.

    The booklet presents a description and illustrates, with photographs, the Eskimo lifestyle and the kinds of activities that occur at a summer fish camp on the Yukon River. Eleven suggested activities are listed for the teacher to present when using the booklet. Activities include studying the map of Alaska; tracing the life cycle of the fish;…

  4. Recycling at Camp.

    ERIC Educational Resources Information Center

    Cummins, William M.

    1988-01-01

    Outlines a Michigan summer camp's efforts to reduce solid waste disposal by recycling cardboard, tin, glass, aluminum, and plastic milk containers. Points out variables affecting the success of such efforts. Discusses Michigan state funding for the development of recycling programs. (SV)

  5. Camp Animal Crackers.

    ERIC Educational Resources Information Center

    McMillan, Erin; And Others

    This guide contains profiles of 16 activities for young children to participate in while attending camp. Each profile contains the theme of the activity, a list of materials needs, a description of the activity itself, and an explanation of the teacher's role in the activity. The activities focus on: (1) songs and dances; (2) dramatic play; (3)…

  6. Saving the Camping Experience.

    ERIC Educational Resources Information Center

    Davies, Jean S.

    1989-01-01

    Discusses developmental encroachment on isolated caves near Pittsford, Vermont. Describes cooperative effort by recreational camps and other agencies to acquire land surrounding cave entrances for conservation. Details successes and hurdles of land acquisition. Describes ongoing work by local campers to maintain and enjoy caves and property. (TES)

  7. Phosphodiesterase 4B in the cardiac L-type Ca²⁺ channel complex regulates Ca²⁺ current and protects against ventricular arrhythmias in mice.

    PubMed

    Leroy, Jérôme; Richter, Wito; Mika, Delphine; Castro, Liliana R V; Abi-Gerges, Aniella; Xie, Moses; Scheitrum, Colleen; Lefebvre, Florence; Schittl, Julia; Mateo, Philippe; Westenbroek, Ruth; Catterall, William A; Charpentier, Flavien; Conti, Marco; Fischmeister, Rodolphe; Vandecasteele, Grégoire

    2011-07-01

    β-Adrenergic receptors (β-ARs) enhance cardiac contractility by increasing cAMP levels and activating PKA. PKA increases Ca²⁺-induced Ca²⁺ release via phosphorylation of L-type Ca²⁺ channels (LTCCs) and ryanodine receptor 2. Multiple cyclic nucleotide phosphodiesterases (PDEs) regulate local cAMP concentration in cardiomyocytes, with PDE4 being predominant for the control of β-AR-dependent cAMP signals. Three genes encoding PDE4 are expressed in mouse heart: Pde4a, Pde4b, and Pde4d. Here we show that both PDE4B and PDE4D are tethered to the LTCC in the mouse heart but that β-AR stimulation of the L-type Ca²⁺ current (ICa,L) is increased only in Pde4b-/- mice. A fraction of PDE4B colocalized with the LTCC along T-tubules in the mouse heart. Under β-AR stimulation, Ca²⁺ transients, cell contraction, and spontaneous Ca²⁺ release events were increased in Pde4b-/- and Pde4d-/- myocytes compared with those in WT myocytes. In vivo, after intraperitoneal injection of isoprenaline, catheter-mediated burst pacing triggered ventricular tachycardia in Pde4b-/- mice but not in WT mice. These results identify PDE4B in the CaV1.2 complex as a critical regulator of ICa,L during β-AR stimulation and suggest that distinct PDE4 subtypes are important for normal regulation of Ca²⁺-induced Ca²⁺ release in cardiomyocytes.

  8. The phosphodiesterase-4 inhibitor rolipram attenuates heroin-seeking behavior induced by cues or heroin priming in rats.

    PubMed

    Lai, Miaojun; Zhu, Huaqiang; Sun, Anna; Zhuang, Dingding; Fu, Dan; Chen, Weisheng; Zhang, Han-Ting; Zhou, Wenhua

    2014-09-01

    Inhibition of phosphodiesterase-4 (PDE4), an enzyme that specifically hydrolyzes cyclic adenosine monophosphate (cAMP) increases intracellular cAMP/cAMP-response element binding protein (CREB) signaling. Activation of this signaling is considered as an important compensatory response that decreases motivational properties of drugs of abuse. However, it is not known whether PDE4 is involved in heroin seeking. Self-administration of heroin (50 μg/kg/infusion) was performed under the fixed ratio 1 (FR1) schedule for 14 d and then drug seeking was extinguished for 10 d. The progressive ratio schedule was used to evaluate the relative motivational value of heroin reinforcement. After training, the conditioned cue or heroin priming (250 μg/kg) was introduced for the reinstatement of heroin-seeking behavior. Pretreatment (i.p.) with rolipram (0.03-0.3 mg/kg), a prototypical, selective PDE4 inhibitor, failed to inhibit heroin self-administration under the FR1 schedule, but decreased the reward values under the progressive ratio schedule in a dose-dependent manner. In addition, rolipram decreased the reinstatement of heroin seeking induced by cues or heroin priming even at the lowest dose (0.03 mg/kg); in contrast, the highest dose (0.3 mg/kg) of rolipram was required to decrease sucrose reinforcement. Finally, the effects of rolipram on heroin-seeking behavior were correlated with the increases in expression of phosphorylated CREB in the nucleus accumbens. The study demonstrated that rolipram inhibited heroin reward and heroin-seeking behavior. The results suggest that PDE4 plays an essential role in mediating heroin seeking and that PDE4 inhibitors may be used as a potential pharmacotherapeutic approach for heroin addiction.

  9. cAMP Regulation of the lactose operon.

    PubMed

    Szeberenyi, Jozsef

    2004-05-01

    Terms to be familiar with before you start to solve the test: lactose operon, adenylate cyclase, cAMP, catabolite activator protein (CAP), expression plasmid, lac operator, lac repressor, lactose, glucose, promoter, cis- and trans-acting factors. PMID:21706723

  10. Multiple Facets of cAMP Signalling and Physiological Impact: cAMP Compartmentalization in the Lung

    PubMed Central

    Oldenburger, Anouk; Maarsingh, Harm; Schmidt, Martina

    2012-01-01

    Therapies involving elevation of the endogenous suppressor cyclic AMP (cAMP) are currently used in the treatment of several chronic inflammatory disorders, including chronic obstructive pulmonary disease (COPD). Characteristics of COPD are airway obstruction, airway inflammation and airway remodelling, processes encompassed by increased airway smooth muscle mass, epithelial changes, goblet cell and submucosal gland hyperplasia. In addition to inflammatory cells, airway smooth muscle cells and (myo)fibroblasts, epithelial cells underpin a variety of key responses in the airways such as inflammatory cytokine release, airway remodelling, mucus hypersecretion and airway barrier function. Cigarette smoke, being next to environmental pollution the main cause of COPD, is believed to cause epithelial hyperpermeability by disrupting the barrier function. Here we will focus on the most recent progress on compartmentalized signalling by cAMP. In addition to G protein-coupled receptors, adenylyl cyclases, cAMP-specific phospho-diesterases (PDEs) maintain compartmentalized cAMP signalling. Intriguingly, spatially discrete cAMP-sensing signalling complexes seem also to involve distinct members of the A-kinase anchoring (AKAP) superfamily and IQ motif containing GTPase activating protein (IQGAPs). In this review, we will highlight the interaction between cAMP and the epithelial barrier to retain proper lung function and to alleviate COPD symptoms and focus on the possible molecular mechanisms involved in this process. Future studies should include the development of cAMP-sensing multiprotein complex specific disruptors and/or stabilizers to orchestrate cellular functions. Compartmentalized cAMP signalling regulates important cellular processes in the lung and may serve as a therapeutic target. PMID:24281338

  11. Spatiotemporal regulation of cAMP signaling controls the human trophoblast fusion

    PubMed Central

    Gerbaud, Pascale; Taskén, Kjetil; Pidoux, Guillaume

    2015-01-01

    During human placentation, mononuclear cytotrophoblasts fuse to form multinucleated syncytia ensuring hormonal production and nutrient exchanges between the maternal and fetal circulation. Syncytial formation is essential for the maintenance of pregnancy and for fetal growth. The cAMP signaling pathway is the major route to trigger trophoblast fusion and its activation results in phosphorylation of specific intracellular target proteins, in transcription of fusogenic genes and assembly of macromolecular protein complexes constituting the fusogenic machinery at the plasma membrane. Specificity in cAMP signaling is ensured by generation of localized pools of cAMP controlled by cAMP phosphodiesterases (PDEs) and by discrete spatial and temporal activation of protein kinase A (PKA) in supramolecular signaling clusters inside the cell organized by A-kinase-anchoring proteins (AKAPs) and by organization of signal termination by protein phosphatases (PPs). Here we present original observations on the available components of the cAMP signaling pathway in the human placenta including PKA, PDE, and PP isoforms as well as AKAPs. We continue to discuss the current knowledge of the spatiotemporal regulation of cAMP signaling triggering trophoblast fusion. PMID:26441659

  12. Chemo-enzymatic synthesis of 2'-O-methoxyethyl ribonucleosides using a phosphodiesterase from Serratia marcescens.

    PubMed

    Marais, Guy; Ghisalba, Oreste

    2005-02-01

    An enzyme able to cleave the 3',5'-phosphate ring of 2'-methoxyethyl cyclic nucleotides (3',5'-cyclic nucleotide phosphodiesterase, EC 3.1.4.17) from Serratia marcescens DSM 30121 was used to deprotect the cyclic phosphate nucleotides after chemical alkylation. The process yielded 2'-O-alkylated nucleosides used as building blocks of antisense oligonucleotides for subsequent potential applications in therapeutics (antisense oligonucleotide synthesis) and diagnostics. The phosphodiesterase from the Gram-negative enteric bacterium S. marcescens was selected on account of the broad substrate range and high activity of the enzyme. The protein was purified by heat-treatment of the crude cell-free extract, followed by column chromatography (gel filtration). It was characterised and showed optimal activity at a broad pH range (pH 6.8-9.4, with a peak at ca. pH 8.5) and at a temperature of 60-65 degrees C. No metal ions were required for activity, although Ba2+ was an activator. Conversion of 2'-O-methoxyethyl cAMP into the corresponding nucleoside derivative on a multi-gram scale was successfully performed in two steps, using the S. marcescens enzyme in conjunction with a commercially available alkaline phosphatase from Escherichia coli.

  13. Phosphodiesterases reduce spontaneous sinoatrial beating but not the 'fight or flight' tachycardia elicited by agonists through Gs-protein-coupled receptors.

    PubMed

    Kaumann, Alberto J

    2011-07-01

    Cyclic AMP (cAMP) steers the generation of basal heart beat in the sinoatrial node. It also induces sinoatrial tachycardia and increased cardiac force, elicited through activation of Gs-protein-coupled receptors (GsPCRs). Phosphodiesterases (PDEs) hydrolyse cAMP. In the heart mainly PDE3 and PDE4 would be expected to limit those functions, and the PDE isoenzymes do indeed reduce basal sinoatrial beating rate and blunt the positive inotropic effects of agonists, mediated by GsPCRs. By contrast, recent evidence shows that GsPCR-mediated sinoatrial tachycardia is not controlled by PDE1-5. A PDE-resistant cAMP pool in sinoatrial cells, generated through activation of GsPCRs, including β(1)- and β(2)-adrenoceptors, appears to guarantee unrestrained tachycardia during fight or flight stress.

  14. Phosphatidylinositol-stimulated phosphorylation of an inhibitory subunit of cGMP phosphodiesterase in vertebrate rod photoreceptors.

    PubMed Central

    Hayashi, F; Lin, G Y; Matsumoto, H; Yamazaki, A

    1991-01-01

    An inhibitory subunit (P gamma) of cGMP phosphodiesterase from vertebrate rod photoreceptors (frog, toad, and bovine) was phosphorylated by cytosolic protein kinase(s) derived from intact frog rod outer segments. The phosphorylation of frog P gamma was stimulated by phosphatidylinositol but not by cAMP or cGMP. One- and two-dimensional gel electrophoresis revealed that 70-80% of P gamma was phosphorylated with 1 mol of phosphate per frog P gamma under optimal conditions. A peptide that derived from an active domain of bovine P gamma was also phosphorylated. Phosphorylation of frog P gamma was inhibited by addition of the peptide to the reaction mixture. Phosphorylation of frog P gamma was also inhibited by addition of transducin subunits or active (P gamma-less) cGMP phosphodiesterase. Okadaic acid, on the other hand, enhanced P gamma phosphorylation, suggesting the presence of protein phosphatase(s) in the cytosolic fraction. These data suggest another mechanism for the regulation of cGMP phosphodiesterase in vertebrate rod photoreceptors. Images PMID:1852003

  15. Object memory enhancement by combining sub-efficacious doses of specific phosphodiesterase inhibitors.

    PubMed

    Bollen, E; Akkerman, S; Puzzo, D; Gulisano, W; Palmeri, A; D'Hooge, R; Balschun, D; Steinbusch, H W M; Blokland, A; Prickaerts, J

    2015-08-01

    The second messengers cGMP and cAMP have a vital role in synaptic plasticity and memory processes. As such, phosphodiesterases inhibitors (PDE-Is), which prevent the breakdown of these cyclic nucleotides, represent a potential treatment strategy in memory decline. Recently it has been demonstrated that cGMP and cAMP signaling act in sequence during memory consolidation, with early cGMP signaling requiring subsequent cAMP signaling. Here, we sought to confirm this relationship, and to evaluate its therapeutic implications. Combining sub-efficacious doses of the cGMP-specific PDE type 5 inhibitor vardenafil (0.1 mg/kg) and cAMP-specific PDE type 4 inhibitor rolipram (0.01 mg/kg) during the early and late memory consolidation phase, respectively, led to improved memory performance in a 24 h interval object recognition task. Similarly, such a sub-efficacious combination treatment enhanced the transition of early-phase long-term potentiation (LTP) to late-phase LTP in hippocampal slices. In addition, both object memory and LTP were improved after administration of two sub-efficacious doses of the dual substrate PDE type 2 inhibitor BAY60 7550 (0.3 mg/kg) at the early and late consolidation phase, respectively. Taken together, combinations of sub-efficacious doses of cAMP- and cGMP-specific PDE-Is have an additive effect on long-term synaptic plasticity and memory formation and might prove a superior alternative to single PDE-I treatment. PMID:25896769

  16. Positive lusitropic effect and diminished myofibrillar sensitivity to calcium produced by cAMP on toad (Bufo arenarum Hensel) ventricle.

    PubMed

    Vila Petroff, M; Vittone, L; Mundiña, C; Chiappe de Cingolani, G; Alicia, M

    1992-01-01

    In intact ventricular strips from toad heart, we studied the relaxant or positive lusitropic effect of different interventions known to increase intracellular cAMP levels. Isoproterenol increased developed tension (DT), maximal rate of contraction (+T), and maximal velocity of relaxation (-T). From 10(-8) to 10(-4)M isoproterenol, -T increased proportionally more than +T being the ratio +T/-T significantly decreased. A single dose of isoproterenol (3 x 10(-8)M) increased cAMP levels from 0.174 +/- 0.022 to 0.329 +/- 0.039 pmoles/mg ww (P < 0.05), increased contractility by 69 +/- 13% and decreased +T/-T by 18.5 +/- 4.55%. Administration of 10(-3)M of dibutyryl cyclic AMP (dcAMP) significantly increased DT and +T and decreased the ratio +T/-T. Similar effects were obtained with milrinone, a specific cAMP phosphodiesterase inhibitor. Papaverine, a non selective phosphodiesterase inhibitor, failed to increase +T but significantly increased -T. In chemically skinned ventricular trabeculae, calcium sensitivity of the myofibrils was significantly increased by 10(-5)M of the phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (IBMX). 10(-3)M dcAMP failed to affect calcium sensitivity of chemically skinned ventricular trabeculae when given alone, but produced a decrease in calcium sensitivity of the myofibrils in the presence of 10(-5)M of either IBMX or papaverine. The results would indicate that the relaxant effect of isoproterenol is mediated in toad ventricle by an increase in intracellular cAMP levels. They furthermore suggest that a decrease in myofilament sensitivity to calcium may be a mechanism by which cAMP produces its relaxant effect.

  17. cAMP and in vivo hypoxia induce tob, ifr1, and fos expression in erythroid cells of the chick embryo.

    PubMed

    Dragon, Stefanie; Offenhäuser, Nina; Baumann, Rosemarie

    2002-04-01

    During avian embryonic development, terminal erythroid differentiation occurs in the circulation. Some of the key events, such as the induction of erythroid 2,3-bisphosphoglycerate (2,3-BPG), carbonic anhydrase (CAII), and pyrimidine 5'-nucleotidase (P5N) synthesis are oxygen dependent (Baumann R, Haller EA, Schöning U, and Weber M, Dev Biol 116: 548-551, 1986; Dragon S and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 280: R870-R878, 2001; Dragon S, Carey C, Martin K, and Baumann R, J Exp Biol 202: 2787-2795, 1999; Dragon S, Glombitza S, Götz R, and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 271: R982-R989, 1996; Dragon S, Hille R, Götz R, and Baumann R, Blood 91: 3052-3058, 1998; Million D, Zillner P, and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 261: R1188-R1196, 1991) in an indirect way: hypoxia stimulates the release of norepinephrine (NE)/adenosine into the circulation (Dragon et al., J Exp Biol 202: 2787-2795, 1999; Dragon et al., Am J Physiol Regulatory Integrative Comp Physiol 271: R982-R989, 1996). This leads via erythroid beta-adrenergic/adenosine A(2) receptor activation to a cAMP signal inducing several proteins in a transcription-dependent manner (Dragon et al., Am J Physiol Regulatory Integrative Comp Physiol 271: R982-R989, 1996; Dragon et al., Blood 91: 3052-3058, 1998; Glombitza S, Dragon S, Berghammer M, Pannermayr M, and Baumann R, Am J Physiol Regulatory Integrative Comp Physiol 271: R973-R981, 1996). To understand how the cAMP-dependent processes are initiated, we screened an erythroid cDNA library for cAMP-regulated genes. We detected three genes that were strongly upregulated (>5-fold) by cAMP in definitive and primitive red blood cells. They are homologous to the mammalian Tob, Ifr1, and Fos proteins. In addition, the genes are induced in the intact embryo during short-term hypoxia. Because the genes are regulators of proliferation and differentiation in other cell types, we suggest that cAMP

  18. Immunomodulatory Effects of Lippia sidoides Extract: Induction of IL-10 Through cAMP and p38 MAPK-Dependent Mechanisms

    PubMed Central

    Rajgopal, Arun; Rebhun, John F.; Burns, Charlie R.; Scholten, Jeffrey D.; Balles, John A.

    2015-01-01

    Abstract Lippia sidoides is an aromatic shrub that grows wild in the northeastern region of Brazil. In local traditional medicine, the aerial portions of this species are used as anti-infectives, antiseptics, spasmolytics, sedatives, hypotensives, and anti-inflammatory agents. In this research, we evaluate the potential immunological properties of Lippia extract through in vitro analysis of its ability to modulate intracellular cyclic adenosine monophosphate (cAMP) levels and interleukin-10 (IL-10) production. These results show that Lippia extract increases intracellular cAMP through the inhibition of phosphodiesterase activity. They also demonstrate that Lippia extract increases IL-10 production in THP-1 monocytes through both an increase in intracellular cAMP and the activation of p38 MAPK. These results suggest that the Lippia-mediated inhibition of phosphodiesterase activity and the subsequent increase in intracellular cAMP may explain some of the biological activities associated with L. sidoides. In addition, the anti-inflammatory activity of L. sidoides may also be due, in part, to its ability to induce IL-10 production through the inhibition of cyclic nucleotide-dependent phosphodiesterase activity and by its activation of the p38 MAPK pathway. PMID:25599252

  19. Immunomodulatory effects of Lippia sidoides extract: induction of IL-10 through cAMP and p38 MAPK-dependent mechanisms.

    PubMed

    Rajgopal, Arun; Rebhun, John F; Burns, Charlie R; Scholten, Jeffrey D; Balles, John A; Fast, David J

    2015-03-01

    Lippia sidoides is an aromatic shrub that grows wild in the northeastern region of Brazil. In local traditional medicine, the aerial portions of this species are used as anti-infectives, antiseptics, spasmolytics, sedatives, hypotensives, and anti-inflammatory agents. In this research, we evaluate the potential immunological properties of Lippia extract through in vitro analysis of its ability to modulate intracellular cyclic adenosine monophosphate (cAMP) levels and interleukin-10 (IL-10) production. These results show that Lippia extract increases intracellular cAMP through the inhibition of phosphodiesterase activity. They also demonstrate that Lippia extract increases IL-10 production in THP-1 monocytes through both an increase in intracellular cAMP and the activation of p38 MAPK. These results suggest that the Lippia-mediated inhibition of phosphodiesterase activity and the subsequent increase in intracellular cAMP may explain some of the biological activities associated with L. sidoides. In addition, the anti-inflammatory activity of L. sidoides may also be due, in part, to its ability to induce IL-10 production through the inhibition of cyclic nucleotide-dependent phosphodiesterase activity and by its activation of the p38 MAPK pathway.

  20. Local regulation of tooth mineralization by sphingomyelin phosphodiesterase 3.

    PubMed

    Khavandgar, Z; Alebrahim, S; Eimar, H; Tamimi, F; McKee, M D; Murshed, M

    2013-04-01

    Sphingomyelin phosphodiesterase 3 (Smpd3) encodes a membrane-bound enzyme that cleaves sphingomyelin to generate several bioactive metabolites. A recessive mutation called fragilitas ossium (fro) in the Smpd3 gene leads to impaired mineralization of bone and tooth extracellular matrix (ECM) in fro/fro mice. In teeth from fro/fro mice at various neonatal ages, radiography and light and electron microscopy showed delayed mantle dentin mineralization and a consequent delay in enamel formation as compared with that in control +/fro mice. These tooth abnormalities progressively improved with time. Immunohistochemistry showed expression of SMPD3 by dentin-forming odontoblasts. SMPD3 deficiency, however, did not affect the differentiation of these cells, as shown by osterix and dentin sialophosphoprotein expression. Using a transgenic mouse rescue model (fro/fro; Col1a1-Smpd3) in which Smpd3 expression is driven by a murine Col1a1 promoter fragment active in osteoblasts and odontoblasts, we demonstrate a complete correction of the tooth mineralization delays. In conclusion, analysis of these data demonstrates that Smpd3 expression in odontoblasts is required for tooth mineralization.

  1. Day Camp Manual: A Study of Mandatory Standards and Desirable Camping Practices for Children's Day Camps. Book V. Revised 1974.

    ERIC Educational Resources Information Center

    Babcock, William

    The final book in a 5-book day camp manual is a questionnaire developed by the Ontario Camping Association to help directors of children's summer day camps check camp operations, principles, and procedures, and to help them determine how and where a camp can be improved. The questionnaire, to be completed by the director with the aid of another…

  2. cAMP controls rod photoreceptor sensitivity via multiple targets in the phototransduction cascade

    PubMed Central

    Astakhova, Luba A.; Samoiliuk, Evgeniia V.; Govardovskii, Victor I.

    2012-01-01

    In early studies, both cyclic AMP (cAMP) and cGMP were considered as potential secondary messengers regulating the conductivity of the vertebrate photoreceptor plasma membrane. Later discovery of the cGMP specificity of cyclic nucleotide–gated channels has shifted attention to cGMP as the only secondary messenger in the phototransduction cascade, and cAMP is not considered in modern schemes of phototransduction. Here, we report evidence that cAMP may also be involved in regulation of the phototransduction cascade. Using a suction pipette technique, we recorded light responses of isolated solitary rods from the frog retina in normal solution and in the medium containing 2 µM of adenylate cyclase activator forskolin. Under forskolin action, flash sensitivity rose more than twofold because of a retarded photoresponse turn-off. The same concentration of forskolin lead to a 2.5-fold increase in the rod outer segment cAMP, which is close to earlier reported natural day/night cAMP variations. Detailed analysis of cAMP action on the phototransduction cascade suggests that several targets are affected by cAMP increase: (a) basal dark phosphodiesterase (PDE) activity decreases; (b) at the same intensity of light background, steady background-induced PDE activity increases; (c) at light backgrounds, guanylate cyclase activity at a given fraction of open channels is reduced; and (d) the magnitude of the Ca2+ exchanger current rises 1.6-fold, which would correspond to a 1.6-fold elevation of [Ca2+]in. Analysis by a complete model of rod phototransduction suggests that an increase of [Ca2+]in might also explain effects (b) and (c). The mechanism(s) by which cAMP could regulate [Ca2+]in and PDE basal activity is unclear. We suggest that these regulations may have adaptive significance and improve the performance of the visual system when it switches between day and night light conditions. PMID:23008435

  3. cAMP controls rod photoreceptor sensitivity via multiple targets in the phototransduction cascade.

    PubMed

    Astakhova, Luba A; Samoiliuk, Evgeniia V; Govardovskii, Victor I; Firsov, Michael L

    2012-10-01

    In early studies, both cyclic AMP (cAMP) and cGMP were considered as potential secondary messengers regulating the conductivity of the vertebrate photoreceptor plasma membrane. Later discovery of the cGMP specificity of cyclic nucleotide-gated channels has shifted attention to cGMP as the only secondary messenger in the phototransduction cascade, and cAMP is not considered in modern schemes of phototransduction. Here, we report evidence that cAMP may also be involved in regulation of the phototransduction cascade. Using a suction pipette technique, we recorded light responses of isolated solitary rods from the frog retina in normal solution and in the medium containing 2 µM of adenylate cyclase activator forskolin. Under forskolin action, flash sensitivity rose more than twofold because of a retarded photoresponse turn-off. The same concentration of forskolin lead to a 2.5-fold increase in the rod outer segment cAMP, which is close to earlier reported natural day/night cAMP variations. Detailed analysis of cAMP action on the phototransduction cascade suggests that several targets are affected by cAMP increase: (a) basal dark phosphodiesterase (PDE) activity decreases; (b) at the same intensity of light background, steady background-induced PDE activity increases; (c) at light backgrounds, guanylate cyclase activity at a given fraction of open channels is reduced; and (d) the magnitude of the Ca(2+) exchanger current rises 1.6-fold, which would correspond to a 1.6-fold elevation of [Ca(2+)](in). Analysis by a complete model of rod phototransduction suggests that an increase of [Ca(2+)](in) might also explain effects (b) and (c). The mechanism(s) by which cAMP could regulate [Ca(2+)](in) and PDE basal activity is unclear. We suggest that these regulations may have adaptive significance and improve the performance of the visual system when it switches between day and night light conditions. PMID:23008435

  4. No ordinary boot camp.

    PubMed

    Tichy, N M

    2001-04-01

    Many companies now run boot camps--comprehensive orientation programs designed to help new hires hit the ground running. They're intense and intimidating, and new employees emerge from them with strong bonds to other recruits and to the organization. But at Trilogy, organizational consultant Noel Tichy discovered one program that's a breed apart. In this article, Tichy gives us a detailed tour of Trilogy's boot camp, Trilogy University, to demonstrate why it's so different--and so effective. Like the best boot camps, it serves as an immersion in both the technical skills new recruits will need for their jobs and Trilogy's corporate culture, which emphasizes risk-taking, teamwork, humility, and a strong customer focus. But this is a new-employee orientation session that's so fundamental to the company as a whole that it's presided over by the CEO and top corporate executives for fully six months of the year. Why? In two three-month sessions, these top executives hone their own strategic thinking about the company as they decide what to teach the new recruits each session. They also find the company's next generation of new products as they judge the innovative ideas the recruits are tasked with developing--making the program Trilogy's main R&D engine. And they pull the company's rising technical stars into mentoring roles for the new recruits, helping to build the next generation of top leadership. After spending months on-site studying Trilogy University, Tichy came away highly impressed by the power of the virtuous teaching cycle the program has set in motion. Leaders of the organization are learning from recruits at the same time that the recruits are learning from the leaders. It's a model, he argues, that other companies would do well to emulate. PMID:11299694

  5. Modulation of Compartmentalised Cyclic Nucleotide Signalling via Local Inhibition of Phosphodiesterase Activity

    PubMed Central

    Brescia, Marcella; Zaccolo, Manuela

    2016-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) are the only enzymes that degrade the cyclic nucleotides cAMP and cGMP, and play a key role in modulating the amplitude and duration of the signal delivered by these two key intracellular second messengers. Defects in cyclic nucleotide signalling are known to be involved in several pathologies. As a consequence, PDEs have long been recognized as potential drug targets, and they have been the focus of intense research for the development of therapeutic agents. A number of PDE inhibitors are currently available for the treatment of disease, including obstructive pulmonary disease, erectile dysfunction, and heart failure. However, the performance of these drugs is not always satisfactory, due to a lack of PDE-isoform specificity and their consequent adverse side effects. Recent advances in our understanding of compartmentalised cyclic nucleotide signalling and the role of PDEs in local regulation of cAMP and cGMP signals offers the opportunity for the development of novel strategies for therapeutic intervention that may overcome the current limitation of conventional PDE inhibitors. PMID:27706091

  6. Renal Epithelial Cyst Formation and Enlargement in vitro: Dependence on cAMP

    NASA Astrophysics Data System (ADS)

    Mangoo-Karim, Roberto; Uchic, Marie; Lechene, Claude; Grantham, Jared J.

    1989-08-01

    Cysts, a common abnormality of kidneys, are collections of urine-like fluid enclosed by a continuous layer of epithelial cells. Renal cysts derive from nephrons and collecting ducts and progressively enlarge as a consequence of epithelial proliferation and transepithelial fluid secretion. The initiation of cyst formation and the factors that control cyst enlargement are unknown. We used an in vitro model of renal cysts to explore the role of the cAMP signal transduction system in the formation and expansion of cysts. MDCK cells, cultured in hydrated-collagen gel, produced polarized monolayered epithelial cysts when intracellular cAMP was increased by prostaglandin E1, arginine vasopressin, cholera toxin, forskolin, or 8-bromoadenosine 3',5'-cyclic monophosphate. All agonists were potentiated by 3-isobutyl-1-methylxanthine, a nucleotide phosphodiesterase inhibitor. The cell proliferation component of cyst enlargement was accelerated by cAMP agonists, as shown by the increased growth of MDCK cells in subconfluent monolayers. The fluid secretion component, reflected by the transepithelial movement of fluid across polarized monolayers of MDCK cells grown on permeable supports, was stimulated by cAMP agonists in the basolateral medium. Chloride levels were higher in the cyst fluid and the secreted fluid than in the bathing medium. We conclude that the development of MDCK cysts is dependent on cAMP. This signal transduction system may be an important modulator of epithelial cell proliferation and transepithelial fluid secretion in the kidney.

  7. Blending Technology with Camp Tradition: Technology Can Simplify Camp Operations.

    ERIC Educational Resources Information Center

    Salzman, Jeff

    2000-01-01

    Discusses uses of technology appropriate for camps, which are service organizations based on building relationships. Describes relationship marketing and how it can be enhanced through use of Web sites, interactive brochures, and client databases. Outlines other technology uses at camp: automated dispensing of medications, satellite tracking of…

  8. Summer Camp as Therapeutic Context: The Camp Logan Program.

    ERIC Educational Resources Information Center

    McCammon, Susan; And Others

    These symposium papers describe various aspects of the Camp Logan, South Carolina, program, a therapeutic summer residential program for children, ages 8-14, who have significant behavior problems. The philosophy and advantages of the therapeutic camping model are discussed, e.g., structure during the summer, controlled though informal…

  9. The Camp Health Manual. An Excellent Reference Written Especially for Organized Camps. Revised.

    ERIC Educational Resources Information Center

    Goldring, David; Middelkamp, J. Neal

    This book is a guide to the diagnosis and care of sick children in organized camping situations. This book presents health care information for the management of medical and surgical problems by the camp counselor, camp director, camp nurse, and camp physician. The chapters are: (1) Camp Standards; (2) The Infirmary; (3) Infirmary Supplies; (4)…

  10. 36 CFR 13.25 - Camping.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... this part. (b) Site time-limits. Camping is authorized for 14 consecutive days in one location. Camping... impacting park resources. (c) Designated campgrounds. Except at designated campgrounds, camping...

  11. Explaining the Value of Camp.

    ERIC Educational Resources Information Center

    Chenery, Mary Faeth

    1994-01-01

    Overviews the philosophy and theory of camp experiences and discusses the special benefits of camps, including experiences that can lead to significant life-changing outcomes, sound educational goals, a sense of psychological and physical safety, and helping children to deal with social problems such as the degradation of the environment and human…

  12. Kids Camping Takes the Challenge.

    ERIC Educational Resources Information Center

    James, Vickie L.; Hohnbaum, Claudia

    2002-01-01

    A Wisconsin Girl Scout camp integrated The Healthy Kids Challenge into its program. The camp evaluated policies related to meals, snacks, physical activities, team building, and self-esteem. Staff inservice training resulted in healthier meals on the same budget and developed ownership of the program. Campers and families had opportunities to…

  13. Encountering Child Abuse at Camp.

    ERIC Educational Resources Information Center

    Durall, John K.

    1997-01-01

    Defines child abuse, including the three categories: physical, sexual, and psychological. Presents characteristics and behaviors of each type of abuse, and long-term effects. Discusses how to handle abuse that occurs at camp, and the effects on the camp. Sidebars present abuse statistics, 15 activities that promote psychological wellness, and 8…

  14. Advances in targeting cyclic nucleotide phosphodiesterases.

    PubMed

    Maurice, Donald H; Ke, Hengming; Ahmad, Faiyaz; Wang, Yousheng; Chung, Jay; Manganiello, Vincent C

    2014-04-01

    Cyclic nucleotide phosphodiesterases (PDEs) catalyse the hydrolysis of cyclic AMP and cyclic GMP, thereby regulating the intracellular concentrations of these cyclic nucleotides, their signalling pathways and, consequently, myriad biological responses in health and disease. Currently, a small number of PDE inhibitors are used clinically for treating the pathophysiological dysregulation of cyclic nucleotide signalling in several disorders, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication and chronic obstructive pulmonary disease. However, pharmaceutical interest in PDEs has been reignited by the increasing understanding of the roles of individual PDEs in regulating the subcellular compartmentalization of specific cyclic nucleotide signalling pathways, by the structure-based design of novel specific inhibitors and by the development of more sophisticated strategies to target individual PDE variants.

  15. Psychiatric aspects of phosphodiesterases: An overview

    PubMed Central

    Murthy, Vasantmeghna S.; Mangot, Ajish G.

    2015-01-01

    Phosphodiesterases (PDE) are exciting new targets in medical sciences. These enzymes are some of the key mediators of cellular functions in the body and hence are attractive sites for drug-induced modulations. With the finding that Tofisopam, a new anxiolytic, inhibits PDEs, the authors were inspired to look into the role of PDE and drugs acting on them in psychiatry. Hence, the review was undertaken. We found several research materials available highlighting the role of PDE in cellular functions and the possible newer etiological mechanisms of neuropsychiatric illnesses such as schizophrenia, depression/anxiety disorders, and cognitive dysfunction involving PDEs. We also found that there are many molecules acting on PDEs, which have the potential to alter the way we treat mental illnesses today. This article is intended to provide an in-depth look at these enzymes so that more cost-effective therapeutic molecules may be synthesized and marketed in India for managing mental illnesses. PMID:26729948

  16. Psychiatric aspects of phosphodiesterases: An overview.

    PubMed

    Murthy, Vasantmeghna S; Mangot, Ajish G

    2015-01-01

    Phosphodiesterases (PDE) are exciting new targets in medical sciences. These enzymes are some of the key mediators of cellular functions in the body and hence are attractive sites for drug-induced modulations. With the finding that Tofisopam, a new anxiolytic, inhibits PDEs, the authors were inspired to look into the role of PDE and drugs acting on them in psychiatry. Hence, the review was undertaken. We found several research materials available highlighting the role of PDE in cellular functions and the possible newer etiological mechanisms of neuropsychiatric illnesses such as schizophrenia, depression/anxiety disorders, and cognitive dysfunction involving PDEs. We also found that there are many molecules acting on PDEs, which have the potential to alter the way we treat mental illnesses today. This article is intended to provide an in-depth look at these enzymes so that more cost-effective therapeutic molecules may be synthesized and marketed in India for managing mental illnesses.

  17. cAMP-stimulated Protein Phosphatase 2A Activity Associated with Muscle A Kinase-anchoring Protein (mAKAP) Signaling Complexes Inhibits the Phosphorylation and Activity of the cAMP-specific Phosphodiesterase PDE4D3*

    PubMed Central

    Dodge-Kafka, Kimberly L.; Bauman, Andrea; Mayer, Nicole; Henson, Edward; Heredia, Lorena; Ahn, Jung; McAvoy, Thomas; Nairn, Angus C.; Kapiloff, Michael S.

    2010-01-01

    The concentration of the second messenger cAMP is tightly controlled in cells by the activity of phosphodiesterases. We have previously described how the protein kinase A-anchoring protein mAKAP serves as a scaffold for the cAMP-dependent protein kinase PKA and the cAMP-specific phosphodiesterase PDE4D3 in cardiac myocytes. PKA and PDE4D3 constitute a negative feedback loop whereby PKA-catalyzed phosphorylation and activation of PDE4D3 attenuate local cAMP levels. We now show that protein phosphatase 2A (PP2A) associated with mAKAP complexes is responsible for reversing the activation of PDE4D3 by catalyzing the dephosphorylation of PDE4D3 serine residue 54. Mapping studies reveal that a C-terminal mAKAP domain (residues 2085–2319) binds PP2A. Binding to mAKAP is required for PP2A function, such that deletion of the C-terminal domain enhances both base-line and forskolin-stimulated PDE4D3 activity. Interestingly, PP2A holoenzyme associated with mAKAP complexes in the heart contains the PP2A targeting subunit B56δ. Like PDE4D3, B56δ is a PKA substrate, and PKA phosphorylation of mAKAP-bound B56δ enhances phosphatase activity 2-fold in the complex. Accordingly, expression of a B56δ mutant that cannot be phosphorylated by PKA results in increased PDE4D3 phosphorylation. Taken together, our findings demonstrate that PP2A associated with mAKAP complexes promotes PDE4D3 dephosphorylation, serving both to inhibit PDE4D3 in unstimulated cells and also to mediate a cAMP-induced positive feedback loop following adenylyl cyclase activation and B56δ phosphorylation. In general, PKA·PP2A·mAKAP complexes exemplify how protein kinases and phosphatases may participate in molecular signaling complexes to dynamically regulate localized intracellular signaling. PMID:20106966

  18. cAMP-stimulated protein phosphatase 2A activity associated with muscle A kinase-anchoring protein (mAKAP) signaling complexes inhibits the phosphorylation and activity of the cAMP-specific phosphodiesterase PDE4D3.

    PubMed

    Dodge-Kafka, Kimberly L; Bauman, Andrea; Mayer, Nicole; Henson, Edward; Heredia, Lorena; Ahn, Jung; McAvoy, Thomas; Nairn, Angus C; Kapiloff, Michael S

    2010-04-01

    The concentration of the second messenger cAMP is tightly controlled in cells by the activity of phosphodiesterases. We have previously described how the protein kinase A-anchoring protein mAKAP serves as a scaffold for the cAMP-dependent protein kinase PKA and the cAMP-specific phosphodiesterase PDE4D3 in cardiac myocytes. PKA and PDE4D3 constitute a negative feedback loop whereby PKA-catalyzed phosphorylation and activation of PDE4D3 attenuate local cAMP levels. We now show that protein phosphatase 2A (PP2A) associated with mAKAP complexes is responsible for reversing the activation of PDE4D3 by catalyzing the dephosphorylation of PDE4D3 serine residue 54. Mapping studies reveal that a C-terminal mAKAP domain (residues 2085-2319) binds PP2A. Binding to mAKAP is required for PP2A function, such that deletion of the C-terminal domain enhances both base-line and forskolin-stimulated PDE4D3 activity. Interestingly, PP2A holoenzyme associated with mAKAP complexes in the heart contains the PP2A targeting subunit B56delta. Like PDE4D3, B56delta is a PKA substrate, and PKA phosphorylation of mAKAP-bound B56delta enhances phosphatase activity 2-fold in the complex. Accordingly, expression of a B56delta mutant that cannot be phosphorylated by PKA results in increased PDE4D3 phosphorylation. Taken together, our findings demonstrate that PP2A associated with mAKAP complexes promotes PDE4D3 dephosphorylation, serving both to inhibit PDE4D3 in unstimulated cells and also to mediate a cAMP-induced positive feedback loop following adenylyl cyclase activation and B56delta phosphorylation. In general, PKA.PP2A.mAKAP complexes exemplify how protein kinases and phosphatases may participate in molecular signaling complexes to dynamically regulate localized intracellular signaling. PMID:20106966

  19. New Members of the Mammalian Glycerophosphodiester Phosphodiesterase Family

    PubMed Central

    Ohshima, Noriyasu; Kudo, Takahiro; Yamashita, Yosuke; Mariggiò, Stefania; Araki, Mari; Honda, Ayako; Nagano, Tomomi; Isaji, Chiaki; Kato, Norihisa; Corda, Daniela; Izumi, Takashi; Yanaka, Noriyuki

    2015-01-01

    The known mammalian glycerophosphodiester phosphodiesterases (GP-PDEs) hydrolyze glycerophosphodiesters. In this study, two novel members of the mammalian GP-PDE family, GDE4 and GDE7, were isolated, and the molecular basis of mammalian GP-PDEs was further explored. The GDE4 and GDE7 sequences are highly homologous and evolutionarily close. GDE4 is expressed in intestinal epithelial cells, spermatids, and macrophages, whereas GDE7 is particularly expressed in gastro-esophageal epithelial cells. Unlike other mammalian GP-PDEs, GDE4 and GDE7 cannot hydrolyze either glycerophosphoinositol or glycerophosphocholine. Unexpectedly, both GDE4 and GDE7 show a lysophospholipase D activity toward lysophosphatidylcholine (lyso-PC). We purified the recombinant GDE4 and GDE7 proteins and show that these enzymes can hydrolyze lyso-PC to produce lysophosphatidic acid (LPA). Further characterization of purified recombinant GDE4 showed that it can also convert lyso-platelet-activating factor (1-O-alkyl-sn-glycero-3-phosphocholine; lyso-PAF) to alkyl-LPA. These data contribute to our current understanding of mammalian GP-PDEs and of their physiological roles via the control of lyso-PC and lyso-PAF metabolism in gastrointestinal epithelial cells and macrophages. PMID:25528375

  20. Streptococcus pyogenes CAMP factor attenuates phagocytic activity of RAW 264.7 cells.

    PubMed

    Kurosawa, Mie; Oda, Masataka; Domon, Hisanori; Saitoh, Issei; Hayasaki, Haruaki; Terao, Yutaka

    2016-02-01

    Streptococcus pyogenes produces molecules that inhibit the function of human immune system, thus allowing the pathogen to grow and spread in tissues. It is known that S. pyogenes CAMP factor increases erythrocytosis induced by Staphylococcus aureus β-hemolysin. However, the effects of CAMP factor for immune cells are unclear. In this study, we investigated the effects of CAMP factor to macrophages. Western blotting analysis demonstrated that all examined strains expressed CAMP factor protein. In the presence of calcium or magnesium ion, CAMP factor was significantly released in the supernatant. In addition, both culture supernatant from S. pyogenes strain SSI-9 and recombinant CAMP factor dose-dependently induced vacuolation in RAW 264.7 cells, but the culture supernatant from Δcfa isogenic mutant strain did not. CAMP factor formed oligomers in RAW 264.7 cells in a time-dependent manner. CAMP factor suppressed cell proliferation via G2 phase cell cycle arrest without inducing cell death. Furthermore, CAMP factor reduced the uptake of S. pyogenes and phagocytic activity indicator by RAW 264.7 cells. These results suggest that CAMP factor works as a macrophage dysfunction factor. Therefore, we conclude that CAMP factor allows S. pyogenes to escape the host immune system, and contribute to the spread of streptococcal infection.

  1. Downregulation of Brain Phosphodiesterase Type IV Measured with 11C-(R)-Rolipram Positron Emission Tomography in Major Depressive Disorder

    PubMed Central

    Fujita, Masahiro; Hines, Christina S.; Zoghbi, Sami S.; Mallinger, Alan G.; Dickstein, Leah P.; Liow, Jeih-San; Zhang, Yi; Pike, Victor W.; Drevets, Wayne C.; Innis, Robert B.; Zarate, Carlos A.

    2012-01-01

    Background Phosphodiesterase type IV (PDE4), an important component of the cyclic adenosine monophosphate (cAMP) cascade, selectively metabolizes cAMP in the brain to the inactive monophosphate. Basic studies suggest that PDE4 mediates the effects of several antidepressants. This study sought to quantify the binding of 11C-(R)-rolipram, a PDE4 inhibitor, as an indirect measure of this enzyme’s activity in the brain of individuals with major depressive disorder (MDD) compared with healthy control subjects. Methods 11C-(R)-Rolipram brain positron emission tomography scans were performed in 28 unmedicated MDD subjects and 25 age- and gender-matched healthy control subjects. Patients were moderately depressed and about one half were treatment-naive. 11C-(R)-Rolipram binding in the brain was measured using arterial 11C-(R)-rolipram levels to correct for the influence of cerebral blood flow. Results Major depressive disorder subjects showed a widespread, approximately 20% reduction in 11C-(R)-rolipram binding (p = .002), which was not caused by different volumes of gray matter. Decreased rolipram binding of similar magnitudes was observed in most brain areas. Rolipram binding did not correlate with the severity of depressive or anxiety symptoms. Conclusions This study is the first to demonstrate that brain levels of PDE4, a critical enzyme that regulates cAMP, are decreased in unmedicated individuals with MDD in vivo. These results are in line with human postmortem and rodent studies demonstrating downregulation of the cAMP cascade in MDD and support the hypothesis that agents such as PDE4 inhibitors, which increase activity within the cAMP cascade, may have antidepressant effects. PMID:22677471

  2. Inhibitors of cyclic nucleotide phosphodiesterase isozymes type-III and type-IV suppress mitogenesis of rat mesangial cells.

    PubMed Central

    Matousovic, K; Grande, J P; Chini, C C; Chini, E N; Dousa, T P

    1995-01-01

    We studied interactions between the mitogen-activated protein kinase (MAPK) signalling pathway and cAMP-protein kinase (PKA) signaling pathway in regulation of mitogenesis of mesangial cells (MC) determined by [3H]thymidine incorporation, with or without added EGF. Forskolin or dibutyryl cAMP strongly (by 60-70%) inhibited [3H]thymidine incorporation into MC. Cilostamide, lixazinone or cilostazol selective inhibitors of cAMP-phosphodiesterase (PDE) isozyme PDE-III, inhibited mitogenesis to similar extent as forskolin and DBcAMP and activated in situ PKA, but without detectable increase in cAMP levels. Cilostamide and cilostazol were more than three times more effective at inhibiting mesangial mitogenesis than rolipram and denbufylline, inhibitors of isozyme PDE-IV, even though PDE-IV was two times more abundant in MC than was PDE-III. On the other hand, when incubated with forskolin, rolipram-enhanced cAMP accumulation was far greater (10-100x) than with cilostamide. EGF increased MAPK activity (+300%); PDE isozyme inhibitors which suppressed mitogenesis also inhibited MAPK. PDE isozyme inhibitors also suppressed PDGF-stimulated MC proliferation. We conclude that cAMP inhibits the mitogen-dependent MAPK-signaling pathway probably by decreasing the activity of Raf-1 due to PKA-catalyzed phosphorylation. Further, we surmise that minor increase in the cAMP pool metabolized by PDE-III is intimately related to regulation of mesangial proliferation. Thus, PDE isozyme inhibitors have the potential to suppress MC proliferation by a focused effect upon signaling pathways. Images PMID:7615811

  3. Perspectives on Camp Administration. Readings for Camp Director Education. Camp Administration Series.

    ERIC Educational Resources Information Center

    Farley, Elizabeth, Ed.

    The publication includes 47 selected readings for camp directors who are interested in reviewing the current status of the profession and who want to be a part of shaping its future. The articles, selected from periodicals directly related to camping and, where appropriate, from related journals, were selected and organized to support the American…

  4. AMPK antagonizes hepatic glucagon-stimulated cyclic AMP signalling via phosphorylation-induced activation of cyclic nucleotide phosphodiesterase 4B

    PubMed Central

    Johanns, M.; Lai, Y.-C.; Hsu, M.-F.; Jacobs, R.; Vertommen, D.; Van Sande, J.; Dumont, J. E.; Woods, A.; Carling, D.; Hue, L.; Viollet, B.; Foretz, M; Rider, M H

    2016-01-01

    Biguanides such as metformin have previously been shown to antagonize hepatic glucagon-stimulated cyclic AMP (cAMP) signalling independently of AMP-activated protein kinase (AMPK) via direct inhibition of adenylate cyclase by AMP. Here we show that incubation of hepatocytes with the small-molecule AMPK activator 991 decreases glucagon-stimulated cAMP accumulation, cAMP-dependent protein kinase (PKA) activity and downstream PKA target phosphorylation. Moreover, incubation of hepatocytes with 991 increases the Vmax of cyclic nucleotide phosphodiesterase 4B (PDE4B) without affecting intracellular adenine nucleotide concentrations. The effects of 991 to decrease glucagon-stimulated cAMP concentrations and activate PDE4B are lost in hepatocytes deleted for both catalytic subunits of AMPK. PDE4B is phosphorylated by AMPK at three sites, and by site-directed mutagenesis, Ser304 phosphorylation is important for activation. In conclusion, we provide a new mechanism by which AMPK antagonizes hepatic glucagon signalling via phosphorylation-induced PDE4B activation. PMID:26952277

  5. ASB16165, a phosphodiesterase 7A inhibitor, reduces cutaneous TNF-alpha level and ameliorates skin edema in phorbol ester 12-O-tetradecanoylphorbol-13-acetate-induced skin inflammation model in mice.

    PubMed

    Kadoshima-Yamaoka, Kumiko; Goto, Megumi; Murakawa, Masao; Yoshioka, Ryosuke; Tanaka, Yoshitaka; Inoue, Hidekazu; Murafuji, Hidenobu; Kanki, Satomi; Hayashi, Yasuhiro; Nagahira, Kazuhiro; Ogata, Atsuto; Nakatsuka, Takashi; Fukuda, Yoshiaki

    2009-06-24

    Possible role of phosphodiesterase 7A (PDE7A) in skin inflammation was examined using ASB16165, a specific inhibitor for PDE7A. Epicutaneous application of phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) to mouse ear resulted in induction of skin edema, and topical treatment with ASB16165 inhibited the induction of skin edema in a dose-dependent manner. The TPA challenge also increased the level of TNF-alpha at the application site, and the ASB16165 treatment reduced the TNF-alpha level in the skin. In addition, ASB16165 suppressed the production of TNF-alpha by human keratinocytes stimulated in vitro with TPA and calcium ionophore. Forskolin, an activator of adenylyl cyclase, as well as dibutyryl cAMP also showed inhibitory effect on the TNF-alpha production in the cells, suggesting involvement of cAMP in TNF-alpha generation. These results demonstrate that PDE7A might regulate TNF-alpha production in keratinocytes in a cAMP-dependent fashion. As immunostaining analysis revealed that PDE7A is expressed in the epidermis and TNF-alpha is known to contribute to the TPA-induced edema, it is possible that the inhibitory effect of ASB16165 on skin edema in mouse TPA-induced dermatitis model is mediated by suppression of TNF-alpha production. This is the first report suggesting the association of PDE7A with the function of keratinocytes. ASB16165 will be useful as an agent for skin inflammation in which TNF-alpha plays a pathogenic role (e.g. psoriasis).

  6. Reactive oxygen species decrease cAMP response element binding protein expression in cardiomyocytes via a protein kinase D1-dependent mechanism that does not require Ser133 phosphorylation.

    PubMed

    Ozgen, Nazira; Guo, Jianfen; Gertsberg, Zoya; Danilo, Peter; Rosen, Michael R; Steinberg, Susan F

    2009-10-01

    Reactive oxygen species (ROS) exert pleiotropic effects on a wide array of signaling proteins that regulate cellular growth and apoptosis. This study shows that long-term treatment with a low concentration of H2O2 leads to the activation of signaling pathways involving extracellular signal-regulated kinase, ribosomal protein S6 kinase, and protein kinase D (PKD) that increase cAMP binding response element protein (CREB) phosphorylation at Ser(133) in cardiomyocytes. Although CREB-Ser(133) phosphorylation typically mediates cAMP-dependent increases in CREB target gene expression, the H2O2-dependent increase in CREB-Ser(133) phosphorylation is accompanied by a decrease in CREB protein abundance and no change in Cre-luciferase reporter activity. Mutagenesis studies indicate that H2O2 decreases CREB protein abundance via a mechanism that does not require CREB-Ser(133) phosphorylation. Rather, the H2O2-dependent decrease in CREB protein is prevented by the proteasome inhibitor lactacystin, by inhibitors of mitogen-activated protein kinase kinase or protein kinase C activity, or by adenoviral-mediated delivery of a small interfering RNA that decreases PKD1 expression. A PKD1-dependent mechanism that links oxidative stress to decreased CREB protein abundance is predicted to contribute to the pathogenesis of heart failure by influencing cardiac growth and apoptosis responses.

  7. Slave Labor Camps of the Third Reich.

    ERIC Educational Resources Information Center

    Stone, Adolf

    1983-01-01

    Describes the ground rules used by Nazi architects in choosing the sites for slave labor camps. While some, like Auschwitz, became extermination camps, others also produced armaments. One camp, Theresienstadt, became a "model" camp to show to reporters and Red Cross representatives. (CS)

  8. American Camping Association Annual Report, 1999.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    Founded in 1910 as the Camp Directors' Association of America, the American Camping Association (ACA) is the largest organization serving the organized camping industry. Over 5,500 members come from all segments of the camp profession. This annual report for 1999 describes ACA activities in support of organizational commitments. These commitments…

  9. Self-Concept Change in Camp Staff.

    ERIC Educational Resources Information Center

    Henderson, Karla A.; Bialeschki, M. Deborah

    The 1981 study ascertained whether the self-concept of 66 camp staff from 2 Wisconsin camps changed more than a control group of 18 college students attending summer school; if differences in self-concept were based on a particular characteristic (age, gender, staff position, years at camp); and in what ways, if any, self-concept of camp staff…

  10. Easter Seal Guide to Special Camping Programs.

    ERIC Educational Resources Information Center

    Crane, Helen B., Ed.

    Intended for organizations having or planning to establish resident resident camping programs for people with special needs, this guide supplements the American Camping Association's Standards. The philosophy, aims, and objectives of specialized camping programs are considered, and the following are discussed: administration, camp site selection,…

  11. 1970s: Camping in Tough Times.

    ERIC Educational Resources Information Center

    Camping Magazine, 1999

    1999-01-01

    In the 1970s, camps were challenged by economic recession, growing administrative demands, and changing attitudes and interests of campers and staff. An illustrative article from 1972, "Facing the Camping Future with Confidence" (Michael F. Buynak), discusses hidden costs in camp operation and the need for camp administration to shift from…

  12. 36 CFR 13.1222 - Camping.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Camping. 13.1222 Section 13.1222 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL... Camp Campground and other designated areas. (b) Camping in Brooks Camp Campground for more than a...

  13. 36 CFR 13.1222 - Camping.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Camping. 13.1222 Section 13.1222 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Katmai National Park and Preserve Brooks Camp Developed Area § 13.1222 Camping. (a) Camping...

  14. 36 CFR 13.1222 - Camping.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Camping. 13.1222 Section 13.1222 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL... Camp Campground and other designated areas. (b) Camping in Brooks Camp Campground for more than a...

  15. 36 CFR 13.1222 - Camping.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Camping. 13.1222 Section 13.1222 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL... Camp Campground and other designated areas. (b) Camping in Brooks Camp Campground for more than a...

  16. 36 CFR 13.1222 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Camping. 13.1222 Section 13.1222 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL... Camp Campground and other designated areas. (b) Camping in Brooks Camp Campground for more than a...

  17. The swing of it: Hammock camping.

    USGS Publications Warehouse

    Marion, Jeff

    2016-01-01

    Hammock camping is dramatically expanding along the Appalachian Trail and raising both questions and concerns among Trail land managers, club members, and backpackers. This article examines some of the advantages and disadvantages of hammock camping, including resource and social impacts. Some Leave No Trace hammock camping practices are included for those using hammocks at well-established campsites and when "pristine-site" camping.

  18. Standards for Day and Resident Camps: The Accreditation Programs of the American Camping Association. 1990 Edition.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    The purpose of this manual is to educate camp directors and camp personnel regarding government-recognized standard practices and procedures followed within the camp industry. These standards also provide a basis for voluntary accreditation of camps by the American Camping Association (ACA) beyond the minimum requirements of licensing. The manual…

  19. Basic Camp Management: An Introduction to Camp Administration. Revised 3rd Edition.

    ERIC Educational Resources Information Center

    Ball, Armand; Ball, Beverly

    This book is the primary text for the Certified Camp Director Program and the Basic Camp Directors Course sponsored by the American Camping Association (Indiana). It provides an orientation for new and prospective camp directors and a quick reference for experienced camp directors. The book covers the following topics: (1) an historical overview…

  20. Camping Out On An Asteroid

    NASA Video Gallery

    An astronaut and a geologist recently spent three days camping out as though they were on an asteroid. They were inside NASA's Space Exploration Vehicle prototype, flying it virtually in a digital ...

  1. The role of ventral striatal cAMP signaling in stress-induced behaviors.

    PubMed

    Plattner, Florian; Hayashi, Kanehiro; Hernández, Adan; Benavides, David R; Tassin, Tara C; Tan, Chunfeng; Day, Jonathan; Fina, Maggy W; Yuen, Eunice Y; Yan, Zhen; Goldberg, Matthew S; Nairn, Angus C; Greengard, Paul; Nestler, Eric J; Taussig, Ronald; Nishi, Akinori; Houslay, Miles D; Bibb, James A

    2015-08-01

    The cAMP and cAMP-dependent protein kinase A (PKA) signaling cascade is a ubiquitous pathway acting downstream of multiple neuromodulators. We found that the phosphorylation of phosphodiesterase-4 (PDE4) by cyclin-dependent protein kinase 5 (Cdk5) facilitated cAMP degradation and homeostasis of cAMP/PKA signaling. In mice, loss of Cdk5 throughout the forebrain elevated cAMP levels and increased PKA activity in striatal neurons, and altered behavioral responses to acute or chronic stressors. Ventral striatum- or D1 dopamine receptor-specific conditional knockout of Cdk5, or ventral striatum infusion of a small interfering peptide that selectively targeted the regulation of PDE4 by Cdk5, produced analogous effects on stress-induced behavioral responses. Together, our results demonstrate that altering cAMP signaling in medium spiny neurons of the ventral striatum can effectively modulate stress-induced behavioral states. We propose that targeting the Cdk5 regulation of PDE4 could be a new therapeutic approach for clinical conditions associated with stress, such as depression. PMID:26192746

  2. The role of ventral striatal cAMP signaling in stress-induced behaviors.

    PubMed

    Plattner, Florian; Hayashi, Kanehiro; Hernández, Adan; Benavides, David R; Tassin, Tara C; Tan, Chunfeng; Day, Jonathan; Fina, Maggy W; Yuen, Eunice Y; Yan, Zhen; Goldberg, Matthew S; Nairn, Angus C; Greengard, Paul; Nestler, Eric J; Taussig, Ronald; Nishi, Akinori; Houslay, Miles D; Bibb, James A

    2015-08-01

    The cAMP and cAMP-dependent protein kinase A (PKA) signaling cascade is a ubiquitous pathway acting downstream of multiple neuromodulators. We found that the phosphorylation of phosphodiesterase-4 (PDE4) by cyclin-dependent protein kinase 5 (Cdk5) facilitated cAMP degradation and homeostasis of cAMP/PKA signaling. In mice, loss of Cdk5 throughout the forebrain elevated cAMP levels and increased PKA activity in striatal neurons, and altered behavioral responses to acute or chronic stressors. Ventral striatum- or D1 dopamine receptor-specific conditional knockout of Cdk5, or ventral striatum infusion of a small interfering peptide that selectively targeted the regulation of PDE4 by Cdk5, produced analogous effects on stress-induced behavioral responses. Together, our results demonstrate that altering cAMP signaling in medium spiny neurons of the ventral striatum can effectively modulate stress-induced behavioral states. We propose that targeting the Cdk5 regulation of PDE4 could be a new therapeutic approach for clinical conditions associated with stress, such as depression.

  3. Human phosphodiesterase 4D7 (PDE4D7) expression is increased in TMPRSS2-ERG-positive primary prostate cancer and independently adds to a reduced risk of post-surgical disease progression

    PubMed Central

    Böttcher, R; Henderson, D J P; Dulla, K; van Strijp, D; Waanders, L F; Tevz, G; Lehman, M L; Merkle, D; van Leenders, G J L H; Baillie, G S; Jenster, G; Houslay, M D; Hoffmann, R

    2015-01-01

    Background: There is an acute need to uncover biomarkers that reflect the molecular pathologies, underpinning prostate cancer progression and poor patient outcome. We have previously demonstrated that in prostate cancer cell lines PDE4D7 is downregulated in advanced cases of the disease. To investigate further the prognostic power of PDE4D7 expression during prostate cancer progression and assess how downregulation of this PDE isoform may affect disease outcome, we have examined PDE4D7 expression in physiologically relevant primary human samples. Methods: About 1405 patient samples across 8 publically available qPCR, Affymetrix Exon 1.0 ST arrays and RNA sequencing data sets were screened for PDE4D7 expression. The TMPRSS2-ERG gene rearrangement status of patient samples was determined by transformation of the exon array and RNA seq expression data to robust z-scores followed by the application of a threshold >3 to define a positive TMPRSS2-ERG gene fusion event in a tumour sample. Results: We demonstrate that PDE4D7 expression positively correlates with primary tumour development. We also show a positive association with the highly prostate cancer-specific gene rearrangement between TMPRSS2 and the ETS transcription factor family member ERG. In addition, we find that in primary TMPRSS2-ERG-positive tumours PDE4D7 expression is significantly positively correlated with low-grade disease and a reduced likelihood of progression after primary treatment. Conversely, PDE4D7 transcript levels become significantly decreased in castration resistant prostate cancer (CRPC). Conclusions: We further characterise and add physiological relevance to PDE4D7 as a novel marker that is associated with the development and progression of prostate tumours. We propose that the assessment of PDE4D7 levels may provide a novel, independent predictor of post-surgical disease progression. PMID:26575822

  4. Identification of two splice variant forms of type-IVB cyclic AMP phosphodiesterase, DPD (rPDE-IVB1) and PDE-4 (rPDE-IVB2) in brain: selective localization in membrane and cytosolic compartments and differential expression in various brain regions.

    PubMed

    Lobban, M; Shakur, Y; Beattie, J; Houslay, M D

    1994-12-01

    In order to detect the two splice variant forms of type-IVB cyclic AMP phosphodiesterase (PDE) activity, DPD (type-IVB1) and PDE-4 (type-IVB2), anti-peptide antisera were generated. One set ('DPD/PDE-4-common'), generated against a peptide sequence found at the common C-terminus of these two PDEs, detected both PDEs. A second set was PDE-4 specific, being directed against a peptide sequence found within the unique N-terminal region of PDE-4. In brain, DPD was found exclusively in the cytosol and PDE-4 exclusively associated with membranes. Both brain DPD and PDE-4 activities, isolated by immunoprecipitation, were cyclic AMP-specific (KmcyclicAMP: approximately 5 microM for DPD; approximately 4 microM for PDE-4) and were inhibited by low rolipram concentrations (K1rolipram approximately 1 microM for both). Transient expression of DPD in COS-1 cells allowed identification of an approx. 64 kDa species which co-migrated on SDS/PAGE with the immunoreactive species identified in both brain cytosol and membrane fractions using the DPD/PDE-4-common antisera. The subunit size observed for PDE-4 (approx. 64 kDa) in brain membranes was similar to that predicted from the cDNA sequence, but that observed for DPD was approx. 4 kDa greater. Type-IV, rolipram-inhibited PDE activity was found in all brain regions except the pituitary, where it formed between 30 and 70% of the PDE activity in membrane and cytosolic fractions when assayed with 1 microM cyclic AMP, PDE-4 formed 40-50% of the membrane type-IV activity in all brain regions save the midbrain (approx. 20%). DPD distribution was highly restricted to certain regions, providing approx. 35% of the type-IV cytosolic activity in hippocampus and 13-21% in cortex, hypothalamus and striatum with no presence in brain stem, cerebellum, midbrain and pituitary. The combined type-IVB PDE activities of DPD and PDE-4 contributed approx. 10% of the total PDE activity in most brain regions except for the pituitary (zero) and the mid

  5. The cAMP Response Element Binding protein (CREB) is activated by Insulin-like Growth Factor-1 (IGF-1) and regulates myostatin gene expression in skeletal myoblast

    SciTech Connect

    Zuloaga, R.; Fuentes, E.N.; Molina, A.; Valdés, J.A.

    2013-10-18

    Highlights: •IGF-1 induces the activation of CREB via IGF-1R/PI3K/PLC signaling pathway. •Calcium dependent signaling pathways regulate myostatin gene expression. •IGF-1 regulates myostatin gene expression via CREB transcription in skeletal myoblast. -- Abstract: Myostatin, a member of the Transforming Growth Factor beta (TGF-β) superfamily, plays an important role as a negative regulator of skeletal muscle growth and differentiation. We have previously reported that IGF-1 induces a transient myostatin mRNA expression, through the activation of the Nuclear Factor of Activated T cells (NFAT) in an IP{sub 3}/calcium-dependent manner. Here we examined the activation of CREB transcription factor as downstream targets of IGF-1 during myoblast differentiation and its role as a regulator of myostatin gene expression. In cultured skeletal myoblast, IGF-1 induced the phosphorylation and transcriptional activation of CREB via IGF-1 Receptor/Phosphatidylinositol 3-Kinase (PI3K)/Phospholipase C gamma (PLC γ), signaling pathways. Also, IGF-1 induced calcium-dependent molecules such as Calmodulin Kinase II (CaMK II), Extracellular signal-regulated Kinases (ERK), Protein Kinase C (PKC). Additionally, we examined myostatin mRNA levels and myostatin promoter activity in differentiated myoblasts stimulated with IGF-1. We found a significant increase in mRNA contents of myostatin and its reporter activity after treatment with IGF-1. The expression of myostatin in differentiated myoblast was downregulated by the transfection of siRNA–CREB and by pharmacological inhibitors of the signaling pathways involved in CREB activation. By using pharmacological and genetic approaches together these data demonstrate that IGF-1 regulates the myostatin gene expression via CREB transcription factor during muscle cell differentiation.

  6. Phosphodiesterase-5 Inhibitors: Action on the Signaling Pathways of Neuroinflammation, Neurodegeneration, and Cognition

    PubMed Central

    Peixoto, Christina Alves; Nunes, Ana Karolina Santana; Garcia-Osta, Ana

    2015-01-01

    Phosphodiesterase type 5 inhibitors (PDE5-Is) have recently emerged as a potential therapeutic strategy for neuroinflammatory, neurodegenerative, and memory loss diseases. Mechanistically, PDE5-Is produce an anti-inflammatory and neuroprotection effect by increasing expression of nitric oxide synthases and accumulation of cGMP and activating protein kinase G (PKG), the signaling pathway of which is thought to play an important role in the development of several neurodiseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), and multiple sclerosis (MS). The aim of this paper was to review present knowledge of the signaling pathways that underlie the use of PDE5-Is in neuroinflammation, neurogenesis, learning, and memory. PMID:26770022

  7. Upregulation of Phosphodiesterase type 5 in the Hyperplastic Prostate

    PubMed Central

    Zhang, Wenhao; Zang, Ning; Jiang, Yaoming; Chen, Ping; Wang, Xinghuan; Zhang, Xinhua

    2015-01-01

    Both erectile dysfunction (ED) and lower urinary tract symptoms (LUTS)/benign prostatic hyperplasia (BPH) are common in the aging male. Numerous clinical trials have demonstrated the efficacy and safety of phosphodiesterase type 5 inhibitors (PDE5-Is) for treating LUTS/BPH with/without ED. However, the influence of BPH on prostatic PDE5 expression has never been studied. A testosterone-induced rat model of BPH was developed and human hyperplastic prostate specimens were harvested during cystoprostatectomy. PDE5, nNOS, eNOS and α1-adrenoreceptor subtypes (α1aARs, α1bARs and α1dARs) were determined with real-time RT-PCR for rat tissues whilst PDE5 and α1-adrenoreceptor subtypes were determined in human samples. PDE5 was further analyzed with Western-blot and histological examination. Serum testosterone was measured with ELISA. The rat BPH model was validated as having a significantly enlarged prostate. PDE5 localized mainly in fibromuscular stroma in prostate. Our data showed a significant and previously undocumented upregulation of PDE5 in both rat and human BPH, along with increased expression of nNOS and α1dARs for rat tissues and α1aARs for human BPH. The upregulation of PDE5 in the hyperplastic prostate could explain the mechanism and contribute to the high effectiveness of PDE5-Is for treating LUTS/BPH. Fibromuscular stroma could be the main target for PDE5-Is within prostate. PMID:26657792

  8. Development of a high throughput screen for allosteric modulators of melanocortin-4 receptor signaling using a real time cAMP assay

    PubMed Central

    Pantel, Jacques; Williams, Savannah Y.; Mi, Dehui; Sebag, Julien; Corbin, Jackie D.; Weaver, C. David; Cone, Roger D.

    2011-01-01

    The melanocortin MC4 receptor is a potential target for the development of drugs for both obesity and cachexia. Melanocortin MC4 receptor ligands known thus far are orthosteric agonists or antagonists, however the agonists, in particular, have generally exhibited unwanted side effects. For some receptors, allosteric modulators are expected to reduce side-effect profiles. To identify allosteric modulators of the melanocortin MC4 receptor, we created HEK293 cell lines coexpressing the human melanocortin MC4 receptor and a modified luciferase-based cAMP sensor. Monitoring luminescence as a readout of real-time intracellular cAMP concentration, we demonstrate this cell line is able to report melanocortin agonist responses, as well as inverse agonist response to the physiological AgRP peptide. Based on the MC4R-GLO cell line, we developed an assay that was shown to meet HTS standards (Z’=0.50). A pilot screen run on the Microsource Spectrum compound library (n= 2,000) successfully identified 62 positive modulators. This screen identified predicted families of compounds: β2AR agonists –the β2AR being endogenously expressed in HEK293 cells-, an adenylyl cyclase activator and finally a distribution of phosphodiesterase (PDE) inhibitors well characterized or recently identified. In this last category, we identified a structural family of coumarin-derived compounds (imperatorin, osthol and prenyletin), along with deracoxib, a drug in veterinary use for its COX2 inhibitory properties. This latter finding unveiled a new off-target mechanism of action for deracoxib as a PDE inhibitor. Overall, these data are the first report of an HTS for allosteric modulators for a Gs protein coupled receptor. PMID:21296065

  9. Selective enhancement of wnt4 expression by cyclic AMP-associated cooperation between rat central astrocytes and microglia.

    PubMed

    Ohnishi, Masatoshi; Urasaki, Tomoka; Ochiai, Hiroyuki; Matsuoka, Kohei; Takeo, Shin; Harada, Tomoki; Ohsugi, Yoshihito; Inoue, Atsuko

    2015-11-13

    The wnt protein family has important members involved in cell differentiation, proliferation and plasticity expression; however, little is known about its biosynthesis processes. On the other hand, an increase in the intracerebral cyclic adenosine 3', 5'-monophosphate (cAMP) level leads to synaptic plasticity via the de novo synthesis of any protein. Here, the effect of dibutyryl cAMP (dbcAMP), a membrane permeability cAMP analog, on the wnt family was investigated in rat primary-cultured glial cells containing astrocytes and microglia. Among wnt3a, 4, 5a, 7a and 11 mRNA, only wnt4 expression was increased by longer treatment (24 h), compared with short treatment (2 h), with dbcAMP in a concentration-dependent manner, and its effect reached statistical significance at 1 mM. In cultures of isolated astrocytes or microglia, wnt4 expression was not affected by 1 mM dbcAMP for 24 h, and microglial wnt4 protein was undetectable even when cells were treated with the drug. Mixed glial cells treated for 24 h with 1 mM dbcAMP showed significantly increased wnt4 protein, as well as mRNA. Immunofluorescence manifested that cells that expressed wnt4 protein were astrocytes, but not microglia. Intraperitoneal injection of 1.25 mg/kg rolipram, a phosphodiesterase (PDE) IV inhibitor that can pass through the blood brain barrier and inhibits cAMP degradation specifically, showed a tendency to increase wnt4 expression in the adult rat brain after 24 h, and the increases in wnt4 mRNA and protein levels reached statistical significance in the hippocampus and striatum, respectively. This is the first finding to help elucidate the selective biosynthesis of central wnt4 through cAMP-stimulated microglia and astrocytes interaction.

  10. Enhanced PDE4B expression augments LPS-inducible TNF expression in ethanol-primed monocytes: relevance to alcoholic liver disease.

    PubMed

    Gobejishvili, Leila; Barve, Shirish; Joshi-Barve, Swati; McClain, Craig

    2008-10-01

    Increased plasma and hepatic TNF-alpha expression is well documented in patients with alcoholic hepatitis and is implicated in the pathogenesis of alcoholic liver disease. We have previously shown that monocytes from patients with alcoholic hepatitis show increased constitutive and LPS-induced NF-kappaB activation and TNF-alpha production. Our recent studies showed that chronic ethanol exposure significantly decreased cellular cAMP levels in both LPS-stimulated and unstimulated monocytes and Kupffer cells, leading to an increase in LPS-inducible TNF-alpha production by affecting NF-kappaB activation and induction of TNF mRNA expression. Accordingly, the mechanisms underlying this ethanol-induced decrease in cellular cAMP leading to an increase in TNF expression were examined in monocytes/macrophages. In this study, chronic ethanol exposure was observed to significantly increase LPS-inducible expression of cAMP-specific phosphodiesterase (PDE)4B that degrades cellular cAMP. Increased PDE4B expression was associated with enhanced NF-kappaB activation and transcriptional activity and subsequent priming of monocytes/macrophages leading to enhanced LPS-inducible TNF-alpha production. Selective inhibition of PDE4 by rolipram abrogated LPS-mediated TNF-alpha expression at both protein and mRNA levels in control and ethanol-treated cells. Notably, PDE4 inhibition did not affect LPS-inducible NF-kappaB activation but significantly decreased NF-kappaB transcriptional activity. These findings strongly support the pathogenic role of PDE4B in the ethanol-mediated priming of monocytes/macrophages and increased LPS-inducible TNF production and the subsequent development of alcoholic liver disease (ALD). Since enhanced TNF expression plays a significant role in the evolution of clinical and experimental ALD, its downregulation via selective PDE4B inhibitors could constitute a novel therapeutic approach in the treatment of ALD. PMID:18687753

  11. Histamine induced elevation of cyclic AMP phosphodiesterase activity in human monocytes.

    PubMed

    Holden, C A; Chan, S C; Norris, S; Hanifin, J M

    1987-10-01

    We have previously reported histamine desensitization of human blood mononuclear leukocytes resulting in reduced cAMP responses to beta-adrenergic agonists, histamine and prostaglandin E1. This heterologous desensitization occurred at low, micromolar histamine concentrations and was accompanied by elevation of cAMP-phosphodiesterase (PDE) activity in these cells. We have now investigated the activity of PDE in the lymphocyte and monocyte fractions of mononuclear leukocytes to determine the site of histamine effect. PDE activity per cell was higher in monocytes (0.075 +/- 0.070 units) than lymphocytes (0.026 +/- 0.08) units). Monocytes responded to 10(-6) M histamine stimulation with a much greater increase in PDE activity (0.354 +/- 0.1 units) than did lymphocytes (0.047 +/- 0.015 units). Histamine receptor studies, using thiazolylethylamine and chlorpheniramine as H1-agonist and antagonist respectively and dimaprit and cimetidine as H2-agonists and antagonists respectively, indicated that the histamine stimulation of PDE activity is mediated predominantly through H1 histamine receptor in the monocytes and the H1 receptor in the lymphocytes. Previously histamine had been thought to increase cyclic AMP by acting on H2 receptors to activate adenylate cyclase. Our studies show that stimulation of H1 or H2 receptors by low histamine concentration can cause the opposite effect i.e. increased catabolism and a net reduction in cAMP levels. The localization of this effect predominantly to monocytes indicates a potentially important mechanism for histamine action on immune regulation. PMID:2891264

  12. Inhibition of phosphodiesterase 2 reverses impaired cognition and neuronal remodeling caused by chronic stress.

    PubMed

    Xu, Ying; Pan, Jianchun; Sun, Jiao; Ding, Lianshu; Ruan, Lina; Reed, Miranda; Yu, Xuefeng; Klabnik, Jonathan; Lin, Dan; Li, Jianxin; Chen, Ling; Zhang, Chong; Zhang, Hanting; O'Donnell, James M

    2015-02-01

    Chronic stress and neuronal vulnerability have recently been recognized as factors contributing to cognitive disorders. One way to modify neuronal vulnerability is through mediation of phosphodiesterase 2 (PDE2), an enzyme that exerts its action on cognitive processes via the control of intracellular second messengers, cGMP and, to a lesser extent, cAMP. This study explored the effects of a PDE2 inhibitor, Bay 60-7550, on stress-induced learning and memory dysfunction in terms of its ramification on behavioral, morphologic, and molecular changes. Bay 60-7550 reversed stress-induced cognitive impairment in the Morris water maze, novel object recognition, and location tasks (object recognition test and/or object location test), effects prevented by treatment with 7-NI, a selective inhibitor of neuronal nitric oxide synthase; MK801, a glutamate receptor (NMDAR) inhibitor; myr-AIP, a CaMKII inhibitor; and KT5823, a protein kinase G inhibitor. Bay 60-7550 also ameliorated stress-induced structural remodeling in the CA1 of the hippocampus, leading to increases in dendritic branching, length, and spine density. However, the neuroplasticity initiated by Bay 60-7550 was not seen in the presence of 7-NI, MK801, myr-AIP, or KT5823. PDE2 inhibition reduced stress-induced extracellular-regulated protein kinase activation and attenuated stress-induced decreases in transcription factors (e.g., Elk-1, TORC1, and CREB phosphorylation) and plasticity-related proteins (e.g., Egr-1 and brain-derived neurotrophic factor). Pretreatment with inhibitors of NMDA, CaMKII, neuronal nitric oxide synthase, and protein kinase G (or protein kinase A) blocked the effects of Bay 60-7550 on cGMP or cAMP signaling. These findings indicate that the effect of PDE2 inhibition on stress-induced memory impairment is potentially mediated via modulation of neuroplasticity-related NMDAR-CaMKII-cGMP/cAMP signaling. PMID:25442113

  13. Inhibition of phosphodiesterase 2 reverses impaired cognition and neuronal remodeling caused by chronic stress

    PubMed Central

    Xu, Ying; Pan, Jianchun; Sun, Jiao; Ding, Lianshu; Ruan, Lina; Reed, Miranda; Yu, Xuefeng; Klabni, Jonathan; Lin, Dan; Li, Jianxin; Chen, Ling; Zhang, Chong; Zhang, Hanting; O’Donnell, James M.

    2014-01-01

    Chronic stress and neuronal vulnerability have recently been recognized as factors contributing to cognitive disorders. One way to modify neuronal vulnerability is through mediation of phosphodiesterase 2 (PDE2), an enzyme that exerts its action on cognitive processes via the control of intracellular second messengers, cGMP and, to a lesser extent, cAMP. This study explored the effects of a PDE2 inhibitor, Bay 60-7550, on stress-induced learning and memory dysfunction in terms of its ramification on behavioral, morphological and molecular changes. Bay 60-7550 reversed stress-induced cognitive impairment in the Morris water maze (MWM), novel object recognition and location tasks (ORT/OLT), effects prevented by treatment with 7-NI, a selective inhibitor of neuronal nitric oxide synthase (nNOS); MK801, a glutamate receptor (NMDAR) inhibitor; myr-AIP, a CaMKII inhibitor; and KT5823, a PKG inhibitor. Bay 60-7550 also ameliorated stress-induced structural remodeling in the CA1 of the hippocampus, leading to increases in dendritic branching, length, and spine density. However, the neuroplasticity initiated by Bay 60-7550 was not seen in the presence of 7-NI, MK801, myr-AIP or KT5823. PDE2 inhibition reduced stress-induced ERK activation and attenuated stress-induced decreases in transcription factors (e.g., Elk-1, TORC1, and pCREB) and plasticity-related proteins (e.g, Egr-1 and BDNF). Pre-treatment with inhibitors of NMDA, CaMKII, nNOS, PKG (or PKA), blocked the effects of Bay 60-7550 on cGMP or cAMP signaling. These findings indicate that the effect of PDE2 inhibition on stress-induced memory impairment is potentially mediated via modulation of neuroplasticity-related, NMDAR-CaMKII-cGMP/cAMP signaling. PMID:25442113

  14. Inhibition of phosphodiesterase 2 reverses impaired cognition and neuronal remodeling caused by chronic stress.

    PubMed

    Xu, Ying; Pan, Jianchun; Sun, Jiao; Ding, Lianshu; Ruan, Lina; Reed, Miranda; Yu, Xuefeng; Klabnik, Jonathan; Lin, Dan; Li, Jianxin; Chen, Ling; Zhang, Chong; Zhang, Hanting; O'Donnell, James M

    2015-02-01

    Chronic stress and neuronal vulnerability have recently been recognized as factors contributing to cognitive disorders. One way to modify neuronal vulnerability is through mediation of phosphodiesterase 2 (PDE2), an enzyme that exerts its action on cognitive processes via the control of intracellular second messengers, cGMP and, to a lesser extent, cAMP. This study explored the effects of a PDE2 inhibitor, Bay 60-7550, on stress-induced learning and memory dysfunction in terms of its ramification on behavioral, morphologic, and molecular changes. Bay 60-7550 reversed stress-induced cognitive impairment in the Morris water maze, novel object recognition, and location tasks (object recognition test and/or object location test), effects prevented by treatment with 7-NI, a selective inhibitor of neuronal nitric oxide synthase; MK801, a glutamate receptor (NMDAR) inhibitor; myr-AIP, a CaMKII inhibitor; and KT5823, a protein kinase G inhibitor. Bay 60-7550 also ameliorated stress-induced structural remodeling in the CA1 of the hippocampus, leading to increases in dendritic branching, length, and spine density. However, the neuroplasticity initiated by Bay 60-7550 was not seen in the presence of 7-NI, MK801, myr-AIP, or KT5823. PDE2 inhibition reduced stress-induced extracellular-regulated protein kinase activation and attenuated stress-induced decreases in transcription factors (e.g., Elk-1, TORC1, and CREB phosphorylation) and plasticity-related proteins (e.g., Egr-1 and brain-derived neurotrophic factor). Pretreatment with inhibitors of NMDA, CaMKII, neuronal nitric oxide synthase, and protein kinase G (or protein kinase A) blocked the effects of Bay 60-7550 on cGMP or cAMP signaling. These findings indicate that the effect of PDE2 inhibition on stress-induced memory impairment is potentially mediated via modulation of neuroplasticity-related NMDAR-CaMKII-cGMP/cAMP signaling.

  15. Hydromania: Summer Science Camp Curriculum.

    SciTech Connect

    Moura, Joan

    1995-07-01

    In 1992, Bonneville Power Administration (BPA) and the US Department of Energy (DOE) began a collaborative pilot project with the Portland Parks and Recreation Community Schools Program and others to provide summer science camps to children in Grades 4--6. Camps run two weeks in duration between late June and mid-August. Sessions are five days per week, from 9 a.m. to 3 p.m. In addition to hands-on science and math curriculum, at least three field trips are incorporated into the educational learning experience. The purpose of the BPA/DOE summer camps is to make available opportunities for fun, motivating experiences in science to students who otherwise would have difficulty accessing them. This includes inner city, minority, rural and low income students. Public law 101-510, which Congress passed in 1990, authorizes DOE facilities to establish collaborative inner-city and rural partnership programs in science and math. A primary goal of the BPA summer hands on science camps is to bring affordable science camp experiences to students where they live. It uses everyday materials to engage students` minds and to give them a sense that they have succeeded through a fun hands-on learning environment.

  16. Prayer Camps and Biomedical Care in Ghana: Is Collaboration in Mental Health Care Possible?

    PubMed Central

    Arias, Daniel; Taylor, Lauren; Ofori-Atta, Angela; Bradley, Elizabeth H.

    2016-01-01

    Background Experts have suggested that intersectoral partnerships between prayer camps and biomedical care providers may be an effective strategy to address the overwhelming shortage of mental health care workers in Africa and other low-income settings. Nevertheless, previous studies have not explored whether the prayer camp and biomedical staff beliefs and practices provide sufficient common ground to enable cooperative relationships. Therefore, we sought to examine the beliefs and practices of prayer camp staff and the perspective of biomedical care providers, with the goal of characterizing interest in—and potential for—intersectoral partnership between prayer camp staff and biomedical care providers. Methods We conducted 50 open-ended, semi-structured interviews with prophets and staff at nine Christian prayer camps in Ghana, and with staff within Ghana’s three public psychiatric hospitals. We used the purposive sampling method to recruit participants and the constant comparative method for qualitative data analysis. Results Prayer camp staff expressed interest in collaboration with biomedical mental health care providers, particularly if partnerships could provide technical support introducing medications in the prayer camp and address key shortcomings in their infrastructure and hygienic conditions. Nevertheless, challenges for collaboration were apparent as prayer camp staff expressed strong beliefs in a spiritual rather than biomedical explanatory model for mental illness, frequently used fasting and chained restraints in the course of treatment, and endorsed only short-term use of medication to treat mental illness—expressing concerns that long-term medication regimens masked underlying spiritual causes of illness. Biomedical providers were skeptical about the spiritual interpretations of mental illness held by faith healers, and were concerned by the use of chains, fasting, and the lack of adequate living facilities for patients in prayer camps

  17. He Sapa Bloketu Waecun: 2008 Summer Science and Cultural Camps

    NASA Astrophysics Data System (ADS)

    Kliche, D. V.; Sanovia, J.; Decker, R.; Bolman, J.

    2008-12-01

    The South Dakota School of Mines, Humboldt State University and Sinte Gleska University with support from the National Science Foundation, sponsored four camps for South Dakota Lakota youth to nurture a geosciences learning community linked to culturally significant sites in the Black Hills. These camps utilized outdoor, experiential learning to integrate indigenous knowledge with contemporary western science. The project resulted in increased awareness among Native and non-Native Americans, young and adult, about the importance of geosciences in their connection and interpretation of nature. The project also motivated participants in learning and becoming active in land and resources protection and the importance of becoming knowledgeable and active in regulatory policies (both Tribal and State). The four camps were scheduled during the month of June, 2008, which is the month of the summer solstice, a sacred time for the Lakota people which signal the Lakota Sundance Ceremony. The timing of the camps was chosen to give the Native American participants the framework to express their connection to Native lands through the understanding of their oral history. For the first time in such camps, middle and high school students were encouraged to have a parent or relative attending with them. The camps proved to be a great success among students and their families. The curriculum and activities helped participants immerse themselves mentally, physically and spiritually into an experience of a life time. We plan to show our results from these camps and emphasize the usefulness of this new approach in teaching science and encouraging the new generation to pursue careers in geosciences.

  18. Cultural behaviour and the invention of traditions: music and musical practices in the early concentration camps, 1933-6/7.

    PubMed

    Fackler, Guido

    2010-01-01

    This article investigates music in the concentration camps before the second world war. For the camp authorities, ordering prisoners to sing songs or play in orchestras was an instrument of domination. But for the prisoners, music could also be an expression of solidarity and survival: inmates could retain a degree of their own agency in the pre-war camps, despite the often unbearable living conditions and harsh treatment by guards. The present article emphasizes this ambiguity of music in the early camps. It illustrates the emergence of musical traditions in the pre-war camps which came to have a significant impact on everyday life in the camps. It helps to overcome the view that concentration camp prisoners were simply passive victims.

  19. Academic Boot Camp for the Writing of Psychology Research Reports

    ERIC Educational Resources Information Center

    Skues, Jason L.; Wise, Lisa

    2014-01-01

    Herein, we describe the implementation of, and responses to, a structured writing workshop in the form of an academic boot camp. Participants were 42 undergraduate psychology students from a medium-sized Australian university who were completing their major assignment for the semester. A majority of the students expressed satisfaction with the…

  20. Youth development and the camp experience.

    PubMed

    Garst, Barry A; Browne, Laurie P; Bialeschki, M Deborah

    2011-01-01

    The organized camp experience has been an important part of the lives of children, youth, and adults for over 150 years. The camp experience is a way for young people to explore and search for an authenticity often missing in other parts of their lives that contributes to their healthy transition into adulthood. Over the past decade, tremendous growth in the volume and rigor of camp-related research has occurred, facilitated by a targeted research agenda conducted by the American Camp Association. This agenda was founded on three national research projects conducted between 2003 and 2007: a study to identify the developmental outcomes of the camp experience, a benchmarking study of the youth development supports and opportunities provided through camp experiences, and a program improvement project directed toward enhancing supports and opportunities provided by camps. The findings from these research projects suggest that camp experiences promote developmental outcomes in both campers and staff and that camps provide the supports and opportunities needed for positive youth development. This article explores the developmental outcomes of the camp experience and the characteristics of the supports and opportunities afforded by camp experiences, including settings, structures, and programs and activities, as a way to provide a clearer understanding of camp as a positive youth development setting. Innovations and opportunities in research related to the provision of quality camp experiences are also considered. PMID:21786411

  1. Structural basis for the design of selective phosphodiesterase 4B inhibitors.

    PubMed

    Fox, David; Burgin, Alex B; Gurney, Mark E

    2014-03-01

    Phosphodiesterase-4B (PDE4B) regulates the pro-inflammatory Toll Receptor -Tumor Necrosis Factor α (TNFα) pathway in monocytes, macrophages and microglial cells. As such, it is an important, although under-exploited molecular target for anti-inflammatory drugs. This is due in part to the difficulty of developing selective PDE4B inhibitors as the amino acid sequence of the PDE4 active site is identical in all PDE4 subtypes (PDE4A-D). We show that highly selective PDE4B inhibitors can be designed by exploiting sequence differences outside the active site. Specifically, PDE4B selectivity can be achieved by capture of a C-terminal regulatory helix, now termed CR3 (Control Region 3), across the active site in a conformation that closes access by cAMP. PDE4B selectivity is driven by a single amino acid polymorphism in CR3 (Leu674 in PDE4B1 versus Gln594 in PDE4D). The reciprocal mutations in PDE4B and PDE4D cause a 70-80 fold shift in selectivity. Our structural studies show that CR3 is flexible and can adopt multiple orientations and multiple registries in the closed conformation. The new co-crystal structure with bound ligand provides a guide map for the design of PDE4B selective anti-inflammatory drugs. PMID:24361374

  2. Phosphodiesterase Inhibition Rescues Chronic Cognitive Deficits Induced by Traumatic Brain Injury

    PubMed Central

    Titus, David J.; Sakurai, Atsushi; Kang, Yuan; Furones, Concepcion; Jergova, Stanislava; Santos, Rosmery; Sick, Thomas J.; Atkins, Coleen M.

    2013-01-01

    Traumatic brain injury (TBI) modulates several cell signaling pathways in the hippocampus critical for memory formation. Previous studies have found that the cAMP-protein kinase A signaling pathway is downregulated after TBI and that treatment with a phosphodiesterase (PDE) 4 inhibitor rolipram rescues the decrease in cAMP. In the present study, we examined the effect of rolipram on TBI-induced cognitive impairments. At 2 weeks after moderate fluid-percussion brain injury or sham surgery, adult male Sprague Dawley rats received vehicle or rolipram (0.03 mg/kg) 30 min before water maze acquisition or cue and contextual fear conditioning. TBI animals treated with rolipram showed a significant improvement in water maze acquisition and retention of both cue and contextual fear conditioning compared with vehicle-treated TBI animals. Cue and contextual fear conditioning significantly increased phosphorylated CREB levels in the hippocampus of sham animals, but not in TBI animals. This deficit in CREB activation during learning was rescued in TBI animals treated with rolipram. Hippocampal long-term potentiation was reduced in TBI animals, and this was also rescued with rolipram treatment. These results indicate that the PDE4 inhibitor rolipram rescues cognitive impairments after TBI, and this may be mediated through increased CREB activation during learning. PMID:23516287

  3. Structures of the Four Subfamilies of Phosphodiesterase-4 Provide Insight into the Selectivity of Their Inhibitors

    SciTech Connect

    Wang, H.; Peng, M; Chen , Y; Geng, J; Robinson, H; Houslay , M; Cai, J; Ke, H

    2007-01-01

    PDE4 (phosphodiesterase-4)-selective inhibitors have attracted much attention as potential therapeutics for the treatment of both depression and major inflammatory diseases, but their practical application has been compromised by side effects. A possible cause for the side effects is that current PDE4-selective inhibitors similarly inhibit isoforms from all four PDE4 subfamilies. The development of PDE4 subfamily-selective inhibitors has been hampered by a lack of structural information. In the present study, we rectify this by providing the crystal structures of the catalytic domains of PDE4A, PDE4B and PDE4D in complex with the PDE4 inhibitor NVP 4-[8-(3-nitrophenyl)-[1,7]naphthyridin-6-yl]benzoic acid as well as the unliganded PDE4C structure. NVP binds in the same conformation to the deep cAMP substrate pocket and interacts with the same residues in each instance. However, detailed structural comparison reveals significant conformational differences. Although the active sites of PDE4B and PDE4D are mostly comparable, PDE4A shows significant displacements of the residues next to the invariant glutamine residue that is critical for substrate and inhibitor binding. PDE4C appears to be more distal from other PDE4 subfamilies, with certain key residues being disordered. Our analyses provide the first structural basis for the development of PDE4 subfamily-selective inhibitors.

  4. Evaluation of Antidepressant and Anxiolytic Activity of Phosphodiesterase 3 Inhibitor - Cilostazol

    PubMed Central

    Patel, Dipesh S.; Anand, Indermeet Singh; Bhatt, Parloop A.

    2012-01-01

    Background: Cyclic nucleotide Phosphodiesterases (PDEs) are ubiquitously distributed in mammalian tissues and play a major role in cell signaling by hydrolyzing cyclic Adenosine Monophosphate (cAMP) and cyclic Guanosine Monophosphate (cGMP). Impairments in signal transduction have been implicated as possible mechanism of reduced plasticity and neuronal survival in major depressive disorders. PDE inhibitors possess a potentially powerful means to manipulate secondary messengers involved in learning, memory and mood. Cilostazol is an antiplatelet agent indicated for the treatment of intermittent claudication with peripheral artery occlusion and for the prevention of ischemic stroke worldwide. Various animal studies have reported neuroprotective, anti apoptotic, cognition and cerebral blood flow improvement properties of cilostazol. Materials and Methods: In this study, the antidepressant and anxiolytic effects of cilostazol were evaluated in mice using behavioral tests sensitive to clinically effective antidepressant compound. Results: Cilostazol, administered intraperitoneally (20 mg/kg), decreased immobility time of mice when subjected to forced swim test and tail suspension test as compared to standard fluoxetine (20 mg/kg). Cilostazol also produced significant decrease in the number of marbles buried as compared to fluoxetine in marble burying model. Conclusion: The present study suggests that cilostazol possesses potential antidepressant and anxiolytic activity, which could be of therapeutic interest for use in patients with depressive disorders. PMID:23162186

  5. Functional Proteomic Profiling of Phosphodiesterases Using SeraFILE Separations Platform

    PubMed Central

    Oka, Amita R.; Kuruc, Matthew P.; Gujarathi, Ketan M.; Roy, Swapan

    2012-01-01

    Functional proteomic profiling can help identify targets for disease diagnosis and therapy. Available methods are limited by the inability to profile many functional properties measured by enzymes kinetics. The functional proteomic profiling approach proposed here seeks to overcome such limitations. It begins with surface-based proteome separations of tissue/cell-line extracts, using SeraFILE, a proprietary protein separations platform. Enzyme kinetic properties of resulting subproteomes are then characterized, and the data integrated into proteomic profiles. As a model, SeraFILE-derived subproteomes of cyclic nucleotide-hydrolyzing phosphodiesterases (PDEs) from bovine brain homogenate (BBH) and rat brain homogenate (RBH) were characterized for cAMP hydrolysis activity in the presence (challenge condition) and absence of cGMP. Functional profiles of RBH and BBH were compiled from the enzyme activity response to the challenge condition in each of the respective subproteomes. Intersample analysis showed that comparable profiles differed in only a few data points, and that distinctive subproteomes can be generated from comparable tissue samples from different animals. These results demonstrate that the proposed methods provide a means to simplify intersample differences, and to localize proteins attributable to sample-specific responses. It can be potentially applied for disease and nondisease sample comparison in biomarker discovery and drug discovery profiling. PMID:23227336

  6. Astro Camp is a blast!

    NASA Technical Reports Server (NTRS)

    2006-01-01

    An Astro Camp counselor and her campers perform a science experiment to learn what types of `fuel' will best propel their 'rockets.' Stennis Space Center's popular series of day camps have campers design, build and test model rockets based on the principles that would be used to build different types of rockets suitable for a mission to the moon or Mars. They learn details like how far they would travel, how long it would take, what supplies they would need and how to survive in that environment.

  7. Going Camping? A Basic Guide to Camping with Students.

    ERIC Educational Resources Information Center

    1977

    Beginning with the "often overlooked areas", this guide to camping trips for secondary students presents initial planning procedures such as: site selection; number of leaders per number of students (2 leaders for each group of 10-12); parental permission forms; transportation arrangements; public school announcements; medical aid expertise and…

  8. Making Camp Environmentally Friendly: How Two Camps Did It.

    ERIC Educational Resources Information Center

    Westerman, Martin; Griner-Johnson, Russ

    1991-01-01

    Describes the efforts of two camps administered by the Brandeis-Bardin Institute (Brandeis, California) in implementing water and energy conservation programs, involving recycling, composting, and landfill savings. Programs were successful in eliminating excess waste and teaching campers to care more about their environments at home and at work.…

  9. The Popeye Domain Containing Genes and cAMP Signaling

    PubMed Central

    Brand, Thomas; Poon, Kar Lai; Simrick, Subreena; Schindler, Roland F.R.

    2016-01-01

    3'-5'-cyclic adenosine monophosphate (cAMP) is a second messenger, which plays an important role in the heart. It is generated in response to activation of G-protein-coupled receptors (GPCRs). Initially, it was thought that protein kinase A (PKA) exclusively mediates cAMP-induced cellular responses such as an increase in cardiac contractility, relaxation, and heart rate. With the identification of the exchange factor directly activated by cAMP (EPAC) and hyperpolarizing cyclic nucleotide-gated (HCN) channels as cAMP effector proteins it became clear that a protein network is involved in cAMP signaling. The Popeye domain containing (Popdc) genes encode yet another family of cAMP-binding proteins, which are prominently expressed in the heart. Loss-of-function mutations in mice are associated with cardiac arrhythmia and impaired skeletal muscle regeneration. Interestingly, the cardiac phenotype, which is present in both, Popdc1 and Popdc2 null mutants, is characterized by a stress-induced sinus bradycardia, suggesting that Popdc proteins participate in cAMP signaling in the sinuatrial node. The identification of the two-pore channel TREK-1 and Caveolin 3 as Popdc-interacting proteins represents a first step into understanding the mechanisms of heart rate modulation triggered by Popdc proteins. PMID:27500161

  10. Camp Counseling: A Great Resume Builder.

    ERIC Educational Resources Information Center

    Fass, Jack R.

    1993-01-01

    Advocates camp counseling as an excellent preparation for human services careers. Camp counseling offers (1) leisure services preparation; (2) leadership training; (3) collaborative skills; and (4) life experience. (KS)

  11. Including People with Disabilities in Camp Programs: A Resource for Camp Directors.

    ERIC Educational Resources Information Center

    Roswal, Glenn M., Ed.; Dowd, Karen J., Ed.; Bynum, Jerry W., Ed.

    Written primarily by camp administrators affiliated with the National Easter Seal Society, this publication is designed to help camp directors meet the challenges of including campers of all abilities in their camp programs. The first section provides an overview of the inclusion concept in general and at camp, and discusses legal and medical…

  12. Correlation between selective inhibition of the cyclic nucleotide phosphodiesterases and the contractile activity in human pregnant myometrium near term.

    PubMed

    Leroy, M J; Cedrin, I; Breuiller, M; Giovagrandi, Y; Ferre, F

    1989-01-01

    The present study was carried out to determine the ability of various pharmacological agents to selectively inhibit each cytosolic form of phosphodiesterase isolated from the longitudinal layer of human myometria near term. Among the drugs tested, zaprinast specifically inhibits the first form of PDE which hydrolyses both substrates (cAMP and cGMP) and is stimulated by the Ca2+-calmodulin complex. A second form of PDE specific for cAMP hydrolysis and Ca2+-calmodulin insensitive is only present during pregnancy. Rolipram is the most potent and selective inhibitor of this second form. It is also the most efficient compound to inhibit in vitro the spontaneous contractions of near term myometria. The double effect of rolipram suggests an important role of the second form of PDE in the mechanisms of contractility during the pregnancy. In addition rolipram or other derivatives might be of a therapeutic interest in the prevention of prematurity in so far as they are devoid of undesirable maternal and fetal side effects.

  13. Sleep At Camp: A Survey.

    ERIC Educational Resources Information Center

    Pravda, Myra

    1997-01-01

    Among 40 camp directors surveyed, the majority believed that campers get enough sleep, but that staff members and directors do not get enough sleep. Addresses how sleep deprivation can affect job performance and offers strategies for helping staff understand the importance of sleep to keep them alert and functioning in their job. Includes…

  14. Winter Wilderness Travel and Camping.

    ERIC Educational Resources Information Center

    Gilchrest, Norman

    Knowledge and skill are needed for safe and enjoyable travel and camping in the wilderness in winter. The beauty of snow and ice, reduced human use, and higher tolerance of animals toward humans make the wilderness attractive during winter. The uniqueness of winter travel presents several challenges that are not present in other seasons. Safety is…

  15. Lyme Disease Comes to Camp.

    ERIC Educational Resources Information Center

    Peterson, Michael

    1989-01-01

    Describes one summer camp's plan for dealing with Lyme disease. Describes the disease and the deer tick. Recommends avoiding tick exposure through clothing, frequent examination, showers, and avoiding high grass and brushy areas, and using chemical insect repellents and chemicals to kill ticks in deer mouse nests. (DHP)

  16. Hazard Tree Management for Camps.

    ERIC Educational Resources Information Center

    Kong, Earl

    2002-01-01

    The principles behind a camp's hazard tree program are, first, identifying and removing those hazards that offer a clear, immediate threat, and then creating a management plan for the other trees. The plan should be written and contain goals and objectives, field evaluations, and treatments. Follow-up evaluations should be done annually and after…

  17. AVTC Hosts TechnoCamp

    ERIC Educational Resources Information Center

    Miner, Brenda

    2006-01-01

    The Area Vo-Tech Center (AVTC) in Russellville, Arkansas, recently hosted its first TechnoCamp to encourage enrollment based on the aptitude and interest level of the students enrolling in the various programs. The center currently offers student enrollment in auto technology, computer engineering, cosmetology, construction technology, drafting…

  18. Day Camp Manual: Program. Book IV.

    ERIC Educational Resources Information Center

    Babcock, William

    Book IV in a 5-book day camp manual discusses the camp program. Section I describes the organization, definition, and elements essential to successful day camp programs. Section II, which addresses the benefits and special considerations of mass programs, includes rainy day contingencies, materials to have on hand, and activity suggestions.…

  19. A Cost Study of Resident Camps, 1985.

    ERIC Educational Resources Information Center

    Ball, Armand, Ed.

    Directors of 87 resident camps affiliated with the American Camping Association responded to a random sample survey and provided data to compile this first collection of camp operating ration tables. Fifty-three tables show average percentage of total, median percentage of total, average in dollars, and median in dollars statistics in 14 income…

  20. Camping Is Your Gift to the World.

    ERIC Educational Resources Information Center

    Thurber, Christopher A.

    2002-01-01

    Many camp professionals wonder how the events of September 11 will affect their camps. Advice is given on dealing with concerns of parents, campers, staff, and directors. Stability is comforting--change only what is absolutely necessary. Compassion and inclusion, the behaviors modeled at camp, are antidotes to misunderstanding and marginalization,…

  1. Camping for Youth with Chronic Illnesses.

    ERIC Educational Resources Information Center

    Burns, Joanna L.; Keller, M. Jean

    1994-01-01

    Camp Fortnight brought 25 British children with cystic fibrosis to experience a 2-week camping program in Texas. Campers (ages 11-15) participated in wilderness experiences, a challenge course, fishing, horseback riding, creative arts, cooking, hiking, outdoor camping, and field trips. Profiles campers and their experiences. (LP)

  2. Preserving the Character of Historic Camps.

    ERIC Educational Resources Information Center

    Stepenoff, Bonnie

    1991-01-01

    Historic buildings, trails, and other camp features form the character of a particular camp and provide an escape to the past. Rehabilitating and preserving old camp structures requires attention to details and maintenance considerations. The U.S. Department of the Interior publishes guidelines for identifying and preserving historical properties.…

  3. Research in Action: Camping and Senior Adults.

    ERIC Educational Resources Information Center

    Chenery, Mary Faeth

    1987-01-01

    To determine the nature of camp for senior citizens, four participant observers (staff members) conducted open-ended interviews with campers at a five-day residential camp in Oregon. Results showed universal satisfaction with the camp experience. Campers valued the wide range of activities, comfortable places to socialize, and staff relations with…

  4. Extension Sustainability Camp: Design, Implementation, and Evaluation

    ERIC Educational Resources Information Center

    Brain, Roslynn; Upton, Sally; Tingey, Brett

    2015-01-01

    Sustainability Camps provide an opportunity for Extension educators to be in the forefront of sustainability outreach and to meet the growing demand for sustainability education. This article shares development, implementation, and evaluation of an Extension Sustainability Camp for youth, grades 4-6. Camp impact was measured via daily pre-and…

  5. DIRECTORY OF CAMPS FOR THE HANDICAPPED.

    ERIC Educational Resources Information Center

    National Easter Seal Society for Crippled Children and Adults, Chicago, IL.

    ONE HUNDRED AND SEVENTY-SEVEN RESIDENT CAMPS IN THE UNITED STATES AND CANADA AND 77 DAY CAMPS IN THE UNITED STATES WHICH SERVE CHILDREN OR ADULTS WITH PHYSICAL, MENTAL, SOCIAL, AND EMOTIONAL HANDICAPS ARE LISTED ALPHABETICALLY BY STATE. FOR EACH CAMP, INFORMATION ON TYPES OF THE HANDICAPPED WHO ARE ACCEPTED, SPECIFIC EXCLUSIONS, AGE RANGE, NUMBER…

  6. 36 CFR 13.1208 - Lake Camp.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Lake Camp. 13.1208 Section 13... § 13.1208 Lake Camp. Leaving a boat, trailer, or vehicle unattended for more than 72 hours at the facilities associated with the Lake Camp launching ramp is prohibited without authorization from...

  7. 43 CFR 423.33 - Camping.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Camping. 423.33 Section 423.33 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE INTERIOR PUBLIC CONDUCT ON BUREAU OF RECLAMATION FACILITIES, LANDS, AND WATERBODIES Rules of Conduct § 423.33 Camping. (a) You may camp on Reclamation...

  8. Involvement of the CDC25 gene product in the signal transmission pathway of the glucose-induced RAS-mediated cAMP signal in the yeast Saccharomyces cerevisiae.

    PubMed

    van Aelst, L; Jans, A W; Thevelein, J M

    1991-02-01

    Addition of glucose or related fermentable sugars to derepressed cells of the yeast Saccharomyces cerevisiae triggers a RAS-protein-mediated cAMP signal, which induces a protein phosphorylation cascade. Yeast strains without a functional CDC25 gene were deficient in basal cAMP synthesis and in the glucose-induced cAMP signal. Addition of dinitrophenol, which in wild-type strains strongly stimulates in vivo cAMP synthesis by lowering intracellular pH, did not enhance the cAMP level. cdc25 disruption mutants, in which the basal cAMP level was restored by the RAS2val19 oncogene or by disruption of the gene (PDE2) coding for the high-affinity phosphodiesterase, were still deficient in the glucose- and acidification-induced cAMP responses. These results indicate that the CDC25 gene product is required not only for basal cAMP synthesis in yeast but also for specific activation of cAMP synthesis by the signal transmission pathway leading from glucose to adenyl cyclase. They also show that intracellular acidification stimulates the pathway at or upstream of the CDC25 protein. When shifted to the restrictive temperature, cells with the temperature sensitive cdc25-5 mutation lost their cAMP content within a few minutes. After prolonged incubation at the restrictive temperature, cells with this mutation, and also those with the temperature sensitive cdc25-1 mutation, arrested at the 'start' point (in G1) of the cell cycle, and subsequently accumulated in the resting state G0. In contrast with cdc25-5 cells, however, the cAMP level did not decrease and normal glucose- and acidification-induced cAMP responses were observed when cdc25-1 cells were shifted to the restrictive temperature.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. The role of ventral striatal cAMP signaling in stress-induced behaviors

    PubMed Central

    Plattner, Florian; Hayashi, Kanehiro; Hernandez, Adan; Benavides, David R.; Tassin, Tara C.; Tan, Chunfeng; Day, Jonathan; Fina, Maggy W.; Yuen, Eunice Y.; Yan, Zhen; Goldberg, Matthew S.; Nairn, Angus C.; Greengard, Paul; Nestler, Eric J.; Taussig, Ronald; Nishi, Akinori; Houslay, Miles D.; Bibb, James A.

    2015-01-01

    The cAMP/PKA signaling cascade is a ubiquitous pathway acting downstream of multiple neuromodulators. We found that the phosphorylation of phosphodiesterase-4 (PDE4) by cyclin-dependent protein kinase 5 (Cdk5) facilitates cAMP degradation and homeostasis of cAMP/PKA signaling. In mice, loss of Cdk5 throughout the forebrain elevated cAMP levels and increased PKA activity in striatal neurons, and altered behavioral responses to acute or chronic stressors. Ventral striatum- or D1 dopamine receptor-specific conditional knockout of Cdk5, or ventral striatum infusion of a small interfering peptide that selectively targets the regulation of PDE4 by Cdk5, all produced analogical effects on stress-induced behavioral responses. Together, our results demonstrate that altering cAMP signaling in medium spiny neurons of the ventral striatum can effectively modulate stress-induced behavioral states. We propose that targeting the Cdk5 regulation of PDE4 could be a new therapeutic approach for clinical conditions associated with stress, such as depression. PMID:26192746

  10. Autosomal-dominant striatal degeneration is caused by a mutation in the phosphodiesterase 8B gene.

    PubMed

    Appenzeller, Silke; Schirmacher, Anja; Halfter, Hartmut; Bäumer, Sebastian; Pendziwiat, Manuela; Timmerman, Vincent; De Jonghe, Peter; Fekete, Klára; Stögbauer, Florian; Lüdemann, Peter; Hund, Margret; Quabius, Elgar Susanne; Ringelstein, E Bernd; Kuhlenbäumer, Gregor

    2010-01-01

    Autosomal-dominant striatal degeneration (ADSD) is an autosomal-dominant movement disorder affecting the striatal part of the basal ganglia. ADSD is characterized by bradykinesia, dysarthria, and muscle rigidity. These symptoms resemble idiopathic Parkinson disease, but tremor is not present. Using genetic linkage analysis, we have mapped the causative genetic defect to a 3.25 megabase candidate region on chromosome 5q13.3-q14.1. A maximum LOD score of 4.1 (Theta = 0) was obtained at marker D5S1962. Here we show that ADSD is caused by a complex frameshift mutation (c.94G>C+c.95delT) in the phosphodiesterase 8B (PDE8B) gene, which results in a loss of enzymatic phosphodiesterase activity. We found that PDE8B is highly expressed in the brain, especially in the putamen, which is affected by ADSD. PDE8B degrades cyclic AMP, a second messenger implied in dopamine signaling. Dopamine is one of the main neurotransmitters involved in movement control and is deficient in Parkinson disease. We believe that the functional analysis of PDE8B will help to further elucidate the pathomechanism of ADSD as well as contribute to a better understanding of movement disorders.

  11. Inhibition of Phosphodiesterase-4 during Pneumococcal Pneumonia Reduces Inflammation and Lung Injury in Mice.

    PubMed

    Tavares, Luciana P; Garcia, Cristiana C; Vago, Juliana P; Queiroz-Junior, Celso M; Galvão, Izabela; David, Bruna A; Rachid, Milene A; Silva, Patrícia M R; Russo, Remo C; Teixeira, Mauro M; Sousa, Lirlândia P

    2016-07-01

    Pneumococcal pneumonia is a leading cause of mortality worldwide. The inflammatory response to bacteria is necessary to control infection, but it may also contribute to tissue damage. Phosphodiesterase-4 inhibitors, such as rolipram (ROL), effectively reduce inflammation. Here, we examined the impact of ROL in a pneumococcal pneumonia murine model. Mice were infected intranasally with 10(5)-10(6) CFU of Streptococcus pneumoniae, treated with ROL in a prophylactic or therapeutic schedule in combination, or not, with the antibiotic ceftriaxone. Inflammation and bacteria counts were assessed, and ex vivo phagocytosis assays were performed. ROL treatment during S. pneumoniae infection decreased neutrophil recruitment into lungs and airways and reduced lung injury. Prophylactic ROL treatment also decreased cytokine levels in the airways. Although modulation of inflammation by ROL ameliorated pneumonia, bacteria burden was not reduced. On the other hand, antibiotic therapy reduced bacteria without reducing neutrophil infiltration, cytokine level, or lung injury. Combined ROL and ceftriaxone treatment decreased lethality rates and was more efficient in reducing inflammation, by increasing proresolving protein annexin A1 (AnxA1) expression, and bacterial burden by enhancing phagocytosis. Lack of AnxA1 increased inflammation and lethality induced by pneumococcal infection. These data show that immunomodulatory effects of phosphodiesterase-4 inhibitors are useful during severe pneumococcal pneumonia and suggest their potential benefit as adjunctive therapy during infectious diseases. PMID:26677751

  12. The c-di-GMP phosphodiesterase BifA regulates biofilm development in Pseudomonas putida.

    PubMed

    Jiménez-Fernández, Alicia; López-Sánchez, Aroa; Calero, Patricia; Govantes, Fernando

    2015-02-01

    We previously showed the isolation of biofilmpersistent Pseudomonas putida mutants that fail to undergo biofilm dispersal upon entry in stationary phase. Two such mutants were found to bear insertions in PP0914, encoding a GGDEF/EAL domain protein with high similarity to Pseudomon asaeruginosa BifA. Here we show the phenotypic characterization of a ΔbifA mutant in P. putida KT2442.This mutant displayed increased biofilm and pellicle formation, cell aggregation in liquid medium and decreased starvation-induced biofilm dispersal relative to the wild type. Unlike its P. aeruginosa counterpart, P. putida BifA did not affect swarming motility. The hyperadherent phenotype of the ΔbifA mutant correlates with a general increase in cyclic diguanylate (c-di-GMP) levels, Congo Red-binding exopolyaccharide production and transcription of the adhesin-encoding lapA gene. Integrity of the EAL motif and a modified GGDEF motif (altered to GGDQF)were crucial for BifA activity, and c-di-GMP depletion by overexpression of a heterologous c-di-GMP phosphodiesterase in the ΔbifA mutant restored wild-type biofilm dispersal and lapA expression.Our results indicate that BifA is a phosphodiesterase involved in the regulation of the c-di-GMP pool and required for the generation of the low c-di-GMP signal that triggers starvation-induced biofilm dispersal.

  13. Autosomal-Dominant Striatal Degeneration Is Caused by a Mutation in the Phosphodiesterase 8B Gene

    PubMed Central

    Appenzeller, Silke; Schirmacher, Anja; Halfter, Hartmut; Bäumer, Sebastian; Pendziwiat, Manuela; Timmerman, Vincent; De Jonghe, Peter; Fekete, Klára; Stögbauer, Florian; Lüdemann, Peter; Hund, Margret; Quabius, Elgar Susanne; Ringelstein, E. Bernd; Kuhlenbäumer, Gregor

    2010-01-01

    Autosomal-dominant striatal degeneration (ADSD) is an autosomal-dominant movement disorder affecting the striatal part of the basal ganglia. ADSD is characterized by bradykinesia, dysarthria, and muscle rigidity. These symptoms resemble idiopathic Parkinson disease, but tremor is not present. Using genetic linkage analysis, we have mapped the causative genetic defect to a 3.25 megabase candidate region on chromosome 5q13.3-q14.1. A maximum LOD score of 4.1 (Θ = 0) was obtained at marker D5S1962. Here we show that ADSD is caused by a complex frameshift mutation (c.94G>C+c.95delT) in the phosphodiesterase 8B (PDE8B) gene, which results in a loss of enzymatic phosphodiesterase activity. We found that PDE8B is highly expressed in the brain, especially in the putamen, which is affected by ADSD. PDE8B degrades cyclic AMP, a second messenger implied in dopamine signaling. Dopamine is one of the main neurotransmitters involved in movement control and is deficient in Parkinson disease. We believe that the functional analysis of PDE8B will help to further elucidate the pathomechanism of ADSD as well as contribute to a better understanding of movement disorders. PMID:20085714

  14. Phosphodiesterase 5 inhibitors augment UT-15C-stimulated ATP release from erythrocytes of humans with pulmonary arterial hypertension

    PubMed Central

    Bowles, Elizabeth A; Moody, Gina N; Yeragunta, Yashaswini; Stephenson, Alan H; Ellsworth, Mary L

    2015-01-01

    Both prostacyclin analogs and phosphodiesterase 5 (PDE5) inhibitors are effective treatments for pulmonary arterial hypertension (PAH). In addition to direct effects on vascular smooth muscle, prostacyclin analogs increase cAMP levels and ATP release from healthy human erythrocytes. We hypothesized that UT-15C, an orally available form of the prostacyclin analog, treprostinil, would stimulate ATP release from erythrocytes of humans with PAH and that this release would be augmented by PDE5 inhibitors. Erythrocytes were isolated and the effect of UT-15C on cAMP levels and ATP release were measured in the presence and absence of the PDE5 inhibitors, zaprinast or tadalafil. In addition, the ability of a soluble guanylyl cyclase inhibitor to prevent the effects of tadalafil was determined. Erythrocytes of healthy humans and humans with PAH respond to UT-15C with increases in cAMP levels and ATP release. In both groups, UT-15C-induced ATP release was potentiated by zaprinast and tadalafil. The effect of tadalafil was prevented by pre-treatment with an inhibitor of soluble guanylyl cyclase in healthy human erythrocytes. Importantly, UT-15C-induced ATP release was greater in PAH erythrocytes than in healthy human erythrocytes in both the presence and the absence of PDE5 inhibitors. The finding that prostacyclin analogs and PDE5 inhibitors work synergistically to enhance release of the potent vasodilator ATP from PAH erythrocytes provides a new rationale for the co-administration of these drugs in this disease. Moreover, these results suggest that the erythrocyte is a novel target for future drug development for the treatment of PAH. PMID:25125498

  15. Chronic lymphocytic leukemia and B and T cells differ in their response to cyclic nucleotide phosphodiesterase inhibitors.

    PubMed

    Meyers, John A; Su, Derrick W; Lerner, Adam

    2009-05-01

    Phosphodiesterase (PDE)4 inhibitors, which activate cAMP signaling by reducing cAMP catabolism, are known to induce apoptosis in B lineage chronic lymphocytic leukemia (CLL) cells but not normal human T cells. The explanation for such differential sensitivity remains unknown. In this study, we report studies contrasting the response to PDE4 inhibitor treatment in CLL cells and normal human T and B cells. Affymetrix gene chip analysis in the three cell populations following treatment with the PDE4 inhibitor rolipram identified a set of up-regulated transcripts with unusually high fold changes in the CLL samples, several of which are likely part of compensatory negative feedback loops. The high fold changes were due to low basal transcript levels in CLL cells, suggesting that cAMP-mediated signaling may be unusually tightly regulated in this cell type. Rolipram treatment augmented cAMP levels and induced ATF-1/CREB serine 63/133 phosphorylation in both B lineage cell types but not T cells. As treatment with the broad-spectrum PDE inhibitor 3-isobutyl-1-methylxanthine induced T cell CREB phosphorylation, we tested a series of family-specific PDE inhibitors for their ability to mimic 3-isobutyl-1-methylxanthine-induced ATF-1/CREB phosphorylation. Whereas PDE3 inhibitors alone had no effect, the combination of PDE3 and PDE4 inhibitors induced ATF-1/CREB serine 63/133 phosphorylation in T cells. Consistent with this observation, PDE3B transcript and protein levels were low in CLL cells but easily detectable in T cells. Combined PDE3/4 inhibition did not induce T cell apoptosis, suggesting that cAMP-mediated signal transduction that leads to robust ATF-1/CREB serine 63/133 phosphorylation is not sufficient to induce apoptosis in this lymphoid lineage.

  16. The ACA Standards Organizer. A Tool for the Preparation of Camps for Visitation for American Camping Association Camp Accreditation and Site Approval.

    ERIC Educational Resources Information Center

    Chenery, Mary Faeth

    This comprehensive guide helps camp administrators prepare for camp accreditation and site approval visits by the American Camping Association (ACA). The introductory sections outline the purpose and administration of the standards program and provide definitions of camping terms. Standards are organized under one section for camp accreditation…

  17. Escherichia coli K-12 YfgF is an anaerobic cyclic di-GMP phosphodiesterase with roles in cell surface remodelling and the oxidative stress response.

    PubMed

    Lacey, Melissa M; Partridge, Jonathan D; Green, Jeffrey

    2010-09-01

    The Escherichia coli K-12 yfgF gene encodes a protein with domains associated with cyclic di-GMP signalling: GGDEF (associated with diguanylate cyclase activity) and EAL (associated with cyclic di-GMP phosphodiesterase activity). Here, it is shown that yfgF is expressed under anaerobic conditions from a class II FNR (regulator of fumarate and nitrate reduction)-dependent promoter. Anaerobic expression of yfgF is greatest in stationary phase, and in cultures grown at 28 degrees C, suggesting that low growth rates promote yfgF expression. Mutation of yfgF resulted in altered cell surface properties and enhanced sensitivity when anaerobic cultures were exposed to peroxides. The purified YfgF GGDEF-EAL (YfgF(GE)) and EAL (YfgF(E)) domains possessed cyclic di-GMP-specific phosphodiesterase activity, but lacked diguanylate cyclase activity. However, the catalytically inactive GGDEF domain was required for YfgF(GE) dimerization and enhanced cyclic di-GMP phosphodiesterase activity in the presence of physiological concentrations of Mg(2+). The cyclic di-GMP phosphodiesterase activity of YfgF(GE) and YfgF(E) was inhibited by the product of the reaction, 5'-phosphoguanylyl-(3'-5')-guanosine (pGpG). Thus, it is shown that the yfgF gene encodes an anaerobic cyclic di-GMP phosphodiesterase that is involved in remodelling the cell surface of E. coli K-12 and in the response to peroxide shock, with implications for integrating three global regulatory networks, i.e. oxygen regulation, cyclic di-GMP signalling and the oxidative stress response.

  18. cAMP/PKA/CREB/GLT1 signaling involved in the antidepressant-like effects of phosphodiesterase 4D inhibitor (GEBR-7b) in rats

    PubMed Central

    Liu, Xu; Guo, Haibiao; Sayed, Mohammad Daud SOM; Lu, Yang; Yang, Ting; Zhou, Dongsheng; Chen, Zhongming; Wang, Haitao; Wang, Chuang; Xu, Jiangping

    2016-01-01

    Objectives GEBR-7b, a potential phosphodiesterase 4D inhibitor, has been shown to have memory-enhancing effects in rodents. However, it is still unknown whether GEBR-7b also has the antidepressant-like effects in rats. Herein, we examined the potential of GEBR-7b to attenuate depression-like behaviors in the rat model of depression induced by chronic unpredictable stress (CUS). Next, we also investigated the alterations of cyclic adenosine monophosphate (cAMP), protein kinase A (PKA) catalytic subunit (PKAca), cAMP response element-binding (CREB), and glutamate transporter 1 (GLT1) levels produced by GEBR-7b in the rats model of depression. Methods Effects of GEBR-7b on CUS (35 days)-induced depression-like behaviors were examined by measuring immobility time in the forced swimming test (FST). Hippocampal cAMP levels were examined by enzyme-linked immunosorbent assay, whereas PKAca, phosphorylation of CREB (pCREB), CREB, and GLT1 in the hippocampus of rats were subjected to Western blot analysis. Results CUS exposure caused a depression-like behavior evidenced by the increased immobility time in FST. Depression-like behavior induced by CUS was accompanied by a significant increased GLT, decreased cAMP, PKAca, pCREB activities in hippocampus. However, repeated GEBR-7b administration significantly reversed CUS-induced depression-like behavior and changes of cAMP/PKA/CREB/GLT1 signaling. No alteration was observed in locomotor activity in open field test. Conclusion These findings indicate that GEBR-7b reversed the depression-like behaviors induced by CUS in rats, which is at least in part mediated by modulating cAMP, PKAca, pCREB, and GLT1 levels in the hippocampus of rats, supporting its neuroprotective potential against behavioral and biochemical dysfunctions induced by CUS. PMID:26855578

  19. The Pseudomonas aeruginosa Chp Chemosensory System Regulates Intracellular cAMP Levels by Modulating Adenylate Cyclase Activity

    PubMed Central

    Fulcher, Nanette B.; Holliday, Phillip M.; Klem, Erich; Cann, Martin J.; Wolfgang, Matthew C.

    2010-01-01

    Summary Multiple virulence systems in the opportunistic pathogen Pseudomonas aeruginosa are regulated by the second messenger signaling molecule adenosine 3’, 5’-cyclic monophosphate (cAMP). Production of cAMP by the putative adenylate cyclase enzyme CyaB represents a critical control point for virulence gene regulation. To identify regulators of CyaB, we screened a transposon insertion library for mutants with reduced intracellular cAMP. The majority of insertions resulting in reduced cAMP mapped to the Chp gene cluster encoding a putative chemotaxis-like chemosensory system. Further genetic analysis of the Chp system revealed that it has both positive and negative effects on intracellular cAMP and that it regulates cAMP levels by modulating CyaB activity. The Chp system was previously implicated in the production and function of type IV pili (TFP). Given that cAMP and the cAMP-dependent transcriptional regulator Vfr control TFP biogenesis gene expression, we explored the relationship between cAMP, the Chp system and TFP regulation. We discovered that the Chp system controls TFP production through modulation of cAMP while control of TFP-dependent twitching motility is cAMP-independent. Overall, our data define a novel function for a chemotaxis-like system in controlling cAMP production and establish a regulatory link between the Chp system, TFP and other cAMP-dependent virulence systems. PMID:20345659

  20. Prostaglandin E2 inhibits NLRP3 inflammasome activation through EP4 receptor and intracellular cAMP in human macrophages

    PubMed Central

    Liu, Yueqin; Martinez-Anton, Asuncion; Qi, Hai-Yan; Logun, Carolea; Alsaaty, Sara; Park, Yong Hwan; Kastner, Daniel L.; Chae, Jae Jin; Shelhamer, James H.

    2015-01-01

    Prostaglandin E2 (PGE2) is a potent lipid mediator involved in maintaining homeostasis but also promotion of acute inflammation or immune suppression in chronic inflammation and cancer. NLRP3 inflammasome plays an important role in host defense. Uncontrolled activation of NLRP3 inflammasome, due to mutations in the NLRP3 gene causes cryopyrin-associated periodic syndromes (CAPS). Here, we showed that NLRP3 inflammasome activation is inhibited by PGE2 in human primary monocyte-derived macrophages. This effect was mediated through prostaglandin E receptor 4 (EP4) and an increase in intracellular cAMP, independently of protein kinase A (PKA) or exchange protein directly activated by cAMP (Epac). A specific agonist of EP4 mimicked, while its antagonist or EP4 knockdown reversed PGE2-mediated NLRP3 inhibition. PGE2 caused an increase in intracellular cAMP. Blockade of adenylate cyclase by its inhibitor reversed PGE2-mediated NLRP3 inhibition. Increase of intracellular cAMP by an activator of adenylate cyclase or an analog of cAMP, or a blockade of cAMP degradation by phosphodiesterase inhibitor decreased NLRP3 activation. PKA or Epac agonists did not mimic and their antagonists did not reverse PGE2-mediated NLRP3 inhibition. In addition, constitutive IL-1β secretion from LPS-primed PBMCs of CAPS patients was substantially reduced by high doses of PGE2. Moreover, blocking cytosolic phospholipase A2α by its inhibitor or siRNA or inhibiting cyclooxygenase 2, resulting in inhibition of endogenous PGE2 production, caused an increase in NLRP3 inflammasome activation. Our results suggest that PGE2 might play a role in maintaining homeostasis during the resolution phase of inflammation and might serve as an autocrine and paracrine regulator. PMID:25917098

  1. Sildenafil, a selective phosphodiesterase type 5 inhibitor, enhances memory reconsolidation of an inhibitory avoidance task in mice.

    PubMed

    Boccia, M M; Blake, M G; Krawczyk, M C; Baratti, C M

    2011-07-01

    Intracellular levels of the second messengers cAMP and cGMP are maintained through a balance between production, carried out by adenyl cyclase (AC) and guanylyl cyclase (GC), and degradation, carried out by phosphodiesterases (PDEs). Recently, PDEs have gained increased attention as potential new targets for cognition enhancement, with particular reference to phosphodiesterase type 5 (PDE5A). It is accepted that once consolidation is completed memory becomes permanent, but it has also been suggested that reactivation (memory retrieval) of the original memory makes it sensitive to the same treatments that affect memory consolidation when given after training. This new period of sensitivity coined the term reconsolidation. Sildenafil (1, 3, and 10mg/kg, ip), a cGMP-PDE5 inhibitor, facilitated retention performance of a one-trial step-through inhibitory avoidance task, when administered to CF-1 male mice immediately after retrieval. The effects of sildenafil (1mg/kg, ip) were time-dependent, long-lasting and inversely correlated with memory age. The administration of sildenafil (1mg/kg, ip) 30 min prior to the 2nd retention test did not affect retention of mice given post-retrieval injections of either vehicle or sildenafil (1mg/kg, ip). Finally, an enhancement of retention was also observed in CF-1 female mice receiving sildenafil (1mg/kg, ip) immediately, but not 180 min after retrieval. In the present paper we reported for the first time that systemic administration of sildenafil after memory reactivation enhances retention performance of the original learning. Our results indirectly point out cGMP, a component of the NO/cGMP/PKG pathway, as a necessary factor for memory reconsolidation.

  2. Purification and properties of the light-activated cyclic nucleotide phosphodiesterase of rod outer segments.

    PubMed

    Miki, N; Baraban, J M; Keirns, J J; Boyce, J J; Bitensky, M W

    1975-08-25

    Frog (Rana catesbiana) rod outer segment disc membranes contain a cyclic nucleotide phosphodiesterase (EC 3.1.4.17) which is activated by light in the presence of ATP. This enzyme is firmly bound to the disc membrane, but can be eluted from the membrane with 10 mM Tris-HCl buffer, pH 7.4 and 2 mM EDTA. The eluted phosphodiesterase has reduced activity, but can be activated approximately 10-fold by polycations such as protamine and polylysine. The eluted phosphodiesterase can no longer be activated by light in the presence of ATP, that is, activation by light apparently depends on the native orientation of phosphodiesterase in relationship to other disc membrane components. The eluted phosphodiesterase was purified to homogeneity as judged by analytical polyacrylamide gel electrophoresis and polyacrylamide gel isoelectric focusing. The over-all purification from intact retina was approximately 925-fold. The purification of phosphodiesterase from the isolated rod outer segment preparation was about 185-fold with a 28% yield. Phosphodiesterase accounts for approximately 0.5% of the disc membrane protein. The eluted phosphodiesterase (inactive form) has a sedimentation coefficient of 12.4 S corresponding to an approximate molecular weight of 240,000. Sodium dodecyl sulfate polyacrylamide gel electrophoresis separates the purified phosphodiesterase into two subunits of 120,000 and 110,000 daltons. With cyclic 3':5'-GMP (cGMP) as substrate the Km for the purified phosphodiesterase is 70 muM. Protamine increases the Vmax without changing the Km for cGMP. The isoelectric point (pI) of the native dimer is 5.7. Limited exposure of the eluted phosphodiesterase (inactive form) to trypsin produces a somewhat greater activation than is obtained with 0.5 mg/ml of protamine. The trypsin-activated phosphodiesterase has a sedimentation coefficient of 7.8 S corresponding to an approximate molecular weight of 170,000. The 110,000-dalton subunit is much less sensitive to trypsin

  3. US SPACE CAMP CALIFORNIA - DAY CAMP GRAND OPENING WITH KEVIN JONES (WHISMAN SCHOOL) AND LUCRETIA

    NASA Technical Reports Server (NTRS)

    1996-01-01

    US SPACE CAMP CALIFORNIA - DAY CAMP GRAND OPENING WITH KEVIN JONES (WHISMAN SCHOOL) AND LUCRETIA SUTHERLIN (MCNAIR SCHOOL). AMES SPONSORED STUDENTS AND RACHAEL QUIRING (STAFF) - AUTOGRAPH SIGNING BY Astronaut Wally Schirra

  4. Central Atlantic Magmatic Province (CAMP): The Palisade Sill Connection

    NASA Astrophysics Data System (ADS)

    Ghatak, A.; Basu, A. R.

    2012-12-01

    The extensively studied 200Ma Central Atlantic Magmatic Province (CAMP) is considered to be the world's largest continental Large Igneous Province (LIP) covering up to 7 X 106 km2. This igneous province has been linked to the ~200Ma Mesozoic opening of the Central Atlantic Ocean. This opening fragmented the CAMP into several segments that occur on four different tectonic plates today. The CAMP related LIP is different from others in that it constitutes almost entirely of dikes and sills with sparse volcanic outflows. The 200 Ma Palisade Sill, exposed along the Hudson River in northeastern North America is an expression of the CAMP magmatism. On the basis of similar ages of eruption, Palisade Sill tholeiites have been correlated to other CAMP exposures in four continents. We provide an isotopic tracer study of the Palisade Sill basalts and relate them to low-Ti (<2 wt %) CAMP related tholeiites from North and South America, western Europe, and West Africa. We report Nd-Sr-Pb isotopic and multiple trace element data of 19 basalts and gabbros, 3 chilled margin basalts, and 4 sandstones spanning the entire length and thickness of the Palisade Sill in New York and New Jersey. These geochemical data are essential to understand the relationship between mantle geodynamic processes involved in the generation of the CAMP tholeiites prior to the formation of the of the Atlantic Ocean crust. The Palisade Sill basalts of this study yield the typical composition of low-Ti CAMP tholeiites with small LREE enrichments (LaN/SmN = 1.7 to 2.3), radiogenic Sr and negative ɛNd(I) values (87Sr/87Sr(I) = 0.70668 to 0.71037; ɛNd(I) = -0.64 to -3.8), and Pb-isotopic ratios (e.g. 206Pb/204Pb = 18.11 to 18.69) above the NHRL and subparallel to it. These geochemical data indicate the Palisade Sill basalts were derived from a slightly enriched OIB-like mantle source. Further, these rocks were derived by ~15% melting of a slightly depleted spinel peridotite with up to 20% contamination by the

  5. Stories from Camp: Understanding the Impact of What We Do.

    ERIC Educational Resources Information Center

    DeGraaf, Don; Glover, Jessie

    2002-01-01

    A study examining the impacts of camp on staff interviewed 29 former seasonal camp staff. All respondents reported positive benefits in their personal and professional lives and the strong influence of camp in shaping career choices. Reflections on camp fell into three categories: uniqueness of camp, making memories for kids, and freedom. (TD)

  6. A Day in the Life of Three Special Needs Camps

    ERIC Educational Resources Information Center

    Winbaum, Stephen

    2006-01-01

    In this article, the author talks about three special needs camps--Camp Kirk, Camp Talisman and Camp Caglewood. Camp Kirk's philosophy is to encourage their children to take risks in a structured setting, like high ropes courses, rock climbing wall, martial arts, and traditional activities like swimming, arts and craft, drama, and others. Once…

  7. Inhibition of phosphodiesterase 4 reduces ethanol intake and preference in C57BL/6J mice.

    PubMed

    Blednov, Yuri A; Benavidez, Jillian M; Black, Mendy; Harris, R Adron

    2014-01-01

    Some anti-inflammatory medications reduce alcohol consumption in rodent models. Inhibition of phosphodiesterases (PDE) increases cAMP and reduces inflammatory signaling. Rolipram, an inhibitor of PDE4, markedly reduced ethanol intake and preference in mice and reduced ethanol seeking and consumption in alcohol-preferring fawn-hooded rats (Hu et al., 2011; Wen et al., 2012). To determine if these effects were specific for PDE4, we compared nine PDE inhibitors with different subtype selectivity: propentofylline (nonspecific), vinpocetine (PDE1), olprinone, milrinone (PDE3), zaprinast (PDE5), rolipram, mesopram, piclamilast, and CDP840 (PDE4). Alcohol intake was measured in C57BL/6J male mice using 24-h two-bottle choice and two-bottle choice with limited (3-h) access to alcohol. Only the selective PDE4 inhibitors reduced ethanol intake and preference in the 24-h two-bottle choice test. For rolipram, piclamilast, and CDP840, this effect was observed after the first 6 h but not after the next 18 h. Mesopram, however, produced a long-lasting reduction of ethanol intake and preference. In the limited access test, rolipram, piclamilast, and mesopram reduced ethanol consumption and total fluid intake and did not change preference for ethanol, whereas CDP840 reduced both consumption and preference without altering total fluid intake. Our results provide novel evidence for a selective role of PDE4 in regulating ethanol drinking in mice. We suggest that inhibition of PDE4 may be an unexplored target for medication development to reduce excessive alcohol consumption.

  8. Treatment of Cognitive Impairment in Schizophrenia: Potential Value of Phosphodiesterase Inhibitors in Prefrontal Dysfunction.

    PubMed

    Duinen, Marlies Van; Reneerkens, Olga A H; Lambrecht, Lena; Sambeth, Anke; Rutten, Bart P F; Os, Jim Van; Blokland, Arjan; Prickaerts, Jos

    2015-01-01

    No pharmacological treatment is available to date that shows satisfactory effects on cognitive symptoms in patients diagnosed with schizophrenia. Phosphodiesterase inhibitors (PDE-Is) improve neurotransmitter signaling by interfering in intracellular second messenger cascades. By preventing the breakdown of cAMP and/or cGMP, central neurotransmitter activity is maintained. Different PDE families exist with distinct characteristics among which substrate specificity and regional distribution. Preclinical data is promising especially with regard to inhibition of PDE2, PDE4, PDE5 and PDE10. In addition, cognitive improvement has been reported in both elderly and/or non-impaired young human subjects after PDE1 or PDE4 inhibition. Moreover, some of these studies show effects on cognitive domains relevant to schizophrenia, in particular memory. The current review incorporates an overview of the distinct molecular characteristics of the different PDE families and their relationship to the neurobiological mechanisms related to cognitive dysfunction in schizophrenia. So far, procognitive effects of only three types of PDE-Is have been assessed in patients diagnosed with schizophrenia inhibiting PDE3, PDE5 and PDE10. However, the limited data available do not allow to draw firm conclusions on the value of PDE-Is as cognitive enhancers in schizophrenia yet. The field is still in its infancy, but nevertheless different PDE-Is seem promising as candidate to optimise neural communication in the prefrontal cortex favouring cognitive functioning in patients diagnosed with schizophrenia, in particular dual inhibitors including PDE1-Is, PDE3-Is and PDE10A-Is.

  9. Selective phosphodiesterase inhibitors improve performance on the ED/ID cognitive task in rats.

    PubMed

    Rodefer, Joshua S; Saland, Samantha K; Eckrich, Samuel J

    2012-03-01

    A number of selective phosphodiesterase (PDE) inhibitors have been demonstrated to improve learning in several rodent models of cognition. Given that schizophrenia is associated with impairments in frontal lobe-dependent cognitive functions (e.g., working memory and cognitive flexibility), we examined whether PDE inhibitors would attenuate cognitive deficits associated with schizophrenia. Persistent suppression of N-methyl-D-aspartate (NMDA) receptor function produces enduring structural changes in neocortical and limbic regions in a pattern similar to changes reported in schizophrenia. This similarity suggests that subchronic treatment with NMDA receptor antagonists (e.g., phencyclidine, PCP) may represent a useful preclinical model of neurobiological and related cognitive deficits associated with schizophrenia. We treated male Long-Evans rats with subchronic PCP (5 mg/kg, ip, BID, 7 d) or saline and then examined the effects of acute treatment with selected doses of PDE inhibitors that have been demonstrated to regulate both intracellular levels of cAMP and/or cGMP, and to improve cognitive function. We used an extradimensional-intradimensional (ED/ID) test of cognitive flexibility similar to those used in humans and non-human primates for assessing executive function. Subchronic treatment with PCP produced a selective impairment on ED shift (EDS) performance without significant impairment on any other discrimination problem when compared to saline-treated control animals. Selected doses of the four PDEIs evaluated (PDE2: BAY 60-7550; PDE4: rolipram; PDE5: sildenafil; PDE10A: papaverine) were able to significantly attenuate this cognitive deficit in EDS performance. This suggests that this rodent model of executive function was sensitive to pro-cognitive effects of intracellular effects resulting from PDE inhibition. Together, these data suggest that inhibition of PDE activity may represent valuable therapeutic targets to improve cognition associated with

  10. Kinetic and Structural Studies of Phosphodiesterase-8A and Implication on the Inhibitor Selectivity

    SciTech Connect

    Wang, H.; Yan, Z; Yang, S; Cai, J; Robinson, H; Ke, H

    2008-01-01

    Cyclic nucleotide phosphodiesterase-8 (PDE8) is a family of cAMP-specific enzymes and plays important roles in many biological processes, including T-cell activation, testosterone production, adrenocortical hyperplasia, and thyroid function. However, no PDE8 selective inhibitors are available for trial treatment of human diseases. Here we report kinetic properties of the highly active PDE8A1 catalytic domain prepared from refolding and its crystal structures in the unliganded and 3-isobutyl-1-methylxanthine (IBMX) bound forms at 1.9 and 2.1 Angstroms resolutions, respectively. The PDE8A1 catalytic domain has a KM of 1.8 eM, Vmax of 6.1 emol/min/mg, a kcat of 4.0 s-1 for cAMP, and a KM of 1.6 mM, Vmax of 2.5 emol/min/mg, a kcat of 1.6 s-1 for cGMP, thus indicating that the substrate specificity of PDE8 is dominated by KM. The structure of the PDE8A1 catalytic domain has similar topology as those of other PDE families but contains two extra helices around Asn685-Thr710. Since this fragment is distant from the active site of the enzyme, its impact on the catalysis is unclear. The PDE8A1 catalytic domain is insensitive to the IBMX inhibition (IC50 = 700 eM). The unfavorable interaction of IBMX in the PDE8A1-IBMX structure suggests an important role of Tyr748 in the inhibitor binding. Indeed, the mutation of Tyr748 to phenylalanine increases the PDE8A1 sensitivity to several nonselective or family selective PDE inhibitors. Thus, the structural and mutagenesis studies provide not only insight into the enzymatic properties but also guidelines for design of PDE8 selective inhibitors.

  11. Program evaluation of Protovation Camp

    NASA Astrophysics Data System (ADS)

    Healy, Laurel Lynell Martin

    The purpose of this program evaluation was to determine the extent to which Protovation Camp utilized the combined resources of multiple institutions to impact student learning in science, technology, engineering, and math. The partnership consisted of multiple institutions: the university, providing graduate students to facilitate inquiry-based lessons; the science center, allowing the use of their facilities and resources; and the elementary school, contributing rising third through fifth grade campers. All of these components were examined. The mixed-methods approach used post hoc quantitative data for campers, which consisted of pre-test and post-test scores on the Test of Science-Related Attitudes (TOSRA), the Draw-A-Scientist Test, and content tests based on the camp activities. Additionally, TOSRA scores and current survey results for the graduate students were used along with qualitative data collected from plusdelta charts to determine the impact of participation in Protovation Camp on teachers and students. Results of the program evaluation indicated that when students were taught inquiry-based lessons that ignite wonder, both their attitudes toward science and their knowledge about science improved. An implication for teacher preparation programs was that practicing inquiry-based lessons on actual students (campers) was an important component for teachers (graduate students) as they prepare to positively impact student learning in their own classrooms. Immediate feedback from the campers in the form of pre-test and post-test scores and from peers on plusdelta charts allowed the graduate students the opportunity to make needed adjustments to improve effectiveness before using the lesson with a new set of campers or later in their own classrooms. Keywords. Teacher preparation, Inquiry-based instruction, STEM instructions, University and museum partnerships

  12. Basic Camp Director Education Course. Student Guide to Home Study. Camp Administration Series.

    ERIC Educational Resources Information Center

    Williams, Teri

    Project STRETCH's (Strategies to Try out Resources to Enhance the Training of Camp directors serving the Handicapped) home study student guide for the basic camp director education course begins with a brief course overview, the desired outcomes of camp director education, instructions on phases I and II of home study, a student needs assessment…

  13. Themes from a Camp Maintenance Network: Camp Maintenance and Property Personnel Share Their Insights and Challenges.

    ERIC Educational Resources Information Center

    Whyman, Wynne

    2003-01-01

    A camp maintenance survey was completed by maintenance personnel from 99 camps. Results highlighted several important considerations: ensuring sufficient maintenance funds for aging infrastructure, including camp/property personnel in decision making, publicizing completed maintenance projects, examining long-term needs of the land, and adopting…

  14. Camp Programs Provide Community Opportunities: Camp Henry Helps Community with Youth Diversion Program.

    ERIC Educational Resources Information Center

    Vander Kooi, Gregory; Astle, Judy Hughes; Jacobs, Jeff

    2001-01-01

    A Michigan program gives juvenile first-offenders in alcohol, drug, or tobacco crimes the option of completing a youth diversion program at a local camp as an alternative to traditional juvenile justice. Considerations for the camp included board support, compatibility with camp philosophy, and staff competence. The program has lower recidivism…

  15. PSYCHOLOGICAL ANALYSIS OF CAMP ACTIVITIES IN SELECTED KENNEDY FOUNDATION SPONSORED CAMPS FOR THE MENTALLY RETARDED.

    ERIC Educational Resources Information Center

    PAINTER, GENEVIEVE

    RECREATIONAL ACTIVITIES OBSERVED AT SIX SUMMER DAY CAMPS (REPRESENTATIVE OF 26 SUCH CAMPS SPONSORED BY THE KENNEDY FOUNDATION) ARE REPORTED. EACH CAMP WAS VISITED AND THE FIRST 25 ACTIVITIES PRESENTED WERE ANALYZED BY ONE OF TWO THEORETICAL MODELS. THE MODEL FOR MEANINGFUL (COGNITIVE) ACTIVITIES WAS USED TO RATE ACTIVITIES IN TERMS OF…

  16. Engineering of Bacillus subtilis Strains To Allow Rapid Characterization of Heterologous Diguanylate Cyclases and Phosphodiesterases

    PubMed Central

    Gao, Xiaohui; Dong, Xiao; Subramanian, Sundharraman; Matthews, Paige M.; Cooper, Caleb A.; Kearns, Daniel B.

    2014-01-01

    Microbial processes, including biofilm formation, motility, and virulence, are often regulated by changes in the available concentration of cyclic dimeric guanosine monophosphate (c-di-GMP). Generally, high c-di-GMP concentrations are correlated with decreased motility and increased biofilm formation and low c-di-GMP concentrations are correlated with an increase in motility and activation of virulence pathways. The study of c-di-GMP is complicated, however, by the fact that organisms often encode dozens of redundant enzymes that synthesize and hydrolyze c-di-GMP, diguanylate cyclases (DGCs), and c-di-GMP phosphodiesterases (PDEs); thus, determining the contribution of any one particular enzyme is challenging. In an effort to develop a facile system to study c-di-GMP metabolic enzymes, we have engineered a suite of Bacillus subtilis strains to assess the effect of individual heterologously expressed proteins on c-di-GMP levels. As a proof of principle, we characterized all 37 known genes encoding predicted DGCs and PDEs in Clostridium difficile using parallel readouts of swarming motility and fluorescence from green fluorescent protein (GFP) expressed under the control of a c-di-GMP-controlled riboswitch. We found that 27 of the 37 putative C. difficile 630 c-di-GMP metabolic enzymes had either active cyclase or phosphodiesterase activity, with agreement between our motility phenotypes and fluorescence-based c-di-GMP reporter. Finally, we show that there appears to be a threshold level of c-di-GMP needed to inhibit motility in Bacillus subtilis. PMID:25085482

  17. AKAP79, PKC, PKA and PDE4 participate in a Gq-linked muscarinic receptor and adenylate cyclase 2 cAMP signalling complex

    PubMed Central

    Shen, Jia X.; Cooper, Dermot M. F.

    2014-01-01

    AC2 (adenylate cyclase 2) is stimulated by activation of Gq-coupled muscarinic receptors through PKC (protein kinase C) to generate localized cAMP in HEK (human embryonic kidney)-293 cells. In the present study, we utilized a sensitive live-cell imaging technique to unravel the proteins that play essential roles in a Gq-coupled muscarinic receptor-mediated cAMP signalling complex. We reveal that, upon agonist binding to the Gq-coupled muscarinic receptor, AKAP79 (A-kinase-anchoring protein 79) recruits PKC to activate AC2 to produce cAMP. The cAMP formed is degraded by PDE4 (phosphodiesterase 4) activated by an AKAP-anchored PKA (protein kinase A). Calcineurin, a phosphatase bound to AKAP79, is not involved in this regulation. Overall, a transient cAMP increase is generated from AC2 by Gq-coupled muscarinic receptor activation, subject to sophisticated regulation through AKAP79, PKC, PDE4 and PKA, which significantly enhances acetylcholine-mediated signalling. PMID:23889134

  18. Sequence analysis and characterization of a 40-kilodalton Borrelia hermsii glycerophosphodiester phosphodiesterase homolog.

    PubMed Central

    Shang, E S; Skare, J T; Erdjument-Bromage, H; Blanco, D R; Tempst, P; Miller, J N; Lovett, M A

    1997-01-01

    We report the purification, molecular cloning, and characterization of a 40-kDa glycerophosphodiester phosphodiesterase homolog from Borrelia hermsii. The 40-kDa protein was solubilized from whole organisms with 0.1% Triton X-100, phase partitioned into the Triton X-114 detergent phase, and purified by fast-performance liquid chromatography (FPLC). The gene encoding the 40-kDa protein was cloned from a B. hermsii chromosomal DNA lambda EXlox expression library and identified by using affinity antibodies generated against the purified native protein. The deduced amino acid sequence included a 20-amino-acid signal peptide encoding a putative leader peptidase II cleavage site, indicating that the 40-kDa protein was a lipoprotein. Based on significant homology (31 to 52% identity) of the 40-kDa protein to glycerophosphodiester phosphodiesterases of Escherichia coli (GlpQ), Bacillus subtilis (GlpQ), and Haemophilus influenzae (Hpd; protein D), we have designated this B. hermsii 40-kDa lipoprotein a glycerophosphodiester phosphodiesterase (Gpd) homolog, the first B. hermsii lipoprotein to have a putative functional assignment. A nonlipidated form of the Gpd homolog was overproduced as a fusion protein in E. coli BL21(DE3)(pLysE) and was used to immunize rabbits to generate specific antiserum. Immunoblot analysis with anti-Gpd serum recognized recombinant H. influenzae protein D, and conversely, antiserum to H. influenzae protein D recognized recombinant B. hermsii Gpd (rGpd), indicating antigenic conservation between these proteins. Antiserum to rGpd also identified native Gpd as a constituent of purified outer membrane vesicles prepared from B. hermsii. Screening of other pathogenic spirochetes with anti-rGpd serum revealed the presence of antigenically related proteins in Borrelia burgdorferi, Treponema pallidum, and Leptospira kirschneri. Further sequence analysis both upstream and downstream of the Gpd homolog showed additional homologs of glycerol metabolism

  19. Cis-Lunar Base Camp

    NASA Technical Reports Server (NTRS)

    Merrill, Raymond G.; Goodliff, Kandyce E.; Mazanek, Daniel D.; Reeves, John D., Jr.

    2012-01-01

    Historically, when mounting expeditions into uncharted territories, explorers have established strategically positioned base camps to pre-position required equipment and consumables. These base camps are secure, safe positions from which expeditions can depart when conditions are favorable, at which technology and operations can be tested and validated, and facilitate timely access to more robust facilities in the event of an emergency. For human exploration missions into deep space, cis-lunar space is well suited to serve as such a base camp. The outer regions of cis-lunar space, such as the Earth-Moon Lagrange points, lie near the edge of Earth s gravity well, allowing equipment and consumables to be aggregated with easy access to deep space and to the lunar surface, as well as more distant destinations, such as near-Earth Asteroids (NEAs) and Mars and its moons. Several approaches to utilizing a cis-lunar base camp for sustainable human exploration, as well as some possible future applications are identified. The primary objective of the analysis presented in this paper is to identify options, show the macro trends, and provide information that can be used as a basis for more detailed mission development. Compared within are the high-level performance and cost of 15 preliminary cis-lunar exploration campaigns that establish the capability to conduct crewed missions of up to one year in duration, and then aggregate mass in cis-lunar space to facilitate an expedition from Cis-Lunar Base Camp. Launch vehicles, chemical propulsion stages, and electric propulsion stages are discussed and parametric sizing values are used to create architectures of in-space transportation elements that extend the existing in-space supply chain to cis-lunar space. The transportation options to cis-lunar space assessed vary in efficiency by almost 50%; from 0.16 to 0.68 kg of cargo in cis-lunar space for every kilogram of mass in Low Earth Orbit (LEO). For the 15 cases, 5-year campaign

  20. Youth Development and the Camp Experience

    ERIC Educational Resources Information Center

    Garst, Barry A.; Browne, Laurie P.; Bialeschki, M. Deborah

    2011-01-01

    The organized camp experience has been an important part of the lives of children, youth, and adults for over 150 years and is a social institution that touches more lives than any other except for schools. Camp is more than a location or a program; it encompasses the affective, cognitive, behavioral, physical, social, and spiritual benefits that…

  1. Summer Camp of Mathematical Modeling in China

    ERIC Educational Resources Information Center

    Tian, Xiaoxi; Xie, Jinxing

    2013-01-01

    The Summer Camp of Mathematical Modeling in China is a recently created experience designed to further Chinese students' academic pursuits in mathematical modeling. Students are given more than three months to research on a mathematical modeling project. Researchers and teams with outstanding projects are invited to the Summer Camp to present…

  2. Tech Camp Unleashes Creativity and Collaboration

    ERIC Educational Resources Information Center

    Bardin, Joe

    2008-01-01

    Each August, teachers from around the state gather for the Arizona K-12 Center's Tech Camp, a week-long immersion in technology for the classroom. The Arizona K-12 Center's mission is to improve teaching and learning in Arizona's schools through high-quality professional development and teacher leadership. The formula Tech Camp follows is a simple…

  3. Science Camp: Just for the Girls

    ERIC Educational Resources Information Center

    Cavanagh, Sean

    2007-01-01

    Research shows that girls tend to lose interest in science and math as they move through the education pipeline--a retreat that often begins during middle school. Summer science camps can be part of reversing that trend, some say. Academic camps are on the rise across the country, including ones to get adolescent girls excited about the…

  4. 1980s: Camp in a Computer Age.

    ERIC Educational Resources Information Center

    Camping Magazine, 1999

    1999-01-01

    Important issues for camps in the 1980s included soaring land values, high insurance rates, the spread of computers, and growing demand for specialized services. An illustrative article from 1985, "Yuppies Are Coming" (Tim Duffield), discusses the willingness of baby-boomer parents to pay for high-quality enrichment experiences at camp and other…

  5. Suicides in the Nazi Concentration Camps.

    ERIC Educational Resources Information Center

    Ryn, Zdzislaw

    1986-01-01

    On the basis of psychiatric interviews with 69 former prisoners of the Auschwitz-Birkenau concentration camp, this paper describes the circumstances, motives, and ways of committing suicide in the camp. The interviews made it clear that thousands of prisoners perished by suicide. The number of committed suicides was larger than that of attempted…

  6. Day Camp Manual: Staff. Book II.

    ERIC Educational Resources Information Center

    Babcock, William

    Book II of a 5-book day camp manual focuses on staff selection and training. Following the philosophy that developing an effective staff is the responsibility of the camp director, the book contains major sections on staff recruitment (sources of staff, problems, selection procedures, job descriptions, personnel practices); guidelines for job…

  7. Why It's Good to Go to Camp

    ERIC Educational Resources Information Center

    Howell, Matthew

    2008-01-01

    In the author's fourth year of undergraduate studies at the University of Waterloo he had the opportunity to explore the benefits attained by children attending a summer camp by way of an academic literature review. The author worked with Dr. Troy Glover who has been commissioned by a group of camping associations to perform a study on the…

  8. Children with Cancer: Positive Benefits of Camp.

    ERIC Educational Resources Information Center

    Winfree, Christy; Williams, Richard; Powell, Gwynn M.

    2002-01-01

    A relatively new method of helping pediatric cancer patients cope with their illness is specially designed summer camps. Camp helps children with cancer address psychological effects of the disease, bodily changes, and self-concept, and helps parents and siblings cope. Sidebars present resources and tips on incorporating children with cancer into…

  9. What Happens to Campers at Camp?

    ERIC Educational Resources Information Center

    Powell, Gwynn M.

    2003-01-01

    A study of 66 children with cancer and 43 siblings attending the Ronald McDonald Camp found that disease-specific camps allow children membership in a community of peers, which enhances self-esteem and social acceptance. A separate, longitudinal study of 38 beginning and experienced campers found that campers' intrapersonal and interpersonal…

  10. Cedar Ridge Camp: Using the Local Environment

    ERIC Educational Resources Information Center

    Burke, Grayson

    2007-01-01

    In 2007 Cedar Ridge Camp opened for its first season as a traditional co-ed summer camp and year-round outdoor education and recreation centre. The mission would centre on creating a program that would encourage personal development and growth through a shared outdoor experience. Cedar Ridge's main goals were to promote the formation of close…

  11. Smart Kids at CAMP-US

    ERIC Educational Resources Information Center

    Sinatra, Richard

    2004-01-01

    This article describes the summer literacy, athletic, and computer program know as CAMP-US. Each year, CAMP-US serves approximately 500 children in 10-day sessions. The first half of each day is devoted to literacy and computer instruction, while the second half is spent engaging in recreational activities such as swimming, soccer, softball,…

  12. Forest Fire: A Crisis Reality for Camp.

    ERIC Educational Resources Information Center

    Brown, Don; Mickelson, Rhonda

    2002-01-01

    Two camp directors were interviewed about evacuations from their camps due to forest fires. Topics covered include descriptions of the events; actions taken; aspects of advance planning that proved helpful; unexpected portions of the experience and resultant changes made in plans; relations with outside agencies, the media, and parents; working…

  13. Dealing with World Issues in Camp.

    ERIC Educational Resources Information Center

    Kujawa, Charles

    1986-01-01

    Discusses dealing with global issues in the camp setting in a way that broadens young people's world views. Topics include the educational advantages of the camp setting, desired outcomes for campers, guidelines for staff, and program ideas for dealing with issues such as environmental awareness, racism, and economic justice. (JHZ)

  14. Using the Internet To Promote Your Camp.

    ERIC Educational Resources Information Center

    Schenk, Tom

    1997-01-01

    Discusses the benefits of camps having a World Wide Web site and tips for choosing a Web provider. Offers recommendations for creating a Web page, including determining a goal, outlining content, choosing a graphic style and colors, testing the Web design, and promoting the Web site in camp brochures. Includes sample Web sites and Web resources.…

  15. Depletion of tyrosyl DNA phosphodiesterase 2 activity enhances etoposide-mediated double-strand break formation and cell killing.

    PubMed

    Kont, Yasemin Saygideger; Dutta, Arijit; Mallisetty, Apurva; Mathew, Jeena; Minas, Tsion; Kraus, Christina; Dhopeshwarkar, Priyanka; Kallakury, Bhaskar; Mitra, Sankar; Üren, Aykut; Adhikari, Sanjay

    2016-07-01

    DNA topoisomerase 2 (Top2) poisons, including common anticancer drugs etoposide and doxorubicin kill cancer cells by stabilizing covalent Top2-tyrosyl-DNA 5'-phosphodiester adducts and DNA double-strand breaks (DSBs). Proteolytic degradation of the covalently attached Top2 leaves a 5'-tyrosylated blocked termini which is removed by tyrosyl DNA phosphodiesterase 2 (TDP2), prior to DSB repair through non-homologous end joining (NHEJ). Thus, TDP2 confers resistance of tumor cells to Top2-poisons by repairing such covalent DNA-protein adducts, and its pharmacological inhibition could enhance the efficacy of Top2-poisons. We discovered NSC111041, a selective inhibitor of TDP2, by optimizing a high throughput screening (HTS) assay for TDP2's 5'-tyrosyl phosphodiesterase activity and subsequent validation studies. We found that NSC111041 inhibits TDP2's binding to DNA without getting intercalated into DNA and enhanced etoposide's cytotoxicity synergistically in TDP2-expressing cells but not in TDP2 depleted cells. Furthermore, NSC111041 enhanced formation of etoposide-induced γ-H2AX foci presumably by affecting DSB repair. Immuno-histochemical analysis showed higher TDP2 expression in a sub-set of different type of tumor tissues. These findings underscore the feasibility of clinical use of suitable TDP2 inhibitors in adjuvant therapy with Top2-poisons for a sub-set of cancer patients with high TDP2 expression. PMID:27235629

  16. A cardiac mitochondrial cAMP signaling pathway regulates calcium accumulation, permeability transition and cell death

    PubMed Central

    Wang, Z; Liu, D; Varin, A; Nicolas, V; Courilleau, D; Mateo, P; Caubere, C; Rouet, P; Gomez, A-M; Vandecasteele, G; Fischmeister, R; Brenner, C

    2016-01-01

    Although cardiac cytosolic cyclic 3′,5′-adenosine monophosphate (cAMP) regulates multiple processes, such as beating, contractility, metabolism and apoptosis, little is known yet on the role of this second messenger within cardiac mitochondria. Using cellular and subcellular approaches, we demonstrate here the local expression of several actors of cAMP signaling within cardiac mitochondria, namely a truncated form of soluble AC (sACt) and the exchange protein directly activated by cAMP 1 (Epac1), and show a protective role for sACt against cell death, apoptosis as well as necrosis in primary cardiomyocytes. Upon stimulation with bicarbonate (HCO3−) and Ca2+, sACt produces cAMP, which in turn stimulates oxygen consumption, increases the mitochondrial membrane potential (ΔΨm) and ATP production. cAMP is rate limiting for matrix Ca2+ entry via Epac1 and the mitochondrial calcium uniporter and, as a consequence, prevents mitochondrial permeability transition (MPT). The mitochondrial cAMP effects involve neither protein kinase A, Epac2 nor the mitochondrial Na+/Ca2+ exchanger. In addition, in mitochondria isolated from failing rat hearts, stimulation of the mitochondrial cAMP pathway by HCO3− rescued the sensitization of mitochondria to Ca2+-induced MPT. Thus, our study identifies a link between mitochondrial cAMP, mitochondrial metabolism and cell death in the heart, which is independent of cytosolic cAMP signaling. Our results might have implications for therapeutic prevention of cell death in cardiac pathologies. PMID:27100892

  17. Chemical Validation of Phosphodiesterase C as a Chemotherapeutic Target in Trypanosoma cruzi, the Etiological Agent of Chagas' Disease▿ †

    PubMed Central

    King-Keller, Sharon; Li, Minyong; Smith, Alyssa; Zheng, Shilong; Kaur, Gurpreet; Yang, Xiaochuan; Wang, Binghe; Docampo, Roberto

    2010-01-01

    Trypanosoma cruzi phosphodiesterase (PDE) C (TcrPDEC), a novel and rather unusual PDE in which, unlike all other class I PDEs, the catalytic domain is localized in the middle of the polypeptide chain, is able to hydrolyze cyclic GMP (cGMP), although it prefers cyclic AMP (cAMP), and has a FYVE-type domain in its N-terminal region (S. Kunz et al., FEBS J. 272:6412-6422, 2005). TcrPDEC shows homology to the mammalian PDE4 family members. PDE4 inhibitors are currently under development for the treatment of inflammatory diseases, such as asthma, chronic pulmonary diseases, and psoriasis, and for treating depression and serving as cognitive enhancers. We therefore tested a number of compounds originally synthesized as potential PDE4 inhibitors on T. cruzi amastigote growth, and we obtained several useful hits. We then conducted homology modeling of T. cruzi PDEC and identified other compounds as potential inhibitors through virtual screening. Testing of these compounds against amastigote growth and recombinant TcrPDEC activity resulted in several potent inhibitors. The most-potent inhibitors were found to increase the cellular concentration of cAMP. Preincubation of cells in the presence of one of these compounds stimulated volume recovery after hyposmotic stress, in agreement with their TcrPDEC inhibitory activity in vitro, providing chemical validation of this target. The compounds found could be useful tools in the study of osmoregulation in T. cruzi. In addition, their further optimization could result in the development of new drugs against Chagas' disease and other trypanosomiases. PMID:20625148

  18. Vasoactive intestinal peptide synergistically stimulates DNA synthesis in mouse 3T3 cells: Role of cAMP, Ca sup 2+ , and protein kinase C

    SciTech Connect

    Zurier, B.B.; Kozma, M.; Sinnett-Smith, J.; Rozengurt, E. )

    1988-05-01

    Vasoactive intestinal peptide synergistically stimulated initiation of DNA synthesis in Swiss 3T3 cells. The peptide stimulated ({sup 3}H)thymidine incorporation in the presence of insulin and either forskolin or an inhibitor of cAMP phosphodiesterase in a concentration-dependent manner. Half-maximal effect was obtained at 1 nM. At mitogenic concentrations, VIP stimulated a marked accumulation (eightfold) of cAMP. In contrast to other growth-promoting neuropeptides, VIP did not induce an increase in cytosolic free Ca{sup 2+} or the activation of protein kinase C. The authors conclude that neuropeptides can modulate long-term cell proliferation through multiple signaling pathways.

  19. The Characterization of Escherichia coli CpdB as a Recombinant Protein Reveals that, besides Having the Expected 3´-Nucleotidase and 2´,3´-Cyclic Mononucleotide Phosphodiesterase Activities, It Is Also Active as Cyclic Dinucleotide Phosphodiesterase

    PubMed Central

    López-Villamizar, Iralis; Cabezas, Alicia; Pinto, Rosa María; Canales, José; Ribeiro, João Meireles; Cameselle, José Carlos; Costas, María Jesús

    2016-01-01

    Endogenous cyclic diadenylate phosphodiesterase activity was accidentally detected in lysates of Escherichia coli BL21. Since this kind of activity is uncommon in Gram-negative bacteria, its identification was undertaken. After partial purification and analysis by denaturing gel electrophoresis, renatured activity correlated with a protein identified by fingerprinting as CpdB (cpdB gene product), which is annotated as 3´-nucleotidase / 2´,3´-cyclic-mononucleotide phosphodiesterase, and it is synthesized as a precursor protein with a signal sequence removable upon export to the periplasm. It has never been studied as a recombinant protein. The coding sequence of mature CpdB was cloned and expressed as a GST fusion protein. The study of the purified recombinant protein, separated from GST, confirmed CpdB annotation. The assay of catalytic efficiencies (kcat/Km) for a large substrate set revealed novel CpdB features, including very high efficiencies for 3´-AMP and 2´,3´-cyclic mononucleotides, and previously unknown activities on cyclic and linear dinucleotides. The catalytic efficiencies of the latter activities, though low in relative terms when compared to the major ones, are far from negligible. Actually, they are perfectly comparable to those of the ‘average’ enzyme and the known, bona fide cyclic dinucleotide phosphodiesterases. On the other hand, CpdB differs from these enzymes in its extracytoplasmic location and in the absence of EAL, HD and DHH domains. Instead, it contains the domains of the 5´-nucleotidase family pertaining to the metallophosphoesterase superfamily, although CpdB lacks 5´-nucleotidase activity. The possibility that the extracytoplasmic activity of CpdB on cyclic dinucleotides could have physiological meaning is discussed. PMID:27294396

  20. cAMP prevents TNF-induced apoptosis through inhibiting DISC complex formation in rat hepatocytes

    SciTech Connect

    Bhattacharjee, Rajesh; Xiang, Wenpei; Wang, Yinna; Zhang, Xiaoying

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer cAMP blocks cell death induced by TNF and actinomycin D in cultured hepatocytes. Black-Right-Pointing-Pointer cAMP blocks NF-{kappa}B activation induced by TNF and actinomycin D. Black-Right-Pointing-Pointer cAMP blocks DISC formation following TNF and actinomycin D exposure. Black-Right-Pointing-Pointer cAMP blocks TNF signaling at a proximal step. -- Abstract: Tumor necrosis factor {alpha} (TNF) is a pleiotropic proinflammatory cytokine that plays a role in immunity and the control of cell proliferation, cell differentiation, and apoptosis. The pleiotropic nature of TNF is due to the formation of different signaling complexes upon the binding of TNF to its receptor, TNF receptor type 1 (TNFR1). TNF induces apoptosis in various mammalian cells when the cells are co-treated with a transcription inhibitor like actinomycin D (ActD). When TNFR1 is activated, it recruits an adaptor protein, TNF receptor-associated protein with death domain (TRADD), through its cytoplasmic death effector domain (DED). TRADD, in turn, recruits other signaling proteins, including TNF receptor-associated protein 2 (TRAF2) and receptor-associated protein kinase (RIPK) 1, to form a complex. Subsequently, this complex combines with FADD and procaspase-8, converts into a death-inducing signaling complex (DISC) to induce apoptosis. Cyclic AMP (cAMP) is a second messenger that regulates various cellular processes such as cell proliferation, gene expression, and apoptosis. cAMP analogues are reported to act as anti-apoptotic agents in various cell types, including hepatocytes. We found that a cAMP analogue, dibutyryl cAMP (db-cAMP), inhibits TNF + ActD-induced apoptosis in rat hepatocytes. The protein kinase A (PKA) inhibitor KT-5720 reverses this inhibitory effect of cAMP on apoptosis. Cytoprotection by cAMP involves down-regulation of various apoptotic signal regulators like TRADD and FADD and inhibition of caspase-8 and caspase-3 cleavage. We also found

  1. Parathyroid hormone stimulates juxtaglomerular cell cAMP accumulation without stimulating renin release

    PubMed Central

    Atchison, Douglas K.; Harding, Pamela; Cecilia Ortiz-Capisano, M.; Peterson, Edward L.

    2012-01-01

    Parathyroid hormone (PTH) is positively coupled to the generation of cAMP via its actions on the PTH1R and PTH2R receptors. Renin secretion from juxtaglomerular (JG) cells is stimulated by elevated intracellular cAMP, and every stimulus that increases renin secretion is thought to do so via increasing cAMP. Thus we hypothesized that PTH increases renin release from primary cultures of mouse JG cells by elevating intracellular cAMP via the PTH1R receptor. We found PTH1R, but not PTH2R, mRNA expressed in JG cells. While PTH increased JG cell cAMP content from (log10 means ± SE) 3.27 ± 0.06 to 3.92 ± 0.12 fmol/mg protein (P < 0.001), it did not affect renin release. The PTH1R-specific agonist, parathyroid hormone-related protein (PTHrP), also increased JG cell cAMP from 3.13 ± 0.09 to 3.93 ± 0.09 fmol/mg protein (P < 0.001), again without effect on renin release. PTH2R receptor agonists had no effect on cAMP or renin release. PTHrP increased cAMP in the presence of both low and high extracellular calcium from 3.31 ± 0.17 to 3.83 ± 0.20 fmol/mg protein (P < 0.01) and from 3.29 ± 0.18 to 3.63 ± 0.22 fmol/mg protein (P < 0.05), respectively, with no effect on renin release. PTHrP increased JG cell cAMP in the presence of adenylyl cyclase-V inhibition from 2.85 ± 0.17 to 3.44 ± 0.14 fmol/mg protein (P < 0.001) without affecting renin release. As a positive control, forskolin increased JG cell cAMP from 3.39 ± 0.13 to 4.48 ± 0.07 fmol/mg protein (P < 0.01) and renin release from 2.96 ± 0.10 to 3.29 ± 0.08 ng ANG I·mg prot−1·h−1 (P < 0.01). Thus PTH increases JG cell cAMP via non-calcium-sensitive adenylate cyclases without affecting renin release. These data suggest compartmentalization of cAMP signaling in JG cells. PMID:22896038

  2. Parathyroid hormone stimulates juxtaglomerular cell cAMP accumulation without stimulating renin release.

    PubMed

    Atchison, Douglas K; Harding, Pamela; Cecilia Ortiz-Capisano, M; Peterson, Edward L; Beierwaltes, William H

    2012-10-15

    Parathyroid hormone (PTH) is positively coupled to the generation of cAMP via its actions on the PTH1R and PTH2R receptors. Renin secretion from juxtaglomerular (JG) cells is stimulated by elevated intracellular cAMP, and every stimulus that increases renin secretion is thought to do so via increasing cAMP. Thus we hypothesized that PTH increases renin release from primary cultures of mouse JG cells by elevating intracellular cAMP via the PTH1R receptor. We found PTH1R, but not PTH2R, mRNA expressed in JG cells. While PTH increased JG cell cAMP content from (log(10) means ± SE) 3.27 ± 0.06 to 3.92 ± 0.12 fmol/mg protein (P < 0.001), it did not affect renin release. The PTH1R-specific agonist, parathyroid hormone-related protein (PTHrP), also increased JG cell cAMP from 3.13 ± 0.09 to 3.93 ± 0.09 fmol/mg protein (P < 0.001), again without effect on renin release. PTH2R receptor agonists had no effect on cAMP or renin release. PTHrP increased cAMP in the presence of both low and high extracellular calcium from 3.31 ± 0.17 to 3.83 ± 0.20 fmol/mg protein (P < 0.01) and from 3.29 ± 0.18 to 3.63 ± 0.22 fmol/mg protein (P < 0.05), respectively, with no effect on renin release. PTHrP increased JG cell cAMP in the presence of adenylyl cyclase-V inhibition from 2.85 ± 0.17 to 3.44 ± 0.14 fmol/mg protein (P < 0.001) without affecting renin release. As a positive control, forskolin increased JG cell cAMP from 3.39 ± 0.13 to 4.48 ± 0.07 fmol/mg protein (P < 0.01) and renin release from 2.96 ± 0.10 to 3.29 ± 0.08 ng ANG I·mg prot(-1)·h(-1) (P < 0.01). Thus PTH increases JG cell cAMP via non-calcium-sensitive adenylate cyclases without affecting renin release. These data suggest compartmentalization of cAMP signaling in JG cells.

  3. Cdk5 Is Required for Memory Function and Hippocampal Plasticity via the cAMP Signaling Pathway

    PubMed Central

    Gao, Jun; Joseph, Nadine; Xie, Zhigang; Zhou, Ying; Durak, Omer; Zhang, Lei; Zhu, J. Julius; Clauser, Karl R.; Carr, Steven A.; Tsai, Li-Huei

    2011-01-01

    Memory formation is modulated by pre- and post-synaptic signaling events in neurons. The neuronal protein kinase Cyclin-Dependent Kinase 5 (Cdk5) phosphorylates a variety of synaptic substrates and is implicated in memory formation. It has also been shown to play a role in homeostatic regulation of synaptic plasticity in cultured neurons. Surprisingly, we found that Cdk5 loss of function in hippocampal circuits results in severe impairments in memory formation and retrieval. Moreover, Cdk5 loss of function in the hippocampus disrupts cAMP signaling due to an aberrant increase in phosphodiesterase (PDE) proteins. Dysregulation of cAMP is associated with defective CREB phosphorylation and disrupted composition of synaptic proteins in Cdk5-deficient mice. Rolipram, a PDE4 inhibitor that prevents cAMP depletion, restores synaptic plasticity and memory formation in Cdk5-deficient mice. Collectively, our results demonstrate a critical role for Cdk5 in the regulation of cAMP-mediated hippocampal functions essential for synaptic plasticity and memory formation. PMID:21984943

  4. Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma.

    PubMed

    Ge, Xuecai; Milenkovic, Ljiljana; Suyama, Kaye; Hartl, Tom; Purzner, Teresa; Winans, Amy; Meyer, Tobias; Scott, Matthew P

    2015-01-01

    Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors. PMID:26371509

  5. Chemoproteomics demonstrates target engagement and exquisite selectivity of the clinical phosphodiesterase 10A inhibitor MP-10 in its native environment.

    PubMed

    Schülke, Jan-Philip; McAllister, Laura A; Geoghegan, Kieran F; Parikh, Vinod; Chappie, Thomas A; Verhoest, Patrick R; Schmidt, Christopher J; Johnson, Douglas S; Brandon, Nicholas J

    2014-12-19

    Phosphodiesterases (PDEs) regulate the levels of the second messengers cAMP and cGMP and are important drug targets. PDE10A is highly enriched in medium spiny neurons of the striatum and is an attractive drug target for the treatment of basal ganglia diseases like schizophrenia, Parkinson's disease, or Huntington's disease. Here we describe the design, synthesis, and application of a variety of chemical biology probes, based on the first clinically tested PDE10A inhibitor MP-10, which were used to characterize the chemoproteomic profile of the clinical candidate in its native environment. A clickable photoaffinity probe was used to measure target engagement of MP-10 and revealed differences between whole cell and membrane preparations. Moreover, our results illustrate the importance of the linker design in the creation of functional probes. Biotinylated affinity probes allowed identification of drug-interaction partners in rodent and human tissue and quantitative mass spectrometry analysis revealed highly specific binding of MP-10 to PDE10A with virtually no off-target binding. The profiling of PDE10A chemical biology probes described herein illustrates a strategy by which high affinity inhibitors can be converted into probes for determining selectivity and target engagement of drug candidates in complex biological matrices from native sources.

  6. Inhibition of human cAMP-phosphodiesterase as a mechanism of the spasmolytic effect of Matricaria recutita L.

    PubMed

    Maschi, Omar; Cero, Esther Dal; Galli, Germana V; Caruso, Donatella; Bosisio, Enrica; Dell'Agli, Mario

    2008-07-01

    Mechanisms underlying the spasmolytic activity of chamomile still remain unclear. Inhibition of cAMP- and cGMP-phosphodiesterases (PDE) is one of the mechanisms operated by spasmolytic drugs. In this study, the effect of chamomile on PDE was investigated. Human platelet cAMP-PDE and recombinant PDE5A1 were assayed in the presence of infusions prepared from sifted flowers and capitula. LC-ESI-MS/MS analysis showed different compositions in infusions made with sifted flowers and capitula. Chamomile inhibited cAMP-PDE activity (IC50 = 17.9-40.5 microg/mL), while cGMP-PDE5 was less affected (-15% at 50 microg/mL). Among the individual compounds tested, only flavonoids showed an inhibitory effect (IC50 = 1.3-14.9 microM), contributing to around 39% of the infusion inhibition; other compounds responsible for cAMP-PDE inhibition still remain unknown. Although experimental evidence supporting the use of chamomile for gastrointestinal minor spasms dates back to the fifties, cAMP-PDE inhibition as a likely mechanism underlying the spasmolytic activity is reported for the first time. PMID:18553893

  7. Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma

    PubMed Central

    Ge, Xuecai; Milenkovic, Ljiljana; Suyama, Kaye; Hartl, Tom; Purzner, Teresa; Winans, Amy; Meyer, Tobias; Scott, Matthew P

    2015-01-01

    Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors. DOI: http://dx.doi.org/10.7554/eLife.07068.001 PMID:26371509

  8. Inhibition of phosphodiesterase-4 reverses the cognitive dysfunction and oxidative stress induced by Aβ25-35 in rats.

    PubMed

    Zhuo, Yeye; Guo, Haibiao; Cheng, Yufang; Wang, Chuang; Wang, Canmao; Wu, Jingang; Zou, Zhengqiang; Gan, Danna; Li, Yiwen; Xu, Jiangping

    2016-08-01

    Phosphodiesterase-4 (PDE4) inhibitors prevent the breakdown of the second messenger cAMP and have been demonstrated to improve learning in several animal models of cognition. In this study, we explored the antioxidative effects of rolipram in Alzheimer's disease (AD) by using bilateral Aβ25-35 injection into the hippocampus of rats, which were used as an AD model. Rats received 3 intraperitoneal (i.p.) doses of rolipram (0.1, 0.5 and 1.25 mg/kg) daily after the injection of Aβ25-35 for 25 days. Chronic administration of rolipram prevented the memory impairments induced by Aβ25-35, as assessed using the passive avoidance test and the Morris water maze test. Furthermore, rolipram significantly reduced the oxidative stress induced by Aβ25-35, as evidenced by the decrease in the levels of reactive oxygen species (ROS) and malondialdehyde (MDA), and restored the reduced GSH levels and superoxide dismutase (SOD) activity. Moreover, western blotting and real-time reverse transcription polymerase chain reaction (RT-PCR) analysis showed that rolipram remarkably upregulated thioredoxin (Trx) and inhibited the inducible nitric oxide synthase/nitric oxide (iNOS/NO) pathway in the hippocampus. These results demonstrated that rolipram improved the learning and memory abilities in an Aβ25-35-induced AD rat model. The mechanism underlying these effects may be due to the noticeable antioxidative effects of rolipram. PMID:26920899

  9. [Phosphodiesterase-5 inhibitors for the treatment of pulmonary arterial hypertension].

    PubMed

    Beltrán-Gámez, Miguel E; Sandoval-Zárate, Julio; Pulido, Tomás

    2015-01-01

    In experimental and clinical cardiology, phosphodiesterase type 5 (PDE-5) inhibitors have brought scientific interest as a therapeutic tool in pulmonary arterial hypertension (PAH) management in recent years. Phosphodiesterases are a superfamily of enzymes that inactivate cyclic adenosine monophosphate and cyclic guanosine monophosphate, the second messengers of prostacyclin and nitric oxide. The rationale for the use of PDE-5 inhibitors in PAH is based on their capacity to overexpresss the nitric oxide pathway pursued inhibition of cyclic guanosine monophosphate hydrolysis. By increasing cyclic guanosine monophosphate levels it promotes vasodilation, antiproliferative and pro-apoptotic effects that may reverse pulmonary vascular remodeling. There is also evidence that these drugs may directly enhance right ventricular contractility through an increase in cyclic adenosine monophosphate mediated by the inhibition of the cyclic guanosine monophosphate -sensitive PDE-3. Sildenafil, tadalafil and vardenafil are 3 specific PDE-5 inhibitors in current clinical use, which share similar mechanisms of action but present some significant differences regarding potency, selectivity for PDE-5 and pharmacokinetic properties. Sildenafil received approval in 2005 by the Food and Drug Administration and the European Medicines Agency and tadalafil in 2009 by the Food and Drug Administration and the European Medicines Agency for the treatment of PAH in patients classified as NYHA/WHO functional class II and III. In Mexico, sildenafil and tadalafil were approved by Comisión Federal de Protección contra Riesgos Sanitarios for this indication in 2010 and 2011, respectively. PMID:26047999

  10. [Phosphodiesterase-5 inhibitors for the treatment of pulmonary arterial hypertension].

    PubMed

    Beltrán-Gámez, Miguel E; Sandoval-Zárate, Julio; Pulido, Tomás

    2015-01-01

    In experimental and clinical cardiology, phosphodiesterase type 5 (PDE-5) inhibitors have brought scientific interest as a therapeutic tool in pulmonary arterial hypertension (PAH) management in recent years. Phosphodiesterases are a superfamily of enzymes that inactivate cyclic adenosine monophosphate and cyclic guanosine monophosphate, the second messengers of prostacyclin and nitric oxide. The rationale for the use of PDE-5 inhibitors in PAH is based on their capacity to overexpresss the nitric oxide pathway pursued inhibition of cyclic guanosine monophosphate hydrolysis. By increasing cyclic guanosine monophosphate levels it promotes vasodilation, antiproliferative and pro-apoptotic effects that may reverse pulmonary vascular remodeling. There is also evidence that these drugs may directly enhance right ventricular contractility through an increase in cyclic adenosine monophosphate mediated by the inhibition of the cyclic guanosine monophosphate -sensitive PDE-3. Sildenafil, tadalafil and vardenafil are 3 specific PDE-5 inhibitors in current clinical use, which share similar mechanisms of action but present some significant differences regarding potency, selectivity for PDE-5 and pharmacokinetic properties. Sildenafil received approval in 2005 by the Food and Drug Administration and the European Medicines Agency and tadalafil in 2009 by the Food and Drug Administration and the European Medicines Agency for the treatment of PAH in patients classified as NYHA/WHO functional class II and III. In Mexico, sildenafil and tadalafil were approved by Comisión Federal de Protección contra Riesgos Sanitarios for this indication in 2010 and 2011, respectively.

  11. "It's not Just Camp!": Understanding the Meaning of Children's Cancer Camps for Children and Families.

    PubMed

    Laing, Catherine M; Moules, Nancy J

    2016-01-01

    The purpose of this philosophical hermeneutic inquiry was to understand the meaning of children's cancer camps for the child with cancer and the family. Six childhood cancer families and 5 cancer camp counselors were interviewed, in order to bring understanding to this topic. Findings from this research revealed that camp means different things for different families, and that much is at play in the cancer camp experience: the healing and developmental power of play, finding acceptance and fit, grief as something to live with versus "get over," storytelling as a means of reshaping and understanding traumatic experiences, and the solidarity of the community as one that creates intense, healing bonds. Children's cancer camps, we conclude, should be considered a necessity, versus a luxury, and could even be thought of as a psychosocial intervention for some children and families. Barriers such as structure of funding and access to resources are present and likely due to the separateness of camps from hospital programs. PMID:25643975

  12. "It's not Just Camp!": Understanding the Meaning of Children's Cancer Camps for Children and Families.

    PubMed

    Laing, Catherine M; Moules, Nancy J

    2016-01-01

    The purpose of this philosophical hermeneutic inquiry was to understand the meaning of children's cancer camps for the child with cancer and the family. Six childhood cancer families and 5 cancer camp counselors were interviewed, in order to bring understanding to this topic. Findings from this research revealed that camp means different things for different families, and that much is at play in the cancer camp experience: the healing and developmental power of play, finding acceptance and fit, grief as something to live with versus "get over," storytelling as a means of reshaping and understanding traumatic experiences, and the solidarity of the community as one that creates intense, healing bonds. Children's cancer camps, we conclude, should be considered a necessity, versus a luxury, and could even be thought of as a psychosocial intervention for some children and families. Barriers such as structure of funding and access to resources are present and likely due to the separateness of camps from hospital programs.

  13. Accreditation Standards for Camp Programs and Services. Revised 1998 Edition.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    The American Camping Association (ACA) accreditation program aims to educate camp owners and directors in the administration of key aspects of camp operation, particularly those related to program quality and the health and safety of campers and staff, and to assist the public in selecting camps that meet industry-accepted and…

  14. Institutionalized Adolescents' Perceptions of a Summer Camp Program

    ERIC Educational Resources Information Center

    Herr, David E.

    1977-01-01

    Describes the use of the facilities of Camp Easter Seal, Virginia, for institutionalized adolescents from different hospitals in Virginia. Also includes the attitudes of the patients toward their camping experience, their camp counselors, and what they learned from their camping experience. (Author/RK)

  15. The History of Organized Camping in the United States.

    ERIC Educational Resources Information Center

    Gibson, H. W.

    H.W. Gibson's life was devoted to camping for boys, from his first job as a Young Men's Christian Association (YMCA) secretary in 1887 through his work as founder of two YMCA camps, state YMCA secretary, author of eight books on the subject, member of camp director associations which preceded the American Camping Association (ACA), and then as…

  16. Site and Facilities: A Resource Book for Camps.

    ERIC Educational Resources Information Center

    Ball, Armand, Ed.; Ball, Beverly, Ed.

    This resource book draws together articles on the development and maintenance of camp sites and facilities. The articles, previously published by "Camping Magazine" and "Journal of Christian Camping," cover (1) site planning and long-range development, including redesigning multiple camp facilities for year-round programs, remodeling and…

  17. Design and Development Issues for Educational Robotics Training Camps

    ERIC Educational Resources Information Center

    Ucgul, Memet; Cagiltay, Kursat

    2014-01-01

    The aim of this study is to explore critical design issues for educational robotics training camps and to describe how these factors should be implemented in the development of such camps. For this purpose, two robotics training camps were organized for elementary school students. The first camp had 30 children attendees, and the second had 22. As…

  18. Alabama State Lodging Tax: A Lesson for All Camps.

    ERIC Educational Resources Information Center

    Sieckmann, Amy

    2003-01-01

    A dispute between an Alabama business conference center and a nearby camp prompted the state's revenue department to charge camps an old lodging tax that had never been applied to them before. The state camping association members worked together to have the tax law and regulations amended so that nonprofit camps were exempt from the tax in most…

  19. Interactions between the subunits of transducin and cyclic GMP phosphodiesterase in Rana catesbiana rod photoreceptors.

    PubMed

    Yamazaki, A; Hayashi, F; Tatsumi, M; Bitensky, M W; George, J S

    1990-07-15

    In bullfrog (Rana catesbiana) rods the activity of cyclic GMP (cGMP) phosphodiesterase was stimulated 10 times by washing disc membranes with an isotonic, GTP-containing buffer. This stimulation was maintained following hydrolysis of GTP and after removal of guanine nucleotides. At least 60-70% of the inhibitory gamma subunit of cGMP phosphodiesterase (P gamma) was physically released from membranes by these washing procedures. When cGMP phosphodiesterase was activated by a hydrolysis-resistant GTP analogue, P gamma was found in the supernatant complexed with the transducin alpha subunit (T alpha) using three chromatography systems. When GTP was used to activate cGMP phosphodiesterase, P gamma was also found in the supernatant complexed with GDP.T alpha. This complex was also isolated using the same three chromatography systems, indicating that P gamma remained tightly bound to T alpha even after bound GTP was hydrolyzed. Interaction with the beta,gamma subunits of transducin, which remained associated with disc membranes, was required for the release of P gamma from the GDP.T alpha complex, which resulted in the deactivation of active cGMP phosphodiesterase. We conclude that during activation of cGMP phosphodiesterase, P gamma is complexed with T alpha (both GTP and GDP forms) in the supernatant and that, following GTP hydrolysis, beta,gamma subunits of transducin are necessary for the release of P gamma from the complex and the resulting inactivation of cGMP phosphodiesterase in frog photoreceptors. PMID:2164007

  20. A Lightning Safety Primer for Camps.

    ERIC Educational Resources Information Center

    Attarian, Aram

    1992-01-01

    Provides the following information about lightning, which is necessary for camp administrators and staff: (1) warning signs of lightning; (2) dangers of lightning; (3) types of lightning injuries; (4) prevention of lightning injury; and (5) helpful training tips. (KS)

  1. Retarded Children at Camp with Normal Children

    ERIC Educational Resources Information Center

    Flax, Norman; Peters, Edward N.

    1969-01-01

    Statistical analysis of data from written forms and scales (designed to measure children's behavior in groups), observations, and interviews indicated that many educalble mentally retarded children can participate successfully in camp activities with normal children. (DR)

  2. Estradiol increases cAMP in the oviductal secretory cells through a nongenomic mechanism.

    PubMed

    Oróstica, María L; Lopez, John; Rojas, Israel; Rocco, Jocelyn; Díaz, Patricia; Reuquén, Patricia; Cardenas, Hugo; Parada-Bustamante, Alexis; Orihuela, Pedro A

    2014-09-01

    In the rat oviduct, estradiol (E2) accelerates egg transport by a nongenomic action that requires previous conversion of E2 to methoxyestrogens via catechol-O-methyltranferase (COMT) and activation of estrogen receptor (ER) with subsequent production of cAMP and inositol triphosphate (IP3). However, the role of the different oviductal cellular phenotypes on this E2 nongenomic pathway remains undetermined. The aim of this study was to investigate the effect of E2 on the levels of cAMP and IP3 in primary cultures of secretory and smooth muscle cells from rat oviducts and determine the mechanism by which E2 increases cAMP in the secretory cells. In the secretory cells, E2 increased cAMP but not IP3, while in the smooth muscle cells E2 decreased cAMP and increased IP3. Suppression of protein synthesis by actinomycin D did not prevent the E2-induced cAMP increase, but this was blocked by the ER antagonist ICI 182 780 and the inhibitors of COMT OR 486, G protein-α inhibitory (Gαi) protein pertussis toxin and adenylyl cyclase (AC) SQ 22536. Expression of the mRNA for the enzymes that metabolizes estrogens, Comt, Cyp1a1, and Cyp1b1 was found in the secretory cells, but this was not affected by E2. Finally, confocal immunofluorescence analysis showed that E2 induced colocalization between ESR1 (ERα) and Gαi in extranuclear regions of the secretory cells. We conclude that E2 differentially regulates cAMP and IP3 in the secretory and smooth muscle cells of the rat oviduct. In the secretory cells, E2 increases cAMP via a nongenomic action that requires activation of COMT and ER, coupling between ESR1 and Gαi, and stimulation of AC.

  3. Requirement of cAMP Signaling for Schwann Cell Differentiation Restricts the Onset of Myelination

    PubMed Central

    Bacallao, Ketty; Monje, Paula V.

    2015-01-01

    Isolated Schwann cells (SCs) respond to cAMP elevation by adopting a differentiated post-mitotic state that exhibits high levels of Krox-20, a transcriptional enhancer of myelination, and mature SC markers such as the myelin lipid galactocerebroside (O1). To address how cAMP controls myelination, we performed a series of cell culture experiments which compared the differentiating responses of isolated and axon-related SCs to cAMP analogs and ascorbate, a known inducer of axon ensheathment, basal lamina formation and myelination. In axon-related SCs, cAMP induced the expression of Krox-20 and O1 without a concomitant increase in the expression of myelin basic protein (MBP) and without promoting axon ensheathment, collagen synthesis or basal lamina assembly. When cAMP was provided together with ascorbate, a dramatic enhancement of MBP expression occurred, indicating that cAMP primes SCs to form myelin only under conditions supportive of basal lamina formation. Experiments using a combination of cell permeable cAMP analogs and type-selective adenylyl cyclase (AC) agonists and antagonists revealed that selective transmembrane AC (tmAC) activation with forskolin was not sufficient for full SC differentiation and that the attainment of an O1 positive state also relied on the activity of the soluble AC (sAC), a bicarbonate sensor that is insensitive to forskolin and GPCR activation. Pharmacological and immunological evidence indicated that SCs expressed sAC and that sAC activity was required for morphological differentiation and the expression of myelin markers such as O1 and protein zero. To conclude, our data indicates that cAMP did not directly drive myelination but rather the transition into an O1 positive state, which is perhaps the most critical cAMP-dependent rate limiting step for the onset of myelination. The temporally restricted role of cAMP in inducing differentiation independently of basal lamina formation provides a clear example of the uncoupling of signals

  4. Requirement of cAMP signaling for Schwann cell differentiation restricts the onset of myelination.

    PubMed

    Bacallao, Ketty; Monje, Paula V

    2015-01-01

    Isolated Schwann cells (SCs) respond to cAMP elevation by adopting a differentiated post-mitotic state that exhibits high levels of Krox-20, a transcriptional enhancer of myelination, and mature SC markers such as the myelin lipid galactocerebroside (O1). To address how cAMP controls myelination, we performed a series of cell culture experiments which compared the differentiating responses of isolated and axon-related SCs to cAMP analogs and ascorbate, a known inducer of axon ensheathment, basal lamina formation and myelination. In axon-related SCs, cAMP induced the expression of Krox-20 and O1 without a concomitant increase in the expression of myelin basic protein (MBP) and without promoting axon ensheathment, collagen synthesis or basal lamina assembly. When cAMP was provided together with ascorbate, a dramatic enhancement of MBP expression occurred, indicating that cAMP primes SCs to form myelin only under conditions supportive of basal lamina formation. Experiments using a combination of cell permeable cAMP analogs and type-selective adenylyl cyclase (AC) agonists and antagonists revealed that selective transmembrane AC (tmAC) activation with forskolin was not sufficient for full SC differentiation and that the attainment of an O1 positive state also relied on the activity of the soluble AC (sAC), a bicarbonate sensor that is insensitive to forskolin and GPCR activation. Pharmacological and immunological evidence indicated that SCs expressed sAC and that sAC activity was required for morphological differentiation and the expression of myelin markers such as O1 and protein zero. To conclude, our data indicates that cAMP did not directly drive myelination but rather the transition into an O1 positive state, which is perhaps the most critical cAMP-dependent rate limiting step for the onset of myelination. The temporally restricted role of cAMP in inducing differentiation independently of basal lamina formation provides a clear example of the uncoupling of signals

  5. PDE4B mediates local feedback regulation of β₁-adrenergic cAMP signaling in a sarcolemmal compartment of cardiac myocytes.

    PubMed

    Mika, Delphine; Richter, Wito; Westenbroek, Ruth E; Catterall, William A; Conti, Marco

    2014-03-01

    Multiple cAMP phosphodiesterase (PDE) isoforms play divergent roles in cardiac homeostasis but the molecular basis for their non-redundant function remains poorly understood. Here, we report a novel role for the PDE4B isoform in β-adrenergic (βAR) signaling in the heart. Genetic ablation of PDE4B disrupted βAR-induced cAMP transients, as measured by FRET sensors, at the sarcolemma but not in the bulk cytosol of cardiomyocytes. This effect was further restricted to a subsarcolemmal compartment because PDE4B regulates β1AR-, but not β2AR- or PGE2-induced responses. The spatially restricted function of PDE4B was confirmed by its selective effects on PKA-mediated phosphorylation patterns. PDE4B limited the PKA-mediated phosphorylation of key players in excitation-contraction coupling that reside in the sarcolemmal compartment, including L-type Ca(2+) channels and ryanodine receptors, but not phosphorylation of distal cytosolic proteins. β1AR- but not β2AR-ligation induced PKA-dependent activation of PDE4B and interruption of this negative feedback with PKA inhibitors increased sarcolemmal cAMP. Thus, PDE4B mediates a crucial PKA-dependent feedback that controls β1AR-dependent cAMP signals in a restricted subsarcolemmal domain. Disruption of this feedback augments local cAMP/PKA signals, leading to an increased intracellular Ca(2+) level and contraction rate.

  6. Standards for Day and Resident Camps: The Accreditation Programs of the American Camping Association. 1990 Edition, Revised 1993.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    This book outlines standards that provide a basis for camp accreditation and site approval by the American Camping Association (ACA), Inc. The purpose of the ACA accreditation programs is to educate camp owners and directors in the administration of key aspects of camp operation, particularly those related to program quality and the health and…

  7. Diabetic nephropathy: Treatment with phosphodiesterase type 5 inhibitors.

    PubMed

    Thompson, Cecil Stanley

    2013-08-15

    The importance of nitric oxide (NO) in vascular physiology is irrefutable; it stimulates the intracellular production of cyclic guanosine monophosphate (cGMP), initiating vascular smooth muscle relaxation. This biochemical process increases the diameter of small arteries, regulating blood flow distribution between arterioles and the microvasculature. The kidney is no exception, since NO predominantly dilates the glomerular afferent arterioles. It is now evident that the vascular production of cGMP can be augmented by inhibitors of phosphodiesterase type 5 (PDE 5), the enzyme which breakdowns this cyclic nucleotide. This has clinical relevance, since diabetic nephropathy (DN) a major microvascular complication of diabetes mellitus and the most common cause of end-stage renal disease, increases intraglomerular capillary pressure, leading to glomerular hypertension. PDE 5 inhibitors may have, therefore, the potential to reduce glomerular hypertension. This review describes the use of PDE 5 inhibitors to improve the metabolic, haemodynamic and inflammatory pathways/responses, all of which are dysfunctional in DN. PMID:23961322

  8. Control of βAR- and N-methyl-D-aspartate (NMDA) Receptor-Dependent cAMP Dynamics in Hippocampal Neurons.

    PubMed

    Chay, Andrew; Zamparo, Ilaria; Koschinski, Andreas; Zaccolo, Manuela; Blackwell, Kim T

    2016-02-01

    Norepinephrine, a neuromodulator that activates β-adrenergic receptors (βARs), facilitates learning and memory as well as the induction of synaptic plasticity in the hippocampus. Several forms of long-term potentiation (LTP) at the Schaffer collateral CA1 synapse require stimulation of both βARs and N-methyl-D-aspartate receptors (NMDARs). To understand the mechanisms mediating the interactions between βAR and NMDAR signaling pathways, we combined FRET imaging of cAMP in hippocampal neuron cultures with spatial mechanistic modeling of signaling pathways in the CA1 pyramidal neuron. Previous work implied that cAMP is synergistically produced in the presence of the βAR agonist isoproterenol and intracellular calcium. In contrast, we show that when application of isoproterenol precedes application of NMDA by several minutes, as is typical of βAR-facilitated LTP experiments, the average amplitude of the cAMP response to NMDA is attenuated compared with the response to NMDA alone. Models simulations suggest that, although the negative feedback loop formed by cAMP, cAMP-dependent protein kinase (PKA), and type 4 phosphodiesterase may be involved in attenuating the cAMP response to NMDA, it is insufficient to explain the range of experimental observations. Instead, attenuation of the cAMP response requires mechanisms upstream of adenylyl cyclase. Our model demonstrates that Gs-to-Gi switching due to PKA phosphorylation of βARs as well as Gi inhibition of type 1 adenylyl cyclase may underlie the experimental observations. This suggests that signaling by β-adrenergic receptors depends on temporal pattern of stimulation, and that switching may represent a novel mechanism for recruiting kinases involved in synaptic plasticity and memory.

  9. Control of βAR- and N-methyl-D-aspartate (NMDA) Receptor-Dependent cAMP Dynamics in Hippocampal Neurons

    PubMed Central

    Chay, Andrew; Zamparo, Ilaria; Koschinski, Andreas; Zaccolo, Manuela; Blackwell, Kim T.

    2016-01-01

    Norepinephrine, a neuromodulator that activates β-adrenergic receptors (βARs), facilitates learning and memory as well as the induction of synaptic plasticity in the hippocampus. Several forms of long-term potentiation (LTP) at the Schaffer collateral CA1 synapse require stimulation of both βARs and N-methyl-D-aspartate receptors (NMDARs). To understand the mechanisms mediating the interactions between βAR and NMDAR signaling pathways, we combined FRET imaging of cAMP in hippocampal neuron cultures with spatial mechanistic modeling of signaling pathways in the CA1 pyramidal neuron. Previous work implied that cAMP is synergistically produced in the presence of the βAR agonist isoproterenol and intracellular calcium. In contrast, we show that when application of isoproterenol precedes application of NMDA by several minutes, as is typical of βAR-facilitated LTP experiments, the average amplitude of the cAMP response to NMDA is attenuated compared with the response to NMDA alone. Models simulations suggest that, although the negative feedback loop formed by cAMP, cAMP-dependent protein kinase (PKA), and type 4 phosphodiesterase may be involved in attenuating the cAMP response to NMDA, it is insufficient to explain the range of experimental observations. Instead, attenuation of the cAMP response requires mechanisms upstream of adenylyl cyclase. Our model demonstrates that Gs-to-Gi switching due to PKA phosphorylation of βARs as well as Gi inhibition of type 1 adenylyl cyclase may underlie the experimental observations. This suggests that signaling by β-adrenergic receptors depends on temporal pattern of stimulation, and that switching may represent a novel mechanism for recruiting kinases involved in synaptic plasticity and memory. PMID:26901880

  10. Control of βAR- and N-methyl-D-aspartate (NMDA) Receptor-Dependent cAMP Dynamics in Hippocampal Neurons.

    PubMed

    Chay, Andrew; Zamparo, Ilaria; Koschinski, Andreas; Zaccolo, Manuela; Blackwell, Kim T

    2016-02-01

    Norepinephrine, a neuromodulator that activates β-adrenergic receptors (βARs), facilitates learning and memory as well as the induction of synaptic plasticity in the hippocampus. Several forms of long-term potentiation (LTP) at the Schaffer collateral CA1 synapse require stimulation of both βARs and N-methyl-D-aspartate receptors (NMDARs). To understand the mechanisms mediating the interactions between βAR and NMDAR signaling pathways, we combined FRET imaging of cAMP in hippocampal neuron cultures with spatial mechanistic modeling of signaling pathways in the CA1 pyramidal neuron. Previous work implied that cAMP is synergistically produced in the presence of the βAR agonist isoproterenol and intracellular calcium. In contrast, we show that when application of isoproterenol precedes application of NMDA by several minutes, as is typical of βAR-facilitated LTP experiments, the average amplitude of the cAMP response to NMDA is attenuated compared with the response to NMDA alone. Models simulations suggest that, although the negative feedback loop formed by cAMP, cAMP-dependent protein kinase (PKA), and type 4 phosphodiesterase may be involved in attenuating the cAMP response to NMDA, it is insufficient to explain the range of experimental observations. Instead, attenuation of the cAMP response requires mechanisms upstream of adenylyl cyclase. Our model demonstrates that Gs-to-Gi switching due to PKA phosphorylation of βARs as well as Gi inhibition of type 1 adenylyl cyclase may underlie the experimental observations. This suggests that signaling by β-adrenergic receptors depends on temporal pattern of stimulation, and that switching may represent a novel mechanism for recruiting kinases involved in synaptic plasticity and memory. PMID:26901880

  11. Inhibitory effects of flavonoids on phosphodiesterase isozymes from guinea pig and their structure-activity relationships.

    PubMed

    Ko, Wun-Chang; Shih, Chwen-Ming; Lai, Ya-Hsin; Chen, Jun-Hao; Huang, Hui-Lin

    2004-11-15

    The structure-activity relationships of flavonoids with regard to their inhibitory effects on phosphodiesterase (PDE) isozymes are little known. The activities of PDE1-5 were measured by a two-step procedure using cAMP with [(3)H]-cAMP or cGMP with [(3)H]-cGMP as substrates. In the present results, PDE1, 5, 2, and 4 isozymes were partially purified from guinea pig lungs in that order, and PDE3 was from the heart. The IC(50) values of PDE1-5 were greater than those reported previously for the reference drugs, vinpocetin, EHNA, milrinone, Ro 20-1724, and zaprinast, by 5-, 5-, 7-, 5-, and 3-fold, respectively. As shown in Table 2, luteolin revealed non-selective inhibition of PDE1-5 with IC(50) values in a range of 10-20 microM, as did genistein except with a low potency on PDE5. Daidzein, an inactive analogue of genistein in tyrosine kinase inhibition, showed selective inhibition of PDE3 with an IC(50) value of around 30 microM, as did eriodictyol with an IC(50) value of around 50 microM. Hesperetin and prunetin exhibited more-selective inhibition of PDE4 with IC(50) values of around 30 and 60 microM, respectively. Luteolin-7-glucoside exhibited dual inhibition of PDE2/PDE4 with an IC(50) value of around 40 microM. Diosmetin more-selectively inhibited PDE2 (IC(50) of 4.8 microM) than PDE1, PDE4, or PDE5. However, biochanin A more-selectively inhibited PDE4 (IC(50) of 8.5 microM) than PDE1 or PDE2. Apigenin inhibited PDE1-3 with IC(50) values of around 10-25 microM. Myricetin inhibited PDE1-4 with IC(50) values of around 10-40 microM. The same was true for quercetin, but we rather consider that it more-selectively inhibited PDE3 and PDE4 (IC(50) of < 10 microM). In conclusion, it is possible to synthesize useful drugs through elucidating the structure-activity relationships of flavonoids with respect to inhibition of PDE isozymes at concentrations used in this in vitro study.

  12. Novel phosphodiesterase 4 inhibitor T-440 reverses and prevents human bronchial contraction induced by allergen.

    PubMed

    Fujii, K; Kohrogi, H; Iwagoe, H; Hamamoto, J; Hirata, N; Goto, E; Kawano, O; Wada, K; Yamagata, S; Ando, M

    1998-01-01

    To study the roles of phosphodiesterase (PDE) 4 in the human airways, we examined the effect of the novel PDE4 inhibitor T-440 in the isolated human bronchus. T-440 inhibited PDE4 extracted from human bronchial smooth muscle. IC50 values for the effect of T-440, rolipram (a PDE4 inhibitor) and theophylline on PDE4 activity of the bronchial tissues were 0.08 microM, 2 microM and > 100 microM, respectively. T-440 (10(-6) M to 10(-5) M) and aminophylline (3.3 x 10(-5) M) significantly reversed the 10(-5) M histamine-induced contraction, the efficacy of 10(-6) M T-440 being almost the same as that of 3.3 x 10(-5) M aminophylline. T-440 (10(-6) M to 10(-5) M) and aminophylline (3.3 x 10(-5) M) significantly reversed the 10(-4) M ACh-induced contraction. But their reversal effects on the ACh-induced contraction were weaker than those on the histamine-induced contraction. T-440 (10(-5) M) significantly reversed the contraction induced by allergen in passively sensitized bronchi. The efficacy of the reversal effect of T-440 (10(-5) M) was significantly higher than that of aminophylline (10(-5) M). T-440 and aminophylline significantly relaxed the basal tension, but pretreatment with T-440 or aminophylline did not significantly prevent histamine- or ACh-induced contraction. In contrast, both T-440 (10(-5) M) and aminophylline (3.3 x 10(-5) M) prevented the contraction induced by allergen, which suggests that PDE4 inhibitor inhibits the release of chemical mediators probably from bronchial mast cells in the allergic response. T-440 (10(-6) M to 10(-5) M) caused the accumulation of cAMP at the concentration that relaxed histamine-induced contraction. Thus selective PDE4 inhibitor is a candidate for the treatment of asthma. PMID:9435174

  13. Inhibitory effects of flavonoids on phosphodiesterase isozymes from guinea pig and their structure-activity relationships.

    PubMed

    Ko, Wun-Chang; Shih, Chwen-Ming; Lai, Ya-Hsin; Chen, Jun-Hao; Huang, Hui-Lin

    2004-11-15

    The structure-activity relationships of flavonoids with regard to their inhibitory effects on phosphodiesterase (PDE) isozymes are little known. The activities of PDE1-5 were measured by a two-step procedure using cAMP with [(3)H]-cAMP or cGMP with [(3)H]-cGMP as substrates. In the present results, PDE1, 5, 2, and 4 isozymes were partially purified from guinea pig lungs in that order, and PDE3 was from the heart. The IC(50) values of PDE1-5 were greater than those reported previously for the reference drugs, vinpocetin, EHNA, milrinone, Ro 20-1724, and zaprinast, by 5-, 5-, 7-, 5-, and 3-fold, respectively. As shown in Table 2, luteolin revealed non-selective inhibition of PDE1-5 with IC(50) values in a range of 10-20 microM, as did genistein except with a low potency on PDE5. Daidzein, an inactive analogue of genistein in tyrosine kinase inhibition, showed selective inhibition of PDE3 with an IC(50) value of around 30 microM, as did eriodictyol with an IC(50) value of around 50 microM. Hesperetin and prunetin exhibited more-selective inhibition of PDE4 with IC(50) values of around 30 and 60 microM, respectively. Luteolin-7-glucoside exhibited dual inhibition of PDE2/PDE4 with an IC(50) value of around 40 microM. Diosmetin more-selectively inhibited PDE2 (IC(50) of 4.8 microM) than PDE1, PDE4, or PDE5. However, biochanin A more-selectively inhibited PDE4 (IC(50) of 8.5 microM) than PDE1 or PDE2. Apigenin inhibited PDE1-3 with IC(50) values of around 10-25 microM. Myricetin inhibited PDE1-4 with IC(50) values of around 10-40 microM. The same was true for quercetin, but we rather consider that it more-selectively inhibited PDE3 and PDE4 (IC(50) of < 10 microM). In conclusion, it is possible to synthesize useful drugs through elucidating the structure-activity relationships of flavonoids with respect to inhibition of PDE isozymes at concentrations used in this in vitro study. PMID:15476679

  14. cAMP inducibility of transcriptional repressor ICER in developing and mature human T lymphocytes.

    PubMed

    Bodor, J; Spetz, A L; Strominger, J L; Habener, J F

    1996-04-16

    Stimulation of the cAMP-dependent signaling pathway exerts an inhibitory effect on the proliferation and effector functions of T cells. The ability of T cells to form high intracellular levels of cAMP is acquired during development in the human thymus and is retained by the majority of mature peripheral T lymphocytes. Here we show that elevated cAMP levels in T cells correlate with the expression of the potent transcriptional repressor ICER (inducible cAMP early repressor) previously described in the hypothalamic-pituitary-gonadal axis. Further, in transcriptional assays in vivo, ICER inhibits calcineurin-mediated expression of the interleukin 2 promoter as well as Tax-mediated transactivation of the human T-lymphotropic virus type I (HTLV-I) promoter. Thus, the induction of ICER in T cells may play an important role in the cAMP-induced quiescence and the persistent latency of HTLV-I.

  15. Allegheny National Forest, CCC Camp ANF1 , The camp’s main ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Allegheny National Forest, CCC Camp ANF-1 , The camp’s main entrance is located at the intersection of Duhring Road (ANF 131) and ANF 124, Pennsylvania, with the interior site road known as Trail Ride Drive., Marienville, Forest County, PA

  16. Project REACH: Camp Goldenrod: Prototype Staff Manual. Appendix R. Camp Staff Training Series.

    ERIC Educational Resources Information Center

    Vinton, Dennis A., Ed.; Farley, Elizabeth M., Ed.

    Resulting from a 3 year project to develop and test competency based programs for camp personnel, the document contains a staff manual for personnel of a residential camp for physically handicapped campers (6 to 18 years old). An introductory section gives an explanation of competency based instruction, describes a module, and provides an overview…

  17. Camping in the Disciplines: Assessing the Effect of Writing Camps on Graduate Student Writers

    ERIC Educational Resources Information Center

    Busl, Gretchen; Donnelly, Kara Lee; Capdevielle, Matthew

    2015-01-01

    In the past ten years, an increasing number of universities have begun organizing writing "camps," or full-week immersion experiences, in an effort to address the increased need to support graduate student writing. Outside of anecdotes and testimonials, we have previously had very little data about these camps' success. This study,…

  18. Different roles of GNAS and cAMP signaling during early and late stages of osteogenic differentiation.

    PubMed

    Zhang, S; Kaplan, F S; Shore, E M

    2012-09-01

    Progressive osseous heteroplasia (POH) and fibrous dysplasia (FD) are genetic diseases of bone formation at opposite ends of the osteogenic spectrum: imperfect osteogenesis of the skeleton occurs in FD, while heterotopic ossification in skin, subcutaneous fat, and skeletal muscle forms in POH. POH is caused by heterozygous inactivating germline mutations in GNAS, which encodes G-protein subunits regulating the cAMP pathway, while FD is caused by GNAS somatic activating mutations. We used pluripotent mouse ES cells to examine the effects of Gnas dysregulation on osteoblast differentiation. At the earliest stages of osteogenesis, Gnas transcripts Gsα, XLαs and 1A are expressed at low levels and cAMP levels are also low. Inhibition of cAMP signaling (as in POH) by 2',5'-dideoxyadenosine enhanced osteoblast differentiation while conversely, increased cAMP signaling (as in FD), induced by forskolin, inhibited osteoblast differentiation. Notably, increased cAMP was inhibitory for osteogenesis only at early stages after osteogenic induction. Expression of osteogenic and adipogenic markers showed that increased cAMP enhanced adipogenesis and impaired osteoblast differentiation even in the presence of osteogenic factors, supporting cAMP as a critical regulator of osteoblast and adipocyte lineage commitment. Furthermore, increased cAMP signaling decreased BMP pathway signaling, indicating that G protein-cAMP pathway activation (as in FD) inhibits osteoblast differentiation, at least in part by blocking the BMP-Smad pathway, and suggesting that GNAS inactivation as occurs in POH enhances osteoblast differentiation, at least in part by stimulating BMP signaling. These data support that differences in cAMP levels during early stages of cell differentiation regulate cell fate decisions. Supporting information available online at http:/www.thieme-connect.de/ejournals/toc/hmr. PMID:22903279

  19. Glycerophosphocholine Metabolism in Higher Plant Cells. Evidence of a New Glyceryl-Phosphodiester Phosphodiesterase

    PubMed Central

    van der Rest, Benoît; Boisson, Anne-Marie; Gout, Elisabeth; Bligny, Richard; Douce, Roland

    2002-01-01

    Glycerophosphocholine (GroPCho) is a diester that accumulates in different physiological processes leading to phospholipid remodeling. However, very little is known about its metabolism in higher plant cells. 31P-Nuclear magnetic resonance spectroscopy and biochemical analyses performed on carrot (Daucus carota) cells fed with GroPCho revealed the existence of an extracellular GroPCho phosphodiesterase. This enzymatic activity splits GroPCho into sn-glycerol-3-phosphate and free choline. In vivo, sn-glycerol-3-phosphate is further hydrolyzed into glycerol and inorganic phosphate by acid phosphatase. We visualized the incorporation and the compartmentation of choline and observed that the major choline pool was phosphorylated and accumulated in the cytosol, whereas a minor fraction was incorporated in the vacuole as free choline. Isolation of plasma membranes, culture medium, and cell wall proteins enabled us to localize this phosphodiesterase activity on the cell wall. We also report the existence of an intracellular glycerophosphodiesterase. This second activity is localized in the vacuole and hydrolyzes GroPCho in a similar fashion to the cell wall phosphodiesterase. Both extra- and intracellular phosphodiesterases are widespread among different plant species and are often enhanced during phosphate deprivation. Finally, competition experiments on the extracellular phosphodiesterase suggested a specificity for glycerophosphodiesters (apparent Km of 50 μm), which distinguishes it from other phosphodiesterases previously described in the literature. PMID:12226504

  20. Phosphodiesterase Inhibition and Regulation of Dopaminergic Frontal and Striatal Functioning: Clinical Implications

    PubMed Central

    Heckman, Pim R. A.; van Duinen, Marlies A.; Bollen, Eva P. P.; Nishi, Akinori; Wennogle, Lawrence P.; Blokland, Arjan

    2016-01-01

    Background: The fronto-striatal circuits are the common neurobiological basis for neuropsychiatric disorders, including schizophrenia, Parkinson’s disease, Huntington’s disease, attention deficit hyperactivity disorder, obsessive-compulsive disorder, and Tourette’s syndrome. Fronto-striatal circuits consist of motor circuits, associative circuits, and limbic circuits. All circuits share 2 common features. First, all fronto-striatal circuits consist of hyper direct, direct, and indirect pathways. Second, all fronto-striatal circuits are modulated by dopamine. Intracellularly, the effect of dopamine is largely mediated through the cyclic adenosine monophosphate/protein kinase A signaling cascade with an additional role for the cyclic guanosine monophosphate/protein kinase G pathway, both of which can be regulated by phosphodiesterases. Phosphodiesterases are thus a potential target for pharmacological intervention in neuropsychiatric disorders related to dopaminergic regulation of fronto-striatal circuits. Methods: Clinical studies of the effects of different phosphodiesterase inhibitors on cognition, affect, and motor function in relation to the fronto-striatal circuits are reviewed. Results: Several selective phosphodiesterase inhibitors have positive effects on cognition, affect, and motor function in relation to the fronto-striatal circuits. Conclusion: Increased understanding of the subcellular localization and unraveling of the signalosome concept of phosphodiesterases including its function and dysfunction in the fronto-striatal circuits will contribute to the design of new specific inhibitors and enhance the potential of phosphodiesterase inhibitors as therapeutics in fronto-striatal circuits. PMID:27037577

  1. Hypothesis: selective phosphodiesterase-5 inhibition improves outcome in preeclampsia.

    PubMed

    Downing, J W; Ramasubramanian, R; Johnson, R F; Minzter, B H; Paschall, R L; Sundell, H W; Engelhardt, B; Lewis, R

    2004-01-01

    The pathogenesis of preeclampsia stems from aberrant changes at the placental interface. The trophoblastic endovascular invasion of tonic spiral arteries that converts them to passive conduits falters. Uteroplacental insufficiency and fetoplacental hypoxemia result. Secondary maternal oxidative stress and an excessive inflammatory response to pregnancy generate the clinical syndrome of preeclampsia. Current treatment focuses on preventing seizures, controlling hypertension, preserving renal function and delivering the baby. We propose that the pathophysiological changes induced by preeclampsia in the placenta parallel those caused by persistent hypoxemia in the lungs at high altitude or with chronic obstructive pulmonary disease. Unrelenting pulmonary hypoxic vasoconstriction induces pulmonary hypertension and cor pulmonale. Inhalation of nitric oxide and phosphodiesterase-5 inhibitors opposes pulmonary hypoxic vasoconstriction, alleviates pulmonary hypertension and improves systemic oxygenation. Notably nitric oxide donor therapy also counters hypoxemic fetoplacental vasoconstriction, a biological response analogous to pulmonary hypoxic vasoconstriction. Fetal oxygenation and nutrition improve. Placental upstream resistance to umbilical arterial blood flow decreases. Fetal right ventricular impedance falls. Heart failure (cor placentale) is avoided. Emergency preterm delivery can be postponed. Other than low dose aspirin and antioxidants vitamins C and E no available therapy specifically targets the underlying disease profile. We hypothesize that, like nitric oxide donation, pharmacological inhibition of placental phosphodiesterase-5 will also protect the fetus but for a longer time. Biological availability of guanosine 3'5'-cyclic monophosphate is boosted due to slowed hydrolysis. Adenosine 3'5'-cyclic monphosphate levels increase in parallel. Cyclic nucleotide accumulation dilates intact tonic spiral arteries and counters hypoxemic fetoplacental vasoconstriction

  2. cAMP signalling meets mitochondrial compartments.

    PubMed

    Lefkimmiatis, Konstantinos

    2014-04-01

    Mitochondria are highly dynamic organelles comprising at least three distinct areas, the OMM (outer mitochondrial membrane), the IMS (intermembrane space) and the mitochondrial matrix. Physical compartmentalization allows these organelles to host different functional domains and therefore participate in a variety of important cellular actions such as ATP synthesis and programmed cell death. In a surprising homology, it is now widely accepted that the ubiquitous second messenger cAMP uses the same stratagem, compartmentalization, in order to achieve the characteristic functional pleiotropy of its pathway. Accumulating evidence suggests that all the main mitochondrial compartments contain segregated cAMP cascades; however, the regulatory properties and functional significance of such domains are not fully understood and often remain controversial issues. The present mini-review discusses our current knowledge of how the marriage between mitochondrial and cAMP compartmentalization is achieved and its effects on the biology of the cell. PMID:24646228

  3. Phosphodiesterase 10A regulates alcohol and saccharin self-administration in rats.

    PubMed

    Logrip, Marian L; Vendruscolo, Leandro F; Schlosburg, Joel E; Koob, George F; Zorrilla, Eric P

    2014-06-01

    A history of stress produces increases in rodent relapse-like alcohol self-administration behavior and regional brain gene expression of phosphodiesterase 10A (PDE10A), a dual-specificity cyclic adenosine monophosphate/cyclic guanosine monophosphate-inhibiting enzyme. Here, we tested the hypothesis that administration of TP-10, a specific PDE10A inhibitor, would reduce alcohol self-administration in conditions predisposing to elevated self-administration. TP-10 administration dose-dependently (0.562, 1.0 mg/kg; subcutaneously) reduced relapse-like alcohol self-administration regardless of stress history enhancement of relapse-like behavior. TP-10 also reduced alcohol self-administration in genetically alcohol-preferring rats, as well as in alcohol-non-dependent and -dependent rats. Effective systemic TP-10 doses did not alter alcohol pharmacokinetics, significantly reduce motor activity or intrabout operant response speed, or promote a conditioned place aversion. TP-10 also reduced saccharin self-administration, suggesting a general role for PDE10A in the self-administration of reinforcing substances. PDE10A inhibition in the dorsolateral striatum, but not the nucleus accumbens, reduced alcohol self-administration. Taken together, the results implicate dorsolateral striatum PDE10A in facilitating alcohol intake and support further investigation of PDE10A systems in the pathophysiology and potential treatment of substance use disorders.

  4. Camp For All Connection: a community health information outreach project*

    PubMed Central

    Huber, Jeffrey T.; Walsh, Teresa J.; Varman, Beatriz

    2005-01-01

    Purpose: The purpose of the Camp For All Connection project is to facilitate access to electronic health information resources at the Camp For All facility. Setting/Participants/Resources: Camp For All is a barrier-free camp working in partnership with organizations to enrich the lives of children and adults with chronic illnesses and disabilities and their families by providing camping and retreat experiences. The camp facility is located on 206 acres in Burton, Texas. The project partners are Texas Woman's University, Houston Academy of Medicine-Texas Medical Center Library, and Camp For All. Brief Description: The Camp For All Connection project placed Internet-connected workstations at the camp's health center in the main lodge and provided training in the use of electronic health information resources. A train-the-trainer approach was used to provide training to Camp For All staff. Results/Outcome: Project workstations are being used by health care providers and camp staff for communication purposes and to make better informed health care decisions for Camp For All campers. Evaluation Method: A post-training evaluation was administered at the end of the train-the-trainer session. In addition, a series of site visits and interviews was conducted with camp staff members involved in the project. The site visits and interviews allowed for ongoing dialog between project staff and project participants. PMID:16059424

  5. Phosphodiesterase 7 Inhibition Induces Dopaminergic Neurogenesis in Hemiparkinsonian Rats

    PubMed Central

    Morales-Garcia, Jose A.; Alonso-Gil, Sandra; Gil, Carmen; Martinez, Ana; Santos, Angel

    2015-01-01

    Parkinson’s disease is characterized by a loss of dopaminergic neurons in a specific brain region, the ventral midbrain. Parkinson’s disease is diagnosed when approximately 50% of the dopaminergic neurons of the substantia nigra pars compacta (SNpc) have degenerated and the others are already affected by the disease. Thus, it is conceivable that all therapeutic strategies, aimed at neuroprotection, start too late. Therefore, an urgent medical need exists to discover new pharmacological targets and novel drugs with disease-modifying properties. In this regard, modulation of endogenous adult neurogenesis toward a dopaminergic phenotype might provide a new strategy to target Parkinson’s disease by partially ameliorating the dopaminergic cell loss that occurs in this disorder. We have previously shown that a phosphodiesterase 7 (PDE7) inhibitor, S14, exerts potent neuroprotective and anti-inflammatory effects in different rodent models of Parkinson’s disease, indicating that this compound could represent a novel therapeutic agent to stop the dopaminergic cell loss that occurs during the progression of the disease. In this report we show that, in addition to its neuroprotective effect, the PDE7 inhibitor S14 is also able to induce endogenous neuroregenerative processes toward a dopaminergic phenotype. We describe a population of actively dividing cells that give rise to new neurons in the SNpc of hemiparkinsonian rats after treatment with S14. In conclusion, our data identify S14 as a novel regulator of dopaminergic neuron generation. Significance Parkinson’s disease is a neurodegenerative disorder characterized by the loss of dopaminergic neurons in the ventral midbrain. Currently, no cure and no effective disease-modifying therapy are available for Parkinson’s disease; therefore, an urgent medical need exists to discover new pharmacological targets and novel drugs for the treatment of this disorder. The present study reports that an inhibitor of the enzyme

  6. Camp Hope: Integrating Tai Chi and Humanbecoming in a Camp for Children.

    PubMed

    Delis, Pamela Coombs; Leger, Robin R; Longton, Fran; Chandler, David Lynn

    2015-10-01

    This article is about Camp Hope, a two-week summer day program for children age 6-12 from low income families and violent neighborhoods and for children who are struggling academically or identified as having behavioral health issues. The camp is operated by House of Peace and Education (HOPE) a 501(c)(3) nonprofit company that began in 1996. The program is designed to offer enrichment activities in a caring community for young children. The theoretical framework behind the vision and policies is Parse's humanbecoming paradigm. In particular, the camp administrators and staff try to see all persons as co-creating of their everchanging humanuniverse process.

  7. 25 CFR 248.9 - Camping and use restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ...-LIEU FISHING SITES § 248.9 Camping and use restrictions. All camping, picnicking, use of alcoholic beverages, setting or use of fires, use of the sites for cleaning of fish, the deposit of any garbage,...

  8. 25 CFR 248.9 - Camping and use restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ...-LIEU FISHING SITES § 248.9 Camping and use restrictions. All camping, picnicking, use of alcoholic beverages, setting or use of fires, use of the sites for cleaning of fish, the deposit of any garbage,...

  9. 25 CFR 248.9 - Camping and use restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ...-LIEU FISHING SITES § 248.9 Camping and use restrictions. All camping, picnicking, use of alcoholic beverages, setting or use of fires, use of the sites for cleaning of fish, the deposit of any garbage,...

  10. 25 CFR 248.9 - Camping and use restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ...-LIEU FISHING SITES § 248.9 Camping and use restrictions. All camping, picnicking, use of alcoholic beverages, setting or use of fires, use of the sites for cleaning of fish, the deposit of any garbage,...

  11. 25 CFR 248.9 - Camping and use restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ...-LIEU FISHING SITES § 248.9 Camping and use restrictions. All camping, picnicking, use of alcoholic beverages, setting or use of fires, use of the sites for cleaning of fish, the deposit of any garbage,...

  12. 8. VIEW EAST, CIVILIAN CONSERVATION CORPS CAMP PARSONS GARAGE, MODERN ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    8. VIEW EAST, CIVILIAN CONSERVATION CORPS CAMP PARSONS GARAGE, MODERN SHED, CIVILIAN CONSERVATION CORPS CAMP PARSONS BLACKSMITH SHOP, AND MODERN SHED. - Parsons Nursery, South side of U.S. Route 219, Parsons, Tucker County, WV

  13. CPSC Warns of Carbon Monoxide Poisoning with Camping Equipment

    MedlinePlus

    ... Warns of Carbon Monoxide Poisoning Hazard with Camping Equipment The U.S. Consumer Product Safety Commission (CPSC) warns ... about the carbon monoxide (CO) hazard with camping equipment. CO can kill you! From 2002–2006, CPSC ...

  14. 3'5'-cyclic adenosine monophosphate-dependent up-regulation of phosphodiesterase type 3A in porcine cumulus cells.

    PubMed

    Sasseville, Maxime; Côté, Nancy; Vigneault, Christian; Guillemette, Christine; Richard, François J

    2007-04-01

    The means by which cumulus cells react to gonadotropin stimulation and regulate the subsequent production and degradation of cAMP are largely unknown. In this article, we report that cyclic nucleotide phosphodiesterase (PDE) type 3A (Pde3a) is transcriptionally regulated in porcine cumulus cells by a cAMP-dependent pathway during in vitro maturation (IVM). cAMP-PDE activity was increased in the cumulus-oocyte complex (COC) after 10 h of IVM, and 78% of this increase was sensitive to a Pde3-specific inhibitor, cilostamide. Although no variation was observed in the oocyte, cilostamide-sensitive cAMP-PDE activity increased in the cumulus cells after IVM. This was supported by Western blotting, which showed that the intensity of a 135-kDa anti-Pde3a immunoreactive band was increased in COC after IVM. The Pde3a mRNA level was up-regulated 28-fold in the COC after 4 h of IVM and remained high up to 12 h. The mRNA up-regulation and increased activity were inhibited by an RNA synthesis inhibitor, alpha-amanitin. The cilostamide-sensitive increase in PDE activity was inhibited by a protein synthesis inhibitor, cycloheximide. Pregnant mare serum gonadotropin (PMSG) caused dose-dependent activation of Pde3. The PMSG-dependent increase in Pde3 activity and Pde3a mRNA were mimicked by the adenylyl cyclase activator forskolin or prostaglandin E2. PMSG-dependent Pde3 activation was inhibited by the protein kinase A-specific inhibitor H89. Collectively, our results show for the first time that degradation of the intracellular cyclic nucleotide by Pde3a is transcriptionally up-regulated via a cAMP-dependent pathway in cumulus cells, suggesting that it has a functional role during the ovulatory gonadotropin surge.

  15. Phosphodiesterase 4B is essential for TH2-cell function and development of airway hyperresponsiveness in allergic asthma

    PubMed Central

    Catherine Jin, S.-L.; Goya, Sho; Nakae, Susumu; Wang, Dan; Bruss, Matthew; Hou, Chiaoyin; Umetsu, Dale; Conti, Marco

    2010-01-01

    Background Cyclic AMP (cAMP) signaling modulates functions of inflammatory cells involved in the pathogenesis of asthma, and type 4 cAMP-specific phosphodiesterases (PDE4s) are essential components of this pathway. Induction of the PDE4 isoform PDE4B is necessary for Toll-like receptor signaling in monocytes and macrophages and is associated with T cell receptor/CD3 in T cells; however, its exact physiological function in the development of allergic asthma remains undefined. Objectives We investigated the role of PDE4B in the development of allergen-induced airway hyperresponsiveness (AHR) and TH2-driven inflammatory responses. Methods Wild-type and PDE4B−/− mice were sensitized and challenged with ovalbumin and AHR measured in response to inhaled methacholine. Airway inflammation was characterized by analyzing leukocyte infiltration and cytokine accumulation in the airways. Ovalbumin-stimulated cell proliferation and TH2 cytokine production were determined in cultured bronchial lymph node cells. Results Mice deficient in PDE4B do not develop AHR. This protective effect was associated with a significant decrease in eosinophils recruitment to the lungs and decreased TH2 cytokine levels in the bronchoalveolar lavage fluid. Defects in T-cell replication, TH2 cytokine production, and dendritic cell migration were evident in cells from the airway-draining lymph nodes. Conversely, accumulation of the TH1 cytokine IFN-γ was not affected in PDE4B−/− mice. Ablation of the orthologous PDE4 gene PDE4A has no impact on airway inflammation. Conclusion By relieving a cAMP-negative constraint, PDE4B plays an essential role in TH2-cell activation and dendritic cell recruitment during airway inflammation. These findings provide proof of concept that PDE4 inhibitors with PDE4B selectivity may have efficacy in asthma treatment. PMID:21047676

  16. STC1 induction by PACAP is mediated through cAMP and ERK1/2 but not PKA in cultured cortical neurons

    PubMed Central

    Holighaus, Yvonne; Weihe, Eberhard; Eiden, Lee E.

    2011-01-01

    The neuroprotective actions of PACAP (pituitary adenylate cyclase-activating polypeptide) in vitro and in vivo suggest that activation of its cognate G protein-coupled receptor PAC1 or downstream signaling molecules, and thus activation of PACAP target genes, could be of therapeutic benefit. Here we show, that cultured rat cortical neurons predominantly expressed the PAC1hop and null variants, activation of which resulted in elevation of the two second messengers cAMP and Ca2+ and expression of the putative neuroprotectant stanniocalcin 1 (STC1). PACAP signaling to the STC1 gene proceeded through the extracellular signal-regulated kinases 1 and 2 (ERK1/2), but not through the cAMP dependent protein kinase (PKA), and was mimicked by the adenylate cyclase activator forskolin. PACAP- and forskolin-mediated activation of ERK1/2 occurred through cAMP, but not PKA. These results suggest that STC1 gene induction proceeds through cAMP and ERK1/2, independently of PKA, the canonical cAMP effector. In contrast, PACAP signaling to the BDNF gene proceeded through PKA, suggesting that two different neuroprotective cAMP pathways co-exist in differentiated cortical neurons. The selective activation of a potentially neuroprotective cAMP dependent pathway different from the canonical cAMP pathway used in many physiological processes, such as memory storage, has implications for pharmacological activation of neuroprotection in vivo. PMID:21975601

  17. Sexual Harassment at Camp: Reducing Liability.

    ERIC Educational Resources Information Center

    Oakleaf, Linda; Grube, Angela Johnson

    2003-01-01

    Employers are responsible for sexual harassment perpetrated by a supervisor. Camps may be responsible for sexual harassment between campers. Steps to reduce liability include providing multiple channels for reporting sexual harassment; having written policies prohibiting sexual harassment and procedures for reporting it; posting these policies and…

  18. E. Coli: Preventing Outbreaks at Camp.

    ERIC Educational Resources Information Center

    McKinney, Mary D.

    1996-01-01

    One strain of E. coli is not usually found in foods, but has been related to consumption of undercooked ground beef. Symptoms are stomach cramps and diarrhea, and 2-7% of infections lead to hemolytic uremic syndrome, which is life threatening. Camps can prevent outbreaks by avoiding uncooked meat on overnight campouts and requiring appropriate…

  19. Seven Habits of Highly Effective Camps.

    ERIC Educational Resources Information Center

    Thurber, Christopher A.

    2002-01-01

    Effective camps share seven habits that are essential elements of success: internal leadership development, explicit expectations for staff, ample camper preparation, personal relationships, supervisors-in-residence, two-way communication flow, and commitment to self-improvement. Three key outcomes for directors, staff, and campers resulting from…

  20. 36 CFR 13.904 - Camping.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Camping. 13.904 Section 13.904 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Denali National Park and Preserve General...

  1. 36 CFR 13.904 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Camping. 13.904 Section 13.904 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Denali National Park and Preserve General...

  2. 36 CFR 13.904 - Camping.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Camping. 13.904 Section 13.904 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Denali National Park and Preserve General...

  3. 36 CFR 13.904 - Camping.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Camping. 13.904 Section 13.904 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Denali National Park and Preserve General...

  4. Teacher Learning in a Digital Writing Camp

    ERIC Educational Resources Information Center

    Zoch, Melody; Myers, Joy; Belcher, Jennifer

    2015-01-01

    This study employed technological pedagogical content knowledge (TPACK) and new literacies frameworks to explore in-service teachers' developing understanding of integrating technology into literacy instruction while concurrently enrolled in a graduate level course and teaching students in a digital writing camp. The inclusion of a…

  5. Developing Leaders for Camp Adventure Programs.

    ERIC Educational Resources Information Center

    Priest, Simon

    1987-01-01

    Discusses effective outdoor leadership as the key to successful adventure programs in camping. Emphasizes unique qualities of adventure tripping and required leadership skills and styles: activity/safety skills, organizational skills, instructional skills, group counseling skills, and experience-based judgment. Provides suggestions for selecting…

  6. Camp Crisis Management: Responding to New Challenges.

    ERIC Educational Resources Information Center

    Evans, Will

    2002-01-01

    Camps should have crisis management plans. Steps to formulating a plan include involving appropriate off-site agencies, identifying potential threats, gathering resources, crafting an appropriate response, training via role-playing, managing incoming and outgoing information, and writing it down. Sidebars present resources, successful response…

  7. 36 CFR 13.1402 - Camping.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Camping. 13.1402 Section 13.1402 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Klondike Gold Rush National Historical Park §...

  8. 36 CFR 13.1402 - Camping.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Camping. 13.1402 Section 13.1402 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Klondike Gold Rush National Historical Park §...

  9. 36 CFR 13.1402 - Camping.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Camping. 13.1402 Section 13.1402 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Klondike Gold Rush National Historical Park §...

  10. The Evening Dance Program at Camp.

    ERIC Educational Resources Information Center

    Cain, Jim

    2002-01-01

    Camp dance programs should be sequenced to maximize campers' comfort levels and create an atmosphere of fun and involvement. Begin with general musical activities that don't involve dancing, such as name that tune. Progress to dances that don't require partners--line dancing, step dancing, and the limbo. Then introduce partnered dancing, using…

  11. 36 CFR 331.9 - Camping.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 3 2012-07-01 2012-07-01 false Camping. 331.9 Section 331.9 Parks, Forests, and Public Property CORPS OF ENGINEERS, DEPARTMENT OF THE ARMY REGULATIONS GOVERNING THE PROTECTION, USE AND MANAGEMENT OF THE FALLS OF THE OHIO NATIONAL WILDLIFE CONSERVATION AREA, KENTUCKY...

  12. 36 CFR 331.9 - Camping.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 3 2014-07-01 2014-07-01 false Camping. 331.9 Section 331.9 Parks, Forests, and Public Property CORPS OF ENGINEERS, DEPARTMENT OF THE ARMY REGULATIONS GOVERNING THE PROTECTION, USE AND MANAGEMENT OF THE FALLS OF THE OHIO NATIONAL WILDLIFE CONSERVATION AREA, KENTUCKY...

  13. 36 CFR 331.9 - Camping.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 3 2011-07-01 2011-07-01 false Camping. 331.9 Section 331.9 Parks, Forests, and Public Property CORPS OF ENGINEERS, DEPARTMENT OF THE ARMY REGULATIONS GOVERNING THE PROTECTION, USE AND MANAGEMENT OF THE FALLS OF THE OHIO NATIONAL WILDLIFE CONSERVATION AREA, KENTUCKY...

  14. 36 CFR 331.9 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 3 2013-07-01 2012-07-01 true Camping. 331.9 Section 331.9 Parks, Forests, and Public Property CORPS OF ENGINEERS, DEPARTMENT OF THE ARMY REGULATIONS GOVERNING THE PROTECTION, USE AND MANAGEMENT OF THE FALLS OF THE OHIO NATIONAL WILDLIFE CONSERVATION AREA, KENTUCKY...

  15. 36 CFR 331.9 - Camping.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 3 2010-07-01 2010-07-01 false Camping. 331.9 Section 331.9 Parks, Forests, and Public Property CORPS OF ENGINEERS, DEPARTMENT OF THE ARMY REGULATIONS GOVERNING THE PROTECTION, USE AND MANAGEMENT OF THE FALLS OF THE OHIO NATIONAL WILDLIFE CONSERVATION AREA, KENTUCKY...

  16. The Gold Mining Camp: A Simulation Game.

    ERIC Educational Resources Information Center

    Stoltman, Joseph P.; Keach, Everett T., Jr.

    This economics simulation game complements the third grade Gold Mining Unit developed by Project Social Studies at the University of Minnesota. The simulation is designed for three purposes: 1) to reinforce the prior learning which occurs in the gold mining camp unit; 2) to involve eight-year-olds in the process of solving simulated economic…

  17. Surgery in a Palestinian refugee camp.

    PubMed

    Cutting, P A; Agha, R

    1992-01-01

    From 1985 to 1987, three Palestinian refugee camps in Lebanon were attacked several times by a Lebanese militia. We present a review of 1276 casualties who were wounded in the refugee camp of Bourj al-Barajneh during two such attacks between December 1985 and April 1987. All were treated in Haifa Hospital (30-40 beds), which had limited equipment, was situated inside the refugee camp and was badly damaged by war. During both attacks, the refugee camp was surrounded and put under siege such that patients could not be evacuated and supplies were not allowed in. The second period lasted almost 6 months resulting in severe shortages of medicines, equipment and food, leading to a rationing of resources and modification of treatment. More than 300 operations were carried out under general anaesthesia, the remainder under local or without anaesthesia. The overall operative mortality was 3.2 per cent. Despite the deprivation, many patients survived severe and complicated wounds because they were quickly brought to the hospital, provided with adequate quantities of fresh blood for transfusion, and sound surgical principles were followed.

  18. 36 CFR 13.25 - Camping.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Camping. 13.25 Section 13.25 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... section, the provisions of § 13.50, or as set forth for specific park areas in subparts H through V...

  19. 36 CFR 13.25 - Camping.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Camping. 13.25 Section 13.25 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... section, the provisions of § 13.50, or as set forth for specific park areas in subparts H through V...

  20. 36 CFR 13.1402 - Camping.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Camping. 13.1402 Section 13.1402 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Klondike Gold Rush National Historical Park §...

  1. 36 CFR 13.1402 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Camping. 13.1402 Section 13.1402 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Klondike Gold Rush National Historical Park §...

  2. Astronomy Camp: Adventures in Scientific Research.

    ERIC Educational Resources Information Center

    Hooper, Eric J.; McCarthy, Donald W.

    1993-01-01

    Explains the nature of the Advanced Astronomy Camp (AAC), which is held at an observatory at the University of Arizona. For one week, students are immersed in doing science and engineering, using modern astronomical equipment under the guidance of scientists and graduate students. (DDR)

  3. Let's Camp (To Be Better Teachers)

    ERIC Educational Resources Information Center

    Kimbrough, Sandy

    2006-01-01

    In this article, the author states that becoming a better physical education teacher can be challenging since it is the time for attending conferences, reading journals, and collaborating with colleagues. She added, that working at a summer camp for children provides a valuable way for physical education teachers not only to earn extra money…

  4. 36 CFR 13.1320 - Camping.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Camping. 13.1320 Section 13.1320 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Kenai Fjords National Park Exit Glacier Developed...

  5. 36 CFR 13.1320 - Camping.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Camping. 13.1320 Section 13.1320 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Kenai Fjords National Park Exit Glacier Developed...

  6. 36 CFR 13.1320 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Camping. 13.1320 Section 13.1320 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Kenai Fjords National Park Exit Glacier Developed...

  7. 36 CFR 13.1320 - Camping.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Camping. 13.1320 Section 13.1320 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Kenai Fjords National Park Exit Glacier Developed...

  8. 36 CFR 13.1320 - Camping.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Camping. 13.1320 Section 13.1320 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Kenai Fjords National Park Exit Glacier Developed...

  9. A Camp Director Remembers World War II.

    ERIC Educational Resources Information Center

    Cohen, Helen Herz

    2003-01-01

    A girl's camp in Maine during World War II had to deal with food rationing and black-market food dealers. Campers picked beans to raise money for refugees, sewed clothes for refugees, and spotted for enemy planes from Mt. Pleasant. An attempt to use a horse-drawn cart for transportation failed, and good help was hard to find. (TD)

  10. 36 CFR 13.904 - Camping.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Camping. 13.904 Section 13.904 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK SYSTEM UNITS IN ALASKA Special Regulations-Denali National Park and Preserve General...

  11. 43 CFR 423.33 - Camping.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... permit issued under 43 CFR part 429; (c) You must not attempt to reserve a campsite for future use by... 43 Public Lands: Interior 1 2014-10-01 2014-10-01 false Camping. 423.33 Section 423.33 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE...

  12. 43 CFR 423.33 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... permit issued under 43 CFR part 429; (c) You must not attempt to reserve a campsite for future use by... 43 Public Lands: Interior 1 2013-10-01 2013-10-01 false Camping. 423.33 Section 423.33 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE...

  13. 43 CFR 423.33 - Camping.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... permit issued under 43 CFR part 429; (c) You must not attempt to reserve a campsite for future use by... 43 Public Lands: Interior 1 2011-10-01 2011-10-01 false Camping. 423.33 Section 423.33 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE...

  14. 36 CFR 13.25 - Camping.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Camping. 13.25 Section 13.25 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... determined by the Superintendent to be interfering with public access or other public interests or...

  15. 43 CFR 423.33 - Camping.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... under 43 CFR part 429; (c) You must not attempt to reserve a campsite for future use by placing... 43 Public Lands: Interior 1 2012-10-01 2011-10-01 true Camping. 423.33 Section 423.33 Public Lands: Interior Regulations Relating to Public Lands BUREAU OF RECLAMATION, DEPARTMENT OF THE INTERIOR...

  16. 36 CFR 13.25 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Camping. 13.25 Section 13.25 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... determined by the Superintendent to be interfering with public access or other public interests or...

  17. CAMP LEJEUNE ENERGY FROM WOOD (CLEW) PROJECT

    EPA Science Inventory

    The paper discusses EPA's Camp Lejeune Energy from Wood (CLEW) project, a demonstration project that converts wood energy to electric power, and provides waste utilization and pollution alleviation. The 1-MWe plant operates a reciprocating engine-generator set on synthetic gas f...

  18. Building Resilient Lives: New Zealand Health Camps

    ERIC Educational Resources Information Center

    Rose, Kirsten

    2009-01-01

    Sometimes it takes a child to raise a whole village. Young people have the gifts and talents to raise their villages. They lack only the confidence of their villages in them to do it. At Te Puna Whaiora Children's Health Camps in New Zealand, the approach to helping families reach their potential is radically changing the lives of many children…

  19. Trends and Issues in Not for Profit Camping.

    ERIC Educational Resources Information Center

    Bialeschki, Deborah; Henderson, Karla; Dahowski, Kate

    A study explored current issues affecting camps run by nonprofit agencies. A literature review revealed areas of concern related to social trends and management practices. A survey of 85 camping professionals and agency executives involved with nonprofit camps examined organizational mission, strategic management issues, and perceived critical…

  20. Creating New Attitudes: Camp Can Shape the Soul.

    ERIC Educational Resources Information Center

    Griffith, Lloyd

    1999-01-01

    Adolescent development can be influenced by two critical factors in camp programming: content of the values that shape and inform the camper and degree of challenge. Camp adventure activities successfully merge recreation and education in an outdoor setting that provides space for the explorations of youth. Camp can play a unique role in guiding…

  1. Transitioning Traditions: Rectifying an Ontario Camp's Indian Council Ring

    ERIC Educational Resources Information Center

    Wilkes, Taylor

    2011-01-01

    Council Ring has always been a very special event, remembered fondly by generations of campers. Taylor Statten Camps (TSC) are not the only camps to cherish such an activity. Across Canada there are dozens of camps that have supported "Indian" assemblies in the past, but a select few still do. Most organizations abandoned them during the 1960s or…

  2. The International Camping Community. A Flame of Friendship.

    ERIC Educational Resources Information Center

    Chamberlain, Richard K.

    1992-01-01

    The International Camping Fellowship (ICF) was organized in 1987 to promote international exchange and collaboration on camping issues. Since its inception, ICF has created a 10-member steering international committee that meets annually to coordinate its work. ICF membership includes more than 140 individuals, camps, and organizations. Lists the…

  3. The Effect of a Camp Experience on Self Esteem.

    ERIC Educational Resources Information Center

    Dorian, Alexia Eve

    Two groups of American adolescents of Greek descent (12-15 year olds N=90 and 16-18 year olds N=166) at an Orthodox Christian Camp in Greece responded to the Rosenberg Self Esteem Scale (1965). Campers took the test on the first day of camp (pretest) and then two weeks later at the camp's conclusion (posttest). All subjects showed a significant…

  4. Illegal Drug Abuse and the Community Camp Strategy in China.

    ERIC Educational Resources Information Center

    Wang, Wen

    1999-01-01

    Provides an initial evaluation of the community camp approach to drug detoxification and rehabilitation utilized in China. Open-ended interviewing schedules were given to two samples from two government sponsored rehabilitation community camps in 1994. Results show that in the two community camps, an average of twelve months' training yielded a…

  5. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 49 Transportation 4 2013-10-01 2013-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  6. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 49 Transportation 4 2014-10-01 2014-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  7. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 49 Transportation 4 2012-10-01 2012-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  8. 7 CFR 502.6 - Hunting, fishing, camping, horseback riding.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 6 2012-01-01 2012-01-01 false Hunting, fishing, camping, horseback riding. 502.6..., MARYLAND § 502.6 Hunting, fishing, camping, horseback riding. The use of BARC grounds for any form of hunting, fishing, camping, or horseback riding is prohibited. Further, the use of these grounds...

  9. 7 CFR 502.6 - Hunting, fishing, camping, horseback riding.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 6 2013-01-01 2013-01-01 false Hunting, fishing, camping, horseback riding. 502.6..., MARYLAND § 502.6 Hunting, fishing, camping, horseback riding. The use of BARC grounds for any form of hunting, fishing, camping, or horseback riding is prohibited. Further, the use of these grounds...

  10. 7 CFR 502.6 - Hunting, fishing, camping, horseback riding.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 6 2014-01-01 2014-01-01 false Hunting, fishing, camping, horseback riding. 502.6..., MARYLAND § 502.6 Hunting, fishing, camping, horseback riding. The use of BARC grounds for any form of hunting, fishing, camping, or horseback riding is prohibited. Further, the use of these grounds...

  11. 7 CFR 502.6 - Hunting, fishing, camping, horseback riding.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 6 2010-01-01 2010-01-01 false Hunting, fishing, camping, horseback riding. 502.6..., MARYLAND § 502.6 Hunting, fishing, camping, horseback riding. The use of BARC grounds for any form of hunting, fishing, camping, or horseback riding is prohibited. Further, the use of these grounds...

  12. 7 CFR 502.6 - Hunting, fishing, camping, horseback riding.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 6 2011-01-01 2011-01-01 false Hunting, fishing, camping, horseback riding. 502.6..., MARYLAND § 502.6 Hunting, fishing, camping, horseback riding. The use of BARC grounds for any form of hunting, fishing, camping, or horseback riding is prohibited. Further, the use of these grounds...

  13. Leadership Skills in Three Chords: Guitars at Camp.

    ERIC Educational Resources Information Center

    Ticknor, Susie

    1992-01-01

    Describes how the guitar can be used to enrich a camp experience. The guitar has an attention-drawing quality, a group-bonding potential, and a mood-setting ability that make it effective in a variety of camp situations. Discusses some common problems with guitar use at camp, and offers recommendations for effective use. (KS)

  14. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 49 Transportation 4 2010-10-01 2010-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  15. Minimum-Impact Camping in the Front Woods.

    ERIC Educational Resources Information Center

    Schatz, Curt

    1994-01-01

    Minimum-impact camping techniques that can be applied to resident camp programs include controlling group size and behavior, designing camp sites, moving groups frequently, proper use of fires, proper disposal of food and human wastes, use of biodegradable soaps, and encouraging staff and camper awareness of impacts on the environment. (LP)

  16. 50 Ways Camps and Campers Can Save the Earth.

    ERIC Educational Resources Information Center

    Henderson, Karla A.

    1991-01-01

    Camp programs have a tremendous potential for providing environmental education and expanding on the existing environmental standards of the American Camping Association. Offers 50 ideas for saving the environment that can be implemented by camp managers, food service managers, office staff, program directors, and campers. (LP)

  17. Camp Health Aide Manual = Manual para trabajadores de salud.

    ERIC Educational Resources Information Center

    Robinson, June Grube; And Others

    This bilingual manual serves as a textbook for migrant Camp Health Aides. Camp Health Aides are members of migrant labor camps enlisted to provide information about health and social services to migrant workers and their families. The manual is divided into 12 tabbed sections representing lessons. Teaching notes printed on contrasting paper…

  18. Emerging Adults' Identity Exploration: Illustrations from inside the "Camp Bubble"

    ERIC Educational Resources Information Center

    Johnson, Sara K.; Goldman, Jane A.; Garey, Anita I.; Britner, Preston A.; Weaver, Shannon E.

    2011-01-01

    The study investigates the experiences of emerging adults who had worked as counselors at overnight summer camps; identity-related issues emerge as most salient in the analysis of in-depth interviews conducted with 12 women and 8 men from 8 camps. Their descriptions portray the identity exploration that took place within the camp context, through…

  19. Summer Camp and Positive Youth Development: Program with Romanian Youth

    ERIC Educational Resources Information Center

    Feenstra, Jennifer S.

    2015-01-01

    A variety of activities are used in camps to help promote positive youth development, improving social skills and self-esteem in campers. I expanded on previous camp research in this study to address the influence camps have on trust, belief in the honesty of others, empowerment, and care for others in youth in Eastern Europe. Since 1999, New…

  20. Integrating Adolescents with Developmental Disabilities in a Residential Summer Camp.

    ERIC Educational Resources Information Center

    Parker, Mitchell S.

    This paper describes Camp Ramah, a residential summer camp in Central Ontario that offers a one month program for adolescents with developmental disabilities. Up to 15 special needs campers, ages 12-25, participate with up to 400 campers and 150 staff in a residential camping experience. Most campers have mild to moderate mental retardation, some…

  1. Trainer Guide: Business and Finance Managerial. Camp Administration Series.

    ERIC Educational Resources Information Center

    Farley, Elizabeth, Ed.

    Suggested ideas on conducting a managerial workshop for camp directors are offered in this trainer's guide. Workshops must be at least one full day of training (6 hours) on each topic to be counted toward the American Camping Association (ACA) Camp Director Certification Program. Suggested topics to be addressed are: (1) basic principles and…

  2. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 49 Transportation 4 2011-10-01 2011-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  3. Youth Camp Safety & Health. Suggested State Statute & Regulations.

    ERIC Educational Resources Information Center

    Center for Disease Control (DHEW/PHS), Atlanta, GA.

    To assist state regulatory agencies in development of comprehensive youth camp safety programs, this publication contains a brief suggested statute that could be used for initiation or modification of any state's youth camp safety programs and it outlines minimal regulations. Various categories of camps are covered--day, primitive, residential,…

  4. 33 CFR 334.910 - Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 33 Navigation and Navigable Waters 3 2013-07-01 2013-07-01 false Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif.; restricted area. 334.910 Section 334.910... AND RESTRICTED AREA REGULATIONS § 334.910 Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine...

  5. 33 CFR 334.910 - Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 3 2011-07-01 2011-07-01 false Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif.; restricted area. 334.910 Section 334.910... AND RESTRICTED AREA REGULATIONS § 334.910 Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine...

  6. 33 CFR 334.910 - Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 33 Navigation and Navigable Waters 3 2014-07-01 2014-07-01 false Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif.; restricted area. 334.910 Section 334.910... AND RESTRICTED AREA REGULATIONS § 334.910 Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine...

  7. 33 CFR 334.910 - Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 3 2012-07-01 2012-07-01 false Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif.; restricted area. 334.910 Section 334.910... AND RESTRICTED AREA REGULATIONS § 334.910 Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine...

  8. 33 CFR 334.910 - Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 3 2010-07-01 2010-07-01 false Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine Corps Base, Camp Pendleton, Calif.; restricted area. 334.910 Section 334.910... AND RESTRICTED AREA REGULATIONS § 334.910 Pacific Ocean, Camp Pendleton Boat Basin, U.S. Marine...

  9. Body Art Comes to Camp: Tattooing and Piercing Are Becoming Mainstream; Does Your Camp Have a Policy?

    ERIC Educational Resources Information Center

    Cameron, Sandy

    2000-01-01

    Tattooing and body piercing are becoming mainstream, especially among the college population that comprises camp staff. Campers often idolize their counselors and want to be like them. Piercings may present a safety hazard. Camps should develop a policy and communicate it to prospective counselors and campers as early as possible. Several camps'…

  10. Histamine receptors on adult rat cardiomyocytes: antagonism of alpha/sub 1/-receptor stimulation of cAMP degradation

    SciTech Connect

    Buxton, I.L.O.; Bowen, S.M.

    1986-03-01

    Incubation of intact cardiomyocytes with the histamine antagonist (/sup 3/H)mepyramine results in rapid reversible binding to a single class of high affinity sites (K/sub D/ = 1.2nM; 50,000 sites/myocyte). In membranes from purified myocytes histamine competition of (/sup 3/H)mepyramine binding (K/sub D/ = 300nM) is not altered by GTP (10..mu..M). Competition of (/sup 3/H)mepyramine binding by H-receptor subtype-selective antagonists suggests the presence of a single class of H/sub 1/-receptors. Incubation of intact myocytes with histamine (luM, H/sub 1/ receptor activation) plus norepinephrine (NE 1uM, alpha/sub 1/ + beta/sub 1/ receptor activation) for 3 min leads to significantly more cAMP accumulation (36.5 pmol/10/sup 6/ myocytes) than NE alone (30 pmol/10/sup 6/ myocytes). Histamine alone does not alter basal cAMP = 10.4 pmol/10/sup 6/ myocytes, or beta/sub 1/ stimulation (isoproternol, 1uM) = 39.6 pmol/10/sup 6/ myocytes. Cyclic AMP accumulation with NE plus prazosin 10nM, (alpha/sub 1/ + beta/sub 1/ + alpha/sub 1/ blockade) is indistinguishable from NE + histamine, (alpha/sub 1/ + beta/sub 1/ + H/sub 1/) stimulation. Histamine competition for (/sup 3/H)prazosin binding suggests that histamine does not block alpha/sub 1/ receptors on the myocyte. These data suggest that H/sub 1/ receptor activation leads to antagonism of the alpha/sub 1/ receptor mediated activation of cAMP phosphodiesterase the authors have recently described.

  11. The Wonders of Phosphodiesterase-5 Inhibitors: A Majestic History

    PubMed Central

    Elhwuegi, AS

    2016-01-01

    The Nobel Prize winning discovery of nitric oxide (NO) in 1986 was the starting point for a new innovation in drug discovery. NO acting as a mediator at different physiological systems is believed to be involved in many physiological and pathological conditions through the formation of the second messenger cyclic guanosine monophosphate (cGMP). cGMP-dependent vasodilation effect of NO is important in regulating pulmonary and systemic pressures, maintaining penis erection, preventing atherosclerosis, preventing platelet aggregation, and protecting and controlling cardiac functions. The main enzyme involved in the termination of cGMP effects is phosphodiesterase enzyme 5 (PDE-5), which is overexpressed in ventricular hypertrophy and heart failure. A milestone in drug discovery was the selective inhibitors of PDE-5 that developed to be a multibillion dollar blockbuster in drug market. PDE-5 inhibitors are approved for the treatment of erectile dysfunctions (EDs), pulmonary hypertension, and benign prostatic hypertrophy. They are also under clinical trials for their cardiac protection against damage induced by ischemia or heart failure. This review article is an update about the pharmacotherapeutics of PDE-5 inhibitors and the majestic history that led to their discovery. The information reported in this review was obtained from the electronic sources of different databases such as PubMed Central, Google Scholar, and Scopus. Keywords used for search included cGMP (mechanisms and functions), EDs (drugs used), nitric oxide, and PDE-5 inhibitors (clinical applications). A total of 165 articles were studied, of which 45 articles were referred to in this review. PMID:27398244

  12. Phosphodiesterase type 5 inhibitor abuse: a critical review.

    PubMed

    Lowe, Gregory; Costabile, Raymond

    2011-06-01

    Abuse of sildenafil has been reported since its introduction in 1999 and commonly documented in combination with illicit drugs among men and women of all ages. Increased risks of sexually transmissible diseases including HIV have been associated with sildenafil use in men who have sex with men. Recognizing the abuse potential of phosphodiesterase type 5 inhibitors (PDE5), we aim to summarize the current knowledge of this abuse. An investigation of EMBASE, PubMed, the Food and Drug Administration (FDA) website, MedWatch, and search engines was performed to evaluate information regarding sildenafil, tadalafil, and vardenafil abuse. The EMBASE search provided 46 articles fitting the search criteria and evaluation led to 21 separate publications with specific information regarding PDE5 abuse. A PubMed search found 10 additional publications. MedWatch reported 44 separate warnings since 2000, most of which reported contamination of herbal products with active drug components. Few reports of abuse were among the 14,818 reports in the FDA AERS for sildenafil. A search for "internet drug store" revealed 6.4 million hits and of 7000 internet pharmacies identified by the Verified Internet Pharmacy Practice Sites Program (VIPPS) only 4% were in proper compliance. The role internet pharmacies play in counterfeit PDE5 or abuse is not well documented; however based on easy access, direct patient marketing, and low advertised cost it is likely this role is underreported. Currently the best recommendation for providers is to recognize the possibility of abuse and to educate patients on risks of this behavior. PMID:21696344

  13. Light and dark active phosphodiesterase regulation in salamander rods

    PubMed Central

    1991-01-01

    We studied the activation of 3',5'-cyclic guanosine monophosphate (cGMP) phosphodiesterase (PDE) by using a cell-permeant enzyme inhibitor. Rods of Ambystoma tigrinum held in a suction electrode were jumped into a stream of 3-isobutyl-1-methylxanthine (IBMX), 0.01-1 mM. Initial transient light-sensitive currents fit the notion that dark and light-activated forms of PDE contributed independently to metabolic activity and were equivalently inhibited by IBMX (apparent Ki 30 microns). Inhibition developed within 50 ms, producing a step decrease of enzyme velocity, which could be offset by activation with flashes or steps of light. The dark PDE activity was equivalent to light activation of enzyme by 1,000 isomerization rod-1s-1, sufficient to hydrolyze the free cGMP pool (1/e) in 0.6 s. Steady light activated PDE in linear proportion to isomerization rate, the range from darkness to current saturation amounting to a 10-fold increase. The conditions for simultaneous onset of inhibitor and illumination to produce no net change of membrane current defined the apparent lifetime of light- activated PDE, TPDE* = 0.9 s, which was independent of both background illumination and current over the range 0-3 x 10(5) isomerization s-1, from 50 to 0 pA. Adaptation was a function of current rather than isomerization: jumps with different proportions of IBMX concentration to steady light intensity produced equal currents, and followed the same course of adaptation in maintained light, despite a 10-fold difference of illumination. Judged from the delay between IBMX- and light-induced currents, the dominant feedback regulatory site comes after PDE on the signal path. The dark active PDE affects the hydrolytic flux and cytoplasmic diffusion of cGMP, as well as the proportional range of the cGMP activity signal in response to light. PMID:1722240

  14. Phosphodiesterase 4D gene polymorphisms in sudden sensorineural hearing loss.

    PubMed

    Chien, Chen-Yu; Tai, Shu-Yu; Wang, Ling-Feng; Hsi, Edward; Chang, Ning-Chia; Wang, Hsun-Mo; Wu, Ming-Tsang; Ho, Kuen-Yao

    2016-09-01

    The phosphodiesterase 4D (PDE4D) gene has been reported as a risk gene for ischemic stroke. The vascular factors are between the hypothesized etiologies of sudden sensorineural hearing loss (SSNHL), and this genetic effect might be attributed for its role in SSNHL. We hypothesized that genetic variants of the PDE4D gene are associated with susceptibility to SSNHL. We conducted a case-control study with 362 SSNHL cases and 209 controls. Three single nucleotide polymorphisms (SNPs) were selected. The genotypes were determined using TaqMan technology. Hardy-Weinberg equilibrium (HWE) was tested for each SNP, and genetic effects were evaluated according to three inheritance modes. We carried out sex-specific analysis to analyze the overall data. All three SNPs were in HWE. When subjects were stratified by sex, the genetic effect was only evident in females but not in males. The TT genotype of rs702553 exhibited an adjusted odds ratio (OR) of 3.83 (95 % confidence interval = 1.46-11.18) (p = 0.006) in female SSNHL. The TT genotype of SNP rs702553 was associated with female SSNHL under the recessive model (p = 0.004, OR 3.70). In multivariate logistic regression analysis, TT genotype of rs702553 was significantly associated with female SSNHL (p = 0.0043, OR 3.70). These results suggest that PDE4D gene polymorphisms influence the susceptibility for the development of SSNHL in the southern Taiwanese female population.

  15. The Wonders of Phosphodiesterase-5 Inhibitors: A Majestic History.

    PubMed

    Elhwuegi, A S

    2016-01-01

    The Nobel Prize winning discovery of nitric oxide (NO) in 1986 was the starting point for a new innovation in drug discovery. NO acting as a mediator at different physiological systems is believed to be involved in many physiological and pathological conditions through the formation of the second messenger cyclic guanosine monophosphate (cGMP). cGMP-dependent vasodilation effect of NO is important in regulating pulmonary and systemic pressures, maintaining penis erection, preventing atherosclerosis, preventing platelet aggregation, and protecting and controlling cardiac functions. The main enzyme involved in the termination of cGMP effects is phosphodiesterase enzyme 5 (PDE-5), which is overexpressed in ventricular hypertrophy and heart failure. A milestone in drug discovery was the selective inhibitors of PDE-5 that developed to be a multibillion dollar blockbuster in drug market. PDE-5 inhibitors are approved for the treatment of erectile dysfunctions (EDs), pulmonary hypertension, and benign prostatic hypertrophy. They are also under clinical trials for their cardiac protection against damage induced by ischemia or heart failure. This review article is an update about the pharmacotherapeutics of PDE-5 inhibitors and the majestic history that led to their discovery. The information reported in this review was obtained from the electronic sources of different databases such as PubMed Central, Google Scholar, and Scopus. Keywords used for search included cGMP (mechanisms and functions), EDs (drugs used), nitric oxide, and PDE-5 inhibitors (clinical applications). A total of 165 articles were studied, of which 45 articles were referred to in this review. PMID:27398244

  16. Summer camps for children with burn injuries: a literature review.

    PubMed

    Maslow, Gary R; Lobato, Debra

    2010-01-01

    The first summer camps for children with burn injuries started over 25 years ago, and as of 2008, there were 60 camps worldwide. This review examines the literature on summer pediatric burn camps. The authors describe common characteristics of burn camp structure, activities, and staffing and then examine the scientific evidence regarding the effect of burn camp programs on campers and camp staff volunteers. A search of Pubmed and Psychinfo databases from 1970 to 2008 for articles related to pediatric burn summer camps identified 17 articles, of which 13 fit the inclusion criteria. Existing literature consists primarily of qualitative studies, suggesting that burn camp can decrease camper isolation, improve self-esteem, and promote coping and social skills. Studies examining volunteer staff at burn camp have consistently found that there are both personal and professional benefits. Quantitative studies of self-esteem have yielded equivocal results. No studies have examined safety or the effect of burn camp on medical or rehabilitation outcomes. For the past 25 years, pediatric summer camps for children with burn injuries have played an important rehabilitation role and provided a strong community that benefits both campers and staff. Future research using more rigorous research methods and examining a broader range of outcomes (eg, safety and medical/rehabilitation outcomes) is recommended. PMID:20644489

  17. PKA-independent cAMP stimulation of white adipocyte exocytosis and adipokine secretion: modulations by Ca2+ and ATP.

    PubMed

    Komai, Ali M; Brännmark, Cecilia; Musovic, Saliha; Olofsson, Charlotta S

    2014-12-01

    We examined the effects of cAMP, Ca(2+) and ATP on exocytosis and adipokine release in white adipocytes by a combination of membrane capacitance patch-clamp recordings and biochemical measurements of adipokine secretion. 3T3-L1 adipocyte exocytosis proceeded even in the complete absence of intracellular Ca(2+) ([Ca(2+)]i; buffered with BAPTA) provided cAMP (0.1 mm) was included in the intracellular (pipette-filling) solution. Exocytosis typically plateaued within ∼10 min, probably signifying depletion of a releasable vesicle pool. Inclusion of 3 mm ATP in combination with elevation of [Ca(2+)]i to ≥700 nm augmented the rate of cAMP-evoked exocytosis ∼2-fold and exocytosis proceeded for longer periods (≥20 min) than with cAMP alone. Exocytosis was stimulated to a similar extent upon substitution of cAMP by the Epac (exchange proteins activated by cAMP) agonist 8-Br-2'-O-Me-cAMP (1 mm included in the pipette solution). Inhibition of protein kinase A (PKA) by addition of Rp-cAMPS (0.5 mm) to the cAMP-containing pipette solution was without effect. A combination of the adenylate cyclase activator forskolin (10 μm) and the phosphodiesterase inhibitor IBMX (200 μm; forsk-IBMX) augmented adiponectin secretion measured over 30 min 3-fold and 2-fold in 3T3-L1 and human subcutaneous adipocytes, respectively. This effect was unaltered by pre-loading of cells with the Ca(2+) chelator BAPTA-AM and 2-fold amplified upon inclusion of the Ca(2+) ionophore ionomycin (1 μm) in the extracellular solution. Adiponectin release was also stimulated by the membrane-permeable Epac agonist 8-Br-2'-O-Me-cAMP-AM but unaffected by inclusion of the membrane-permeable PKA inhibitor Rp-8-Br-cAMPS (200 μm). The adipokines leptin, resistin and apelin were present in low amounts in the incubation medium (1-6% of measured adiponectin). Adipsin was secreted in substantial quantities (50% of adiponectin concentration) but release of this adipokine was unaffected by forsk

  18. PKA-independent cAMP stimulation of white adipocyte exocytosis and adipokine secretion: modulations by Ca2+ and ATP

    PubMed Central

    Komai, Ali M; Brännmark, Cecilia; Musovic, Saliha; Olofsson, Charlotta S

    2014-01-01

    We examined the effects of cAMP, Ca2+ and ATP on exocytosis and adipokine release in white adipocytes by a combination of membrane capacitance patch-clamp recordings and biochemical measurements of adipokine secretion. 3T3-L1 adipocyte exocytosis proceeded even in the complete absence of intracellular Ca2+ ([Ca2+]i; buffered with BAPTA) provided cAMP (0.1 mm) was included in the intracellular (pipette-filling) solution. Exocytosis typically plateaued within ∼10 min, probably signifying depletion of a releasable vesicle pool. Inclusion of 3 mm ATP in combination with elevation of [Ca2+]i to ≥700 nm augmented the rate of cAMP-evoked exocytosis ∼2-fold and exocytosis proceeded for longer periods (≥20 min) than with cAMP alone. Exocytosis was stimulated to a similar extent upon substitution of cAMP by the Epac (exchange proteins activated by cAMP) agonist 8-Br-2′-O-Me-cAMP (1 mm included in the pipette solution). Inhibition of protein kinase A (PKA) by addition of Rp-cAMPS (0.5 mm) to the cAMP-containing pipette solution was without effect. A combination of the adenylate cyclase activator forskolin (10 μm) and the phosphodiesterase inhibitor IBMX (200 μm; forsk–IBMX) augmented adiponectin secretion measured over 30 min 3-fold and 2-fold in 3T3-L1 and human subcutaneous adipocytes, respectively. This effect was unaltered by pre-loading of cells with the Ca2+ chelator BAPTA-AM and 2-fold amplified upon inclusion of the Ca2+ ionophore ionomycin (1 μm) in the extracellular solution. Adiponectin release was also stimulated by the membrane-permeable Epac agonist 8-Br-2′-O-Me-cAMP-AM but unaffected by inclusion of the membrane-permeable PKA inhibitor Rp-8-Br-cAMPS (200 μm). The adipokines leptin, resistin and apelin were present in low amounts in the incubation medium (1–6% of measured adiponectin). Adipsin was secreted in substantial quantities (50% of adiponectin concentration) but release of this adipokine was unaffected by forsk–IBMX. We

  19. Phospholipid turnover and ultrastructural correlates during spontaneous germinal vesicle breakdown of the bovine oocyte: Effects of a cyclic AMP phosphodiesterase inhibitor

    SciTech Connect

    Homa, S.T.; Webster, S.D.; Russell, R.K. )

    1991-08-01

    The turnover of (32P)orthophosphate in bovine oocyte phospholipids was studied during the early stages of spontaneous meiotic maturation, and during inhibition of this process by the cAMP phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (IBMX). Radioactive lipids were separated by TLC and the meiotic stage was determined cytogenetically. Ultrastructure of the nuclear membrane was examined using transmission EM. During the commitment period to meiotic resumption, which precedes germinal vesicle breakdown (GVBD), small localized convolutions appeared in the intact nuclear membrane. This was accompanied by a decrease in (32P)phosphatidic acid (PA) and an increase in (32P)-phosphatidylcholine (PC). This was followed by extensive convolutions, and subsequent dissociation, of the nuclear membrane, concomitant with a tremendous surge in (32P)PC and (32P)phosphatidylethanolamine (PE). The cAMP-mediated maintenance of meiotic arrest involved retention of entire nuclear envelope integrity and total inhibition of the surge in (32P)PC and (32P)PE which accompanied GVBD. The increase in (32P)phosphatidylinositol (PI) associated with all stages of early meiotic resumption was unaffected by IBMX. Microinjection of heparin inhibited GVBD, and injection of inositol 1,4,5-trisphosphate (IP3) overrode IBMX-maintained meiotic arrest in almost 40% of the oocytes. The results suggest that there may be several functions for phospholipid turnover in the regulation of spontaneous meiotic resumption in the bovine oocyte. The first precedes the commitment period, and involves IP3 generation to serve as the primary signal for meiotic resumption. The second occurs concomitant with the commitment period, is unaffected by the level of intracellular cAMP, and is associated with the general turnover of phospholipid.

  20. Improvement of spatial memory function in APPswe/PS1dE9 mice after chronic inhibition of phosphodiesterase type 4D.

    PubMed

    Sierksma, A S R; van den Hove, D L A; Pfau, F; Philippens, M; Bruno, O; Fedele, E; Ricciarelli, R; Steinbusch, H W M; Vanmierlo, T; Prickaerts, J

    2014-02-01

    Phosphodiesterase type 4 inhibitors (PDE4-Is) have received increasing attention as cognition-enhancers and putative treatment strategies for Alzheimer's disease (AD). By preventing cAMP breakdown, PDE4-Is can enhance intracellular signal transduction and increase the phosphorylation of cAMP response element-binding protein (CREB) and transcription of proteins related to synaptic plasticity and associated memory formation. Unfortunately, clinical development of PDE4-Is has been seriously hampered by emetic side effects. The new isoform-specific PDE4D-I, GEBR-7b, has shown to have beneficial effects on memory at non-emetic doses. The aim of the current study was to investigate chronic cognition-enhancing effects of GEBR-7b in a mouse model of AD. To this extent, 5-month-old (5M) APPswe/PS1dE9 mice received daily subcutaneous injections with GEBR-7b (0.001 mg/kg) or vehicle for a period of 3 weeks, and were tested on affective and cognitive behavior at 7M. We demonstrated a cognition-enhancing potential in APPswe/PS1dE9 mice as their spatial memory function at 7M in the object location test was improved by prior GEBR-7b treatment. APPswe/PS1dE9 mice displayed lower levels of CREB phosphorylation, which remained unaltered after chronic GEBR-7b treatment, and higher levels of tau in the hippocampus. Hippocampal brain-derived neurotrophic factor levels and synaptic densities were not different between experimental groups and no effects were observed on hippocampal GSK3β and tau phosphorylation or Aβ levels. In conclusion, GEBR-7b can enhance spatial memory function in the APPswe/PS1dE9 mouse model of AD. Although the underlying mechanisms of its cognition-enhancing potential remain to be elucidated, PDE4D inhibition appears an interesting novel therapeutic option for cognitive deficits in AD.