We have previously reported on the presence of pacemaker neuronal cells uniquely responsive to CO2. These cells originated in the areas of the nucleus ambiguus and nucleus tractus solitarius, 2 mm rostral to the obex, and are obtained from fetal rats at 19 days of gestation (J. Neurosci. Res., 33:590-597, 1992). We believe these cells to be central chemoreceptors likely involved in the generation of breathing. Last year we reported on the unique CO2 dose response of these pacemaker cells and their catecholamine responses. In the present study, we have further characterized these cells by examining their response to indomethacin, morphine, naloxone, and to a placental extract from sheep which inhibits breathing in the unanesthetized fetal sheep preparation. Indomethacin (50 μmol solution; n=16) increased the spike frequency of the pacemaker cells from 336±41 to 384±65 per minute (p<0.01). Morphine (5μmol solution; n=54), similar to its action in the fetal sheep, stimulated firing of the pacemaker cells from 205±25 to 272±29 spikes per minute (p<0.01); this action was invariably inhibited by naloxone (5μmol solution; n=28). Finally, the placental extract (n=22) consistently increased the activity of the pacemaker cells from 301±35 to 452±52 spikes per minute (p<0.01). In all the above, irregular beating cells responded poorly and silent cells did not respond. We conclude that 1) the responses to indomethacin and morphine mimic those observed in the unanesthetized fetal sheep preparation; and 2) the stimulatory response to placental extract, like that of hypoxia we observed previously in the isolated neuronal cell culture, resembles the response in the midbrain transected fetal sheep, in which the “hypoxia centre” has been freed from its rostral inhibitory connections. We speculate that, as in hypoxia, the placental extract inhibits central breathing output in vivo through susceptible neuromediators, such as adenosine for example.