Localizing Effects of Leptin on Upper Airway and Respiratory Control during Sleep

Abstract

Study objectives: Obesity hypoventilation and obstructive sleep apnea are common complications of obesity linked to defects in respiratory pump and upper airway neural control. Leptin-deficient ob/ob mice have impaired ventilatory control and inspiratory flow limitation during sleep, which are both reversed with leptin. We aimed to localize central nervous system (CNS) site(s) of leptin action on respiratory and upper airway neuroventilatory control.

Methods: We localized the effect of leptin to medulla versus hypothalamus by administering intracerbroventricular leptin (10 μg/2 μL) versus vehicle to the lateral (n = 14) versus fourth ventricle (n = 11) of ob/ob mice followed by polysomnographic recording. Analyses were stratified for effects on respiratory (nonflow-limited breaths) and upper airway (inspiratory flow limitation) functions. CNS loci were identified by (1) leptin-induced signal transducer and activator of transcription 3 (STAT3) phosphorylation and (2) projections of respiratory and upper airway motoneurons with a retrograde transsynaptic tracer (pseudorabies virus).

Results: Both routes of leptin administration increased minute ventilation during nonflow-limited breathing in sleep. Phrenic motoneurons were synaptically coupled to the nucleus of the solitary tract, which also showed STAT3 phosphorylation, but not to the hypothalamus. Inspiratory flow limitation and obstructive hypopneas were attenuated by leptin administration to the lateral but not to the fourth cerebral ventricle. Upper airway motoneurons were synaptically coupled with the dorsomedial hypothalamus, which exhibited STAT3 phosphorylation.

Conclusions: Leptin relieves upper airway obstruction in sleep apnea by activating the forebrain, possibly in the dorsomedial hypothalamus. In contrast, leptin upregulates ventilatory control through hindbrain sites of action, possibly in the nucleus of the solitary tract.

Keywords: dorsomedial hypothalamus; nucleus of the solitary tract; ob/ob mouse; obesity hypoventilation syndrome; sleep apnea syndrome.

© 2016 Associated Professional Sleep Societies, LLC.

Figures

Figure 1

The effects of intracerebroventricular administration of leptin (10 μg/2 μL in 5 mM Tris HCl) or vehicle (2 μL of 5 mM Tris HCl) on minute ventilation (VE), tidal volume (VT), and respiratory rate (RR) during nonflow-limited breathing in nonrapid eye movement (NREM) and rapid eye movement (REM) sleep. (A–C) Lateral ventricle administration. (D–F) Fourth ventricle administration. *, †, and ‡, P < 0.05, < 0.01, and < 0.001, respectively, for the effect of leptin.

Figure 2

The effects of intracerebroventricular administration of leptin (10 μg/2 μL in 5 mM Tris HCl) or vehicle (2 μL of 5 mM Tris HCl) on maximal inspiratory flow (VImax), minute ventilation (VE), tidal volume (VT), and respiratory rate (RR) during flow-limited breathing in nonrapid eye movement (NREM) and rapid eye movement REM sleep. (A–D) Lateral ventricle administration. (E–H) Fourth ventricle administration. * and †, P < 0.05 and < 0.01, respectively, for the effect of leptin.

Figure 3

A representative trace of rapid eye movement sleep in the ob/ob mouse treated with vehicle (2 μl of 5 mM Tris-HCl, pH 8.0). Right panel, Compressed recording of electroencephalogram (EEG), nuchal electromyogram (EMG), neck pulse oximetry (SpO2), respiratory flow and effort are shown. Obstructive hypopnea was defined as a discrete period of reduced airflow due to inspiratory limitation flanked by nonflow-limited breaths accompanied by increased effort and oxyhemoglobin desaturation > 3%. Left panel, The shaded area is decompressed. EXPIR denotes expiration and INSP denotes inspiration. Flow limited breaths are denoted with *, and increased effort is labeled with †. a.u., arbitrary units.

Figure 4

The effects of intracerebroventricular administration of leptin (10 μg/2 μL in 5 mM Tris HCl) or vehicle (2 μL of 5 mM Tris HCl) on the oxygen desaturation index (ODI) with oxyhemoglobin desaturations of ≥ 3%, 4%, and 5% (ODI3, ODI4, and ODI5 respectively) during total sleep time, nonrapid eye movement (NREM), and rapid eye movement (REM) sleep. (A–C) Lateral ventricle administration. (D–F) Fourth ventricle administration. *P < 0.05 for the effect of leptin.

Figure 5

Relationships between leptin signaling and respiratory neurons in the brain. The dorsomedial hypothalamus (DMH) (A) and the nucleus of the tractus solitarius (NTS) (B) show positive leptin signaling determined by dark brown staining of the neuron nuclei for phosphorylated signal transducer and activator of transcription 3 (STAT3) 40 min after leptin administration to the lateral ventricle. The Bartha strain of pseudorabies virus (PRV) was topically applied to the inferior surface of the diaphragm (C,D) or injected in the genioglossus muscle (E,F) and neurons involved in respiratory control or the upper airway function were detected in the DMH (C,E) or the medulla (D,F) by positive immunofluorescence of PRV 72 h after injection. CC, corpus callosum; XII, hypoglossal nucleus; 3V, third ventricle.