Abstract-Connexin43 (Cx43) is a major determinant of the electrical properties of the myocardium. Closure of gap junctions causes rapid slowing of propagation velocity (), but the precise effect of a reduction in Cx43 levels due to genetic manipulation has only partially been clarified. In this study, morphological and electrical properties of synthetic strands of cultured neonatal ventricular myocytes from Cx43 ϩ/ϩ (wild type, WT) and Cx ϩ/Ϫ (heterozygote, HZ) mice were compared. Quantitative immunofluorescence analysis of Cx43 demonstrated a 43% reduction of Cx43 expression in the HZ versus WT mice. Cell dimensions, connectivity, and alignment were independent of genotype. Measurement of electrical properties by microelectrodes and optical mapping showed no differences in action potential amplitude or minimum diastolic potential between WT and HZ. However, maximal upstroke velocity of the transmembrane action potential, dV/dt max , was increased and action potential duration was reduced in HZ versus WT. was similar in the two genotypes. Computer simulation of propagation and dV/dt max showed a relatively small dependence of on gap junction coupling, thus explaining the lack of observed differences in between WT and HZ. Importantly, the simulations suggested that the difference in dV/dt max is due to an upregulation of I Na in HZ versus WT. Thus, heterozygote-null mutation of Cx43 produces a complex electrical phenotype in synthetic strands that is characterized by both changes in ion channel function and cell-to-cell coupling. The lack of changes in in this tissue is explained by the dominating role of myoplasmic resistance and the compensatory increase of dV/dt max . Key Words: synthetic cardiac strands Ⅲ neonatal mouse cardiomyocytes Ⅲ connexin43 expression Ⅲ conduction velocity P ropagation of the cardiac impulse is a complex process that depends on the active electrical properties of myocyte membranes and the passive properties of the cellular network. The connexin proteins form the electrical connections between myocytes, and are therefore important determinants of the myocardial electrical properties. The most abundant connexin in ventricular myocardium of both mice and humans is connexin43 (Cx43). 1 The overall electrical conductance between cardiac cells can be changed in two basic ways: (1) by affecting gap junctional conductance (drugs 2,3 and myocardial ischemia 4 ), or (2) by changing connexin expression. 5 Computer simulations 6 and experimental studies 7 have shown that a large reduction in gap junctional conductance can result in reduction of to values of a few centimeters per second, an extreme degree of slowing that cannot be achieved with suppression of current flow through ion channels. 8 More moderate changes in gap junctional conductance due to a change of connexin expression have been shown to occur in ventricular hypertrophy and failure 9,10 and after application of mediators of hypertrophy 11 and mechanical stretch. 12 Mice with a targeted deletion of Cx43 have reduced expression o...

Abstract-The aim of the present study was to morphologically and electrically characterize synthetic strands of mouse ventricular myocytes. Linear strands of mouse ventricular myocytes with widths of 34.7Ϯ4.4 m (W 1 ), 57.9Ϯ2.5 m (W 2 ), and 86.4Ϯ3.6 m (W 3 ) and a length of 10 mm were produced on glass coverslips with a photolithographic technique. Action potentials (APs) were measured from individual cells within the strands with cell-attached microelectrodes. Impulse propagation and AP upstrokes were measured with multisite optical mapping (RH237).Immunostaining was performed to assess cell-cell connections and myofibril arrangement with polyclonal antisera against connexin43 and N-cadherins and monoclonal antibodies against cardiac myosin. Light microscopy and myosin staining showed dense growth of well-developed elongated myocytes with lengths of 34.2Ϯ4.2 m (W 1 ), 36.9Ϯ5.8 m (W 2 ), and 43.7Ϯ6.9 m (W 3 ), and length/width ratios of 3.9Ϯ0.2. Gap junctions were distributed around the cell borders (3 to 4 junctions/m 2 cell area). Each cell was connected by gap junctions to 6.5Ϯ1.1 neighboring cells. AP duration shortened with time in culture (action potential duration at 50% repolarization: day 4, 103Ϯ34 ms; day 8, 16Ϯ3 ms; PϽ0.01). Minimum diastolic potential and AP amplitude were 71Ϯ5 and 97.2Ϯ7.6 mV, respectively. Conduction velocity and the maximum dV/dt of the AP upstroke were 43.9Ϯ13.6 cm/s and 196Ϯ67 V/s, respectively. Thus, neonatal ventricular mouse myocytes can be grown in continuous synthetic strands. Gap junction distribution is similar to the neonatal pattern observed in the hearts of larger mammals. Conduction velocity is in the range observed in adult mice and in the higher range for mammalian species probably due to the higher dV/dt max . This technique will permit the study of propagation, AP, and structure-function relations at cellular resolution in genetically modified mice. (Circ Res. 2000;87:467-473.)

SCW's occurring at increased levels of [Ca(2+)](i) in normoxic or ischemic conditions are mostly confined to two or three cells in the ventricular myocardium. Spatio-temporal summation of changes in membrane potential caused by individual SCW's may underlie the generation of triggered electrical ectopic impulses.