Chemosensitive Phox2b-expressing neurons are crucial for hypercapnic ventilatory response in the nucleus tractus solitarius

The nucleus tractus solitarius (NTS) neurons have been considered to function as central respiratory chemoreceptors. However, the common molecular marker defined for these neurons remains unknown. The present study investigated whether paired-like homeobox 2b (Phox2b)-expressing NTS neurons are recruited in hypercapnic ventilatory response (HCVR) and whether these neurons exhibit intrinsic chemosensitivity. HCVR was assessed using whole body plethysmography and neuronal chemosensitivity was examined by patch clamp recordings in brainstem slices or dissociated neurons from Phox2b-EGFP transgenic mice. Injection of the neurotoxin substance P-saporin (SSP-SAP) into NTS destroyed Phox2b-expressing neurons. Minute ventilation and tidal volume were both reduced by 13% during exposure to 8% CO2 in inspired air when similar to 13% of the Phox2b-expressing neurons were eliminated. However, a loss of similar to 18% of these neurons was associated with considerable decreases in minute ventilation by >= 18% and in tidal volume by >= 22% when challenged by >= 4% CO2. In both cases, breathing frequency was unaffected. Most CO2-activated neurons were immunoreactive to Phox2b. In brainstem slices, similar to 43% of Phox2b-expressing neurons from Phox2b-EGFP mice displayed a sustained or transient increase in firing rate during physiological acidification (pH7.0 or 8% CO2). Such a response was also present in dissociated neurons in favour of an intrinsic property. In voltage clamp recordings, a background K+ channel-like current was found in a subgroup of Phox2b-expressing neurons. Thus, the respiratory deficits caused by injection of SSP-SAP into the NTS are attributable to proportional lesions of CO2/H+-sensitive Phox2b-expressing neurons.