Endocannabinoid Signaling Methods and Protocols

This second edition provides new and updated chapters detailing all major elements of the ECB system. Chapters guide readers through identification of drug targets, electrophysiology, computational chemistry, and machine learning. Written in the successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls.

 Comprehensive and cutting-edge, Endocannabinoid Signaling: Methods and Protocols, Second Edition is a valuable resource for all researchers interested in learning more about this important and developing field.

Need for Methods to Investigate Endocannabinoid Signaling.- Extraction and Simultaneous Quantification of Endocannabinoids and Endocannabinoid-like Lipids in Biological Tissues.- Measuring the Content of Endocannabinoid-like Compounds in Biological Fluids: A Critical Overview of Sample Preparation Methodologies.- LC-MS/MS Analysis of AEA and 2-AG.- Analysis of Omega-3 Fatty Acid Derived N-Acylethanolamines in Biological Matrices.- Endocannabinoid-Binding Receptors as Drug Targets.- Assay of CB1 Receptor Binding.- The Displacement Binding Assay Using Human Cannabinoid CB2 Receptor-Transfected Cells.- Fluorescence-based Assay for TRPV1 Channels.- DNA-Protein-Interaction (DPI)-ELISA Assay for PPAR-? Receptor Binding.- Scintillation Proximity Assay (SPA)-Based Radioligand Binding for PPARa, PPAR? and PPARd Receptors.- A Homogeneous Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Cofactor Recruitment Assay for PPARa andPPAR?.- The Cyclic AMP Assay Using Human Cannabinoid CB2 Receptor-Transfected Cells.- Assay of GTP?S Binding in Autoradiography.- Cellular Assay to Study ß-Arrestin Recruitment by the Cannabinoid Receptors 1 and 2.- Endocannabinoid Metabolism and Transport as Drug Targets.- Assay of NAT Activity.- Radiometric Assay of NAPE-PLD activity.- A Fluorescence-based NAPE-PLD Activity Assay.- Radiometric Assay of FAAH Activity.- Fluorimetric Assay of FAAH activity.- Assay of NAAA Activity.- Assay of DAGLa/ß Activity.- Assay of Monoacylglycerol Lipase Activity.- Radiometric Assay of ABHD2 Activity.- Oxygenation of Anandamide by Lipoxygenases.- Assay of Endocannabinoid Oxidation by Cytochrome P450.- Assays of Endocannabinoid Uptake.- Assessing Gene Expression of the Endocannabinoid System Components by Real-Time Quantitative Reverse Transcription Polymerase Chain Reaction.- Bioinformatics of the Endocannabinoid System: Study of DNA methylation at Rat Cnr1 gene promoter.- DNA Methylation Analysis of Cnr1 Gene Promoter.- Western Blotting of the Endocannabinoid System.- Quantitation of Plasma Membrane (G Protein-Coupled) Receptor Trafficking in Cultured Cells.- Assessing CB1 Expression in the Brain by Immunohistochemical Methods: Light, Confocal and Electron Microscopy.- Measuring Endocannabinoid System Interaction with Biomembranes.- STORM Super-Resolution Imaging of CB1 Receptors in Tissue Preparations.- Visualization of Endocannabinoids in the Cell.- Electrophysiology of Endocannabinoid Signalling.- Machine Learning and Computational Chemistry for the Endocannabinoid System.- A Virtual Reverse Screening Approach to Target Type 2 Cannabinoid Receptor.