title: Herpesvirus-mediated delivery of a genetically encoded fluorescent Ca(2+) sensor to canine cardiomyocytes.
abstract: We report the development and application of a pseudorabies virus-based system for delivery of troponeon, a fluorescent Ca(2+) sensor to adult canine cardiomyocytes. The efficacy of transduction was assessed by calculating the ratio of fluorescently labelled and nonlabelled cells in cell culture. Interaction of the virus vector with electrophysiological properties of cardiomyocytes was evaluated by the analysis of transient outward current (I(to)), kinetics of the intracellular Ca(2+) transients, and cell shortening. Functionality of transferred troponeon was verified by FRET analysis. We demonstrated that the transfer efficiency of troponeon to cultured adult cardiac myocytes was virtually 100%. We showed that even after four days neither the amplitude nor the kinetics of the I(to) current was significantly changed and no major shifts occurred in parameters of [Ca(2+)](i) transients. Furthermore, we demonstrated that infection of cardiomyocytes with the virus did not affect the morphology, viability, and physiological attributes of cells.
authors.affiliation: Department of Pharmacology and Pharmacotherapy, University of Szeged, Hungary.
authors.forename: János
authors.initials: J
authors.lastname: Prorok
authors.forename: Péter P
authors.initials: PP
authors.lastname: Kovács
authors.forename: Attila A
authors.initials: AA
authors.lastname: Kristóf
authors.forename: Norbert
authors.initials: N
authors.lastname: Nagy
authors.forename: Dóra
authors.initials: D
authors.lastname: Tombácz
authors.forename: Judit S
authors.initials: JS
authors.lastname: Tóth
authors.forename: Balázs
authors.initials: B
authors.lastname: Ordög
authors.forename: Norbert
authors.initials: N
authors.lastname: Jost
authors.forename: László
authors.initials: L
authors.lastname: Virág
authors.forename: Julius G
authors.initials: JG
authors.lastname: Papp
authors.forename: András
authors.initials: A
authors.lastname: Varró
authors.forename: András
authors.initials: A
authors.lastname: Tóth
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Boldogkoi
mesh_terms: D000818:Animals; D002118:Calcium / Q000032:analysis* / Q000378:metabolism; D048430:Cell Shape; D002470:Cell Survival; D002478:Cells, Cultured; D004285:Dogs; D031541:Fluorescence Resonance Energy Transfer; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics; D011558:Herpesvirus 1, Suid / Q000235:genetics*; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics*; D008856:Microscopy, Fluorescence; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism / Q000502:physiology* / Q000821:virology; D018408:Patch-Clamp Techniques; D019209:Troponin C / Q000096:biosynthesis / Q000235:genetics*
chemical_list: D008164:Luminescent Proteins; D019209:Troponin C; D002118:Calcium