Receptor and Ion Channel Detection in the Brain provides state-of-the-art and up-to-date methodological information on molecular, neuroanatomical and functional techniques that are currently used to study neurotransmitter receptors and ion channels in the brain. The chapters have been contributed by world-wide recognized neuroscientists who explain in an easy and detailed way well established and tested protocols embracing molecular, cellular, subcellular, anatomical and electrophysiological aspects of the brain. This comprehensive and practical manual is presented in a simple, step-by-step manner for laboratory use, and also offers unambiguous detail and key implementation advice that proves essential for successful results and facilitate choosing the best method for the target proteins under study. This work serves as a useful guide for young researchers and students in training as well as for neurologists and established scientists who wish to extend their repertoire of techniques.
Preface PART I. MOLECULAR TECHNIQUES 1. Production of high-quality antibodies for the study of receptors and ion channels in brain Masahiko Watanabe (Hokkaido University School of Medicine, Japan) 2. Co-immunoprecipitation from brain Xavier Morató, Dasiel O. Borroto-Escuela, Kjell Fuxe,Víctor Fernández-Dueñas and Francisco Ciruela (Universitat de Barcelona, Spain) 3. Sub-synaptic membrane fractionation Paula Canas & Rodrigo Cunha (University of Coimbra, Portugal) 4. Investigation of neurotransmitter receptors in brain slices using cell surface biotinylation Elek Molnar (University of Bristol, Bristol, UK) 5. Single nanoparticle tracking of surface ion channels and receptors in brain cells. Juan Varela, Julien Dupuis, and Laurent Groc (University de Bordeaux, France) 6. Radioligand binding detection of receptors in brain membranes Fuencisla Pilar-Cuéllar, Alvaro Díaz, Emilio Garro-Martínez, Alicia Martín, Beatriz Romero y Elsa M Valdizán (University of Cantabria, Santander, Spain) 7. Recombinant Alphavirus-Mediated Expression of Ion Channels and Receptors in the Brain Markus U. Ehrengruber & Kenneth Lundstrom (University of Zurich, Switzerland) 8. Fluorescent ligands and TR-FRET to study receptor-receptor interactions in the brain Víctor Fernández-Dueñas, Thierry Durrouxand Francisco Ciruela (Universitat de Barcelona, Spain) 9. In situ Proximity Ligation Assay to study and understand the distribution and balance of GPCR homo- and heteroreceptor complexes in the brain Dasiel O. Borroto-Escuela, Miles Woolfenden, Luca Pinton, Antonio Jimenez-Beristein, Julia Oflijan, Manuel Narvaez, Michael Di Palma, Kristin Feltmann, Stefano Sartini, Patrizia Ambrogini, Francisco Ciruela, Riccardo Cuppini and Kjell Fuxe (Karolinska Institutet, Sweeden) PART II. NEUROANATOMICAL TECHNIQUES 10. Fluorescent in situ hybridization for sensitive and specific labeling Miwako Yamasaki and Masahiko Watanabe (Hokkaido University School of Medicine, Japan) 11. Autoradiographic visualization of G protein-coupled receptors in brain Rebeca Vidal, Raquel Linge, María Josefa Castillo, Josep Amigó, Elsa M Valdizán, Elena Castro (University of Cantabria, Santander, Spain) 12. Analysis of the expression profile and regional distribution of neurotransmitter receptors and ion channels in the central nervous system using histoblots Elek Molnar (University of Bristol, UK) 13. Immunohistochemistry for ion channels and their interacting molecules: Tips for improving antibody accessibility Kohtarou Konno & Masahiko Watanabe (Hokkaido University School of Medicine, Japan) 14. Localization of GFP-tagged proteins at the electron microscope Gil-Perotin, S, Cebrián-Silla, A, Herranz-Pérez, V, García-Belda, P, Gil-García, S, Fil, M, Lee JS, Nachury MV, García-Verdugo JM(Universidad de Valencia, Valencia, Spain) 15. Pre-embedding methods in the localization of receptors and ion channels Rafael Luján (Universidad de Castilla-La Mancha, Albacete, Spain) 16. Post-embedding immunohistochemistry in the localization of receptors and ion channels Rafael Luján & Masahiko Watanabe (UCLM, Albacete, Spain) 17. High-Resolution localization of membrane proteins by SDS-digested freeze-fracture replica labelling (SDS-FRL) Harumi Harada, Ryuichi Shigemoto (Institute of Science and Technology IST, Austria) 18. Application of virus vectors for anterograde tract-tracing and single-neuron labeling studies Hiroyuki Hioki, Hisashi Nakamura, Takahiro Furuta (Department of Morphological Brain Science, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan) 19. Analysis of synaptic connections using viral vectors at the electron microscopic level Takahiro Furata, Keiko Okamoto-Furuta, Hiroyuki Hioki(Department of Morphological Brain Science, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan 20. Morphological and neurochemical characterization of electrophysiologically identified cells Yoshiyuki Kubota (Division of Cerebral Circuitry, National Institute for Physiological Sciences, 5-1 Myodaiji-Higashiyama, Okazaki, Aichi 444-8787, Japan) PART III. FUNCTIONAL TECHNIQUES 21. Using electrophysiology to study synaptic and extrasynaptic ionotropic receptors in hippocampal neurons Ian D. Coombs & David Soto (IDIBELL, Spain) 22. Biophysical methods to analyze direct G-protein regulation of neuronal voltage-gated calcium channels Norbert Weiss and Michel De Waard(Academy of Sciences of the Czech Republic, Czech Republic) 23. Electrophysiological recordings in behaving animals Agnès Gruart and José M. Delgado-García (UPO, Spain) 24. Voltammetry in behaving animals Kendra D. Bunner and George V. Rebec (Indiana University, USA) 25. In vivo brain microdialysis of monoamines Jorge E. Ortega, J. Javier Meana, Luis F. Callado (UPV, Spain) 26. Calcium transients in single dendrites and spines of pyramidal neurons in vitro Rhiannon M. Meredith and Martine R. Groen(VU University, The Netherlands) 27. Dynamic recording of membrane potential from hippocampal neurons by using a FRET-based voltage biosensor Víctor Fernández-Dueñas, Xavier Morató, Thomas Knöpfel and Francisco Ciruela (Universitat de Barcelona, Spain) 28. Determination of GPCR-mediated cAMP accumulation in rat striatal synaptosomes Jaume Taura, Víctor Fernández-Dueñas & Francisco Ciruela (Universitat de Barcelona, Spain) 29. GPCR-mediated MAPK/ERK cascade activation in brain slices. Maricel Gómez-Soler, Víctor Fernández-Dueñas & Francisco Ciruela (Universitat de Barcelona, Spain) Index
Receptor and Ion Channel Detection in the Brain provides state-of-the-art and up-to-date methodological information on molecular, neuroanatomical and functional techniques that are currently used to study neurotransmitter receptors and ion channels in the brain. The chapters have been contributed by world-wide recognized neuroscientists who explain in an easy and detailed way well established and tested protocols embracing molecular, cellular, subcellular, anatomical and electrophysiological aspects of the brain. This comprehensive and practical manual is presented in a simple, step-by-step manner for laboratory use, and also offers unambiguous detail and key implementation advice that proves essential for successful results and facilitate choosing the best method for the target proteins under study. This work serves as a useful guide for young researchers and students in training as well as for neurologists and established scientists who wish to extend their repertoire of techniques.
Preface PART I. MOLECULAR TECHNIQUES 1. Production of high-quality antibodies for the study of receptors and ion channels in brain Masahiko Watanabe (Hokkaido University School of Medicine, Japan) 2. Co-immunoprecipitation from brain Xavier Morató, Dasiel O. Borroto-Escuela, Kjell Fuxe,Víctor Fernández-Dueñas and Francisco Ciruela (Universitat de Barcelona, Spain) 3. Sub-synaptic membrane fractionation Paula Canas & Rodrigo Cunha (University of Coimbra, Portugal) 4. Investigation of neurotransmitter receptors in brain slices using cell surface biotinylation Elek Molnar (University of Bristol, Bristol, UK) 5. Single nanoparticle tracking of surface ion channels and receptors in brain cells. Juan Varela, Julien Dupuis, and Laurent Groc (University de Bordeaux, France) 6. Radioligand binding detection of receptors in brain membranes Fuencisla Pilar-Cuéllar, Alvaro Díaz, Emilio Garro-Martínez, Alicia Martín, Beatriz Romero y Elsa M Valdizán (University of Cantabria, Santander, Spain) 7. Recombinant Alphavirus-Mediated Expression of Ion Channels and Receptors in the Brain Markus U. Ehrengruber & Kenneth Lundstrom (University of Zurich, Switzerland) 8. Fluorescent ligands and TR-FRET to study receptor-receptor interactions in the brain Víctor Fernández-Dueñas, Thierry Durrouxand Francisco Ciruela (Universitat de Barcelona, Spain) 9. In situ Proximity Ligation Assay to study and understand the distribution and balance of GPCR homo- and heteroreceptor complexes in the brain Dasiel O. Borroto-Escuela, Miles Woolfenden, Luca Pinton, Antonio Jimenez-Beristein, Julia Oflijan, Manuel Narvaez, Michael Di Palma, Kristin Feltmann, Stefano Sartini, Patrizia Ambrogini, Francisco Ciruela, Riccardo Cuppini and Kjell Fuxe (Karolinska Institutet, Sweeden) PART II. NEUROANATOMICAL TECHNIQUES 10. Fluorescent in situ hybridization for sensitive and specific labeling Miwako Yamasaki and Masahiko Watanabe (Hokkaido University School of Medicine, Japan) 11. Autoradiographic visualization of G protein-coupled receptors in brain Rebeca Vidal, Raquel Linge, María Josefa Castillo, Josep Amigó, Elsa M Valdizán, Elena Castro (University of Cantabria, Santander, Spain) 12. Analysis of the expression profile and regional distribution of neurotransmitter receptors and ion channels in the central nervous system using histoblots Elek Molnar (University of Bristol, UK) 13. Immunohistochemistry for ion channels and their interacting molecules: Tips for improving antibody accessibility Kohtarou Konno & Masahiko Watanabe (Hokkaido University School of Medicine, Japan) 14. Localization of GFP-tagged proteins at the electron microscope Gil-Perotin, S, Cebrián-Silla, A, Herranz-Pérez, V, García-Belda, P, Gil-García, S, Fil, M, Lee JS, Nachury MV, García-Verdugo JM(Universidad de Valencia, Valencia, Spain) 15. Pre-embedding methods in the localization of receptors and ion channels Rafael Luján (Universidad de Castilla-La Mancha, Albacete, Spain) 16. Post-embedding immunohistochemistry in the localization of receptors and ion channels Rafael Luján & Masahiko Watanabe (UCLM, Albacete, Spain) 17. High-Resolution localization of membrane proteins by SDS-digested freeze-fracture replica labelling (SDS-FRL) Harumi Harada, Ryuichi Shigemoto (Institute of Science and Technology IST, Austria) 18. Application of virus vectors for anterograde tract-tracing and single-neuron labeling studies Hiroyuki Hioki, Hisashi Nakamura, Takahiro Furuta (Department of Morphological Brain Science, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan) 19. Analysis of synaptic connections using viral vectors at the electron microscopic level Takahiro Furata, Keiko Okamoto-Furuta, Hiroyuki Hioki(Department of Morphological Brain Science, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan 20. Morphological and neurochemical characterization of electrophysiologically identified cells Yoshiyuki Kubota (Division of Cerebral Circuitry, National Institute for Physiological Sciences, 5-1 Myodaiji-Higashiyama, Okazaki, Aichi 444-8787, Japan) PART III. FUNCTIONAL TECHNIQUES 21. Using electrophysiology to study synaptic and extrasynaptic ionotropic receptors in hippocampal neurons Ian D. Coombs & David Soto (IDIBELL, Spain) 22. Biophysical methods to analyze direct G-protein regulation of neuronal voltage-gated calcium channels Norbert Weiss and Michel De Waard(Academy of Sciences of the Czech Republic, Czech Republic) 23. Electrophysiological recordings in behaving animals Agnès Gruart and José M. Delgado-García (UPO, Spain) 24. Voltammetry in behaving animals Kendra D. Bunner and George V. Rebec (Indiana University, USA) 25. In vivo brain microdialysis of monoamines Jorge E. Ortega, J. Javier Meana, Luis F. Callado (UPV, Spain) 26. Calcium transients in single dendrites and spines of pyramidal neurons in vitro Rhiannon M. Meredith and Martine R. Groen(VU University, The Netherlands) 27. Dynamic recording of membrane potential from hippocampal neurons by using a FRET-based voltage biosensor Víctor Fernández-Dueñas, Xavier Morató, Thomas Knöpfel and Francisco Ciruela (Universitat de Barcelona, Spain) 28. Determination of GPCR-mediated cAMP accumulation in rat striatal synaptosomes Jaume Taura, Víctor Fernández-Dueñas & Francisco Ciruela (Universitat de Barcelona, Spain) 29. GPCR-mediated MAPK/ERK cascade activation in brain slices. Maricel Gómez-Soler, Víctor Fernández-Dueñas & Francisco Ciruela (Universitat de Barcelona, Spain) Index
Preface.- Production of high-quality antibodies for the study of receptors and ion channels in brain.- Co-immunoprecipitation from brain.- Sub-synaptic membrane fractionation.- Investigation of neurotransmitter receptors in brain slices using cell surface biotinylation.- Single nanoparticle tracking of surface ion channels and receptors in brain cells.- Radioligand binding detection of receptors in brain membranes.- Recombinant Alphavirus-Mediated Expression of Ion Channels and Receptors in the Brain.- Fluorescent ligands and TR-FRET to study receptor-receptor interactions in the brain.- In situ Proximity Ligation Assay to study and understand the distribution and balance of GPCR homo- and heteroreceptor complexes in the brain.- Fluorescent in situ hybridization for sensitive and specific labeling.- Autoradiographic visualization of G protein-coupled receptors in brain.- Analysis of the expression profile and regional distribution of neurotransmitter receptors and ion channels in the central nervous system using histoblots.- Immunohistochemistry for ion channels and their interacting molecules: Tips for improving antibody accessibility.- Localization of GFP-tagged proteins at the electron microscope.- Pre-embedding methods in the localization of receptors and ion channels.- Post-embedding immunohistochemistry in the localization of receptors and ion channels.- High-Resolution localization of membrane proteins by SDS-digested freeze-fracture replica labelling (SDS-FRL).- Application of virus vectors for anterograde tract-tracing and single-neuron labeling studies.- Analysis of synaptic connections using viral vectors at the electron microscopic level.- Morphological and neurochemical characterization of electrophysiologically identified cells.- Using electrophysiology to study synaptic and extrasynaptic ionotropic receptors in hippocampal neurons.- Biophysical methods to analyze direct G-protein regulation of neuronal voltage-gated calcium channels.-Electrophysiological recordings in behaving animals.- Voltammetry in behaving animals.- In vivo brain microdialysis of monoamines.- Calcium transients in single dendrites and spines of pyramidal neurons in vitro.- Dynamic recording of membrane potential from hippocampal neurons by using a FRET-based voltage biosensor.- Determination of GPCR-mediated cAMP accumulation in rat striatal synaptosomes.- GPCR-mediated MAPK/ERK cascade activation in brain slices.- Index.
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