Previous studies demonstrated that strain H129 of herpes simplex virus-1 undergoes anterograde transneuronal transport in mice and primates after peripheral or central injection. In this study, H129 was used in rats to identify CNS regions that receive relayed viscero-sensory inputs from the stomach wall. We also examined whether transneuronal viral transport in this model is exclusively anterograde. H129 or an established retrograde transneuronal viral tracer, pseudorabies virus (PRV), was injected into the ventral stomach wall in intact rats or in rats with previous subdiaphragmatic vagal sensory deafferentation. Rats were perfused with fixative 3-5 d later, and tissues were processed for immunocytochemical detection of transported virus. In intact rats, H129 was transported transneuronally via vagal and spinal viscerosensory neurons to postsynaptic target cells in the medullary dorsal vagal complex and thoracic dorsal horn, respectively, with subsequent transport to discrete regions of the medullary and pontine reticular formation, cerebellum, parabrachial nucleus, periaqueductal gray, thalamus, hypothalamus, amygdala, bed nucleus of the stria terminalis, and other central sites. Comparison of labeling patterns in intact and vagal deafferented rats indicated that H129 also produced first-order retrograde infection of autonomic neurons that project directly to virus injection sites, similar to PRV. Unlike PRV, however, H129 was not transported transneuronally in the retrograde direction from infected neurons to central sources of presynaptic input. We conclude that transneuronal transport of H129 occurs exclusively in the anterograde direction to reveal CNS regions that receive direct and relayed viscerosensory signals.