Current diagnostic methods for OSA and CSA are costly, unreliable, and the therapeutic options are limited, with varying efficacy across patients. As the prevalence of sleep apnea and cardiovascular disease continue to rise, identifying innovative therapies through advances in biomedicine and personalized medicine has become increasingly critical. This review discusses the mechanistic links between sleep apnea and subsequent cardiovascular outcomes from a technical perspective, focusing on innovations currently applied to the diagnosis and treatment of sleep apnea, and opportunities for further advancement in the field.

BackgroundCurrent therapeutic strategies for acute myocardial infarction rely on reperfusion and pharmacological management, which are typically administered hours after an event. Activation of the cardiac cholinergic efferents superior to the heart soon after a coronary occlusion has shown promise as a potential therapy to reduce arrhythmias and improve ventricular function. We tested whether selective activation of cholinergic neurons within the intrinsic cardiac ganglia (ICG) would also reduce arrhythmias and improve oxygenation of ischemic border zone tissue after an acute coronary occlusion.MethodsDesigner Receptors Exclusively Activated by Designer Drugs (DREADDs) were selectively expressed in cholinergic neurons of the ICG via pericardial sac injections of an HM3Dq DREADDs virus in transgenic rats that expressed Cre recombinase in cholinergic neurons. Cholinergic ICG neurons were activated using the synthetic DREADDs ligand clozapine-N-oxide (CNO). Heart rate reductions after intraperitoneal injection of CNO confirmed downstream effect of DREADDs-mediated cholinergic ICG activation in ECG telemetry studies. The effect of cholinergic ICG activation on PR interval, arrhythmia burden, ischemic border zone tissue oxygenation and epicardial NADH fluorescence 20 min after ligation of the left anterior descending coronary artery (LAD) was then studied in excised perfused hearts of DREADDS-expressing rats and rats that did not receive the HM3Dq DREADDs virus.ResultsLAD ligation resulted in a well defined ischemic zone that encompassed a large portion of the left ventricle, where pO2 in the center of the ischemic zone typically dropped to 0 mmHg within 10 s. Subsequent DREADDs-mediated cholinergic ICG activation prolonged the PR interval from 39.13 ± 6.17 ms to 42.46 ± 6.87 ms and lowered the incidence of arrhythmia from 0.9398 ± 0.5063 min−1 to .5727 ± 0.3103 min−1. DREADDs-mediated cholinergic ICG activation also increased ischemic border zone pO2 from 42.13 ± 49.82 mmHg to 82.25 ± 66.87 mmHg and NADH fluorescence trended lower in the ischemic zone, indicating increased mitochondrial oxidation. These effects were blocked when the muscarinic antagonist atropine was administered before CNO.ConclusionResults indicate that selective stimulation of cholinergic ICG neurons could improve local delivery of oxygen to the ischemic border zone soon after a coronary occlusion and reduce arrhythmia burden through a muscarinic-dependent mechanism, supporting further studies of the intrinsic cardiac cholinergic network as a therapeutic target for early intervention before reperfusion therapy to activate cardioprotective pathways.

Rationale. There is no effective pharmacotherapy for obesity hypoventilation syndrome (OHS). Intranasal leptin augments the hypercapnic ventilatory response (HCVR), attenuates upper airway obstruction, and increases ventilation during sleep in diet-induced obese (DIO) mice. Respiratory effects of leptin can be attenuated by serotonergic antagonists. Objectives. To establish if serotonergic innervation of the hypoglossal motoneurons (XII MN) mediates effects of leptin on OHS. Methods. We examined effects of intranasal leptin on the HCVR, sleep architecture, arousal latency, flow limited (obstructed) and non-flow limited breathing, genioglossus muscle (GG) activity and metabolic rate across sleep/wake states in the presence and absence of serotonergic neurons innervating XII MN in DIO Sert-flp mice expressing FlpO recombinase in the serotonergic neurons. These mice were transfected into the XII MN with retrograde adeno-associated virus carrying either FlpO-dependent caspase or control yellow fluorescent protein (YFP). Measurements and Main Results. Control YFP virus was densely localized to the serotonergic neurons of the medullary raphe (MR), but not the dorsal raphe (DR), and these neurons were ablated by caspase. Leptin enhanced the HCVR, increased arousal latency in males, but not in females, and these effects were abolished by caspase. Neither leptin nor caspase affected sleep architecture or metabolic rate. Leptin increased GG activity awake and during NREM sleep, attenuated pharyngeal obstruction and increased minute ventilation in NREM and REM sleep. All effects of leptin were abolished by the FlpO-dependent caspase. Conclusions: Leptin treats OHS by stimulating MR serotonergic neurons, which project to XII MN and stimulate pharyngeal muscles during sleep.

Background:The primary cause of death associated with opioids is opioid-induced respiratory depression (OIRD). Naloxone is used to reverse OIRD, but this drug is a competitive antagonist of μ-opioid receptor (MOR) and reverses analgesia, which limits its therapeutic use. Alternative non-opioid receptor antagonist-based approaches to OIRD treatment and prevention are needed. The aim of this study was to evaluate if setmelanotide (SET) is capable of reversing OIRD in a mouse model. Methods:C57BL/6J male and female mice and Sprague-Dawley rats were given IP morphine or fentanyl and then treated 15 min later with either SET or vehicle VEH (IP) in a random order. Breathing was recorded by barometric plethysmography, and pain sensitivity was measured by the tail-flick test. Results:In mice with OIRD, SET induced a 3-fold reduction of the apnea index, and decreased apnea duration as compared to the VEH treatment. SET increased respiratory rate and did not affect opioid-induced analgesia. Photostimulation of MC4R+ ChR2-expressing fibers in the parafacial region of MC4R-Cre mice elicited short-latency excitatory postsynaptic current in rostral ventral respiratory group (rVRG) pre-motoneurons projecting to the phrenic nucleus in the C3-C4 ventral horns of the spinal cord. Fentanyl inhibited the activity of rVRG neurons and SET reversed this effect. Conclusions:SET effectively treated OIRD by increasing respiratory rate and inducing a significant decrease in the number of apneas without decreasing analgesia.

Significance:Cardiac panoramic optical mapping is a powerful approach for studying action potential dispersion and mapping arrhythmia triggers and propagation pathways over the entire surface of the heart. However, tissue type (muscle, connective tissue, and infarct scar) is also important for interpreting mapping data and is difficult to identify using optical mapping data alone. Aim:Panoramically map transmembrane potential and tissue type from the surface of infarcted hearts for correlative analysis of cardiac structure and function. Approach:We developed a multimodal panoramic imaging system to map epicardial tissue type (determined by collagen content) using a line-scan hyperspectral camera and a precision stage to translate and rotate the heart while illuminating the epicardial surface with UV light. Transmembrane potential was subsequently optically mapped by imaging a potentiometric probe with four high speed CMOS cameras position around the heart. The epicardial surface was reconstructed for each heart using images acquired every 3.6 deg of rotation, onto which hyperspectral and optical mapping data were texture mapped. All cameras were registered to one coordinate frame using a calibration procedure. Results:This system combines, for the first time, high-resolution hyperspectral imaging with optical mapping for quantitative correlative tissue structure-function analyses. It was used to study excitation wave propagation and action potentials across the surface of perfused rat hearts having a four-week-old infarct. The spectral band of collagen fluorescence (400 to 520 nm) revealed infarcted and border zone tissue. PVCs and reentrant activity were observed in 3 of 4 hearts at S1-S2 pacing intervals between 80 and 65 msec (S1 = 150 msec). PVCs originated near the infarct border and propagated around the infarct. Using the integral of spectral intensity from 400 to 435 nm, a k-means clustering algorithm classified each mapped site as either healthy, border zone, or infarcted tissue. Average action potential duration within those tissue types was longest for infarcted tissue, shorter for border zone tissue, and shortest for healthy tissue, a preliminary result that is consistent with the effect of an infarct on ventricular electrophysiology. Conclusions:This work demonstrates that panoramic hyperspectral mapping of tissue type and transmembrane potential is a powerful approach that enables functional mapping data to be analyzed within the context of local tissue type (healthy, infarct, and border) in living hearts.

Osteoarthritis (OA), a degenerative joint disease, is associated with increased systemic inflammation, chronic pain, and cardiovascular dysfunction. Epidemiological evidence establishes that OA increases the risk of cardiovascular disease (CVD) threefold, yet the causal role of OA's contributions remains underexamined. We assessed cardiac function longitudinally following destabilization of the medial meniscus (DMM) surgery to induce osteoarthritis in mice. DMM-mice exhibited significant, sexually dimorphic alterations in echocardiographic parameters. Female DMM mice developed impaired relaxation with altered E/A ratios, increased E/e' ratios, and prolonged intraventricular relaxation time with no change in ejection fraction, while male DMM mice showed progressive systolic dysfunction with decreasing ejection fraction, increased E/e' ratio, and prolonged intraventricular contraction time. Transcriptomic profiles and biochemical analyses demonstrated divergent cellular responses involving fibrosis and oxidative stress in female mice, whereas autophagic and apoptotic responses were observed in male mice. Using a tumor necrosis factor 2 (TNFR2) agonist shown to reduce systemic inflammation, we investigated its potential therapeutic role in the context of OA-induced cardiovascular dysfunction. TNFR2 agonism proved to be effective both prophylactically and therapeutically for female diastolic dysfunction. While prophylactic and therapeutic administration delayed male systolic dysfunction, the efficacy declined over time. Our findings demonstrate evidence of a novel sexually dimorphic model of OA-induced CVD that recapitulates the sexually dimorphic pattern of patient phenotypes and a promising new therapeutic approach to CVD.

Abstract Introduction Obstructive sleep apnea (OSA) affects many people throughout the world and is a leading cause of cardiovascular morbidity and mortality. One of the major causes of OSA is loss of genioglossus (GG) muscle tone during sleep. Emergent pharmacotherapy has limitations and new drugs with better potential to treat OSA are required. We have previously shown that diet-induced obese (DIO) mice develop OSA and melanocortin 4 receptor (MC4R) agonist setmelanotide (SET) could be used for OSA treatment. We have also shown MC4R agonists act in the retrotrapezoid nucleus (RTN) to augment hypercapnic ventilatory response (HCVR). We hypothesized that SET treats OSA by increasing GG muscle tone and that effects of setmelanotide on sleep and breathing can be localized to the MC4R on the RTN neurons. Methods We transfected DIO MC4R-Cre mice with adeno-associated virus (AAV) harboring excitatory designer receptor exclusive activated by designer drugs (DREADD) and DIO MC4R-flox mice with AAV-Cre-recombinase (icre) into the RTN. Transgenic and C57BL/6J mice were headmounted with EEG and GG and nuchal EMG electrodes. Sleep and HCVR studies (8% CO2 challenge) were performed in the plethysmography chamber. Results SET (1 mg/kg, i.p) significantly increased GG muscle activity during NREM sleep (SET: 0.86 ± 0.08 vs. Veh: 0.48 ± 0.04 a.u.; p < 0.005), REM sleep (SET: 0.38 ± 0.05 vs. Veh: 0.27 ± 0.04 a.u.; p < 0.05) and during HCVR (SET: 2.04 ± 0.25 vs. Veh: 1.25 ± 0.11 a.u.; p < 0.05). Experiments in MC4R-Cre mice showed that RTN-MC4R neurons activation by DREADD ligands increased sleep efficiency (Veh: 49.69 ± 0.02% vs. J60: 58.07 ± 0.03% of the light phase). Experiments in MC4R-flox mice showed that MC4R knockdown in RTN abolished effects of SET on HCVR (control, SET: 0.29 ± 0.02 vs. Veh: 0.25 ± 0.03; p < 0.05; MC4R knockdown, SET: 0.17 ± 0.03 vs. Veh: 0.19 ± 0.04; p > 0.05). Conclusion Our study suggests SET targets RTN-MC4R neurons to augment HCVR and treats OSA by improving GG muscle activity. Furthermore, RTN-MC4R neurons activation can increase total sleep time. Support (if any) R01 HL174409

Thirty-nine highly acknowledged experts in the areas of the physiology and the evolution of the vagus nerve and of vertebrate social behavior (many whose works have been cited in the polyvagal theory [PVT] literature as supporting the theory) were invited by the first author to participate as co-authors of this article. They were asked to evaluate the PVT and comment upon an overview of the theory written by its author (Porges, 2025a). All those invited, save one, accepted and co-authored the paper. The dissenting scholar was "unfamiliar with the PVT." This article specifically appraises--based upon the current state of knowledge of autonomic function and vertebrate evolution--several major elements of the PVT, as described in Porges (2025a) and elsewhere. These include: 1) the validity of PVT assumptions that respiratory sinus arrhythmia is a direct measure of the extent of central vagal drive to the heart; 2) PVT characterizations regarding the neuroanatomy and functions of two major brainstem vagal nuclei, the ventrally situated Nucleus Ambiguus and the Dorsal Motor Nucleus of the vagus nerve; 3) PVT assertions regarding the evolution of the vagus nerve; 4) PVT claims about the specificity of mammalian social behavior in relation to nonmammalian vertebrates, and 5) PVT interpretations of earlier seminal physiological literature. All co-authors agree that major tenets of the PVT are not supported by past or current knowledge and, in several instances, are inconsistent with the broader evidence base. Since the topics addressed constitute fundamental premises of the PVT, we conclude that the PVT is untenable, because it is not defensible based on existing neurophysiological and evolutionary evidence. The psychological elements composing the superstructure of the PVT are primarily derived from earlier psychological literature and are neither clarified nor strengthened by PVT constructs that lack evidence. This article does not intend to address alternative explanations about relations between vagal function and psychological processes, although such explanations do exist.

Opioid addiction and misuse are a serious national crisis that affects public health, as well as social and economic welfare. Mortality due to opioid misuse is further exasperated by the combination of opioids with non-opioid respiratory depressants such as xylazine that are resistant to mu opioid receptor antagonists such as naloxone. This study tested the hypothesis that oxytocin can mitigate the severe opioid induced respiratory depression (OIRD) and mortality induced by high doses of fentanyl or the combination of fentanyl with xylazine. Our results show OXT can improve survival and respiratory function in both male and female rats with opioid induced respiratory depression caused by fentanyl, as well as a combination of fentanyl and xylazine. The improvement in respiratory function by OXT post fentanyl-xylazine was significantly greater than the recovery using only naloxone. Chemogenetic activation of OXT receptor positive neurons in the ventral respiratory group (VRG) provided similar benefits to that of OXT administration in reversing OIRD. These results indicate OXT is a promising therapeutic target for reversing OIRD and the respiratory depression that occurs with the combination of opioids and xylazine, a situation where naloxone is only partially effective. Additional translational benefits of OXT include it can be repurposed as it is already a FDA approved drug for other uses, has a high safety profile, and is unlikely to induce the withdrawal or reversal of analgesia that occurs with naloxone.

Chronic low-grade inflammation is increasingly recognized as a key driver of heart failure (HF) progression; however, the direct contribution of systemic inflammatory disorders such as osteoarthritis (OA) remains unclear. Here, we establish a murine model of OA-induced HF using destabilization of the medial meniscus (DMM) to induce systemic inflammation and sex-specific cardiac remodeling. Longitudinal echocardiography revealed that females develop diastolic dysfunction with preserved ejection fraction, resembling HFpEF, whereas males exhibit progressive systolic impairment, consistent with a transitional HFmrEF-to-HFrEF phenotype. Morphometric and histological analyses confirmed concentric hypertrophy in females and eccentric remodeling in males. Transcriptomic profiling identified distinct molecular programs—females upregulated extracellular matrix, cytoskeletal, and calcium-handling genes, while males showed enrichment of inflammatory and immune signaling pathways. Immunoblot analyses further validated these sex-specific molecular signatures: females displayed increased ANP, BNP, Sirt1, and AMPK expression, consistent with metabolic resilience and fibrotic remodeling, whereas males exhibited elevated p38 MAPK, NF-κB, LC3B, and cleaved caspase-3, reflecting heightened inflammation, autophagy, and apoptosis. Both sexes demonstrated downregulation of mitochondrial and lipid metabolic proteins, indicating convergent energetic stress. Collectively, these findings identify OA as a systemic inflammatory driver of heart failure, delineate the molecular and proteomic basis of sex-dependent cardiac remodeling, and introduce a translational preclinical model that recapitulates the clinical heterogeneity of HFpEF and HFmrEF/HFrEF, providing a foundation for mechanistic and therapeutic exploration. ### Competing Interest Statement The authors have declared no competing interest. National Institute of Neurological Disorders and Stroke, R01 NS124123

Obstructive sleep apnoea (OSA) is a prevalent cardiorespiratory disorder associated with significant neurocognitive consequences. Despite the higher prevalence of OSA in men, there is a strong association between OSA and Alzheimer's disease (AD), which disproportionately affects women. This study aimed to investigate the impact of chronic intermittent hypoxia (CIH), a hallmark of OSA, on cognitive function and AD markers in ovariectomized, female rats. At 8 weeks of age, 16 Sprague-Dawley rats underwent ovariectomy and were exposed to CIH for 26 weeks. Cognitive function was assessed using the Morris water maze, revealing significant deficits in spatial learning (P < 0.0001) and memory (P = 0.008) in CIH-exposed rats, compared to controls. Analysis of hippocampal tissue showed increased total tau protein (P = 0.0078), indicative of AD pathology. Additionally, CIH-exposed rats exhibited respiratory dysfunction characterized by increased frequency of apnoeas (P = 0.0328). These findings provide preclinical evidence of the association between OSA, cognitive decline and AD pathology in females, emphasizing the importance of sex-specific research in understanding and addressing these pathophysiological interconnections.

Diagnoses of prediabetes and metabolic syndromes, such as metabolic-associated steatotic liver disease (MASLD), are increasing at an alarming rate worldwide, often simultaneously. A significant consequence of these is high risk of cardiovascular disease, highlighting the need for cardiac-specific therapeutics for intervention during the prediabetic stage. Recent studies have demonstrated that chemogenetic activation of the cardiac parasympathetic system through hypothalamic oxytocin (OXT) neurons provides cardioprotective effects in heart disease models by targeting excitatory neurotransmission to brainstem cardiac vagal neurons. In a prediabetic rat model, we hypothesized that stimulating this neural network would offer cardioprotection. To test this, we induced prediabetes through prolonged high-fat, high-fructose feeding. We stereotactically injected viral vectors into the paraventricular nucleus (PVN) of the hypothalamus in neonatal rats to express designer receptors exclusively activated by designer drugs (i.e. Designer Receptors Exclusively Activated by Designer Drugs; DREADDs) in a subset of PVN OXT neurons, chronically activated using the designer drug clozapine N-oxide (CNO). Treated animals exhibited improved cardiac diastolic function and reduced left ventricular fibrosis compared to non-treated diseased animals after 4 weeks of CNO, with no change to systemic insulin resistance, hyperinsulinaemia, and elevated triglycerides. Transcriptional analysis of left ventricular tissue indicates the preservation of pathways involved in metabolism (Pdk4, Pdp1 and Hk2) and ion handling (Sln, Atp1a2 and Atp1a4) in treated compared to diseased animals. These findings underscore the benefits of stimulating the OXT network to counteract prediabetic cardiomyopathy, independent of systemic prediabetes. OXT neurons and their downstream networks appear to be a promising therapeutic target for activating protective parasympathetic-mediated cellular pathways within the heart during prediabetic cardiomyopathy. KEY POINTS: Male rats fed a long-term high-fat, high-fructose diet develop prediabetes, marked by systemic insulin resistance and hyperinsulinaemia, along with characteristics of metabolic-associated steatotic liver disease. Animals exhibit prediabetic cardiomyopathy marked by diastolic dysfunction, interstitial fibrosis, a thickened left ventricular wall and tachycardia with reduced heart rate variability. Designer Receptors Exclusively Activated by Designer Drugs (DREADDs)-mediated chemogenetic activation of oxytocin-expressing neurons in the paraventricular nucleus of the hypothalamus improves cardiac diastolic dysfunction, reduces fibrosis and restores autonomic balance without impacting systemic insulin resistance or hyperinsulinaemia. Transcriptomic analysis of the left ventricle suggests that activation of hypothalamic oxytocin neurons influences cardiac metabolism and ion handling, potentially serving as mechanisms of protection. Our data demonstrate for the first time that paraventricular oxytocin neuron activation is a cardiac-specific approach to improve prediabetic cardiomyopathy.

Autonomic imbalance-particularly reduced activity from brainstem parasympathetic cardiac vagal neurons (CVNs)-is a major characteristic of many cardiorespiratory diseases. Therapeutic approaches to selectively enhance CVN activity have been limited by the lack of defined, translationally relevant targets. Previous studies have identified an important excitatory synaptic pathway from oxytocin (OXT) neurons in the paraventricular nucleus of the hypothalamus to brainstem CVNs, suggesting that OXT could provide a key selective excitation of CVNs. In clinical studies, intranasal OXT has been shown to increase parasympathetic cardiac activity, improve autonomic balance, and reduce obstructive event durations and oxygen desaturations in obstructive sleep apnea patients. However, the mechanisms by which activation of hypothalamic OXT neurons, or intranasal OXT, enhance brainstem parasympathetic cardiac activity remain unclear. CVNs are located in two cholinergic brainstem nuclei: nucleus ambiguus (NA) and dorsal motor nucleus of the vagus (DMNX). In this study, we characterize the colocalization of OXT receptors (OXTRs) in both CVNs and non-CVN cholinergic neurons in the male and female mouse NA and DMNX nuclei. We found that OXT receptors are highly expressed in CVNs in the DMNX, but not in the NA. OXT increases the firing of DMNX CVN, with no effect on NA CVNs. Selective chemogenetic excitation of OXTR+ CVNs in the DMNX-achieved by a combination of Cre- and flp-dependent DREADD expression-evoked a rapid and sustained bradycardia. These findings suggest that activation of DMNX CVNs expressing OXTR with oxytocin may represent a novel translational therapeutic target for restoring autonomic balance in cardiorespiratory disorders.

Diagnoses of prediabetes and metabolic syndromes, such as metabolic-associated steatotic liver disease, are increasing at an alarming rate worldwide, often simultaneously. A significant consequence of these is high risk of cardiovascular disease, highlighting the need for cardiac-specific therapeutics for intervention during the prediabetic stage. Recent studies have demonstrated that chemogenetic activation of the cardiac parasympathetic system through hypothalamic oxytocin (OXT) neurons provides cardioprotective effects in heart disease models by targeting excitatory neurotransmission to brainstem cardiac vagal neurons. In a prediabetic rat model, we hypothesized that stimulating this neural network would offer cardioprotection. To test this, we induced prediabetes through prolonged high-fat, high-fructose feeding. We stereotactically injected viral vectors into the paraventricular nucleus (PVN) of the hypothalamus in neonatal rats to express designer receptors exclusively activated by designer drugs (DREADDs) in a subset of PVN OXT neurons, and chronically activated them using the designer drug clozapine-n-oxide (CNO). Treated animals exhibited improved cardiac diastolic function and reduced left ventricular fibrosis compared to non-treated diseased animals after 4 weeks of CNO, with no change to systemic insulin resistance, hyperinsulinemia, and elevated triglycerides. Transcriptional analysis of left ventricular tissue indicates the preservation of pathways involved in metabolism (Pdk4, Pdp1, Hk2) and ion handling (Sln, Atp1a2, Atp1a4) in treated compared to diseased animals. These findings underscore the benefits of stimulating the OXT network to counteract prediabetic cardiomyopathy, independent of systemic prediabetes. OXT neurons and their downstream networks appear to be a promising therapeutic target for activating protective parasympathetic-mediated cellular pathways within the heart during prediabetic cardiomyopathy.

A balance of cholinergic and catecholaminergic activation is necessary to maintain heart health. Interrogating the interaction between these pathways can be done using optogenetics through selective expression of channelrhodopsin-2 (ChR2) in cardiac autonomic neurons. Such cardiac applications of optogenetics allow for the study of the intrinsic release of neurotransmitters in a spatiotemporal manner. This method illustrates an ex vivo approach for specific optogenetic stimulation of cardiac neurons in perfused mouse hearts. Transgenic mice were bred to express ChR2 in either choline acetyltransferase (ChAT) or tyrosine hydroxylase (TH) neurons throughout the body. A micro-LED (465 nm) encased in a silicone elastomer was prepared for stimulating the neurons of the right atrium that innervate the sinoatrial node. The micro-LED was connected to a function generator set to pulse waves at 10 Hz with a 30 ms pulse width. Hearts with confirmed expression were excised and retrogradely perfused on a Langendorff system circulating Krebs-Henseleit solution. Electrocardiogram (ECG), temperature, and coronary flow rate were recorded using the LabChart software. Once the heart stabilized, the micro-LED was placed on the right atrium and tested for optimal heart rate response. An application of this approach combines the intrinsic release of cholinergic neurotransmitter (acetylcholine) during optogenetic activation of a ChAT-ChR2 mouse heart simultaneously with increasing exogenous catecholaminergic neurotransmitter (norepinephrine) added to the perfusate. The resulting changes in heart rate during the simultaneous cholinergic and catecholaminergic activation are presented. This method describes a valuable experimental approach for investigating the kinetics of sudden intrinsic autonomic neuron activation in perfused hearts and the interactions between cardiac cholinergic and catecholaminergic activity.

Weight loss medications are emerging candidates for pharmacotherapy of sleep-disordered breathing (SDB). A melanocortin 4 receptor (MC4R) agonist, setmelanotide (Set), is used to treat obesity caused by abnormal melanocortin and leptin signaling. We hypothesized that Set can treat SDB in mice with diet-induced obesity. We performed a proof-of-concept randomized crossover trial of a single dose of Set versus vehicle and a 2-week daily Set versus vehicle trial, examined colocalization of Mc4r mRNAs with the markers of CO2-sensing neurons Phox2b and neuromedin B in the brainstem, and expressed Cre-dependent designer receptors exclusively activated by designer drugs (DREADDs) or caspase in obese Mc4r-Cre mice. Set increased minute ventilation across sleep/wake states, enhanced the hypercapnic ventilatory response (HCVR), and abolished apneas during sleep. Phox2b+ neurons in the nucleus of the solitary tract (NTS) and the parafacial region expressed Mc4r. Chemogenetic stimulation of the MC4R+ neurons in the parafacial region, but not in the NTS, augmented HCVR without any changes in metabolism. Caspase elimination of the parafacial MC4R+ neurons abolished effects of Set on HCVR. Parafacial MC4R+ neurons projected to the respiratory premotor neurons retrogradely labeled from C3-C4. In conclusion, MC4R agonists enhance the HCVR and treat SDB by acting on the parafacial MC4R+ neurons.