Comparison of Herpes Simplex Virus Reactivation in Ganglia In Vivo and in Explants Demonstrates Quantitative and Qualitative Differences

Abstract

ABSTRACT The in vivo ganglionic environment directs the latent herpes simplex virus transcriptional program. Since stress-driven perturbations in sensory neurons are thought to play a critical role in the transition from latency to reactivation, a primary concern in the selection of a valid model of the molecular interactions leading to reactivation is the faithful recapitulation of these environments. In this study reactivation of latently infected ganglia excised and cultured in vitro (explanted) is compared to reactivation occurring in latently infected ganglia in vivo following hyperthermic stress. Three notable points emerged. (i) Neurons in explanted ganglia exhibited marked morphological changes within 2 to 3 h postexplant. DNA fragmentation in neuronal nuclei was detected at 3 h, and atypical expression of cell cycle- and stress-regulated proteins such as geminin, cdk2, cdk4, and cytochrome c became apparent at 2 to 48 h. These changes were associated with axotomy and explant and not with the initiation or progression of reactivation and were not observed in ganglia following in vivo hyperthermic stress. (ii) Despite these differences, during the first 22 h primary reactivation events were restricted to a very small number of neurons in vivo and in explanted ganglia. This suggests that at any given time only a few latently infected neurons are competent to reactivate or that the probability of reactivation occurring in any particular neuron is very low. Importantly, the marked changes detected in explanted ganglia were not correlated with increased reactivation, demonstrating that these changes were not associated with the reactivation process per se. (iii) Secondary spread of virus was evident in explanted ganglia within 36 h, an event not observed in vivo. We conclude that explant reactivation may provide an ancillary system for selected studies of the early events in reactivation. However, clear signs of neuronal degeneration within 2 to 3 h postexplant indicate that these ganglia are undergoing major physiological changes not associated with the reactivation process. This ongoing neurodegeneration could alter even the early virus-host interactions in reactivation, and thus caution in the extrapolation of results obtained in explants to the in vivo interactions initiating reactivation is warranted.