Dorsal Raphe Serotonin Neurons Mediate CO₂-Induced Arousal from Sleep

Abstract

Arousal from sleep in response to CO is a critical protective phenomenon. Dysregulation of CO-induced arousal contributes to morbidity and mortality from prevalent diseases, such as obstructive sleep apnea and sudden infant death syndrome. Despite the critical nature of this protective reflex, the precise mechanism for CO-induced arousal is unknown. Because CO is a major regulator of breathing, prevailing theories suggest that activation of respiratory chemo- and mechano-sensors is required for CO-induced arousal. However, populations of neurons that are not involved in the regulation of breathing are also chemosensitive. Among these are serotonin (5-HT) neurons in the dorsal raphe nucleus (DRN) that comprise a component of the ascending arousal system. We hypothesized that direct stimulation of these neurons with CO could cause arousal from sleep independently of enhancing breathing. Dialysis of CO-rich acidified solution into DRN, but not medullary raphe responsible for modulating breathing, caused arousal from sleep. Arousal was lost in mice with a genetic absence of 5-HT neurons, and with acute pharmacological or optogenetic inactivation of DRN 5-HT neurons. Here we demonstrate that CO can cause arousal from sleep directly, without requiring enhancement of breathing, and that chemosensitive 5-HT neurons in the DRN critically mediate this arousal. Better understanding mechanisms underlying this protective reflex may lead to interventions to reduce disease-associated morbidity and mortality. Although CO-induced arousal is critical to a number of diseases, the specific mechanism is not well understood. We previously demonstrated that serotonin (5-HT) neurons are important for CO-induced arousal, as mice without 5-HT neurons do not arouse to CO Many have interpreted this to mean that medullary 5-HT neurons that regulate breathing are important in this arousal mechanism. Here we found that direct application of CO-rich aCSF to the dorsal raphe nucleus, but not the medullary raphe, causes arousal from sleep, and that this arousal was lost with genetic ablation or acute inhibition of 5-HT neurons. We propose that 5-HT neurons in the dorsal raphe nucleus can be activated directly by CO to cause arousal independently of respiratory activation.

Effect of monoamine reuptake inhibition and α1 blockade on respiratory arrest and death following electroshock‐induced seizures in mice

“…31 Frozen brains were cryostat sectioned (30 μm; Leica), and placed into free-floating wells or mounted on slides. 31 Frozen brains were cryostat sectioned (30 μm; Leica), and placed into free-floating wells or mounted on slides.…”

“…31 Frozen brains were cryostat sectioned (30 μm; Leica), and placed into free-floating wells or mounted on slides. 31 Only mice in which the cannula was verified in the lateral ventricle were included in analysis. Mounted sections were observed with an epifluorescent microscope (Zeiss).…”

A companion to the preclinical common data elements for physiologic data in rodent epilepsy models. A report of the TASK3 Physiology Working Group of the ILAE/AES Joint Translational Task Force

“…In this commonly used method, animals are not restrained, but movement is restricted by the use of a relatively small chamber to keep the volume small with respect to the animal's size. This allows for measurements in freely behaving animals attached to tethers for EEG, electromyography (EMG), and electrocardiography (ECG) measurements . Because the data depend on measuring pressure waves, movements associated with seizures during the ictal phase might impair the breathing measurements.…”

“…Because the data depend on measuring pressure waves, movements associated with seizures during the ictal phase might impair the breathing measurements. The method is suitable for measuring breathing during the pre‐ and postictal phases and allows for successful assessment of the effects of a variety of variables such as sleep state, time of day, sex, and genetic, optogenetic, and pharmacologic manipulations in a number of seizure/epilepsy models . For more details on the procedure see Appendix and Ref.…”

“…For more details see Appendix and Refs. . Forced oscillation technique (FOT) . Measurements are typically obtained by analyzing pressure and volume signals acquired in reaction to a predefined, small amplitude, oscillatory airflow waveform (also referred to as perturbation or input signal) applied at the subject's airway opening.…”