A, ventral view of the rat brainstem showing the location of the serotonergic neurons that reside close to the ventral medullary surface (tryptophan-hydroxylase, TpOHase, immunoreactivity in green; calibration bar: 1.5 mm). The blood vessels are red because they have been filled with a resin. The superficial serotonergic neurons are close to blood vessels and reside in regions of the ventral medullary surface assumed to contain CRCs because experimental acidification of these regions increases breathing. We added the white ovals that outline the retrotrapezoid nucleus. B, typical serotonin neuron in culture that responds briskly to extracellular acidification. A and B reprinted with slight modification from Respiration Physiology and Neurobiology, Volume 168, Corcoran et al., Medullary serotonin neurons and central CO2 chemoreception, pp. 49-58, Copyright (2009), with permission from Elsevier. C. the lateral B3 group of serotonergic neurons is insensitive to hypercapnia in anesthetized rats. C1, example of a serotonergic neuron (tryptophan-hydroxylase-immunoreactive) filled with biotinamide after electrophysiological study in vivo (calibration bar: 20 microns). C2, location of 24 biochemically identified serotonergic neurons found unresponsive to hypercapnia in vivo (red dots). These neurons were located in the lateral band of serotonergic neurons shown in panel A at the level of the RTN (pyr, pyramidal tract; RPa, raphe pallidus; TpOHase, tryptophan hydroxylase). C3 example of a single identified lateral B3 group serotonergic neuron. The neuron was unaffected by increasing end-expiratory CO2 by 5% from just below the apneic threshold. The activity of the respiratory controller, monitored at the level of the phrenic nerve (iPND) was robustly activated by CO2 as expected. C1-3 from (Mulkey et al., 2004). D, raphe obscurus serotonergic neurons driven by the respiratory controller. These recordings were obtained in a coronal slice of neonate medulla oblongata that generates a respiratory-like activity (the “breathing slice”) (Smith et al., 1991). In this slice, the activity of the residual respiratory network was monitored by the mass discharge of the hypoglossal motor neurons (integral XII). In the lower two traces the membrane potential of the serotonergic cell was deliberately hyperpolarized to reveal the excitatory drive potential synchronized with the inspiratory phase. Reproduced with permission from the Journal of Neuroscience from (Ptak et al., 2009). E, putative raphe obscurus serotonergic neurons recorded in a conscious cat. Most cells (21/27) did not respond to hypercapnia. Small subsets of putative serotonergic neurons, one of which is illustrated here, were activated by CO2 but only while the cats were awake. Reproduced with permission from the Journal of Neuroscience from (Veasey et al., 1995). F, the release of serotonin contributes to the activity of the respiratory controller in vitro. The figure shows the mass activity of the preBötzinger region of the ventral respiratory column in the “breathing slice” (ipreBot, an indication of the inspiratory phase of the breathing cycle; top trace) and an inspiratory neuron that was recorded intracellularly (lower trace). Superfusion of the slice with a serotonin receptor 2A antagonist slowed the fictive breathing rate and the amplitude of the respiratory bursts indicating that the ongoing release of serotonin was contributing to the activity of the network under these in vitro conditions. Reproduced with permission from the Journal of Neuroscience from (Pena and Ramirez, 2002). G, putative role of serotonergic neurons in central chemoreception. Left: a small subset of serotonergic neurons, the location of which needs to be clarified, may be central respiratory chemoreceptors, i.e. may be acid-sensitive in vivo and may activate the respiratory controller among other targets. Right, most serotonergic neurons recorded so far in adult mammals in vivo were not CO2-responsive. A few serotonergic neurons were CO2–activated, either because of an intrinsic acid-sensitivity or because they receive excitatory inputs from the respiratory controller. The serotonergic system at large activates the breathing network and regulates its response to CO2. RTN neurons receive an excitatory input from serotonergic cells (Mulkey et al., 2007a). In theory, this input could originate from pH-sensitive (left) or pH-insensitive serotonergic cells (right).