Stereotaxic microinjection of herpes simplex virus (HSV) into the mouse olfactory bulb resulted in infection of neurons of the piriform cortex. Neurons infected with the wildtype HSV showed no evident phosphorylation of c-Jun N-terminal protein kinase (JNK)/c-Jun. In contrast, neurons infected with a US3 gene-disrupted mutant of the L1BR1 virus displayed phosphorylated JNK/c-Jun in a nuclear staining fashion. Induction of neuronal apoptosis by the wildtype HSV was partially suppressed when compared with that of the L1BR1 virus. A US3-rescued isolate of the L1B(-)11 virus behaved as did the wildtype virus. Collectively, the US3 protein kinase of HSV plays a role in attenuating the virus-induced activation of the JNK signal transduction pathway in the central nervous system and may contribute, at least in part, to controlling neuronal apoptosis.