Contribution of retrotrapezoid nucleus and carotid bodies to asphyxia‐induced arousal in rats

Abstract

Arousal in response to asphyxia is a life‐saving reflex that helps restore normal breathing and blood gas homeostasis. The carotid bodies (CB) are essential to hypoxia‐induced arousal whereas CNS serotonergic neurons and the lateral parabrachial nucleus contribute to CO2‐induced arousal. Here we wished to test whether the retrotrapezoid nucleus (RTN) is also implicated in CO2‐induced arousal. Our hypothesis was that RTN contributes to arousal elicited by hypercapnia but not hypoxia. The rationale is as follows. RTN is an important pluricellular CO2 detector that mediates the bulk of the hypercapnic ventilatory reflex (HCVR). RTN is likely absent since birth in central congenital hypoventilation syndrome, CCHS, a developmental disease in which the HCVR and asphyxia‐induced arousal are both greatly reduced. To test our hypothesis, we studied four groups of adult Sprague‐Dawley rats. In one group, we nearly completely destroyed RTN (“RTN lesion” group; n=8) with microinjections of saporin conjugated with substance‐P (2.4 ng/injection). The cognate control group (“RTN control”; n=7) received saline. The CBs were ablated in a third rat cohort (“CBx rats”; n=7). The fourth group (“CB control rats”; n=7) underwent sham surgery. Four weeks later, rats were placed in a plethysmograph chamber where breathing frequency and amplitude, EEG and EMG were recorded. Using mass flow controllers, the gas mixture perfusing the chamber was intermittently changed for exactly 1 min from normoxia (21% O2/balance N2) back to normoxia (control for flow interruption), from normoxia to hypercapnia (15% CO2/21% O2/balance N2), or from normoxia to hypoxia (10% O2, balance N2). Sleep survival curves representing the cumulative probability for SWS to persist after onset of the gas change were obtained. Two‐way ANOVA for repeated measures was used for statistical analysis of these curves. Sleep architecture was assessed by measuring the proportion of time the rats spent in SWS, REM sleep or quiet waking and the number of sleep stage transitions. Based on these measurements, sleep was unaffected by CBx or RTN lesion. However, the probability of SWS to persist 20 sec after the beginning of stimulus was significantly higher in RTN lesion rats compared to control (73.6 ± 14% vs. 54.8 ± 18 % at 20 sec from the beginning of stimulus). The corresponding figures at the 30 s time‐point were 57.8 ± 12 vs. 9.6 ± 13% and, at 40 sec, 40.1 ± 11 vs. 0 %. By contrast, RTN lesion had no detectable effect on hypoxia‐induced arousal. The arousal deficits elicited by CBx were different. CBx‐rats exposed to hypoxia had a significantly probability of SWS to persist compared to CB Control rats (89.9 ± 9 vs. 67.1 ± 9 % at 20 sec; 81.5 ± 15 vs. 56.4 ± 14 % at 30 sec, and 70.8 ± 16 vs. 39.9 ± 11 at 40 sec after the beginning of stimulus). In summary, we confirm the importance of the CBs to hypoxia‐induced arousal and demonstrate that arousal to hypercapnia is selectively reduced after RTN lesion. Neither CBx nor RTN lesions eliminated the arousal elicited by hypoxia or hypercapnia, however. Additional O2 and CO2 sensors (peripheral or central) therefore likely contribute to asphyxia‐induced arousal.Support or Funding InformationNational Institutes of Health (HL074011 and HL28785 to P.G.G.)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.