Neuromedin B-expressing neurons in the retrotrapezoid nucleus regulate respiratory homeostasis and promote stable breathing in adult mice.

Respiratory chemoreceptor activity encoding arterial PCO 2 and PO 2 is a critical determinant of ventilation. Currently, the relative importance of several putative chemoreceptor mechanisms for maintaining eupneic breathing and respiratory homeostasis is debated. Transcriptomic and anatomical evidence suggest that bombesin-related peptide Neuromedin-B ( Nmb ) expression identifies chemoreceptor neurons in the retrotrapezoid nucleus (RTN) that mediate the hypercapnic ventilatory response, but functional support is missing. In this study, we generated a transgenic Nmb -Cre mouse and used Cre-dependent cell ablation and optogenetics to test the hypothesis that RTN Nmb neurons are necessary for the CO 2 -depedent drive to breathe in adult male and female mice. Selective ablation of ∼95% of RTN Nmb neurons causes compensated respiratory acidosis due to alveolar hypoventilation, as well as profound breathing instability and respiratory-related sleep disruption. Following RTN Nmb lesion, mice were hypoxemic at rest and were prone to severe apneas during hyperoxia, suggesting that oxygen-sensitive mechanisms, presumably the peripheral chemoreceptors, compensate for the loss of RTN Nmb neurons. Interestingly, ventilation following RTN Nmb -lesion was unresponsive to hypercapnia, but behavioral responses to CO 2 (freezing and avoidance) and the hypoxia ventilatory response were preserved. Neuroanatomical mapping shows that RTN Nmb neurons are highly collateralized and innervate the respiratory-related centers in the pons and medulla with a strong ipsilateral preference. Together, this evidence suggests RTN Nmb neurons are dedicated to the respiratory effects of arterial PCO 2 /pH and maintain respiratory homeostasis in intact conditions and suggest that malfunction of these neurons could underlie the etiology of certain forms of sleep-disordered breathing in humans. Significance Statement: Respiratory chemoreceptors stimulate neural respiratory motor output to regulate arterial PCO 2 and PO 2 , thereby maintaining optimal gas exchange. Neurons in the retrotrapezoid nucleus (RTN) that express the bombesin-related peptide Neuromedin-B are proposed to be important in this process, but functional evidence has not been established. Here, we developed a transgenic mouse model and demonstrated that RTN neurons are fundamental for respiratory homeostasis and mediate the stimulatory effects of CO 2 on breathing. Our functional and anatomical data indicate that Nmb -expressing RTN neurons are an integral component of the neural mechanisms that mediate CO 2 -dependent drive to breathe and maintain alveolar ventilation. This work highlights the importance of the interdependent and dynamic integration of CO 2 - and O 2 -sensing mechanisms in respiratory homeostasis of mammals.