In this study, we characterized the pathways that generate the trigeminal blink reflex in the guinea pig. Blinks were evoked by stimulation of the supraorbital branch of the trigeminal nerve and measured by recording electromyographic activity in the lid-closing orbicularis oculi muscle (OOemg) and, in one case, lid position. Blinks evoked by stimulation of the supraorbital nerve consisted of two bursts of muscle activity ipsilateral to the side of stimulation. The first, R1, had a latency of 6.9 ms and the second, R2, had a latency of 17.25 ms. Increasing stimulus intensity to 3 times threshold for evoking an ipsilateral blink elicited an R1 and R2 response contralaterally, with latencies of 9.2 ms and 19.25 ms, respectively. We investigated the causes for this bipartite response that is seen in the guinea pig, as well as other mammals including humans. The two-component response could arise from different populations of afferents, or from different central circuits, or a combination of these two causes. Multiunit recording in the trigeminal ganglion and simultaneous measurement of the OOemg showed that activation of A beta afferents alone was sufficient to elicit both the R1 and the R2 responses, but that activation of A delta afferents could enhance both responses. Different neural circuits, however, produce the R1 and R2 responses. Transganglionic tracing with wheatgerm agglutin or choleragenoid subunit of cholera toxin bound to HRP revealed that primary afferents from the supraorbital branch of the trigeminal nerve terminated densely in the dorsal horn of spinal cord segment C1 and in the caudalis-interpolaris border region of the spinal trigeminal nucleus. Injections of HRP into the orbicularis oculi motoneuron region of the facial nucleus showed that both of these regions projected to the facial nucleus. Hemisections at the level of C1 eliminated the R2 blink response, but not the R1 response, evoked by stimulation of the supraorbital branch of the trigeminal nerve. Subsequent hemisections at the level of the obex eliminated the R1 response. Microinjections of the GABAB agonist baclofen into the spinal trigeminal nucleus at the level of the obex abolished the R1 but not the R2 response. Thus, the spinal trigeminal nucleus produces the R1 component, whereas the R2 component originates in the C1 region of the spinal cord.