Effects of the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and protein kinase C (PKC) inhibitors on cell-cell communication were studied in a normal rat liver cell line, clone 9. Communication was observed and quantitated with microspectrofluorometric and image analysis techniques following scrape-loading of the cells with lucifer yellow. Lucifer yellow migrated as far as ten cells away from the scraped edge in control populations. Two minute TPA (25–50 μg/ml) treatment inhibited dye movement such that the dye remained mainly in the cells at the cut edge. The TPA-induced inhibition of cell-cell communication could be partially blocked by 15 min treatment of the cell populations with the PKC inhibitors trifluoperazine (30 μg/ml), staurosporine (2 × 10−8 or 2 × 10−6 M), sangivamycin (15 or 200 μM), or a PKC inhibitor peptide (20 μg/ml) scraped in at the same time as lucifer yellow. Normal communication was observed in cultures treated only with PKC inhibitors. Lower concentrations of TPA (50 ng/ml-1 μg/ml) used for 2 min did not inhibit dye communication. Our results demonstrate the phorbol ester-induced interruption of cell-cell communication. The inhibition of PKC by inhibitors eliminates the effect of TPA on communication. Our data are consistent with a role of PKC in the control of junctional communication. © 1993 Wiley-Liss, Inc.