Targeted Retrograde Chemogenetic Approach to Treat Sleep Apnea

Abstract

IntroductionObstructive Sleep Apnea (OSA) is a prevalent condition and a major cause of morbidity and mortality. The tongue and specifically genioglossus (GG) muscle have been implicated in the pathogenesis of upper airway (UA) obstruction during sleep. Nasal positive airway pressure can treat OSA but poor adherence severely limits its effectiveness. There is an urgent need for pharmacotherapy that reverses neuromuscular defects in upper airway function. We have previously shown that diet induced obese C57BL/6J (DIO) mice have OSA, similar to humans. We have also recently demonstrated that Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) deployed by intracerebral injection can be expressed in hypoglossal motor neurons and that activation of these DREADDs by specific ligand increased GG activity and improved pharyngeal patency. The goal of our novel approach is to avoid intracranial injection and improve specificity of DREADD delivery to hypoglossal motor neurons innervating genioglossus muscle. We hypothesize that tongue injecting retrograde adeno‐associated virus type 9 (AAV9) carrying excitatory DREADDs will (1) deploy DREADDs in hypoglossal motor neurons; (2) improve UA patency and (3) and treat sleep apnea in DIO mice.MethodsDREADDs (AAV9‐hSyn‐hM3(Gq)‐mCherry n=10) or Control AAV9‐hSyn‐GFP (n=5) were delivered bilaterally by tongue injection in DIO mice. Six weeks after gene delivery, mice were injected with specific ligand (J60) and saline intraperitoneally (IP) one week apart in a random order: we performed GG electromyography (EMGGG), UA dynamic MRI, positron emission tomography (PET) scan and sleep studies.ResultsDREADDs or control virus were expressed in the tongue and hypoglossal nucleus in all mice. In DREADD‐transfected mice, administration of J60 resulted in (1) 7.05 fold increase in EMGGG activity (p<0.001); (2) pharyngeal dilation at the rim of the soft palate (0.46 ±0.03mm2 to 1.16±0.05mm2, p<0.05) by MRI; (3) increased tongue metabolic activity (2.21± 0.28 to 4.14 ± 0.31 g/ml, p<0.001) by PET scan; (4) increased minute ventilation (22.53 ± 10.39 to 60.18 ± 33.28 mL/min) and maximal inspiratory flow (1.58 ± 0.58 to 4.11 ± 1.98 mL/sec) during sleep, treating sleep apnea. No changes were seen following j60 injection in mice injected with control virus or saline injection.ConclusionTongue injection of retrograde AAV9 carrying DREADDs successfully deployed DREADDs in hypoglossal motor neurons. Activation of these DREADDs by specific ligand (J60) treated OSA.Support or Funding InformationRO1HL138932, AHA 19CDA34660245, ATS Foundation Unrestricted Grant(A) Whole body PET scan in mouse treated with retrograde excitatory DREADD after Saline (left) or J60 (right). Note the increase in tongue metabolic activity marked by the intense uptake after J60 (right). (B) Plotted Standardized Uptake Values (SUVs) in these animals. *, p < 0.001.Figure 1(A) Representative genioglossal electromyography (EMGGG), recorded at baseline (left) and after J60 administration (right). Note the increase in both phasic and tonic EMG activity. (B) Plotted EMG responses to J60 or saline in the same DREADD treated animals. *, p < 0.001.Figure 2