In Rana catesbeiana the upper airways are used for two distinct yet highly coordinated ventilatory behaviours: buccal ventilation and lung inflation cycles. How these behaviours are generated and coordinated is unknown. The purpose of this study was to identify putative rhythmogenic brainstem loci involved in these ventilatory behaviours. We surveyed the isolated postmetamorphic brainstem to determine sites where local depolarization, produced by microinjecting the non-NMDA glutamate receptor agonist, AMPA, augmented the ventilatory motor patterns. Two sites were identified: a caudal site, at the level of cranial nerve (CN) X, where AMPA injections caused increased buccal burst frequency but abolished lung bursts, and a rostral site, between the levels of CN VIII and IX, where injections increased the frequency of both types of ventilatory bursts. These two sites were further examined using GABA microinjections to locally inhibit cells. GABA injected into the caudal site suppressed the buccal rhythm but the lung rhythm continued, albeit at a different frequency. When GABA was injected into the rostral site the lung bursts were abolished but the buccal rhythm continued. When the two sites were physically separated by transection, both rostral and caudal brainstem sections were capable of rhythmogenesis. The results suggest the respiratory network within the amphibian brainstem is composed of at least two distinct but interacting oscillators, the buccal and lung oscillators. These putative oscillators may provide a promising experimental model for studying coupled oscillators in vertebrates. www.jphysiol.org unpublished observations). Finally, hypercapnic challenge in preparations from postmetamorphic animals causes lung burst frequency to increase but has no effect on buccal frequency (Torgerson et al. 1997b). Transection studies have so far been suggestive but ultimately inconclusive in determining whether lung and buccal rhythm generating circuits are spatially separated. In premetamorphic animals, transection studies indicate that the only region of the brainstem capable of rhythmogenesis resides caudal to CN IX (Gdovin et al. 1999;Torgerson et al. 2001b). In postmetamorphic animals, brainstem sections were capable of rhythmogenesis as long as they included the region between CN VII and IX, suggesting that this region alone was essential for rhythmogenesis (Torgerson et al. 2001b). Thus, transection studies to date have failed to demonstrate the presence of multiple rhythmogenic brainstem sites in the same animal. Here we report the results of a drug microinjection and transection study. Our rationale was to use drug microinjections to identify important sites for rhythmogenesis and then use transection to determine whether rhythmogenesis persisted after the sites were physically separated. Similar techniques have been used previously to identify brain regions important for respiration and other behaviours (e.g. Smith et al. 1991;Coles & Dick, 1996;Ramirez et al. 1998;Solomon et al. 1999;McCrimmon et al. 2000;...
