Fate of intercellular junctions in isolated adult rat cardiac cells

Freeze fracture and thin section techniques have revealed morphological changes in gap junctions and intercalated discs of adult rat myocytes following enzymatic dissociation. Cell separation leaves behind small vesicular remnants of formerly adjacent cells connected to the intact cell by gap junctions; in contrast, desmosomes cleave at the region of intercellular contact. Apparently, the next step in gap junction breakdown is internalization of the remnants. In thin section, lanthanum penetration reveals that the cleft of some apparently internalized gap junctions is in contact with the sarcolemma, while that of others is not. In freeze fracture replicas, cytoplasmic gap junctions frequently possess hexagonally packed domains of E-face pits separated by smooth regions that may correspond to separations of membranes of internalized junctions found in thin section. Study of material maintained overnight at 37°C showed no surface junctional remnants; topologies of cytoplasmic gap junctions were generally complex, and concentric membrane vesicles were common. These observations suggest that enzymatic dissociation initiates a progressive, defined sequence of junctional internalization that begins with attached cell remnants and may have as the last determinable step the separation into single membranes inside the intact cell.