Designer Receptors Exclusively Activated by Designer Drugs Approach to Treatment of Sleep-disordered Breathing.

Rationale: Obstructive sleep apnea is recurrent upper airway obstruction caused by a loss of upper airway muscle tone during sleep. The main goal of our study was to determine if designer receptors exclusively activated by designer drugs (DREADD) could be used to activate the genioglossus muscle as a potential novel treatment strategy for sleep apnea. We have previously shown that the prototypical DREADD ligand clozapine-N-oxide increased pharyngeal diameter in mice expressing DREADD in the hypoglossal nucleus. However, the need for direct brainstem viral injections and clozapine-N-oxide toxicity diminished translational potential of this approach, and breathing during sleep was not examined.Objectives: Here, we took advantage of our model of sleep-disordered breathing in diet-induced obese mice, retrograde properties of the adeno-associated virus serotype 9 (AAV9) viral vector, and the novel DREADD ligand J60.Methods: We administered AAV9-hSyn-hM3(Gq)-mCherry or control AAV9 into the genioglossus muscle of diet-induced obese mice and examined the effect of J60 on genioglossus activity, pharyngeal patency, and breathing during sleep.Measurements and Main Results: Compared with control, J60 increased genioglossus tonic activity by greater than sixfold and tongue uptake of 2-deoxy-2-[18F]fluoro-d-glucose by 1.5-fold. J60 increased pharyngeal patency and relieved upper airway obstruction during non-REM sleep.Conclusions: We conclude that following intralingual administration of AAV9-DREADD, J60 can activate the genioglossus muscle and improve pharyngeal patency and breathing during sleep.

Hox-mediated endodermal identity patterns pharyngeal muscle formation in the chordate pharynx.

The chordate pharynx, possessing gill slits and the endostyle, is a complex of multiple tissues that are highly organized along the anterior-posterior (AP) axis. Although Hox genes show AP coordinated expression in the pharyngeal endoderm, tissue-specific roles of these factors for establishing the regional identities within this tissue have not been demonstrated. Here, we show that Hox1 is essential for the establishment of AP axial identity of the endostyle, a major structure of the pharyngeal endoderm, in the ascidian Ciona intestinalis We found that knockout of Hox1 causes posterior-to-anterior transformation of the endostyle identity, and that Hox1 represses Otx expression and anterior identity, and vice versa. Furthermore, alteration of the regional identity of the endostyle disrupts the formation of body wall muscles, suggesting that the endodermal axial identity is essential for coordinated pharyngeal development. Our results demonstrate an essential role of Hox genes in establishment of the AP regional identity in the pharyngeal endoderm and reveal crosstalk between endoderm and mesoderm during development of chordate pharynx.

Restoration of pharyngeal dilator muscle force in dystrophin-deficient (mdx) mice following co-treatment with neutralizing interleukin-6 receptor antibodies and urocortin 2.

NEW FINDINGS: What is the central question of this study? We previously reported impaired upper airway dilator muscle function in the mdx mouse model of Duchenne muscular dystrophy (DMD). Our aim was to assess the effect of blocking interleukin-6 receptor signalling and stimulating corticotrophin-releasing factor receptor 2 signalling on mdx sternohyoid muscle structure and function. What is the main finding and its importance? The interventional treatment had a positive inotropic effect on sternohyoid muscle force, restoring mechanical work and power to wild-type values, reduced myofibre central nucleation and preserved the myosin heavy chain type IIb fibre complement of mdx sternohyoid muscle. These data might have implications for development of pharmacotherapies for DMD with relevance to respiratory muscle performance. The mdx mouse model of Duchenne muscular dystrophy shows evidence of impaired pharyngeal dilator muscle function. We hypothesized that inflammatory and stress-related factors are implicated in airway dilator muscle dysfunction. Six-week-old mdx (n = 26) and wild-type (WT; n = 26) mice received either saline (0.9% w/v) or a co-administration of neutralizing interleukin-6 receptor antibodies (0.2 mg kg-1 ) and corticotrophin-releasing factor receptor 2 agonist (urocortin 2; 30 µg kg-1 ) over 2 weeks. Sternohyoid muscle isometric and isotonic contractile function was examined ex vivo. Muscle fibre centronucleation and muscle cellular infiltration, collagen content, fibre-type distribution and fibre cross-sectional area were determined by histology and immunofluorescence. Muscle chemokine content was examined by use of a multiplex assay. Sternohyoid peak specific force at 100 Hz was significantly reduced in mdx compared with WT. Drug treatment completely restored force in mdx sternohyoid to WT levels. The percentage of centrally nucleated muscle fibres was significantly increased in mdx, and this was partly ameliorated after drug treatment. The areal density of infiltrates and collagen content were significantly increased in mdx sternohyoid; both indices were unaffected by drug treatment. The abundance of myosin heavy chain type IIb fibres was significantly decreased in mdx sternohyoid; drug treatment preserved myosin heavy chain type IIb complement in mdx muscle. The chemokines macrophage inflammatory protein 2, interferon-γ-induced protein 10 and macrophage inflammatory protein 3α were significantly increased in mdx sternohyoid compared with WT. Drug treatment significantly increased chemokine expression in mdx but not WT sternohyoid. Recovery of contractile function was impressive in our study, with implications for Duchenne muscular dystrophy. The precise molecular mechanisms by which the drug treatment exerts an inotropic effect on mdx sternohyoid muscle remain to be elucidated.

Desipramine Increases Genioglossus Activity and Reduces Upper Airway Collapsibility during Non-REM Sleep in Healthy Subjects.

RATIONALE: Obstructive sleep apnea is a state-dependent disease. One of the key factors that triggers upper airway collapse is decreased pharyngeal dilator muscle activity during sleep. To date, there have not been effective methods to reverse pharyngeal hypotonia pharmacologically in sleeping humans. OBJECTIVES: We tested the hypothesis that administration of desipramine 200 mg prevents the state-related reduction in genioglossus activity that occurs during sleep and thereby decreases pharyngeal collapsibility. METHODS: We conducted a placebo-controlled, double-blind, crossover trial with 10 healthy participants. Participants received active treatment or placebo in randomized order 2 hours before sleep in the physiology laboratory. MEASUREMENTS AND MAIN RESULTS: Genioglossus activity during wakefulness and sleep, genioglossus muscle responsiveness to negative epiglottic pressure, and upper airway collapsibility during passive and active conditions were compared between on- and off-drug states. Desipramine abolished the normal reduction of genioglossus activity from wakefulness to non-REM sleep that occurred on the placebo night. Specifically, tonic (median, 96% [86-120] vs. 75% [50-92] wakefulness; P = 0.01) but not phasic genioglossus activity was higher with desipramine compared with placebo. Upper airway collapsibility was also reduced with desipramine compared with placebo (-10.0 cm H2O [-15.2 to -5.8] vs. -8.1 cm H2O [-10.4 to -6.3]; P = 0.037). CONCLUSIONS: Desipramine reduces the state-related drop in tonic genioglossus muscle activity that occurs from wakefulness to non-REM sleep and reduces airway collapsibility. These data provide a rationale for a new pharmacologic therapy for obstructive sleep apnea. Clinical trial registered with www.clinicaltrials.gov (NCT02428478).

The response to high CO2 levels requires the neuropeptide secretion component HID-1 to promote pumping inhibition.

Carbon dioxide (CO2) is a key molecule in many biological processes; however, mechanisms by which organisms sense and respond to high CO2 levels remain largely unknown. Here we report that acute CO2 exposure leads to a rapid cessation in the contraction of the pharynx muscles in Caenorhabditis elegans. To uncover the molecular mechanisms underlying this response, we performed a forward genetic screen and found that hid-1, a key component in neuropeptide signaling, regulates this inhibition in muscle contraction. Surprisingly, we found that this hid-1-mediated pathway is independent of any previously known pathways controlling CO2 avoidance and oxygen sensing. In addition, animals with mutations in unc-31 and egl-21 (neuropeptide secretion and maturation components) show impaired inhibition of muscle contraction following acute exposure to high CO2 levels, in further support of our findings. Interestingly, the observed response in the pharynx muscle requires the BAG neurons, which also mediate CO2 avoidance. This novel hid-1-mediated pathway sheds new light on the physiological effects of high CO2 levels on animals at the organism-wide level.

The ß2 -adrenoceptor agonist terbutaline recovers rat pharyngeal dilator muscle force decline during severe hypoxia.

RATIONALE: Obstructive sleep apnoea syndrome (OSAS) is a debilitating condition characterized by recurrent occlusions of the pharyngeal airway during sleep accompanied by arterial hypoxaemia. Upper airway muscle dysfunction is implicated in the pathophysiology of OSAS. Pharmacological agents that improve muscle contractile and endurance properties may have therapeutic value. AIM: We tested the hypothesis that the ß(2) -adrenoceptor agonist terbutaline improves rat sternohyoid muscle performance especially during hypoxic stress. METHODS: Isometric contractile and endurance properties were examined ex vivo in Krebs solution at 35°C. Muscles were incubated in tissue baths under hyperoxic (95% O(2) /5% CO(2)) conditions in the absence (control) or presence of the ß(2) -adrenoceptor agonist terbutaline (1 µM). In additional experiments under hypoxic (95% N(2) /5% CO(2)) conditions, the effects of terbutaline were examined in the presence of the ß-adrenoceptor antagonist propranolol (1 µM). RESULTS: Hypoxia significantly impaired sternohyoid force production. Terbutaline completely recovered hypoxic depression of force, an effect that was blocked by co-application with propranolol. CONCLUSION: The ß(2) -adrenoceptor agonist terbutaline completely recovers hypoxic depression of upper airway muscle force. ß(2) -adrenoceptor agonists warrant investigation in animal models of OSAS reporting upper airway and diaphragm muscle dysfunction.

Transient receptor potential melastatin (TRPM) channels mediate clozapine-induced phenotypes in Caenorhabditis elegans.

The molecular mechanisms of action of antipsychotic drugs (APDs) are not fully understood. Here, we characterize phenotypes of missense and knockout mutations in the Caenorhabditis elegans transient receptor potential melastatin (TRPM) channel ortholog gtl-2, a candidate APD target identified in a genome-wide RNAi (RNA interference) screen for Suppressors of Clozapine-induced Larval Arrest (scla genes). We then employ the developmental phenotypes of gtl-2(lf) mutants to validate our previous gtl-2(RNAi) result. GTL-2 acts in the excretory canal cell to regulate Mg(2+) homeostasis. Using exc (excretory canal abnormal) gene mutants, we demonstrate that excretory canal cell function is necessary for clozapine-induced developmental delay and lethality. Moreover, cell-specific promoter-driven expression studies reveal that GTL-2 function in the excretory canal cell is important for its role in the SCLA phenotype. We then investigate the mechanism by which GTL-2 function in the excretory canal cell impacts clozapine-induced phenotypes. gtl-2(lf) mutations cause hypermagnesemia, and we show that exposure of the wild-type strain to high Mg(2+) phenocopies gtl-2(lf) with respect to suppression of clozapine-induced developmental delay and lethality. Our results suggest that GTL-2 TRPM channel function in the excretory canal cell is important for clozapine's developmental effects. TRP channels are expressed in mammalian brain and are implicated in the pathogenesis of mental illnesses but have not been previously implicated in APD action.

Ultrasound-Guided Botulinum Neurotoxin Injection for Alleviating Cricopharyngeus Muscle Spasticity: A Cadaveric Feasibility Study with Nerve Ending Analysis.

Botulinum neurotoxin (BNT) injection into the cricopharyngeus muscle (CPM) under ultrasound (US) guidance is a minimally invasive technique performed to relieve cricopharyngeal dysphagia by reducing CPM spasticity. This technique is basically accessible only to both lateral sides of the CPM. This cadaveric study aimed to evaluate whether US-guided injection could effectively deliver BNT to abundant areas of gross nerve endings within the CPM. We utilized a newly modified Sihler's staining method to identify regions with abundant neural endings within the CPM while preserving the three-dimensional morphology of the muscle in 10 sides of 5 fresh cadavers. A mixture of 0.2 mL dye was injected into the 16 sides of CPM under US guidance in 8 cadavers. Nerve endings were abundant in posterolateral areas of the CPM; the injected dye was identified at the posterolateral area on 12 sides (12/16 side, 75%) without diffusion into the posterior cricoarytenoid muscle. The injection failed on four sides (two sides of the prevertebral fascia and two sides of the esophagus below the CPM). These results suggest that US-guided injection could be a feasible technique as it can deliver BNT to the most abundant nerve distribution areas within the CPM in most cases.

Phasic genioglossus and palatoglossus muscle activity during recovery from sevoflurane anesthesia: a prospective observational study in children.

BACKGROUND: Inhalational anesthetic effects on upper airway muscle activity in children are largely unknown. The authors tested the hypothesis that phasic inspiratory genioglossus and palatoglossus activity increases during recovery from sevoflurane anesthesia in a dose-dependent manner in children. METHODS: Sixteen children, aged 2.0 to 6.9 yr, scheduled for elective urological surgery were studied. Electromyogram recordings were acquired using intramuscular needle electrodes during spontaneous ventilation. After a 15-min period of equilibration, electromyogram activity was recorded over 30 s at each of three end-tidal concentrations, 1.5, 1.0, and 0.5 minimum alveolar concentration (MAC), administered in sequence. RESULTS: Phasic genioglossus activity was noted in four children at 1.5 MAC, five at 1.0 MAC, and six children at 0.5 MAC sevoflurane. Phasic palatoglossus activity was noted in 4 children at 1.5 MAC, 6 at 1.0 MAC, and 10 children at 0.5 MAC sevoflurane. Both the proportion of children exhibiting phasic activity, and the magnitude of phasic activity increased during recovery from anesthesia. For the genioglossus, decreasing the depth of sevoflurane anesthesia from 1.5 to 1.0 MAC increased phasic activity by approximately 35% and a further decrease to 0.5 MAC more than doubled activity (median [range] at 1.5 and 0.5 MAC: 2.7 µV [0 to 4.0 µV] and 8.6 µV [3.2 to 17.6], respectively; P = 0.029). A similar dose-related increase was recorded at the palatoglossus (P = 0.0002). CONCLUSIONS: Genioglossus and palatoglossus activity increases during recovery from sevoflurane anesthesia in a dose-dependent manner over the clinical range of sevoflurane concentrations in children.

Effects of pharyngeal water stimulation on swallowing behaviors in healthy humans.

The aims of the present study were to determine whether the interval between swallows and the electromyographic (EMG) burst patterns of the suprahyoid muscles is affected by peripheral inputs during swallowing. Eighteen normal adults were asked to perform repetitive voluntary swallowing as quickly as possible, and three variables of swallowing were measured and evaluated, i.e., the swallowing intervals and the time interval between the onset and peak (rising time) and between the peak and offset (falling time) of the suprahyoid EMG burst. During recording, pharyngeal fluid infusion was applied with distilled water or 0.3 M NaCl solution at a very slow infusion rate (0.2 mL/min). The former and latter were used to activate and inhibit the excitation of water-sensitive receptors in the pharynx, respectively. The swallowing interval was significantly shorter during infusion of water than during infusion of NaCl solution. The rising time was also significantly shorter during infusion of water than during infusion of NaCl solution. There was a linear positive correlation between these values and facilitatory effects: the longer either the swallowing interval or rising time with infusion of 0.3 M NaCl solution, the stronger the facilitation of swallowing by the activation of water receptors. Facilitatory effects on the swallowing interval and rising time showed a linear correlation. It is suggested that weak liquid stimulation changed sensory inputs into the swallowing center and synchronously modulated the swallowing interval and time interval between the onset and peak of the EMG burst.

Effects of genistein and estrogen on the genioglossus in rats exposed to chronic intermittent hypoxia may be HIF-1α dependent.

OBJECTIVES: Chronic intermittent hypoxia (CIH) is a frequent feature of OSAHS. The present study was designed to evaluate the effects of genistein and estrogen on genioglossus contractile and regeneration properties in CIH rats and investigate the involvement of HIF-1α. METHODS: Ovariectomized female rats were exposed to CIH for 5 weeks. Genistein and estrogen were administered by intraperitoneal injection. The genioglossus myoblasts of rat were also isolated and cultured in vitro, and the HIF-1α shRNA lentivirus was used. RESULTS: Muscle fatigue resistance and myogenic regeneration were significantly decreased after CIH but were partially reversed by estrogen and genistein treatment. The effect of estrogen was more powerful than that of genistein. Compared with control group, RT-PCR and western blotting showed higher levels of HIF-1α mRNA and protein in the CIH group, but estrogen and genistein treatment reduced the levels of HIF-1α mRNA and protein in rats exposed to CIH. In genioglossus myoblasts, the expression of HIF-1α was up-regulated under hypoxia rather than normoxia and decreased over time under both hypoxia and normoxia during myogenic differentiation. HIF-1α knockdown relieved myogenesis inhibition under hypoxia. CONCLUSION: We concluded that genistein and estrogen may inhibit the overexpression of HIF-1α induced by CIH and improve the endurance and regeneration of the genioglossus muscle.

Tempol ameliorates pharyngeal dilator muscle dysfunction in a rodent model of chronic intermittent hypoxia.

Respiratory muscle dysfunction is implicated in the pathophysiology of obstructive sleep apnea syndrome (OSAS), an oxidative stress disorder prevalent in men. Pharmacotherapy for OSAS is an attractive option, and antioxidant treatments may prove beneficial. We examined the effects of chronic intermittent hypoxia (CIH) on breathing and pharyngeal dilator muscle structure and function in male and female rats. Additionally, we tested the efficacy of antioxidant treatment in preventing (chronic administration) or reversing (acute administration) CIH-induced effects in male rats. Adult male and female Wistar rats were exposed to alternating cycles of normoxia and hypoxia (90 s each; Fi(O(2)) = 5% O(2) at nadir; Sa(O(2)) ∼ 80%) or sham treatment for 8 h/d for 9 days. Tempol (1 mM, superoxide dismutase mimetic) was administered to subgroups of sham- and CIH-treated animals. Breathing was assessed by whole-body plethysmography. Sternohyoid muscle contractile and endurance properties were examined in vitro. Muscle fiber type and cross-sectional area and the activity of key metabolic enzymes were determined. CIH decreased sternohyoid muscle force in male rats only. This was not attributable to fiber transitions or alterations in oxidative or glycolytic enzyme activity. Muscle weakness after CIH was prevented by chronic Tempol supplementation and was reversed by acute antioxidant treatment in vitro. CIH increased normoxic ventilation in male rats only. Sex differences exist in the effects of CIH on the respiratory system, which may contribute to the higher prevalence of OSAS in male subjects. Antioxidant treatment may be beneficial as an adjunct OSAS therapy.

REM sleep-like episodes of motoneuronal depression and respiratory rate increase are triggered by pontine carbachol microinjections in in situ perfused rat brainstem preparation.

Hypoglossal nerve activity (HNA) controls the position and movements of the tongue. In persons with compromised upper airway anatomy, sleep-related hypotonia of the tongue and other pharyngeal muscles causes increased upper airway resistance, or total upper airway obstructions, thus disrupting both sleep and breathing. Hypoglossal nerve activity reaches its nadir, and obstructive episodes are longest and most severe, during rapid eye movement stage of sleep (REMS). Microinjections of a cholinergic agonist, carbachol, into the pons have been used in vivo to investigate the mechanisms of respiratory control during REMS. Here, we recorded inspiratory-modulated phrenic nerve activity and HNA and microinjected carbachol (25-50 nl, 10 mm) into the pons in an in situ perfused working heart-brainstem rat preparation (WHBP), an ex vivo model previously validated for studies of the chemical and reflex control of breathing. Carbachol microinjections were made into 40 sites in 33 juvenile rat preparations and, at 24 sites, they triggered depression of HNA with increased respiratory rate and little change of phrenic nerve activity, a pattern akin to that during natural REMS in vivo. The REMS-like episodes started 151 ± 73 s (SD) following microinjections, lasted 20.3 ± 4.5 min, were elicited most effectively from the dorsal part of the rostral nucleus pontis oralis, and were prevented by perfusion of the preparation with atropine. The WHBP offers a novel model with which to investigate cellular and neurochemical mechanisms of REMS-related upper airway hypotonia in situ without anaesthesia and with full control over the cellular environment.

Botulinum toxin improves dysphagia associated with multiple sclerosis.

OBJECTIVE: To evaluate the efficacy of botulinum neurotoxin type A (BoNT/A) for severe oro-pharyngeal dysphagia associated with multiple sclerosis (MS). PATIENTS AND METHODS: BoNT/A was injected percutaneously into the hyperactive cricopharyngeal muscle of 14 dysphagic MS patients under electromyographic control. Patients were evaluated by videofluoroscopic and electromyographic examinations and by the Penetration/Aspiration Scale (PAS), at week 1, 4, 12, 16, 18, and 24 after BoNT/A injection. RESULTS: All patients showed a significant improvement in all the swallowing outcome measures. CONCLUSION: No specific treatment for oro-pharyngeal dysphagia related to MS has been described to date. Our preliminary findings suggest a potential benefit from BoNT/A treatment in MS patients with dysphagia associated with upper esophageal sphincter hyperactivity.

Botulinum toxin type A: an effective treatment to restore phonation in laryngectomized patients unable to voice.

We evaluated the efficacy of Botulinum toxin type A (BTXA) as an alternative to surgical intervention to facilitate phonation in 34 laryngectomized patients (31 males and 3 women) who were unable to produce tracheoesophageal voice because of spasm of the middle and inferior pharyngeal constrictor muscles (PCM). EMG was recorded to confirm activity in these muscles during attempted vocalization. Parapharyngeal nerve block (Carbocaine 2%, 5 cc) was used to demonstrate short-term fluent voice after relaxation of the pharyngeal constrictor muscle. At a later occasion, 100 U of Botox (Allergan) in ten patients and 50 U in two patients were injected unilaterally at one location in the PCM percutaneously under EMG guidance. All patients then underwent a voice therapy program. In 11 out of 12 patients an improvement of phonation was evident after 24-48 h and it was long lasting. This result was also seen in a patient previously myotomized without improvement. Only one patient needed to be reinjected every 3 months. At a follow-up after 3 months the EMG recorded in four patients showed a low-amplitude or complete absence of activity in the treated muscle. No side effects developed. BTX therapy, especially when associated with the speech therapy, is efficacious in restoring voice to laryngectomees who are unable to voice because of spasm of the PCMs. Our results confirm previous reports. This method is our approach of choice in managing PCM spasm because it is non-invasive, not painful, has few or no side effects, and is frequently long-lasting.

Superoxide scavengers improve rat pharyngeal dilator muscle performance.

Obstructive sleep apnea is a common disorder associated with upper airway muscle dysfunction. Agents that improve respiratory muscle performance may be useful as an adjunct therapy. The aim of this study was to examine the effects of antioxidants on rat pharyngeal dilator muscle performance. Adult male Wistar rats were killed humanely and isometric contractile properties of isolated sternohyoid muscle strips were examined in physiological salt solution at 35 degrees C in vitro. Muscle strips were incubated in tissue baths under hyperoxic (95%O(2)/5%CO(2)) or hypoxic (95%N(2)/5%CO(2)) conditions in the absence (control) or presence of the antioxidants: N-acetylcysteine (10 mM), Tiron (10 mM), or Tempol (10 mM). Force-frequency relationship was determined in response to supramaximal stimulation (10-100 Hz in increments of 10-20 Hz, train duration: 300 ms). Isometric force was also recorded during repetitive muscle stimulation (40 Hz, 300 ms every 2 s for 2 min). Under hyperoxic conditions, Tiron and Tempol, but not N-acetylcysteine, significantly increased sternohyoid muscle force and caused a left-shift in the force-frequency relationship. In addition, Tempol had a significant positive inotropic effect over the initial 90 seconds of repeated muscle activation. Hypoxia caused a significant decrease in sternohyoid muscle force. Under hypoxic conditions, Tempol-incubated muscles generated significantly higher forces compared with control muscles and showed improved performance in the early phase of the fatigue trial. This study illustrates that superoxide scavengers increase upper airway muscle force and that this effect persists under hypoxic conditions. We conclude that antioxidant treatment may be beneficial as a therapy in obstructive sleep apnea.

An electrophysiological approach to the diagnosis of neurogenic dysphagia: implications for botulinum toxin treatment.

OBJECTIVES: Botulinum toxin (BTX) injection into the cricopharyngeal (CP) muscle has been proposed for the treatment of neurogenic dysphagia due to CP hyperactivity. The aim was to determine whether an electrophysiological method exploring oropharyngeal swallowing could guide treatment and discriminate responders from non-responders, based on the association of CP dysfunction with other electrophysiological abnormalities of swallowing. METHODS: Patients with different neurological disorders were examined: Parkinson disease, progressive supranuclear palsy, multiple system atrophy-Parkinson variant, multiple system atrophy cerebellar variant, stroke, multiple sclerosis and ataxia telangiectasia. All patients presented with clinical dysphagia, and with complete absence of CP muscle inhibition during the hypopharyngeal phase of swallowing. Each patient underwent clinical and electrophysiological investigations before and after treatment with BTX into the CP muscle of one side (15 units of Botox). Clinical and electrophysiological procedures were performed in a blind manner by two different investigators. The following electrophysiological measures were analysed: (1) duration of EMG activity of suprahyoid/submental muscles (SHEMG-D); (2) duration of laryngopharyngeal mechanogram (LPM-D); (3) duration of the inhibition of the CP muscle EMG activity (CPEMG-ID); and (4) interval between onset of EMG activity of suprahyoid/submental muscles and onset of laryngopharyngeal mechanogram (I-SHEMG-LPM). RESULTS: Two months after treatment, 50% of patients showed a significant improvement. Patients with prolonged or reduced SHEMG-D values and prolonged I-SHEMG-LPM values did not respond to BTX. Therefore, values for which BTX had no effect (warning values) were identified. CONCLUSIONS: This electrophysiological method can recognise swallowing abnormalities which may affect the outcome of the therapeutic approach to dysphagia with BTX treatment.

Neostigmine/glycopyrrolate administered after recovery from neuromuscular block increases upper airway collapsibility by decreasing genioglossus muscle activity in response to negative pharyngeal pressure.

BACKGROUND: Reversal of residual neuromuscular blockade by acetylcholinesterase inhibitors (e.g., neostigmine) improves respiratory function. However, neostigmine may also impair muscle strength. We hypothesized that neostigmine administered after recovery of the train-of-four (TOF) ratio impairs upper airway integrity and genioglossus muscle function. METHODS: We measured, in 10 healthy male volunteers, epiglottic and nasal mask pressures, genioglossus electromyogram, air flow, respiratory timing, and changes in lung volume before, during (TOF ratio: 0.5), and after recovery of the TOF ratio to unity, and after administration of neostigmine 0.03 mg/kg IV (with glycopyrrolate 0.0075 mg/kg). Upper airway critical closing pressure (Pcrit) was calculated from flow-limited breaths during random pharyngeal negative pressure challenges. RESULTS: Pcrit increased significantly after administration of neostigmine/glycopyrrolate compared with both TOF recovery (mean ± SD, by 27 ± 21%; P = 0.02) and baseline (by 38 ± 17%; P = 0.002). In parallel, phasic genioglossus activity evoked by negative pharyngeal pressure decreased (by 37 ± 29%, P = 0.005) compared with recovery, almost to a level observed at a TOF ratio of 0.5. Lung volume, respiratory timing, tidal volume, and minute ventilation remained unchanged after neostigmine/glycopyrrolate injection. CONCLUSION: Neostigmine/glycopyrrolate, when administered after recovery from neuromuscular block, increases upper airway collapsibility and impairs genioglossus muscle activation in response to negative pharyngeal pressure. Reversal with acetylcholinesterase inhibitors may be undesirable in the absence of neuromuscular blockade.

Effect of Venlafaxine on Apnea-Hypopnea Index in Patients With Sleep Apnea: A Randomized, Double-Blind Crossover Study.

BACKGROUND: One of the key mechanisms underlying OSA is reduced pharyngeal muscle tone during sleep. Data suggest that pharmacologic augmentation of central serotonergic/adrenergic tone increases pharyngeal muscle tone. RESEARCH QUESTION: We hypothesized that venlafaxine, a serotonin-norepinephrine reuptake inhibitor, would improve OSA severity. STUDY DESIGN AND METHODS: In this mechanistic, randomized, double-blind, placebo-controlled crossover trial, 20 patients with OSA underwent two overnight polysomnograms ≥ 4 days apart, receiving either 50 mg of immediate-release venlafaxine or placebo before bedtime. Primary outcomes were the apnea-hypopnea index (AHI) and peripheral oxygen saturation (Spo2) nadir, and secondary outcomes included sleep parameters and pathophysiologic traits with a view toward understanding the impact of venlafaxine on mechanisms underlying OSA. RESULTS: Overall, there was no significant difference between venlafaxine and placebo regarding AHI (mean reduction, -5.6 events/h [95% CI, -12.0 to 0.9]; P = .09) or Spo2 nadir (median increase, +1.0% [-0.5 to 5]; P = .11). Venlafaxine reduced total sleep time, sleep efficiency, and rapid eye movement (REM) sleep, while increasing non-REM stage 1 sleep (Pall < .05). On the basis of exploratory post hoc analyses venlafaxine decreased ("improved") the ventilatory response to arousal (-30%; P = .049) and lowered ("worsened") the predicted arousal threshold (-13%; [P = .02]; ie, more arousable), with no effects on other pathophysiologic traits (Pall ≥ .3). Post hoc analyses further suggested effect modification by arousal threshold (P = .002): AHI improved by 19% in patients with a high arousal threshold (-10.9 events/h [-3.9 to -17.9]) but tended to increase in patients with a low arousal threshold (+7 events/h [-2.0 to 16]). Other predictors of response were elevated AHI and less collapsible upper airway anatomy at baseline (|r| > 0.5, P ≤ .02). INTERPRETATION: In unselected patients, venlafaxine simultaneously worsened and improved various pathophysiologic traits, resulting in a zero net effect. Careful patient selection based on pathophysiologic traits, or combination therapy with drugs countering its alerting effects, may produce a more robust response. TRIAL REGISTRY: ClinicalTrials.gov; No.: NCT02714400; URL: www.clinicaltrials.gov.

Characterization of the effects of methylmercury on Caenorhabditis elegans.

The rising prevalence of methylmercury (MeHg) in seafood and in the global environment provides an impetus for delineating the mechanism of the toxicity of MeHg. Deleterious effects of MeHg have been widely observed in humans and in other mammals, the most striking of which occur in the nervous system. Here we test the model organism, Caenorhabditis elegans (C. elegans), for MeHg toxicity. The simple, well-defined anatomy of the C. elegans nervous system and its ready visualization with green fluorescent protein (GFP) markers facilitated our study of the effects of methylmercuric chloride (MeHgCl) on neural development. Although MeHgCl was lethal to C. elegans, induced a developmental delay, and decreased pharyngeal pumping, other traits including lifespan, brood size, swimming rate, and nervous system morphology were not obviously perturbed in animals that survived MeHgCl exposure. Despite the limited effects of MeHgCl on C. elegans development and behavior, intracellular mercury (Hg) concentrations (