Figure 2.

RTN chemoreceptors express Phox2b. A, Identification of a putative RTN chemoreceptor. This neuron was recorded below the caudal end of the facial motor nucleus in vivo. Graded levels of CO₂ were added to the breathing mixture to quantify the steady-state relationship between the firing rate of the cell and end-expiratory CO₂. The inset illustrates the activity of the cell at a higher resolution. Note the slight tendency of the cell to slow during inspiration and postinspiration. AP, Arterial pressure. B, Perievent activity histogram triggered on the integrated phrenic discharge (iPND). This analysis reveals that the cell is only mildly entrained by the central respiratory network. The averaged CO₂ trace is flat, because the vagus nerves were cut, and this procedure uncoupled the central respiratory rhythm from the ventilation cycle. C, Mean relationship between RTN neuronal discharge and end-expiratory CO₂ at steady state (18 neurons). The graph also illustrates the relationship between the average PND per unit of time (neural equivalent of minute · volume ventilation, mvPND, normalized for the maximum discharge observed at 10% CO₂) and end-expiratory CO₂ at steady state (8 rats). D, Example of one RTN chemoreceptor neuron (small arrows) labeled in vivo with biotinamide [Phox2b immunoreactivity revealed with Alexa 488 (green), biotinamide with Cy-3 (red); colocalization shown in yellow]. E, Computer-assisted mapping of the sites at which biotinamide-labeled chemoreceptor neurons were found (N = 18). Black dots represent the cells that were Phox2b-ir, and open circles signify Phox2b-negative cells. Abbreviations: RPa, Raphe pallidus; 7, facial motor nucleus; py, pyramidal tract; sp5, spinal trigeminal tract. The three coronal sections represent from top to bottom bregma levels −11.0, −11.3, and −11.6. Scale bars: D, 50 μm; E, 1 mm.