Heather Jameson

Serotonergic modulation of the trigeminocardiac reflex neurotransmission to cardiac vagal neurons in the nucleus ambiguus.

Stimulation of the trigeminal nerve evokes a dramatic decrease in heart rate and blood pressure, and this reflex has generally been termed the trigeminocardiac reflex. A subset of the trigeminocardiac reflex is the diving reflex in which the nasal mucosa is stimulated with water or air-borne chemical irritants. Activation of the diving reflex evokes a pronounced bradycardia, mediated by increased parasympathetic cardiac activity, and is the most powerful autonomic reflex. However, exaggeration of this protective response could be detrimental and has been implicated in Sudden Infant Death Syndrome (SIDS). Despite the importance and strength of the trigeminocardiac reflex, there is little information about the cellular mechanisms and brain stem pathways that constitute this reflex. To address these issues, stimulation of trigeminal afferent fibers and the evoked excitatory postsynaptic currents were recorded in cardiac vagal neurons (CVNs) in an in vitro brain stem slice preparation. This synaptic pathway is robust and activation of the trigeminal pathway often evoked action potentials in CVNs. Application of the serotonin (5-HT) reuptake inhibitor citalopram significantly enhanced these responses. Consistent with the hypothesis this pathway is endogenously modulated by 5-HT receptors the 5-HT1A receptor antagonist, WAY 100635 inhibited, whereas the 5-HT2A/C receptor antagonist, ketanserin facilitated the excitatory neurotransmission to CVNs. The 5-HT1A receptor agonist 8-hydroxy-2-(dipropylamino)tetralin hydrobromide increased, whereas the 5-HT2 receptor agonist, alpha-methylserotonin maleate salt inhibited this reflex pathway. These results indicate stimulation of trigeminal fibers evokes a powerful excitatory and polysynaptic pathway to CVNs, and this pathway is endogenously modulated and differentially enhanced and depressed, by 5-HT1A and 5-HT2 receptors, respectively.

Mapping and Identification of GABAergic Neurons in Transgenic Mice Projecting to Cardiac Vagal Neurons in the Nucleus Ambiguus Using Photo-Uncaging

The neural control of heart rate is determined primarily by the activity of preganglionic parasympathetic cardiac vagal neurons (CVNs) originating in the nucleus ambiguus (NA) in the brain stem. GABAergic inputs to CVNs play an essential role in determining the activity of these neurons including a robust inhibition during each inspiratory burst. The origin of GABAergic innervation has yet to be determined however. A transgenic mouse line expressing green florescent protein (GFP) in GABAergic cells was used in conjunction with caged glutamate to identify both clusters and individual GABAergic neurons that evoke inhibitory GABAergic synaptic responses in CVNs. Transverse slices were taken with CVNs patch-clamped in the whole cell configuration. Sections containing both the pre-Botzinger complex as well as the calamus scriptorius were divided into approximately 90 quadrants, each 200 x 200 microm and were sequentially photostimulated. Inhibitory post synaptic currents (IPSCs) were recorded in CVNs after a 5-ms photostimulation of 50 microM caged glutamate. The four areas that contained GABAergic cells projecting to CVNs were 200 microm medial, 400 microm medial, 200 microm ventral, and 1,200 microm dorsal and 1,000 microm medial to patched CVNs. Once foci of GABAergic cells projecting to CVNs were determined, photostimulation of individual GABAergic neurons was conducted. The results from this study suggest that GABAergic cells located in four specific areas project to CVNs, and that these cells can be individually identified and stimulated using photouncaging to recruit GABAergic neurotransmission to CVNs.

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.

Purinergic P2X Receptors Mediate Excitatory Transmission to Cardiac Vagal Neurons in the Nucleus Ambiguus After Hypoxia

Challenges such as hypoxia elicit a powerful response from both the central cardiovascular and respiratory neuronal networks. Recent work indicates that purinergic neurotransmission in the brain stem is an important modulator of central respiratory network responses to hypoxia. This study tests whether alterations in purinergic neurotransmission extend beyond respiratory responses to hypoxia and also mediates respiratory inputs to cardiac vagal neurons. To examine central cardiorespiratory responses to hypoxia, we used an in vitro medullary slice that allows simultaneous examination of rhythmic respiratory-related activity and synaptic neurotransmission to cardioinhibitory vagal neurons. Here we show that P2X receptor activation mediates respiratory-related excitatory neurotransmission to parasympathetic cardiac vagal neurons, the dominant control of heart rate. These data demonstrate a critical functional role for adenosine 5′-triphosphate–mediated purinergic signaling in facilitating respiratory-related excitatory neurotransmission to cardiac vagal neurons after hypoxia.

Visualization of oxytocin release that mediates paired pulse facilitation in hypothalamic pathways to brainstem autonomic neurons

Recent work has shown that oxytocin is involved in more than lactation and uterine contraction. The paraventricular nucleus of the hypothalamus (PVN) contains neuroendocrine neurons that control the release of hormones, including vasopressin and oxytocin. Other populations of PVN neurons do not release hormones, but rather project to and release neurotransmitters onto other neurons in the CNS involved in fluid retention, thermoregulation, sexual behavior and responses to stress. Activation of oxytocin receptors can be cardioprotective and reduces the adverse cardiovascular consequences of anxiety and stress, yet how oxytocin can affect heart rate and cardiac function is unknown. While anatomical work has shown the presence of peptides, including oxytocin, in the projections from the PVN to parasympathetic nuclei, electrophysiological studies to date have only demonstrated release of glutamate and activation of fast ligand gated receptors in these pathways. In this study, using rats, we directly show, using sniffer CHO cells that express oxytocin receptors and the Ca2+ indicator R-GECO, that optogenetic activation of channelrhodopsin-2 (ChR2) expressing PVN fibers in the brainstem activates oxytocin receptors in the dorsomotor nucleus of the vagus (DMNV). We also demonstrate that while a single photoactivation of PVN terminals only activates glutamatergic receptors in brainstem cardiac vagal neurons (CVNs), neurons that dominate the neural control of heart rate, both the paired pulse facilitation, and sustained enhancement of glutamate release in this pathway is mediated by activation of oxytocin receptors. Our results provide direct evidence that a pathway from the PVN likely releases oxytocin and enhances short-term plasticity of this critical autonomic connection.

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.

Oxytocin neuron activation prevents hypertension that occurs with chronic intermittent hypoxia/hypercapnia in rats.

Hypertension is a common outcome associated with obstructive sleep apnea (OSA), a prevalent yet poorly treated cardiovascular disease. Recent studies showed oxytocin (OXT), released from hypothalamic paraventricular nucleus (PVN) neurons, activates cardiac vagal neurons in the dorsal motor nucleus of the vagus (DMNX) and may blunt cardiovascular responses to stress. This study tests whether the release of OXT from PVN fibers in the DMNX is diminished with chronic intermittent hypoxia-hypercapnia (CIH/H) exposure, an animal model of OSA, and whether activation of PVN OXT neurons restores OXT release in the DMNX and prevents the hypertension resulting from CIH/H. To assess OXT release from PVN fibers, Chinese hamster ovarian (CHO) cells were engineered to be highly sensitive to OXT by stable expression of the human recombinant OXT receptor and the calcium indicator R-GECO1. PVN fibers in the DMNX were selectively photoactivated in vitro by expression of channelrhodopsin. The release of OXT onto CHO cells in the DMNX was blunted in rats exposed to 21 days of CIH/H. Chronic activation of PVN OXT neurons in vivo, using designer receptors exclusively activated by designer drugs, restored the release of OXT onto CHO cells in the DMNX. Chronic PVN OXT neuron activation in vivo also prevented the hypertension that occurred in conscious unrestrained telemetry-equipped sham rats exposed to 3 wk of CIH/H. These results demonstrate that chronic activation of OXT neurons restores the release of OXT from PVN fibers in the DMNX and prevents the hypertension that occurs with 3 wk of CIH/H exposure.

Prenatal nicotine exposure enhances the trigeminocardiac reflex via serotonin receptor facilitation in brainstem pathways

In this study we used a rat model for prenatal nicotine exposure to test whether clinically relevant concentrations of brain nicotine and cotinine are passed from dams exposed to nicotine to her pups, whether this changes the trigeminocardiac reflex (TCR), and whether serotonergic function in the TCR brainstem circuitry is altered. Pregnant Sprague-Dawley dams were exposed to 6 mg·kg(-1)·day(-1) of nicotine via osmotic minipumps for the duration of pregnancy. Following birth dams and pups were killed, blood was collected, and brain nicotine and cotinine levels were measured. A separate group of prenatal nicotine-exposed pups was used for electrophysiological recordings. A horizontal brainstem slice was obtained by carefully preserving the trigeminal nerve with fluorescent identification of cardiac vagal neurons (CVNs) in the nucleus ambiguus. Stimulation of the trigeminal nerve evoked excitatory postsynaptic current in CVNs. Our data demonstrate that prenatal nicotine exposure significantly exaggerates both the TCR-evoked changes in heart rate in conscious unrestrained pups, and the excitatory neurotransmission to CVNs upon trigeminal afferent nerve stimulation within this brainstem reflex circuit. Application of the 5-HT1A receptor antagonist WAY 100635 (100 μM) and 5-HT2A/C receptor antagonist ketanserin (10 μM)significantly decreased neurotransmission, indicating an increased facilitation of 5-HT function in prenatal nicotine-exposed animals. Prenatal nicotine exposure enhances activation of 5-HT receptors and exaggerates the trigeminocardiac reflex.

The Lateral Paragigantocellular Nucleus Modulates Parasympathetic Cardiac Neurons: A Mechanism for Rapid Eye Movement Sleep-Dependent Changes in Heart Rate

Rapid eye movement (REM) sleep is generally associated with a withdrawal of parasympathetic activity and heart rate increases; however, episodic vagally mediated heart rate decelerations also occur during REM sleep. This alternating pattern of autonomic activation provides a physiological basis for REM sleep-induced cardiac arrhythmias. Medullary neurons within the lateral paragigantocellular nucleus (LPGi) are thought to be active after REM sleep recovery and play a role in REM sleep control. In proximity to the LPGi are parasympathetic cardiac vagal neurons (CVNs) within the nucleus ambiguus (NA), which are critical for controlling heart rate. This study examined brain stem pathways that may mediate REM sleep-related reductions in parasympathetic cardiac activity. Electrical stimulation of the LPGi evoked inhibitory GABAergic postsynaptic currents in CVNs in an in vitro brain stem slice preparation in rats. Because brain stem cholinergic mechanisms are involved in REM sleep regulation, we also studied the role of nicotinic neurotransmission in modulation of GABAergic pathway from the LGPi to CVNs. Application of nicotine diminished the GABAergic responses evoked by electrical stimulation. This inhibitory effect of nicotine was prevented by the alpha7 nicotinic receptor antagonist alpha-bungarotoxin. Moreover, hypoxia/hypercapnia (H/H) diminished LPGi-evoked GABAergic current in CVNs, and this inhibitory effect was also prevented by alpha-bungarotoxin. In conclusion, stimulation of the LPGi evokes an inhibitory pathway to CVNs, which may constitute a mechanism for the reduced parasympathetic cardiac activity and increase in heart rate during REM sleep. Inhibition of this pathway by nicotinic receptor activation and H/H may play a role in REM sleep-related and apnea-associated bradyarrhythmias.