Distinctive gap junction channel types connect WB cells, a clonal cell line derived from rat liver

Gap junctions, dye coupling, and junctional conductance were studied in a cell line (WB) that is derived from rat liver and displays a phenotype similar to ''oval'' cells. In freeze-fracture replicas, two distinctive particle sizes were detected in gap junctional plaques. Immunocytochemical studies indicated punctate staining at membrane appositions using antibodies to connexin43 and to a brain gap junction-associated antigen (34 kDa). No staining was observed using antibodies prepared against rat liver gap junction proteins (connexins 32 and 26). Pairs of WB cells were electrically and dye coupled. Junctional conductance (g(j)) between cell pairs averaged ~10 nS; occasionally, g(j) was low enough that unitary junctional conductances (γ(j)) could be detected. Using a CsCl-containing electrode solution, distinctive γ(j) values were recorded: ~20-30 pS, ~80-90 pS, and the sum of the other sizes. The largest γ(j) events were apparently due to random coincident openings or closures of the smaller channels. Several treatments reduced g(j). Frequency distributions of γ(j) were unaltered by 2 mM halothane or 3.5 heptanol, but the sizes of intermediate and largest events were reduced slightly by 100 nM phorbol ester, and the relative frequency of the largest events was increased by 10 μM glutaraldehyde. We conclude that the distinctive γ(j) values represent openings and closures of two distinct types of gap junction channels rather than substates of a single channel type; these unitary conductances may correspond to the dual immunoreactivity and to the two particle sizes seen in freeze fracture.