Latent herpes simplex virus infection of the superior cervical autonomic ganglion was reactivated in vivo by postganglionic neurectomy. Two methods were used to demonstrate viral reactivation: (i) recovery of infectious herpes simplex virus in ganglion homogenates and (ii) acceleration of virus expression in ganglion explants in culture. Both the percentage of mice exhibiting reactivated ganglion infection and the viral titers detected in ganglia increased when neurectomized mice were treated with cyclophosphamide. Antithymocyte serum treatment prolonged the time course over which neurectomy-induced virus could be detected, but neither antithymocyte serum nor cyclophosphamide reactivated herpes simplex virus in the absence of neurectomy. These results demonstrate that postganlionic neurectomy provides a specific stimulus for herpes simplex virus reactivation and that cell-mediated immune defense are involved in the highly efficient elimination of reactivated virus from the ganglion in vivo.