Phosphodiesterase Methods and Protocols

Phosphodiesterase Methods and Protocols

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Adenosine 3 ,5 -cyclic monophosphate (cAMP) and guanosine 3 ,5 -cyclic monophosphate are ubiquitous nucleotides that have been described as the first and second messengers. In concert with intracellular calcium and IP3, they play a major role in the control of intracellular signaling, which orchestrates normal and pathophysiological responses. Downstream from the cyclic nucleotide synthesis by adenylyl and guanylyl cyclases, the multigenic family of cyclic nucleotide phosphodiesterases (PDEs), by specifically hydrolyzing cyclic nucleotides, controls cAMP and cGMP levels to maintain a basal state. Their critical role in intracellular sign- ing has recently designated them as new therapeutical targets. Several leading pharmaceutical companies are searching and developing new therapeutic agents that would potently and selectively inhibit PDE isozymes, notably PDE4 and PDE5. Nevertheless, the precise mechanism and the contribution of the various PDE isozymes in modulating intracellular signaling remain to be established. The aim of Phosphodiesterase Methods and Protocols is to provide a palette of a variety of conceptual and technical approaches designed to solve qu- tions concerning the role of PDEs, and ultimately of their different variants, in physiological functions as well as their implications in several more

Product details

  • Hardback | 336 pages
  • 160 x 236.2 x 30.5mm | 748.44g
  • Humana Press Inc.
  • Totowa, NJ, United States
  • English
  • 2005 ed.
  • 1 Illustrations, color; 92 Illustrations, black and white; 336 p. 93 illus., 1 illus. in color.
  • 1588293149
  • 9781588293145

Review quote

"...interesting and thought provoking, but technically challenging...excellent guide to current research techniques in the field of phosphodiesterase." - Doody's Health Sciences Book Review Journalshow more

Back cover copy

The multigenic family of cyclic nucleotide phosphodiesterases (PDEs) is now known to offer many new therapeutic possibilities through their influence over intracellular signaling, though their precise cell mechanisms to modulating that process remain uncertain. In Phosphodiesterase Methods and Protocols, a panel of research leaders in the PDE field describes new concepts and techniques for investigating the role of PDEs in orchestrating normal and pathophysiological responses. Presented in step-by-step detail, these readily reproducible methods allow the measurement of cyclic nucleotide variations in living cells, as well as their visualization in a spatiotemporal manner, the localization and characterization of their activities in tissues and living cells, and the assessment of targeted PDEs in creating specific tools and drugs. Additional chapters discuss the generation of PDE4 knockout mice to demonstrate not only the potential role of targeted PDEs, but also their use in further studies of the central role of PDE regulation in intracellular signaling control. These techniques offer clinicians a way to find new therapies for numerous pathologies whose molecular origins are unknown and whose treatment is symptomatic. Alterations of intracellular signaling related to PDE deregulation in such pathologies as inflammation, neurodegeneration, and cancer may explain the difficulties observed in their prevention and treatment. The protocols follow the successful Methods in Molecular Biology(TM) series format, each offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls. Path-breaking and highly practical, Phosphodiesterase Methods and Protocols offers cell biologists, molecular biologists, pharmacologists, and medicinal chemists a broad ranging survey of the advanced tools and concepts needed to understand the role of PDEs in physiological functions, their implications in several critical pathologies, and the opportunities they offer for new drug more

Table of contents

Study of Cyclic Adenosine Monophosphate Microdomains in Cells Marco Mongillo, Anna Terrin, Sandrine Evellin, Valentina Lissandron, and Manuela Zaccolo High-Resolution Measurements of Cyclic Adenosine Monophosphate Signals in 3D Microdomains Jeffrey W. Karpen and Thomas C. Rich Cygnets: In Vivo Characterization of Novel cGMP Indicators and In Vivo Imaging of Intracellular cGMP Akira Honda, Carolyn L. Sawyer, Sharon M. Cawley, and Wolfgang R. G. Dostmann High-Throughput Screening of Phosphodiesterase Activity in Living Cells Thomas C. Rich and Jeffrey W. Karpen Assessment of Phosphodiesterase Isozyme Contribution in Cell and Tissue Extracts Therese Keravis, Rima Thaseldar-Roumie, and Claire Lugnier Localization of the Cyclic Guanosine 39,59-Monophosphate- Hydrolyzing Phosphodiesterase Type 9 in Rat Brain by Nonradioactive In Situ Hybridization Wilma C. G. van Staveren and Marjanne Markerink-van Ittersum Determination of Ca2+/Calmodulin-Stimulated Phosphodiesterase Activity in Intact Cells Chen Yan Adenovirus-Mediated Overexpression of Murine Cyclic Nucleotide Phosphodiesterase 3B Faiyaz Ahmad, Linda Harndahl, Yan Tang, Lena Stenson Holst, and Vincent C. Manganiello Identification of Promoter Elements in the 5'-Flanking Region of Murine Cyclic Nucleotide Phosphodiesterase 3B Gene Hanguan Liu, Jing Rong Tang, Eva Degerman, and Vincent C. Manganiello Purification of PDE6 Isozymes From Mammalian Retina Dana C. Pentia, Suzanne Hosier, Rachel A. Collupy, Beverly A. Valeriani, and Rick H. Cote Cyclic Guanosine 59-Monophosphate Binding to Regulatory GAF Domains of Photoreceptor Phosphodiesterase Rick H. Cote Renaturation of the Catalytic Domain of PDE4A Expressed in Escherichia coli as Inclusion Bodies Wito Richter, Thomas Hermsdorf, and Dietrich Dettmer Determining the SubunitStructure of Phosphodiesterases Using Gel Filtration and Sucrose Density Gradient Centrifugation Wito Richter Crystallization of Cyclic Nucleotide Phosphodiesterases Hengming Ke, Qing Huai, and Robert X. Xu Generation of PDE4 Knockout Mice by Gene Targeting S.-L. Catherine Jin, Anne M. Latour, and Marco Conti Immunoprecipitation of PDE2 Phosphorylated and Inactivated by an Associated Protein Kinase J. Kelley Bentley Investigation of Extracellular Signal-Regulated Kinase 2 Mitogen-Activated Protein Kinase Phosphorylation and Regulation of Activity of PDE4 Cyclic Adenosine Monophosphate-Specific Phosphodiesterases Elaine V. Hill, Miles D. Houslay, and George S. Baillie Radiolabeled Ligand Binding to the Catalytic or Allosteric Sites of PDE5 and PDE11 James L. Weeks II, Mitsi A. Blount, Alfreda Beasley, Roya Zoraghi, Melissa K. Thomas, Konjeti Raja Sekhar, Jackie D. Corbin, and Sharron H. Francis Analysis of Dimerization Determinants of the PDE6 Catalytic Subunits Khakim G. Muradov, Kimberly K. Boyd, and Nikolai O. Artemyev Interaction Between Catalytic and Inhibitory Subunits of PDE6 Nikolai O. Artemyev Purification, Reconstitution on Lipid Vesicles, and Assays of PDE6 and Its Activator G Protein, Transducin Theodore G. Wensel, Feng He, and Justine A. Malinski Indexshow more