Retrograde transynaptic pseudorabies virus infection of central autonomic circuits in neonatal rats

Pseudorabies virus (PRV) is widely used to map synaptically-linked neural circuits in adult animals. The present study sought to determine whether PRV has similar utility in neonatal rats, and whether central PRV infection in neonates elicits astrocytic and microglia/macrophage responses similar to those that contribute to specific transynaptic neuronal infection in adult rats. Retrograde transneuronal infection of autonomic circuits was examined 24-64 h after injection of an attenuated strain of PRV (PRV-Bartha) into the ventral stomach wall of 1-day-old rats. Brain and spinal cord sections were processed for immunocytochemical detection of PRV. Alternate sections were processed for immunolocalization of glial fibrillary acidic protein (GFAP) to identify fibrous astrocytes, or for an antigen associated with the complement C3bi receptor (OX42) to identify microglia. As in adult rats, the number and distribution of infected CNS neurons in neonatal rats increased progressively with advancing post-inoculation survival. Infected CNS neurons initially were restricted to the thoracic intermediolateral cell column and the dorsal motor nucleus of the vagus. Longer survival times led to retrograde transynaptic infection of additional neurons in the thoracic spinal cord, nucleus of the solitary tract, ventrolateral medulla, and caudal raphe nuclei. At the longest post-inoculation intervals, infected neurons also were observed in the area postrema and in certain autonomic-related regions of the rostral brainstem, hypothalamus, and amygdala. Quantitative analysis of immunolabeling in the dorsal vagal complex demonstrated that regions containing neurons at early stages of viral infection displayed increased astrocytic GFAP immunostaining; conversely, areas containing neurons at later stages of infection were characterized by a significant loss of GFAP staining and a parallel increase of OX42 microglia/macrophage immunolabeling. We conclude that PRV is effectively transported through synaptically-linked CNS circuits in neonatal rats, and that spatiotemporally-ordered responses by non-neuronal cells may contribute to the synaptic specificity of transneuronal viral transport.