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1.
Artigo em Inglês | MEDLINE | ID: mdl-34486396

RESUMO

In this review we will briefly summarize the evidence that autonomic imbalance, and more specifically, reduced parasympathetic activity to the heart generates and/or maintains many cardiorespiratory diseases; and discuss mechanisms and sites, from myocytes to the brain, that are potential translational targets for restoring parasympathetic activity and improving cardiorespiratory health.

2.
Int J Mol Sci ; 22(16)2021 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-34445224

RESUMO

The tightly localized noradrenergic neurons (NA) in the locus coeruleus (LC) are well recognized as essential for focused arousal and novelty-oriented responses, while many children with autism spectrum disorder (ASD) exhibit diminished attention, engagement and orienting to exogenous stimuli. This has led to the hypothesis that atypical LC activity may be involved in ASD. Oxytocin (OXT) neurons and receptors are known to play an important role in social behavior, pair bonding and cognitive processes and are under investigation as a potential treatment for ASD. However, little is known about the neurotransmission from hypothalamic paraventricular (PVN) OXT neurons to LC NA neurons. In this study, we test, in male and female rats, whether PVN OXT neurons excite LC neurons, whether oxytocin is released and involved in this neurotransmission, and whether activation of PVN OXT neurons alters novel object recognition. Using "oxytocin sniffer cells" (CHO cells that express the human oxytocin receptor and a Ca indicator) we show that there is release of OXT from hypothalamic PVN OXT fibers in the LC. Optogenetic excitation of PVN OXT fibers excites LC NA neurons by co-release of OXT and glutamate, and this neurotransmission is greater in males than females. In male, but not in female animals, chemogenetic activation of PVN OXT neurons increases attention to novel objects.


Assuntos
Atenção , Locus Cerúleo/metabolismo , Neurônios/metabolismo , Ocitocina/metabolismo , Núcleo Hipotalâmico Paraventricular/metabolismo , Caracteres Sexuais , Transmissão Sináptica , Animais , Células CHO , Cricetulus , Feminino , Humanos , Masculino , Ocitocina/genética , Ratos , Receptores de Ocitocina/genética , Receptores de Ocitocina/metabolismo
3.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201760

RESUMO

Obstructive sleep apnea (OSA) is recurrent obstruction of the upper airway due to the loss of upper airway muscle tone during sleep. OSA is highly prevalent, especially in obesity. There is no pharmacotherapy for OSA. Previous studies have demonstrated the role of leptin, an adipose-tissue-produced hormone, as a potent respiratory stimulant. Leptin signaling via a long functional isoform of leptin receptor, LEPRb, in the nucleus of the solitary tract (NTS), has been implicated in control of breathing. We hypothesized that leptin acts on LEPRb positive neurons in the NTS to increase ventilation and maintain upper airway patency during sleep in obese mice. We expressed designer receptors exclusively activated by designer drugs (DREADD) selectively in the LEPRb positive neurons of the NTS of Leprb-Cre-GFP mice with diet-induced obesity (DIO) and examined the effect of DREADD ligand, J60, on tongue muscle activity and breathing during sleep. J60 was a potent activator of LEPRb positive NTS neurons, but did not stimulate breathing or upper airway muscles during NREM and REM sleep. We conclude that, in DIO mice, the stimulating effects of leptin on breathing during sleep are independent of LEPRb signaling in the NTS.


Assuntos
Neurônios/metabolismo , Receptores de Droga/metabolismo , Receptores para Leptina/metabolismo , Síndromes da Apneia do Sono/fisiopatologia , Núcleo Solitário/citologia , Animais , Eletromiografia , Leptina/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios/efeitos dos fármacos , Obesidade/etiologia , Obesidade/fisiopatologia , Sono REM , Núcleo Solitário/metabolismo
4.
Sleep ; 44(6)2021 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-33624805

RESUMO

STUDY OBJECTIVES: Obesity leads to obstructive sleep apnea (OSA), which is recurrent upper airway obstruction during sleep, and obesity hypoventilation syndrome (OHS), hypoventilation during sleep resulting in daytime hypercapnia. Impaired leptin signaling in the brain was implicated in both conditions, but mechanisms are unknown. We have previously shown that leptin stimulates breathing and treats OSA and OHS in leptin-deficient ob/ob mice and leptin-resistant diet-induced obese mice and that leptin's respiratory effects may occur in the dorsomedial hypothalamus (DMH). We hypothesized that leptin receptor LepRb-deficient db/db mice have obesity hypoventilation and that restoration of leptin signaling in the DMH will increase ventilation during sleep in these animals. METHODS: We measured arterial blood gas in unanesthetized awake db/db mice. We subsequently infected these animals with Ad-LepRb or control Ad-mCherry virus into the DMH and measured ventilation during sleep as well as CO2 production after intracerebroventricular (ICV) infusions of phosphate-buffered saline or leptin. RESULTS: Awake db/db mice had elevated CO2 levels in the arterial blood. Ad-LepRb infection resulted in LepRb expression in the DMH neurons in a similar fashion to wildtype mice. In LepRb-DMH db/db mice, ICV leptin shortened REM sleep and increased inspiratory flow, tidal volume, and minute ventilation during NREM sleep without any effect on the quality of NREM sleep or CO2 production. Leptin had no effect on upper airway obstruction in these animals. CONCLUSION: Leptin stimulates breathing and treats obesity hypoventilation acting on LepRb-positive neurons in the DMH.


Assuntos
Leptina , Receptores para Leptina , Animais , Hipotálamo/metabolismo , Leptina/metabolismo , Camundongos , Camundongos Obesos , Receptores para Leptina/genética , Receptores para Leptina/metabolismo , Sono
5.
Methods Mol Biol ; 2191: 309-321, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32865752

RESUMO

Optogenetic technology has enabled unparalleled insights into cellular and organ physiology by providing exquisite temporal and spatial control of biological pathways. Here, an optogenetic approach is presented for selective activation of the intrinsic cardiac nervous system in excised perfused mouse hearts. The breeding of transgenic mice that have selective expression of channelrhodopsin in either catecholaminergic or cholinergic neurons is described. An approach for perfusing hearts excised from those animals, recording the ECG to measure heart rate changes, and an illumination technique using a custom micro-LED light source to activate channelrhodopsin is explained. We have used these methods in ongoing studies of the kinetics of autonomic control of cardiac electrophysiology and contractility, demonstrating the proven utility of optogenetic technology to enable unparalleled spatiotemporal anatomic-functional probing of the intrinsic cardiac nervous system.


Assuntos
Channelrhodopsins/genética , Coração/fisiologia , Interneurônios/metabolismo , Optogenética/métodos , Potenciais de Ação/genética , Animais , Humanos , Interneurônios/patologia , Camundongos , Camundongos Transgênicos , Estimulação Luminosa/métodos
6.
JACC Clin Electrophysiol ; 6(14): 1827-1840, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33357580

RESUMO

OBJECTIVES: This study sought to investigate the shift of leading pacemaker locations in healthy and failing mammalian hearts over the entire range of physiological heart rates (HRs), and to molecularly characterize spatial regions of spontaneous activity. BACKGROUND: A normal heartbeat originates as an action potential in a group of pacemaker cells known as the sinoatrial node (SAN), located near the superior vena cava. HRs and the anatomical site of origin of pacemaker activity in the adult heart are known to dynamically change in response to various physiological inputs, yet the mechanism of this pacemaker shift is not well understood. METHODS: Optical mapping was applied to ex vivo rat and human isolated right atrial tissues, and HRs were modulated with acetylcholine and isoproterenol. RNA sequencing was performed on tissue areas that elicited spontaneous activity, and comparisons were made to neighboring myocardial tissues. RESULTS: Functional and molecular evidence identified and confirmed the presence of 2 competing right atrial pacemakers localized near the superior vena cava and the inferior vena cava-the superior SAN (sSAN) and inferior SAN (iSAN), respectively-which preferentially control the fast and slow HRs. Both of these regions were evident in non-failing rat and human hearts and maintained spontaneous activity in the rat heart when physically separated from one another. Molecular analysis of these 2 pacemaker regions revealed unique but similar transcriptional profiles, suggesting iSAN dominance when the sSAN is silent. CONCLUSIONS: The presence of 2 spatially distinct dominant pacemakers, sSAN and iSAN, in the mammalian heart clarifies previous identification of migrating pacemakers and corresponding changes in P-wave morphology in mammalian species.


Assuntos
Nó Sinoatrial , Veia Cava Superior , Potenciais de Ação , Animais , Átrios do Coração , Frequência Cardíaca , Humanos , Ratos
7.
Artigo em Inglês | MEDLINE | ID: mdl-33006920

RESUMO

BACKGROUND: Left ventricular (LV) electrical maladaptation to increased heart rate in failing myocardium contributes to morbidity and mortality. Recently, cardiac cholinergic neuron activation reduced loss of contractile function resulting from chronic trans-aortic constriction (TAC) in rats. We hypothesized that chronic activation of cardiac cholinergic neurons would also reduce TAC-induced derangement of cardiac electrical activity. METHODS: We investigated electrophysiological rate adaptation in TAC rat hearts with and without daily chemogenetic activation of hypothalamic oxytocin neurons for downstream cardiac cholinergic neuron stimulation. Sprague Dawley rat hearts were excised, perfused, and optically mapped under dynamic pacing after 16 weeks of TAC with or without 12 weeks of daily chemogenetic treatment. Action potential duration (APD60) and conduction velocity (CV) maps were analyzed for regional rate adaptation to dynamic pacing. RESULTS: At lower pacing rates, untreated TAC induced elevated LV epicardial APD60. Fitted APD60 steady state (APDss) was reduced in treated TAC hearts. At higher pacing rates, treatment heterogeneously reduced APD60 compared to untreated TAC hearts. Variance of conduction loss was reduced in treated hearts compared to untreated hearts during fast pacing. However, CV was markedly reduced in both treated and untreated TAC hearts throughout dynamic pacing. At 150msec pacing cycle length, APD60 v. diastolic interval (DI) dispersion was reduced in treated hearts compared to untreated hearts. CONCLUSIONS: Chronic activation of cardiac cholinergic neurons improved electrophysiological adaptation to increases in pacing rate during development of TAC-induced heart failure. This provides insight into the electrophysiological benefits of cholinergic stimulation as a treatment for heart failure patients.

8.
eNeuro ; 7(5)2020.
Artigo em Inglês | MEDLINE | ID: mdl-32855199

RESUMO

We asked whether the physiological and morphologic properties of hypoglossal motor neurons (CNXII MNs) that innervate protruder or retractor tongue muscles are disrupted in neonatal LgDel mice that carry a heterozygous deletion parallel to that associated with DiGeorge/22q11.2 deletion syndrome (22q11.2DS). Disrupted coordination of tongue movement in LgDel mouse pups may contribute to suckling, feeding, and swallowing (S/F/S) disruptions that parallel pediatric dysphagia in infants and toddlers with 22q11.2DS. Using an in vitro rhythmically active medullary slice preparation, we found spontaneous firing as well as IPSC frequency differed significantly in neonatal LgDel versus wild-type (WT) protruder and retractor CNXII MNs that were identified by retrograde tracing from their target muscles. In response to respiration-related activity, initiation and decay of transiently increased firing in WT protruder MNs is delayed in LgDel, accompanied by altered excitatory/inhibitory (E/I) balance. In addition, LgDel retractor MNs have a transient increase in firing with diminished IPSC frequency that is not seen in WT. There were no significant differences in cell body volume of either XII class in WT and LgDel Sholl analysis showed the total numbers of dendritic intersections (at 50- and 90-µm radii from the cell soma) were significantly greater for LgDel versus WT retractor MNs. Thus, the physiological, synaptic and cellular properties of distinct classes of CNXII MNs that coordinate tongue movement in neonatal WT mice are altered in LgDel Such changes could contribute to sub-optimal coordination of S/F/S that underlies pediatric dysphagia in 22q11.2DS.


Assuntos
Transtornos de Deglutição , Síndrome de DiGeorge , Animais , Criança , Síndrome de DiGeorge/complicações , Síndrome de DiGeorge/genética , Modelos Animais de Doenças , Humanos , Bulbo , Camundongos , Neurônios Motores
9.
Sleep Med ; 74: 242-247, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32862007

RESUMO

BACKGROUND: Activation of the oxytocin network has shown benefits in animal models of Obstructive Sleep Apnea (OSA) as well as other cardiorespiratory diseases. We sought to determine if nocturnal intranasal oxytocin administration could have beneficial effects in reducing the duration and/or frequency of obstructive events in obstructive sleep apnea subjects. METHODS: Two sequential standard "in-lab" polysomnogram (PSG) sleep studies were performed in patients diagnosed with OSA that were randomly assigned to initially receive either placebo or oxytocin (40 i.u.) administered intranasally in this double blinded randomized placebo controlled study. Changes in cardiorespiratory events during sleep, including apnea and hypopnea durations and frequency, risk of event-associated bradycardias, arterial oxygen desaturation and respiratory rate were assessed in 2 h epochs following sleep onset. Oxytocin significantly decreased the duration of obstructive events, as well as the oxygen desaturations and incidence of bradycardia that were associated with these events. Notably, oxytocin increased respiratory rate during non-obstructive periods. There were no significant changes in sleep architecture and no adverse effects were reported. CONCLUSIONS: Oxytocin administration can benefit OSA subjects by reducing the duration and adverse consequences of obstructive events. Oxytocin could also be beneficial in situations involving respiratory depression as oxytocin increased respiratory rate. Additional studies are needed to further understand the mechanisms by which oxytocin promotes these changes in cardiorespiratory function. The long-term efficacy and optimal dose of intranasal oxytocin treatment should also be determined in OSA subjects. ClinicalTrials.gov NCT03148899.


Assuntos
Ocitocina , Apneia Obstrutiva do Sono , Humanos , Oxigênio , Polissonografia , Taxa Respiratória , Apneia Obstrutiva do Sono/tratamento farmacológico
10.
Am J Respir Cell Mol Biol ; 63(4): 502-509, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32603263

RESUMO

Respiratory depression is the main cause of morbidity and mortality associated with opioids. Obesity increases opioid-related mortality, which is mostly related to comorbid obstructive sleep apnea. Naloxone, a µ-opioid receptor blocker, is an effective antidote, but it reverses analgesia. Like humans with obesity, mice with diet-induced obesity hypoventilate during sleep and develop obstructive sleep apnea, which can be treated with intranasal leptin. We hypothesized that intranasal leptin reverses opioid-induced sleep-disordered breathing in obese mice without decreasing analgesia. To test this hypothesis, mice with diet-induced obesity were treated with morphine at 10 mg/kg subcutaneously and with leptin or placebo intranasally. Sleep and breathing were recorded by barometric plethysmography, and pain sensitivity was measured by the tail-flick test. Excitatory postsynaptic currents were recorded in vitro from hypoglossal motor neurons after the application of the µ-opioid receptor agonist [D-Ala2, N-MePhe4, Gly-ol]-enkephalin and leptin. Morphine dramatically increased the frequency of apneas and greatly increased the severity of hypoventilation and obstructive sleep apnea. Leptin decreased the frequency of apneas, improved obstructive sleep apnea, and completely reversed hypoventilation, whereas morphine analgesia was enhanced. Our in vitro studies demonstrated that [D-Ala2, N-MePhe4, Gly-ol]-enkephalin reduced the frequency of excitatory postsynaptic currents in hypoglossal motoneurons and that application of leptin restored excitatory synaptic neurotransmission. Our findings suggest that intranasal leptin may prevent opioid respiratory depression during sleep in patients with obesity receiving opioids without reducing analgesia.


Assuntos
Analgésicos Opioides/efeitos adversos , Leptina/administração & dosagem , Respiração/efeitos dos fármacos , Síndromes da Apneia do Sono/induzido quimicamente , Síndromes da Apneia do Sono/prevenção & controle , Sono/efeitos dos fármacos , Administração Intranasal/métodos , Analgesia/métodos , Animais , Modelos Animais de Doenças , Encefalinas/farmacologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Morfina/farmacologia , Neurônios Motores/efeitos dos fármacos , Neurônios Motores/metabolismo , Receptores Opioides mu/metabolismo , Síndromes da Apneia do Sono/metabolismo , Transmissão Sináptica/efeitos dos fármacos
11.
JACC Basic Transl Sci ; 5(5): 484-497, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32478209

RESUMO

This work shows long-term restoration of the hypothalamic oxytocin (OXT) network preserves OXT release, reduces mortality, cardiac inflammation, fibrosis, and improves autonomic tone and cardiac function in a model of heart failure. Intranasal administration of OXT in patients mimics the short-term changes seen in animals by increasing parasympathetic-and decreasing sympathetic-cardiac activity. This work provides the essential translational foundation to determine if approaches that mimic paraventricular nucleus (PVN) OXT neuron activation, such as safe, noninvasive, and well-tolerated intranasal administration of OXT, can be beneficial in patients with heart failure.

12.
Am J Physiol Heart Circ Physiol ; 319(1): H3-H12, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32412778

RESUMO

Heart failure (HF) is characterized by autonomic imbalance with sympathetic hyperactivity and loss of parasympathetic tone. Intracardiac ganglia (ICG) neurons represent the final common pathway for vagal innervation of the heart and strongly regulate cardiac functions. This study tests whether ICG cholinergic neuron activation mitigates the progression of cardiac dysfunction and reduces mortality that occurs in HF. HF was induced by transaortic constriction (TAC) in male transgenic Long-Evans rats expressing Cre recombinase within choline acetyltransferase (ChAT) neurons. ChAT neurons were selectively activated by expression and activation of excitatory designer receptors exclusively activated by designer receptors (DREADDs) by clozapine-N-oxide (TAC + treatment and sham-treated groups). Control animals expressed DREADDs but received saline (sham and TAC groups). A separate set of animals were telemetry instrumented to record blood pressure (BP) and heart rate (HR). Acute activation of ICG neurons resulted in robust reductions in BP (∼20 mmHg) and HR (∼100 beats/min). All groups of animals were subjected to weekly echocardiography and treadmill stress tests from 3 to 6 wk post-TAC/sham surgery. Activation of ICG cholinergic neurons reduced the left ventricular systolic dysfunction (reductions in ejection fraction, fractional shortening, stroke volume, and cardiac output) and cardiac autonomic dysfunction [reduced HR recovery (HRR) post peak effort] observed in TAC animals. Additionally, activation of ICG ChAT neurons reduced mortality by 30% compared with untreated TAC animals. These data suggest that ICG cholinergic neuron activation reduces cardiac dysfunction and improves survival in HF, indicating that ICG neuron activation could be a novel target for treating HF.NEW & NOTEWORTHY Intracardiac ganglia form the final common pathway for the parasympathetic innervation of the heart. This study has used a novel chemogenetic approach within transgenic ChAT-Cre rats [expressing only Cre-recombinase in choline acetyl transferase (ChAT) neurons] to selectively increase intracardiac cholinergic parasympathetic activity to the heart in a pressure overload-induced heart failure model. The findings from this study confirm that selective activation of intracardiac cholinergic neurons lessens cardiac dysfunction and mortality seen in heart failure, identifying a novel downstream cardiac-selective target for increasing cardioprotective parasympathetic activity in heart failure.


Assuntos
Neurônios Colinérgicos/fisiologia , Insuficiência Cardíaca/fisiopatologia , Coração/inervação , Função Ventricular , Animais , Sistema Nervoso Autônomo/fisiopatologia , Pressão Sanguínea , Colina O-Acetiltransferase/genética , Colina O-Acetiltransferase/metabolismo , Neurônios Colinérgicos/efeitos dos fármacos , Neurônios Colinérgicos/metabolismo , Clozapina/análogos & derivados , Clozapina/farmacologia , Coração/fisiopatologia , Insuficiência Cardíaca/etiologia , Frequência Cardíaca , Masculino , Ratos , Ratos Long-Evans , Obstrução do Fluxo Ventricular Externo/complicações
13.
Front Neurol ; 11: 4, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32082240

RESUMO

Disrupted development of oropharyngeal structures as well as cranial nerve and brainstem circuits may lead to feeding and swallowing difficulties in children with 22q11. 2 deletion syndrome (22q11DS). We previously demonstrated aspiration-based dysphagia during early postnatal life in the LgDel mouse model of 22q11DS along with disrupted oropharyngeal morphogenesis and divergent differentiation and function of cranial motor and sensory nerves. We now ask whether feeding and swallowing deficits persist in adult LgDel mice using methods analogous to those used in human patients to evaluate feeding and swallowing dysfunction. Compared to wild-type mice, videofluoroscopic swallow study revealed that LgDel mice have altered feeding and swallowing behaviors, including slower lick rates, longer inter-lick intervals, and longer pharyngeal transit times with liquid consistency. Transoral endoscopic assessment identified minor structural anomalies of the palate and larynx in one-third of the LgDel mice examined. Video surveillance of feeding-related behaviors showed that LgDel mice eat and drink more frequently. Furthermore, LgDel animals engage in another oromotor behavior, grooming, more frequently, implying that divergent craniofacial and cranial nerve structure and function result in altered oromotor coordination. Finally, LgDel mice have significantly increased lung inflammation, a potential sign of aspiration-based dysphagia, consistent with results from our previous studies of early postnatal animals showing aspiration-related lung inflammation. Thus, oromotor dysfunction, feeding, and swallowing difficulties and their consequences persist in the LgDel 22q11DS mouse model. Apparently, postnatal growth and/or neural plasticity does not fully resolve deficits due to anomalous hindbrain, craniofacial, and cranial nerve development that prefigure perinatal dysphagia in 22q11DS. This new recognition of persistent challenges with feeding and swallowing may provide opportunities for improved therapeutic intervention for adolescents and adults with 22q11DS, as well as others with a history of perinatal feeding and swallowing disorders.

14.
Sleep ; 43(6)2020 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-31832664

RESUMO

Obstructive sleep apnea (OSA) is a common disorder characterized by repetitive sleep-related losses of upper airway patency that occur most frequently during rapid eye movement (REM) sleep. Hypoglossal motoneurons play a key role in regulating upper airway muscle tone and patency during sleep. REM sleep activates GABA and glycine neurons in the ventral medulla (VM) to induce cortical desynchronization and skeletal muscle atonia during REM sleep; however, the role of this brain region in modulating hypoglossal motor activity is unknown. We combined optogenetic and chemogenetic approaches with in-vitro and in-vivo electrophysiology, respectfully, in GAD2-Cre mice of both sexes to test the hypothesis that VM GABA/glycine neurons control the activity of hypoglossal motoneurons and tongue muscles. Here, we show that there is a pathway originating from GABA/glycine neurons in the VM that monosynaptically inhibits brainstem hypoglossal motoneurons innervating both tongue protruder genioglossus (GMNs) and retractor (RMNs) muscles. Optogenetic activation of ChR2-expressing fibers induced a greater postsynaptic inhibition in RMNs than in GMNs. In-vivo chemogenetic activation of VM GABA/glycine neurons produced an inhibitory effect on tongue electromyographic (EMG) activity, decreasing both the amplitude and duration of inspiratory-related EMG bursts without any change in respiratory rate. These results indicate that activation of GABA/glycine neurons from the VM inhibits tongue muscles via a direct pathway to both GMNs and RMNs. This inhibition may play a role in REM sleep associated upper airway obstructions that occur in patients with OSA.


Assuntos
Glicina , Nervo Hipoglosso , Animais , Feminino , Humanos , Masculino , Camundongos , Neurônios Motores , Sono REM , Ácido gama-Aminobutírico
15.
Front Physiol ; 10: 16, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30745877

RESUMO

The balance of sympathetic and parasympathetic tone provides exquisite control of heart rate and contractility and has also been shown to modulate coronary flow and inflammation. Understanding how autonomic balance is altered by cardiac disease is an active area of research, and developing new ways to control this balance provides insights into disease therapies. However, achieving acute neuron-specific stimulation of autonomic neurons can be difficult in experiments that measure the acute effects of nerve stimulation on the heart. Conventional electrical and pharmacological approaches can be spatially and temporally non-selective. Cell-specific expression of light-activated channels (channelrhodopsin, ChR2) is a powerful approach that enables control of the timing and distribution of cellular stimulation using light. We present such an optogenetic approach where parasympathetic cardiac neurons are selectively photoactivated at high temporal precision to initiate cholinergic-mediated slowing of heart rate. Mice were crossbred to express ChR2 in peripheral cholinergic neurons using Cre-Lox recombination driven by a choline acetyltransferase (ChAT) promoter. Hearts from adult mice were excised, perfused, and the epicardium was illuminated (peak 460-465 nm) to photoactivate ChR2. In one set of studies, hearts were illuminated using a large-field LED light source. In other studies, a micro LED was placed on the right atrium to selectively illuminate the junction of the superior vena cava (SVC) and right atrium. The ECG was acquired before, during, and after tissue illumination to measure changes in heart rate. Upon illumination, hearts exhibited sudden and dramatic reductions in heart rate with restoration of normal heart rate after cessation of illumination. Delays in atrioventricular conduction were also observed. Heart rate reductions at the highest irradiance levels were similar to heart rate reductions caused by application of bethanechol (10 µM) or acetylcholine (800 µM). Atropine (50 nM) completely blocked the effect of ChR2 photoactivation, confirming cholinergic mediation. Optogenetic activation of intrinsic parasympathetic neurons reduced heart rate in an immediate, dose-dependent fashion, resembling the slowing of sinus rate in response to acetylcholine. Our results demonstrate a new approach for controlling parasympathetic modulation of cardiac function by selectively activating the endogenous release of acetylcholine from intrinsic cardiac cholinergic neurons. Key Message: Optogenetic photoactivation of intrinsic cardiac neurons provides immediate, tissue-specific stimulation with minimal cross-reactivity. Our results demonstrate that selective expression of channelrhodopsin within cardiac cholinergic neurons enables photoactivated release of acetylcholine, thereby instantaneously slowing sinus rate and altering atrioventricular conduction. This provides for in-depth examination of the endogenous interplay between cardiac autonomic neurons and the functional outcomes of downstream post-synaptic receptor activation.

16.
Am J Respir Crit Care Med ; 199(6): 773-783, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30309268

RESUMO

RATIONALE: Leptin treats upper airway obstruction and alveolar hypoventilation in leptin-deficient ob/ob mice. However, obese humans and mice with diet-induced obesity (DIO) are resistant to leptin because of poor permeability of the blood-brain barrier. We propose that intranasal leptin will bypass leptin resistance and treat sleep-disordered breathing in obesity. OBJECTIVES: To assess if intranasal leptin can treat obesity hypoventilation and upper airway obstruction during sleep in mice with DIO. METHODS: Male C57BL/6J mice were fed with a high-fat diet for 16 weeks. A single dose of leptin (0.4 mg/kg) or BSA (vehicle) were administered intranasally or intraperitoneally, followed by either sleep studies (n = 10) or energy expenditure measurements (n = 10). A subset of mice was treated with leptin daily for 14 days for metabolic outcomes (n = 20). In a separate experiment, retrograde viral tracers were used to examine connections between leptin receptors and respiratory motoneurons. MEASUREMENTS AND MAIN RESULTS: Acute intranasal, but not intraperitoneal, leptin decreased the number of oxygen desaturation events in REM sleep, and increased ventilation in non-REM and REM sleep, independently of metabolic effects. Chronic intranasal leptin decreased food intake and body weight, whereas intraperitoneal leptin had no effect. Intranasal leptin induced signal transducer and activator of transcription 3 phosphorylation in hypothalamic and medullary centers, whereas intraperitoneal leptin had no effect. Leptin receptor-positive cells were synaptically connected to respiratory motoneurons. CONCLUSIONS: In mice with DIO, intranasal leptin bypassed leptin resistance and significantly attenuated sleep-disordered breathing independently of body weight.


Assuntos
Leptina/metabolismo , Absorção Nasal/fisiologia , Obesidade/complicações , Síndromes da Apneia do Sono/complicações , Síndromes da Apneia do Sono/fisiopatologia , Sono/fisiologia , Animais , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais
17.
Front Neurol ; 9: 962, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30487776

RESUMO

Obstructive Sleep Apnea (OSA) is a prevalent condition and a major cause of morbidity and mortality in Western Society. The loss of motor input to the tongue and specifically to the genioglossus muscle during sleep is associated with pharyngeal collapsibility and the development of OSA. We applied a novel chemogenetic method to develop a mouse model of sleep disordered breathing Our goal was to reversibly silence neuromotor input to the genioglossal muscle using an adeno-associated viral vector carrying inhibitory designer receptors exclusively activated by designer drugs AAV5-hM4Di-mCherry (DREADD), which was delivered bilaterally to the hypoglossal nucleus in fifteen C57BL/6J mice. In the in vivo experiment, 4 weeks after the viral administration mice were injected with a DREADD ligand clozapine-N-oxide (CNO, i.p., 1mg/kg) or saline followed by a sleep study; a week later treatments were alternated and a second sleep study was performed. Inspiratory flow limitation was recognized by the presence of a plateau in mid-respiratory flow; oxyhemoglobin desaturations were defined as desaturations >4% from baseline. In the in vitro electrophysiology experiment, four males and three females of 5 days of age were used. Sixteen-nineteen days after DREADD injection brain slices of medulla were prepared and individual hypoglossal motoneurons were recorded before and after CNO application. Positive mCherry staining was detected in the hypoglossal nucleus in all mice confirming successful targeting. In sleep studies, CNO markedly increased the frequency of flow limitation n NREM sleep (from 1.9 ± 1.3% after vehicle injection to 14.2 ± 3.4% after CNO, p < 0.05) and REM sleep (from 22.3% ± 4.1% to 30.9 ± 4.6%, respectively, p < 0.05) compared to saline treatment, but there was no significant oxyhemoglobin desaturation or sleep fragmentation. Electrophysiology recording in brain slices showed that CNO inhibited firing frequency of DREADD-containing hypoglossal motoneurons. We conclude that chemogenetic approach allows to silence hypoglossal motoneurons in mice, which leads to sleep disordered breathing manifested by inspiratory flow limitation during NREM and REM sleep without oxyhemoglobin desaturation or sleep fragmentation. Other co-morbid factors, such as compromised upper airway anatomy, may be needed to achieve recurrent pharyngeal obstruction observed in OSA.

18.
Brain Res ; 1679: 33-38, 2018 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-29162453

RESUMO

Both orexin neurons in the lateral hypothalamus and spinally-projecting pre-sympathetic neurons (PSNs) in the paraventricular nucleus of the hypothalamus (PVN) play an important role in the regulation of cardiovascular function under normal conditions and during cardiovascular challenges such as hypoxia and/or hypercapnia. We have previously established, using selective optogenetic excitation of orexin neurons and pathways, there is a heterogeneous neurotransmission from orexin neurons to PSNs in the PVN. This study was undertaken to test whether this pathway is altered by acute exposure to hypoxia alone and/or combined hypoxia and hypercapnia (H/H). To test this hypothesis, we selectively expressed channelrhodopsin-2 (ChR2) in orexin neurons in the lateral hypothalamus and photoactivated ChR2-expressing fibers to evoke postsynaptic currents in spinally-projecting PSNs in an in vitro slice preparation in rats. In accordance with previously published data, two subpopulations of spinally-projecting PSNs were established, including those with glutamatergic or GABAergic inputs from orexin neurons. Hypoxia alone did not alter the peak amplitude of either glutamatergic or GABAergic neurotransmission, however, H/H significantly inhibited both glutamatergic and GABAergic neurotransmission from orexin neurons to SPNs. In conclusion, H/H may modulate cardiovascular function by affecting heterogeneous pathways from orexin neurons to spinally-projecting PSNs in the PVN.


Assuntos
Hipercapnia/fisiopatologia , Hipóxia/fisiopatologia , Neurônios/fisiologia , Núcleo Hipotalâmico Paraventricular/citologia , Transmissão Sináptica/fisiologia , 6-Ciano-7-nitroquinoxalina-2,3-diona/farmacologia , Animais , Antagonistas de Aminoácidos Excitatórios/farmacologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/genética , Feminino , Antagonistas GABAérgicos/farmacologia , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Hipercapnia/patologia , Hipóxia/patologia , Técnicas In Vitro , Masculino , Neurônios/efeitos dos fármacos , Optogenética , Orexinas/genética , Orexinas/metabolismo , Piridazinas/farmacologia , Ratos , Ratos Transgênicos , Rodopsina/genética , Rodopsina/metabolismo , Transmissão Sináptica/efeitos dos fármacos , Transmissão Sináptica/genética , Fatores de Tempo
19.
Am J Physiol Lung Cell Mol Physiol ; 313(5): L825-L833, 2017 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-28798255

RESUMO

Activation of oxytocin receptors has shown benefits in animal models of obstructive sleep apnea (OSA). We tested if nocturnal oxytocin administration could have beneficial effects in OSA patients. Eight patients diagnosed with OSA were administered intranasal oxytocin (40 IU). Changes in cardiorespiratory events during sleep, including apnea and hypopnea durations and frequency, risk of event-associated arousals, and heart rate variability, were assessed. Oxytocin significantly increased indexes of parasympathetic activity, including heart rate variability, total sleep time, and the postpolysommogram sleep assessment score, an index of self-reported sleep satisfaction. Although the apnea-hypopnea index was not significantly changed with oxytocin administration, when apnea and hypopnea events were compared independently, the frequency of hypopneas, but not apneas, was significantly (P ≤ 0.005) decreased with oxytocin treatment. Both apneas and hypopneas were significantly shortened in duration with oxytocin treatment. Oxytocin treatment significantly decreased the percent of apnea and hypopnea events that were accompanied with an arousal. Oxytocin administration has the potential to restore cardiorespiratory homeostasis and reduce some clinically important (objective and patient-reported) adverse events that occur with OSA. Additional studies are needed to further understand the mechanisms by which oxytocin promotes these changes in cardiorespiratory and autonomic function in OSA patients.


Assuntos
Nível de Alerta/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , Ocitocina/farmacologia , Apneia Obstrutiva do Sono/tratamento farmacológico , Adolescente , Adulto , Encéfalo/efeitos dos fármacos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Ocitocina/administração & dosagem , Apneia Obstrutiva do Sono/fisiopatologia , Adulto Jovem
20.
Neuroscience ; 359: 1-7, 2017 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-28687307

RESUMO

DiGeorge/22q11.2 Deletion Syndrome (22q11DS) is a common genetic microdeletion syndrome that underlies several neurodevelopmental disorders including autism, attention deficit/hyperactivity disorder, and schizophrenia. In addition to cognitive impairments, those with 22q11DS have disrupted feeding and swallowing from birth onward. This perinatal dysphagia significantly compromises nutritional status, impairs appropriate weight gain, and can lead to life threatening aspiration-based infections. Appropriately timed excitation and inhibition of brainstem hypoglossal motor neurons, which innervate tongue muscles, is essential for proper feeding and swallowing. In this study we have examined changes in hypoglossal motor neuron function in the LgDel mouse model of 22q11DS. Hypoglossal motor neurons from LgDel mouse pups have action potentials with afterhyperpolarizations, mediated by a large conductance charybdotoxin-sensitive Ca-activated K current, that are significantly shorter in duration and greater in magnitude than those in wild-type pups. In addition, the amplitude, but not frequency, of glutamatergic excitatory glutamatergic postsynaptic currents (EPSCs) is diminished, and GABAergic, but not glycinergic, neurotransmission to hypoglossal motor neurons was reduced in LgDel animals. These observations provide a foundation for understanding the neurological changes in hypoglossal motor neuron function and their contribution to swallowing abnormalities that occur in DiGeorge/22q11.2 Deletion Syndrome.


Assuntos
Síndrome de DiGeorge/fisiopatologia , Nervo Hipoglosso/fisiopatologia , Bulbo/fisiopatologia , Potenciais da Membrana , Neurônios Motores/fisiologia , Animais , Feminino , Masculino , Camundongos Endogâmicos C57BL , Potenciais Sinápticos
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