INHIBITION OF GAP JUNCTIONAL INTERCELLULAR COMMUNICATION BY BARBITURATES IN LONG-TERM PRIMARY CULTURED RAT HEPATOCYTES IS CORRELATED WITH LIVER-TUMOR PROMOTING ACTIVITY

Rodent liver tumor formation can be promoted by certain barbiturates a nd this may involve their ability to inhibit hepatocyte gap junctional intercellular communication (GJIC), In order to address the mechanism s and specificity of action of barbiturates on hepatocyte gap junction s, we have compared the effects of liver tumor-promoting barbiturates (phenobarbital, sodium barbital and amobarbital: PB, SE and AB, respec tively) and a non-liver tumor-promoting barbiturate (barbituric acid: BA) on primary cultured rat hepatocyte GJIC and connexin32 (Cx32) expr ession after short (1-24 h) and long (2-14 days) treatment, GJIC was e valuated by fluorescent dye microinjection (dye-coupling); Cx32 expres sion was monitored by Northern blot, Western blot and immunohistochemi stry. Both parameters were maintained at high levels over 14 days by c oculture of the cells with WB-F344 rat liver epithelial cells in the p resence of dexamethasone. Treatment with PB (2 mM) for 1 h sharply red uced dye-coupling from similar to 90-30%, but the cells fully recovere d by 24 h, No inhibition was seen with the other barbiturates over thi s 1-day treatment period, Longer treatments (2-14 days) with the promo ters PB, SE and AB, however, gradually reduced hepatocyte dye-coupling to similar to 30-50%, The non-promoter, BA, did not affect hepatocyte GJIC, These decreases in hepatocyte dye-coupling occurred without cha nges in Cx32 or gap junction expression, Dye-coupling of WB-F344 cells and expression of their predominant gap junction protein, connexin43 (Cx43), were also not affected, Thus, the inhibition of GJIC was speci fic to liver tumor promoting barbiturates in hepatocytes, was time-dep endent and was not due to altered Cx32 expression.