CHANGES IN THE EXPRESSION AND DISTRIBUTION OF CONNEXIN-43 IN ISOLATEDCULTURED ADULT GUINEA-PIG CARDIOMYOCYTES

In the present study, we have investigated the changes in the expressi on and distribution of the principal gap-junction channel protein in v entricular muscle, connexin 43 (Cx43), during the first 2 weeks of cul turing adult guinea pig cardiomyocytes at low density to prevent forma tion of cellular contacts. In freshly isolated cardiomyocytes, immunor eactive Cx43 occupied 6.5 +/- 0.4% of the pixel area of the cell, with 85% being localized to dense particles at the step-like end projectio ns of the myocytes (intercalated disk regions) and 15% being within th e sarcoplasm or along the lateral surface of the myocytes (''nondisk'' distribution). During the myocytes' first 48 h in culture, immunoreac tive Cx43 decreased by 27.5% from control values, to 4.7 +/- 0.5% of t he cells' pixel area (P < 0.01), Cx43 particles also redistributed: af ter 48 h in culture similar to 90% of the immunoreactive Cx43 was loca lized in the sarcoplasm and nondisk regions of the myocyte. After 7 da ys, immunoreactive Cx43 only occupied 50% of the cells' control pixel area (P < 0.01) and was nearly uniform in its punctate pattern through out the sarcoplasm. This distribution remained the same during the 2nd week in culture. Changes in myosin light chain staining during 8 days in culture largely paralleled those in Cx43 staining. Laser confocal microscopic analysis of double-immunolabeled myocytes that had been in culture for 24-48 h showed colocalization of Cx43 with clathrin in si milar to 30% of the sarcoplasmic Cx43 particles. Thus it is demonstrat ed that the expression of Cx43 decreases significantly during the firs t 48 h in culture after myocyte isolation and that Cx43 also undergoes substantial redistribution but for the next 2 weeks remains more or l ess unchanged and at relatively high levels (similar to 50%). These da ta indicate that cardiomyocytes in isolation maintain their ability to reconnect with each other for up to at least 2 weeks. This is the fir st time that this property has been investigated in cultured adult ven tricular cardiomyocytes. (C) 1996 Academic Press, Inc.