title: Cell-to-cell coupling in engineered pairs of rat ventricular cardiomyocytes: relation between Cx43 immunofluorescence and intercellular electrical conductance.
abstract: Gap junctions are composed of connexin (Cx) proteins, which mediate intercellular communication. Cx43 is the dominant Cx in ventricular myocardium, and Cx45 is present in trace amounts. Cx43 immunosignal has been associated with cell-to-cell coupling and electrical propagation, but no studies have directly correlated Cx43 immunosignal to electrical cell-to-cell conductance, g(j), in ventricular cardiomyocyte pairs. To assess the correlation between Cx43 immunosignal and g(j), we developed a method to determine both parameters from the same cell pair. Neonatal rat ventricular cardiomyocytes were seeded on micropatterned islands of fibronectin. This allowed formation of cell pairs with reproducible shapes and facilitated tracking of cell pair locations. Moreover, cell spreading was limited by the fibronectin pattern, which allowed us to increase cell height by reducing the surface area of the pattern. Whole cell dual voltage clamp was used to record g(j) of cell pairs after 3-5 days in culture. Fixation of cell pairs before removal of patch electrodes enabled preservation of cell morphology and offline identification of patched pairs. Subsequently, pairs were immunostained, and the volume of junctional Cx43 was quantified using confocal microscopy, image deconvolution, and three-dimensional reconstruction. Our results show a linear correlation between g(j) and Cx43 immunosignal within a range of 8-50 nS.
authors.affiliation: Department of Physiology, University of Bern, Bern, Switzerland.
authors.forename: Megan L
authors.initials: ML
authors.lastname: McCain
authors.forename: Thomas
authors.initials: T
authors.lastname: Desplantez
authors.forename: Nicholas A
authors.initials: NA
authors.lastname: Geisse
authors.forename: Barbara
authors.initials: B
authors.lastname: Rothen-Rutishauser
authors.forename: Helene
authors.initials: H
authors.lastname: Oberer
authors.forename: Kevin Kit
authors.initials: KK
authors.lastname: Parker
authors.forename: Andre G
authors.initials: AG
authors.lastname: Kleber
mesh_terms: D000818:Animals; D002450:Cell Communication / Q000502:physiology*; D002478:Cells, Cultured; D018031:Connexin 43 / Q000378:metabolism*; D004553:Electric Conductivity; D017629:Gap Junctions / Q000378:metabolism*; D006352:Heart Ventricles / Q000166:cytology / Q000378:metabolism*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D051381:Rats
chemical_list: D018031:Connexin 43
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