In order to determine the localization of the glycinergic neurones responsible for the hyperpolarization of the rat trigeminal motoneurones during paradoxical sleep, we developed a new double immunohistochemical method combining the b subunit of the cholera toxin (CTb), a very sensitive retrograde tracer, with glycine immunohistochemistry. After iontophoretic injections of CTb into the trigeminal motor nucleus (Mo5), a large number of double-labelled cells was observed bilaterally in the parvocellular reticular nucleus alpha, dorsolateral to the descending branch of the facial nerve. A moderate number of double-labelled neurones was found in the ipsilateral parvocellular reticular nucleus at the level of the facial nucleus, and bilaterally in the raphe magnus and the gigantocellular reticular alpha nuclei. These results suggest that the glycinergic neurones hyperpolarizing the trigeminal motoneurons during paradoxical sleep might be localized in the parvocellular reticular nucleus alpha.