In the brain tissue of 21 mice infected with herpes simplex virus type 1 (HSV-1) strain F we determined the expression of immunologic nitric oxide synthase (iNOS) as a potential mediator of neuronal injury with a semiquantitative reverse transcription polymerase chain reaction. Viral burden in brain tissue was quantitated with a dilutional polymerase chain reaction assay. Viral burden and iNOS-expression peaked at day 7 following infection. Thereafter viral burden declined to a low baseline value at 6 months following infection, whereas iNOS-expression was still 4-fold increased compared to baseline levels. In experimental herpes simplex virus encephalitis iNOS, as one potent mediator of neuronal injury, is upregulated in the acute and chronic disease. In future, in addition to antiviral treatment, inhibitors of iNOS might offer new therapeutic strategies in herpes simplex virus encephalitis.