Olga Dergacheva

Respiratory modulation of premotor cardiac vagal neurons in the brainstem

The respiratory and cardiovascular systems are highly intertwined, both anatomically and physiologically. Respiratory and cardiovascular neurons are often co-localized in the same brainstem regions, and this is particularly evident in the ventral medulla which contains presympathetic neurons in the rostral ventrolateral medulla, premotor parasympathetic cardioinhibitory neurons in the nucleus ambiguus, and the ventral respiratory group, which includes the pre-Botzinger complex. Anatomical studies of respiratory and cardiovascular neurons have demonstrated that many of these neurons have projections and axon collateral processes which extend into their neighboring cardiorespiratory regions providing an anatomical substrate for cardiorespiratory interactions. As other reports in this Special Issue of Respiratory Physiology & Neurobiology focus on interactions between the respiratory network and baroreceptors, neurons in the nucleus tractus solitarius, presympathetic neurons and sympathetic activity, this report will focus on the respiratory modulation of parasympathetic activity and the neurons that generate parasympathetic activity to the heart, cardiac vagal neurons.

Prenatal Nicotine Exposure Recruits an Excitatory Pathway to Brainstem Parasympathetic Cardioinhibitory Neurons during Hypoxia/Hypercapnia in the Rat: Implications for Sudden Infant Death Syndrome

Maternal cigarette smoking and prenatal nicotine exposure increase the risk for sudden infant death syndrome (SIDS) by 2- to 4-fold, yet despite adverse publicity, nearly one of four pregnant women smoke tobacco. Infants who succumb to SIDS typically experience a severe bradycardia that precedes or is accompanied by centrally mediated life-threatening apneas and gasping. Although the causes of the apnea and bradycardia prevalent in SIDS victims are unknown, it has been hypothesized that these fatal events are exaggerated cardiorespiratory responses to hypoxia or hypercapnia. Changes in heart rate are primarily determined by the activity of cardiac vagal neurons (CVNs) in the brainstem. In this study, we tested whether hypoxia/hypercapnia evokes synaptic pathways to CVNs and whether these cardiorespiratory interactions are altered by prenatal exposure to nicotine. Spontaneous rhythmic inspiratory-related activity was recorded from the hypoglossal rootlet of 700- to 800-μm medullary sections. CVNs were identified in this preparation by retrograde fluorescent labeling, and excitatory synaptic inputs to CVNs were isolated and studied using patch-clamp electrophysiologic techniques. Hypoxia/hypercapnia did not elicit an increase in excitatory neurotransmission to CVNs in unexposed animals, but in animals that were exposed to nicotine in the prenatal period, hypoxia/hypercapnia recruited an excitatory neurotransmission to CVNs. This study establishes a likely neurochemical mechanism for the exaggerated decrease in heart rate in response to hypoxia/hypercapnia that occurs in SIDS victims.

Differential Control of Central Cardiorespiratory Interactions by Hypercapnia and the Effect of Prenatal Nicotine

Hypercapnia evokes a strong cardiorespiratory response including gasping and a pronounced bradycardia; however, the mechanism responsible for these survival responses initiated in the brainstem is unknown. To examine the effects of hypercapnia on the central cardiorespiratory network, we used an in vitro medullary slice that allows simultaneous examination of rhythmic respiratory-related activity and inhibitory synaptic neurotransmission to cardioinhibitory vagal neurons (CVNs). Hypercapnia differentially modulated inhibitory neurotransmission to CVNs; whereas hypercapnia selectively depressed spontaneous glycinergic IPSCs in CVNs without altering respiratory-related increases in glycinergic neurotransmission, it decreased both spontaneous and inspiratory-associated GABAergic IPSCs. Because maternal smoking is the highest risk factor for sudden infant death syndrome (SIDS) and prenatal nicotine exposure is proposed to be the link between maternal smoking and SIDS, we examined the cardiorespiratory responses to hypercapnia in animals exposed to nicotine in the prenatal and perinatal period. In animals exposed to prenatal nicotine, hypercapnia evoked an exaggerated depression of GABAergic IPSCs in CVNs with no significant change in glycinergic neurotransmission. Hypercapnia altered inhibitory neurotransmission to CVNs at both presynaptic and postsynaptic sites. Although the results obtained in this study in vitro cannot be extrapolated with certainty to in vivo responses, the results of this study provide a likely neurochemical mechanism for hypercapnia-evoked bradycardia and the dysregulation of this response with exposure to prenatal nicotine, creating a higher risk for SIDS.

Hypocretin-1 (orexin A) prevents the effects of hypoxia/hypercapnia and enhances the GABAergic pathway from the lateral paragigantocellular nucleus to cardiac vagal neurons in the nucleus ambiguus

Hypocretins (orexins) are hypothalamic neuropeptides that play a crucial role in regulating sleep/wake states and autonomic functions including parasympathetic cardiac activity. We have recently demonstrated stimulation of the lateral paragigantocellular nucleus (LPGi), the nucleus which is thought to play a role in rapid eye movement (REM) sleep control, activates an inhibitory pathway to preganglionic cardiac vagal neurons in the nucleus ambiguus (NA). In this study we test the hypothesis that hypocretin-1 modulates the inhibitory neurotransmission to cardiac vagal neurons evoked by stimulation of the LPGi using whole-cell patch-clamp recordings in an in vitro brain slice preparation from rats. Activation of hypocretin-1 receptors produced a dose-dependent and long-term facilitation of GABAergic postsynaptic currents evoked by electrical stimulation of the LPGi. Hypoxia/hypercapnia diminished LPGi-evoked GABAergic current in cardiac vagal neurons and this inhibition by hypoxia/hypercapnia was prevented by pre-application of hypocretin-1. The action of hypocretin-1 was blocked by the hypocretin-1 receptor antagonist SB-334867. Facilitation of LPGi-evoked GABAergic current in cardiac vagal neurons under both normal condition and during hypoxia/hypercapnia could be the mechanism by which hypocretin-1 affects parasympathetic cardiac function and heart rate during REM sleep. Furthermore, our findings indicate a new potential mechanism that might be involved in the cardiac arrhythmias, bradycardia, and sudden cardiac death that can occur during sleep.

5-Hydroxytryptamine 1A/7 and 4α Receptors Differentially Prevent Opioid-Induced Inhibition of Brain Stem Cardiorespiratory Function

Opioids evoke respiratory depression, bradycardia, and reduced respiratory sinus arrhythmia, whereas serotonin (5-HT) agonists stimulate respiration and cardiorespiratory interactions. This study tested whether serotonin agonists can prevent the inhibitory effects of opioids on cardiorespiratory function. Spontaneous and rhythmic inspiratory-related activity and &ggr;-aminobutyric acid (GABA) neurotransmission to premotor parasympathetic cardioinhibitory neurons in the nucleus ambiguus were recorded simultaneously in an in vitro thick slice preparation. The &mgr;-opioid agonist fentanyl inhibited respiratory frequency. The 5-hydroxytryptamine 1A/7 receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin increased respiratory frequency by itself and also prevented the fentanyl-induced respiratory depression. The 5-hydroxytryptamine 4&agr; agonist BIMU-8 did not by itself change inspiratory activity but prevented the &mgr;-opioid–mediated respiratory depression. Both spontaneous and inspiratory-evoked GABAergic neurotransmission to cardiac vagal neurons were inhibited by fentanyl. 8-Hydroxy-2-(di-n-propylamino)tetralin inhibited spontaneous but not inspiratory-evoked GABAergic activity to parasympathetic cardiac neurons. However, 8-hydroxy-2-(di-n-propylamino)tetralin differentially altered the opioid-mediated depression of inspiratory-evoked GABAergic activity but did not change the opioid-induced reduction in spontaneous GABAergic neurotransmission. In contrast, BIMU-8 did not alter GABAergic neurotransmission to cardiac vagal neurons by itself but prevented the fentanyl depression of both spontaneous and inspiratory-elicited GABAergic neurotransmission to cardiac vagal neurons. In the presence of tetrodotoxin, the inhibition of GABAergic inhibitory postsynaptic currents with fentanyl is prevented by coapplication of BIMU-8, indicating that BIMU-8 acts at presynaptic GABAergic terminals to prevent fentanyl-induced depression. These results suggest that activation of 5-hydroxytryptamine receptors, particularly 5-hydroxytryptamine 4&agr; agonists, may be a useful therapeutic approach in preventing opioid-evoked cardiorespiratory depression.

Prenatal nicotine exposure alters the nicotinic receptor subtypes that modulate excitation of parasympathetic cardiac neurons in the nucleus ambiguus from primarily α3β2 and/or α6βX to α3β4

Nicotinic receptors play an essential role in central cardiorespiratory function, however, the types of nicotinic receptors responsible for activating cardiac vagal neurons in the nucleus ambiguus that control heart rate are unknown. This study tests whether α-conotoxin MII and α-conotoxin AuIB sensitive nicotinic receptors are involved in augmentation of glutamatergic neurotransmission and changes in holding current in cardiac vagal neurons, and whether exposure to nicotine in the prenatal period alters these responses. The nicotinic agonist cytisine significantly increased the holding current and amplitude of glutamatergic mEPSCs. In unexposed animals α-conotoxin MII (100 nM) significantly reduced the increase in mEPSC amplitude and change in holding current evoked by cytisine. However, in animals prenatally exposed to nicotine, α-conotoxin MII blunted but did not block the increase in mEPSC amplitude but blocked the increase in holding current evoked by cytisine. In unexposed animals, α-conotoxin AuIB (10 μM) blocked the cytisine evoked increase in mEPSC amplitude and inhibited but did not abolish the increase in holding current. In contrast, in animals exposed to nicotine, α-conotoxin AuIB blunted the increase in mEPSC amplitude, and completely abolished the cytisine evoked increase in holding current. These data demonstrate that the prenatal nicotine exposure alters the nicotinic receptors involved in excitation of cardiac vagal neurons.

Chronic intermittent hypoxia–hypercapnia blunts heart rate responses and alters neurotransmission to cardiac vagal neurons

Chronic intermittent hypoxia–hypercapnia (CIHH) in adult rats evoked hypertension and blunted the heart rate responses to acute hypoxia–hypercapnia (H–H). CIHH induced an increase in spontaneous inhibitory and decreased excitatory neurotransmission to cardiac vagal neurons. CIHH completely abolished acute H–H evoked inhibition of GABAergic while facilitating glycinergic neurotransmission to cardiac vagal neurons of nucleus ambiguus. These changes with CIHH inhibit cardiac vagal neurons to result in diminished cardioprotective vagal activity to the heart, characteristic of obstructive sleep apnoea.

5-HT2 receptor subtypes mediate different long-term changes in gabaergic activity to parasympathetic cardiac vagal neurons in the nucleus ambiguus

Serotonin (5-HT), and in particular 5-HT(2) receptors, play an important role in cardiorespiratory function within the brainstem. In addition, abnormalities in the 5-HT system have been implicated in many cardiorespiratory disorders, including sudden infant death syndrome. However, little is known about the mechanisms of action of 5-HT(2) receptors in altering the activity of parasympathetic cardiac neurons in the brainstem. In this study we examined the effects of activation of different subtypes of 5-HT(2) receptors on spontaneous and respiratory-evoked GABAergic neurotransmission to cardioinhibitory vagal neurons within the nucleus ambiguus as well as rhythmic fictive inspiratory-related activity in rats. A single application of alpha-Me-5-hydroxytryptamine maleate (alpha-Me-5-HT), a 5-HT(2) receptor agonist, did not significantly alter the frequency of spontaneous or respiratory-evoked GABAergic inhibitory postsynaptic currents (IPSCs) in cardiac vagal neurons. However, repetitive successive applications of alpha-Me-5-HT elicited a long-lasting (>/=1 h) decrease in the frequency of spontaneous as well as inspiratory-related GABAergic IPSCs to cardiac vagal neurons. This study demonstrates multiple, but not single applications of the 5-HT(2) receptor agonist alpha-Me-5-HT caused a long-lasting inhibition of both spontaneous and fictive inspiratory-related GABAergic neurotransmission to CVNs, which can be prevented by the 5-HT(2B) receptor antagonist SB204741, but persisted with the 5-HT(2A/2C) receptor antagonist ketanserin. The 5-HT(2) receptor agonist alpha-Me-5-HT also reversibly and transiently excited central fictive inspiratory activity, which was abolished by ketanserin, but was unaffected by the 5-HT(2B) receptor antagonist SB204741.

The Role of 5-HT3 and Other Excitatory Receptors in Central Cardiorespiratory Responses to Hypoxia: Implications for Sudden Infant Death Syndrome

Although brainstem serotonergic (5-HT) systems are involved in the protective responses to hypoxia, abnormalities of 5-HT function are strongly implicated in SIDS, and the neurochemical mechanisms by which 5-HT receptors influence brainstem cardiorespiratory responses to hypoxia remains unclear. This study focuses on the role of excitatory neurotransmission, including 5-HT3 signaling, to cardiac vagal neurons (CVNs) that dominate the control of heart rate. Excitatory synaptic inputs to CVNs, located in the nucleus ambiguus (NA), were recorded simultaneously with respiratory activity in in vitro brainstem slices. During control conditions excitatory inputs to CVNs were blocked by application of NMDA and AMPA/kainate glutamatergic receptor antagonists, whereas the 5-HT3 and purinergic receptor antagonists ondansetron and pyridoxal-phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS), respectively, had no effect. However, during hypoxia ondansetron inhibited excitatory neurotransmission to CVNs. In recovery from hypoxia, spontaneous and respiratory-related excitatory events were blocked by glutamatergic and purinergic receptor blockers, respectively, whereas ondancetron had no effect. These results demonstrate that hypoxia recruits a 5-HT pathway to CVNs that activates 5-HT3 receptors on CVNs to maintain parasympathetic cardiac activity during hypoxia. Exaggeration of this 5-HT neurotransmission could increase the incidence of bradycardia and risk of sudden infant death during hypoxia.

Action of κ and Δ opioid agonists on premotor cardiac vagal neurons in the nucleus ambiguus

Both enkephalin and dynorphin containing fibers are in close proximity to neurons in the nucleus ambiguus, including cardiac vagal neurons. Microinjection of Delta and kappa agonists into the nucleus ambiguus have been shown to evoke decreases in heart rate. Yet little is known about the mechanisms by which Delta and kappa opioid receptors alter the activity of cardiac vagal neurons. This study tests whether kappa and Delta opioid agonists can alter the activity of cardiac vagal neurons by modulating likely opioid targets including voltage gated calcium currents, and both glycinergic and GABA) neurotransmission to cardiac vagal neurons. Cardiac vagal neurons were identified in vitro by a fluorescent tracer and studied using patch clamp techniques. Neither the kappa agonist spiradoline or the Delta agonist [D-Pen(2), D-Pen(5)]enkephalin (DPDPE) modulated the voltage gated calcium currents in cardiac vagal neurons. DPDPE also did not alter either glycinergic or GABAergic synaptic neurotransmission. Spiradoline did not change GABAergic synaptic inputs, but did significantly inhibit glycinergic synaptic inputs to cardiac vagal neurons. At a concentration of 1 microM, spiradoline inhibited the amplitude of glycinergic events, and at a concentration of 5 microM, spiradoline inhibited both glycinergic amplitude and frequency. Spiradoline also inhibited both the amplitude and frequency of glycinergic miniature inhibitory post-synaptic currents, indicating kappa agonists likely act at both presynaptic and postsynaptic sites to inhibit glycinergic neurotransmission to cardiac vagal neurons.