The antiviral effect of human interferons alpha and beta was inhibited in dose-dependent manner by submillimolar concentrations of neomycin, known to block phosphoinositide hydrolysis and therefore the diacylglycerol formation. On the contrary, the synthetic permeant diacylglycerols (1-oleoyl-2-acetyl-sn or rac-glycerol) were able to induce an interferon-like antiviral state when tested against the vesicular stomatitis virus and herpes simplex type I virus. Hidaka's compound H-8 (1.2 microM), expected to inhibit cAMP- and cGMP-dependent protein kinases, did not modify the antiviral effect of interferon. Our data suggest that the phosphoinositide pathway is involved in transducing the interferon antiviral signal, but, since the exogenous phospholipase C (0.1-1 U/ml) failed to induce an antiviral state, this pathway, although implicated, seems not the only one.