Electrical propagation in synthetic ventricular myocyte strands from germline connexin43 knockout mice.
To characterize the role of connexin43 (Cx43) as a determinant of cardiac propagation, we synthesized strands and pairs of ventricular myocytes from germline Cx43-/- mice. The amount of Cx43, Cx45, and Cx40 in gap junctions was analyzed by immunohistochemistry and confocal microscopy. Intercellular electrical conductance, gj, was measured by the dual-voltage clamp technique (DVC), and electrical propagation was assessed by multisite optical mapping of transmembrane potential using a voltage-sensitive dye. Compared with wild-type (Cx43+/+) strands, immunoreactive signal for Cx43 was reduced by 46% in Cx43+/- strands and was absent in Cx43-/- strands. Cx45 signal was reduced by 46% in Cx43+/- strands and to the limit of detection in Cx43-/- strands, but total Cx45 protein levels measured in immunoblots of whole cell homogenates were equivalent in all genotypes. Cx40 was detected in 2% of myocytes. Intercellular conductance, gj, was reduced by 32% in Cx43+/- cell pairs and by 96% in Cx43-/- cell pairs. The symmetrical dependence of gj on transjunctional voltage and properties of single-channel recordings indicated that Cx45 was the only remaining connexin in Cx43-/- cells. Propagation in Cx43-/- strands was very slow (2.1 cm/s versus 52 cm/s in Cx43+/+) and highly discontinuous, with simultaneous excitation within and long conduction delays (2 to 3 ms) between individual cells. Propagation was abolished by 1 mmol/L heptanol, indicating residual junctional coupling. In summary, knockout of Cx43 in ventricular myocytes leads to very slow conduction dependent on the presence of Cx45. Electrical field effect transmission does not contribute to propagation in synthetic strands.
Beauchamp
Philippe
P
Department of Physiology, University of Bern, Switzerland.
Choby
Cécile
C
Desplantez
Thomas
T
de Peyer
Karin
K
Green
Karen
K
Yamada
Kathryn A
KA
Weingart
Robert
R
Saffitz
Jeffrey E
JE
Kléber
André G
AG
D000200:Action Potentials; D000818:Animals; D002478:Cells, Cultured / Q000502:physiology; D018031:Connexin 43 / Q000235:genetics / Q000502:physiology*; D017630:Connexins / Q000032:analysis / Q000502:physiology; D004553:Electric Conductivity; D017629:Gap Junctions / Q000737:chemistry / Q000502:physiology; D018095:Germ-Line Mutation; D006352:Heart Ventricles / Q000166:cytology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D046912:Multiprotein Complexes; D032383:Myocytes, Cardiac / Q000502:physiology*; D018408:Patch-Clamp Techniques; D000097003:Gap Junction alpha-5 Protein
D018031:Connexin 43; D017630:Connexins; D046912:Multiprotein Complexes; D000097003:Gap Junction alpha-5 Protein; C082920:connexin 45