We characterized the role of neurons within the midline of the medulla oblongata on phrenic and hypoglossal nerve responses to hypercapnia during early-development. Studies were performed on decorticate or anesthetized; vagotomized and mechanically ventilated 14-20 day old piglets. Reversible withdrawal of midline neuronal activity was induced by microinjections of lidocaine (2%, 300 nl; n = 10) and lesioning was caused by microinjections of the neurotoxic agent, ibotenic acid (n = 12), at the same sites. At any given end-tidal CO2, peak phrenic and hypoglossal activities after lidocaine were significantly lower than in the control period (P < 0.01). Similarly, 1-2 h after injections of ibotenic acid, both phrenic and hypoglossal nerve responses to CO2 were significantly lower than in the control period (P < 0.01). The results indicate for the first time that the medullary midline neurons are required for full expression of ventilatory responses to hypercapnia and raise the possibility that dysfunction of these nuclei may contribute to respiratory instability during early postnatal life.