Electrophysiology and 3D-imaging reveal properties of human intracardiac neurons and increased excitability with atrial fibrillation.

Altered autonomic input to the heart plays a major role in atrial fibrillation (AF). Autonomic neurons termed ganglionated plexi (GP) are clustered on the heart surface to provide the last point of neural control of cardiac function. To date the properties of GP neurons in humans are unknown. Here we have addressed this knowledge gap in human GP neuron structure and physiology in patients with and without AF. Human right atrial GP neurons embedded in epicardial adipose tissue were excised during open heart surgery performed on both non-AF and AF patients and then characterised physiologically by whole cell patch clamp techniques. Structural analysis was also performed after fixation at both the single cell and at the entire GP levels via three-dimensional confocal imaging. Human GP neurons were found to exhibit unique properties and structural complexity with branched neurite outgrowth. Significant differences in excitability were revealed between AF and non-AF GP neurons as measured by lower current to induce action potential firing, a reduced occurrence of low action potential firing rates, decreased accommodation and increased synaptic density. Visualisation of entire GPs showed almost all neurons are cholinergic with a small proportion of noradrenergic and dual phenotype neurons. Phenotypic distribution differences occurred with AF including decreased cholinergic and dual phenotype neurons, and increased noradrenergic neurons. These data show both functional and structural differences occur between GP neurons from patients with and without AF, highlighting that cellular plasticity occurs in neural input to the heart that could alter autonomic influence on atrial function. KEY POINTS: The autonomic nervous system plays a critical role in regulating heart rhythm and the initiation of AF; however, the structural and functional properties of human autonomic neurons in the autonomic ganglionated plexi (GP) remain unknown. Here we perform the first whole cell patch clamp electrophysiological and large tissue confocal imaging analysis of these neurons from patients with and without AF. Our data show human GP neurons are functionally and structurally complex. Measurements of action potential kinetics show higher excitability in GP neurons from AF patients as measured by lower current to induce action potential firing, reduced low firing action potential rates, and decreased action potential accommodation. Confocal imaging shows increased synaptic density and noradrenergic phenotypes in patients with AF. Both functional and structural differences occur in GP neurons from patients with AF that could alter autonomic influence on atrial rhythm.

Intrinsic cardiac ganglia and acetylcholine are important in the mechanism of ischaemic preconditioning.

This study aimed to investigate the role of the intrinsic cardiac nervous system in the mechanism of classical myocardial ischaemic preconditioning (IPC). Isolated perfused rat hearts were subjected to 35-min regional ischaemia and 60-min reperfusion. IPC was induced as three cycles of 5-min global ischaemia-reperfusion, and provided significant reduction in infarct size (IS/AAR = 14 ± 2% vs control IS/AAR = 48 ± 3%, p < 0.05). Treatment with the ganglionic antagonist, hexamethonium (50 µM), blocked IPC protection (IS/AAR = 37 ± 7%, p < 0.05 vs IPC). Moreover, the muscarinic antagonist, atropine (100 nM), also abrogated IPC-mediated protection (IS/AAR = 40 ± 3%, p < 0.05 vs IPC). This indicates that intrinsic cardiac ganglia remain intact in the Langendorff preparation and are important in the mechanism of IPC. In a second group of experiments, coronary effluent collected following IPC, from ex vivo perfused rat hearts, provided significant cardioprotection when perfused through a naïve isolated rat heart prior to induction of regional ischaemia-reperfusion injury (IRI) (IS/ARR = 19 ± 2, p < 0.05 vs control effluent). This protection was also abrogated by treating the naïve heart with hexamethonium, indicating the humoral trigger of IPC induces protection via an intrinsic neuronal mechanism (IS/AAR = 46 ± 5%, p < 0.05 vs IPC effluent). In addition, a large release in ACh was observed in coronary effluent was observed following IPC (IPCeff = 0.36 ± 0.03 µM vs C eff = 0.04 ± 0.04 µM, n = 4, p < 0.001). Interestingly, however, IPC effluent was not able to significantly protect isolated cardiomyocytes from simulated ischaemia-reperfusion injury (cell death = 45 ± 6%, p = 0.09 vs control effluent). In conclusion, IPC involves activation of the intrinsic cardiac nervous system, leading to release of ACh in the ventricles and induction of protection via activation of muscarinic receptors.

Vagal stimulation targets select populations of intrinsic cardiac neurons to control neurally induced atrial fibrillation.

Mediastinal nerve stimulation (MNS) reproducibly evokes atrial fibrillation (AF) by excessive and heterogeneous activation of intrinsic cardiac (IC) neurons. This study evaluated whether preemptive vagus nerve stimulation (VNS) impacts MNS-induced evoked changes in IC neural network activity to thereby alter susceptibility to AF. IC neuronal activity in the right atrial ganglionated plexus was directly recorded in anesthetized canines (n = 8) using a linear microelectrode array concomitant with right atrial electrical activity in response to: 1) epicardial touch or great vessel occlusion vs. 2) stellate or vagal stimulation. From these stressors, post hoc analysis (based on the Skellam distribution) defined IC neurons so recorded as afferent, efferent, or convergent (afferent and efferent inputs) local circuit neurons (LCN). The capacity of right-sided MNS to modify IC activity in the induction of AF was determined before and after preemptive right (RCV)- vs. left (LCV)-sided VNS (15 Hz, 500 µs; 1.2× bradycardia threshold). Neuronal (n = 89) activity at baseline (0.11 ± 0.29 Hz) increased during MNS-induced AF (0.51 ± 1.30 Hz; P < 0.001). Convergent LCNs were preferentially activated by MNS. Preemptive RCV reduced MNS-induced changes in LCN activity (by 70%) while mitigating MNS-induced AF (by 75%). Preemptive LCV reduced LCN activity by 60% while mitigating AF potential by 40%. IC neuronal synchrony increased during neurally induced AF, a local neural network response mitigated by preemptive VNS. These antiarrhythmic effects persisted post-VNS for, on average, 26 min. In conclusion, VNS preferentially targets convergent LCNs and their interactive coherence to mitigate the potential for neurally induced AF. The antiarrhythmic properties imposed by VNS exhibit memory.

Vagus nerve stimulation mitigates intrinsic cardiac neuronal remodeling and cardiac hypertrophy induced by chronic pressure overload in guinea pig.

Our objective was to determine whether chronic vagus nerve stimulation (VNS) mitigates pressure overload (PO)-induced remodeling of the cardioneural interface. Guinea pigs (n = 48) were randomized to right or left cervical vagus (RCV or LCV) implant. After 2 wk, chronic left ventricular PO was induced by partial (15-20%) aortic constriction. Of the 31 animals surviving PO induction, 10 were randomized to RCV VNS, 9 to LCV VNS, and 12 to sham VNS. VNS was delivered at 20 Hz and 1.14 ± 0.03 mA at a 22% duty cycle. VNS commenced 10 days after PO induction and was maintained for 40 days. Time-matched controls (n = 9) were evaluated concurrently. Echocardiograms were obtained before and 50 days after PO. At termination, intracellular current-clamp recordings of intrinsic cardiac (IC) neurons were studied in vitro to determine effects of therapy on soma characteristics. Ventricular cardiomyocyte sizes were assessed with histology along with immunoblot analysis of selected proteins in myocardial tissue extracts. In sham-treated animals, PO increased cardiac output (34%, P < 0.004), as well as systolic (114%, P < 0.04) and diastolic (49%, P < 0.002) left ventricular volumes, a hemodynamic response prevented by VNS. PO-induced enhancements of IC synaptic efficacy and muscarinic sensitivity of IC neurons were mitigated by chronic VNS. Increased myocyte size, which doubled in PO (P < 0.05), was mitigated by RCV. PO hypertrophic myocardium displayed decreased glycogen synthase (GS) protein levels and accumulation of the phosphorylated (inactive) form of GS. These PO-induced changes in GS were moderated by left VNS. Chronic VNS targets IC neurons accompanying PO to obtund associated adverse cardiomyocyte remodeling.

Central-peripheral neural network interactions evoked by vagus nerve stimulation: functional consequences on control of cardiac function.

Using vagus nerve stimulation (VNS), we sought to determine the contribution of vagal afferents to efferent control of cardiac function. In anesthetized dogs, the right and left cervical vagosympathetic trunks were stimulated in the intact state, following ipsilateral or contralateral vagus nerve transection (VNTx), and then following bilateral VNTx. Stimulations were performed at currents from 0.25 to 4.0 mA, frequencies from 2 to 30 Hz, and a 500-µs pulse width. Right or left VNS evoked significantly greater current- and frequency-dependent suppression of chronotropic, inotropic, and lusitropic function subsequent to sequential VNTx. Bradycardia threshold was defined as the current first required for a 5% decrease in heart rate. The threshold for the right vs. left vagus-induced bradycardia in the intact state (2.91 ± 0.18 and 3.47 ± 0.20 mA, respectively) decreased significantly with right VNTx (1.69 ± 0.17 mA for right and 3.04 ± 0.27 mA for left) and decreased further following bilateral VNTx (1.29 ± 0.16 mA for right and 1.74 ± 0.19 mA for left). Similar effects were observed following left VNTx. The thresholds for afferent-mediated effects on cardiac parameters were 0.62 ± 0.04 and 0.65 ± 0.06 mA with right and left VNS, respectively, and were reflected primarily as augmentation. Afferent-mediated tachycardias were maintained following ß-blockade but were eliminated by VNTx. The increased effectiveness and decrease in bradycardia threshold with sequential VNTx suggest that 1) vagal afferents inhibit centrally mediated parasympathetic efferent outflow and 2) the ipsilateral and contralateral vagi exert a substantial buffering capacity. The intact threshold reflects the interaction between multiple levels of the cardiac neural hierarchy.

Vagus nerve stimulation mitigates intrinsic cardiac neuronal and adverse myocyte remodeling postmyocardial infarction.

This paper aims to determine whether chronic vagus nerve stimulation (VNS) mitigates myocardial infarction (MI)-induced remodeling of the intrinsic cardiac nervous system (ICNS), along with the cardiac tissue it regulates. Guinea pigs underwent VNS implantation on the right cervical vagus. Two weeks later, MI was produced by ligating the ventral descending coronary artery. VNS stimulation started 7 days post-MI (20 Hz, 0.9 ± 0.2 mA, 14 s on, 48 s off; VNS-MI, n = 7) and was compared with time-matched MI animals with sham VNS (MI n = 7) vs. untreated controls (n = 8). Echocardiograms were performed before and at 90 days post-MI. At termination, IC neuronal intracellular voltage recordings were obtained from whole-mount neuronal plexuses. MI increased left ventricular end systolic volume (LVESV) 30% (P = 0.027) and reduced LV ejection fraction (LVEF) 6.5% (P < 0.001) at 90 days post-MI compared with baseline. In the VNS-MI group, LVESV and LVEF did not differ from baseline. IC neurons showed depolarization of resting membrane potentials and increased input resistance in MI compared with VNS-MI and sham controls (P < 0.05). Neuronal excitability and sensitivity to norepinephrine increased in MI and VNS-MI groups compared with controls (P < 0.05). Synaptic efficacy, as determined by evoked responses to stimulating input axons, was reduced in VNS-MI compared with MI or controls (P < 0.05). VNS induced changes in myocytes, consistent with enhanced glycogenolysis, and blunted the MI-induced increase in the proapoptotic Bcl-2-associated X protein (P < 0.05). VNS mitigates MI-induced remodeling of the ICNS, correspondingly preserving ventricular function via both neural and cardiomyocyte-dependent actions.

Morphological pattern of intrinsic nerve plexus distributed on the rabbit heart and interatrial septum.

Although the rabbit is routinely used as the animal model of choice to investigate cardiac electrophysiology, the neuroanatomy of the rabbit heart is not well documented. The aim of this study was to examine the topography of the intrinsic nerve plexus located on the rabbit heart surface and interatrial septum stained histochemically for acetylcholinesterase using pressure-distended whole hearts and whole-mount preparations from 33 Californian rabbits. Mediastinal cardiac nerves entered the venous part of the heart along the root of the right cranial vein (superior caval vein) and at the bifurcation of the pulmonary trunk. The accessing nerves of the venous part of the heart passed into the nerve plexus of heart hilum at the heart base. Nerves approaching the heart extended epicardially and innervated the atria, interatrial septum and ventricles by five nerve subplexuses, i.e. left and middle dorsal, dorsal right atrial, ventral right and left atrial subplexuses. Numerous nerves accessed the arterial part of the arterial part of the heart hilum between the aorta and pulmonary trunk, and distributed onto ventricles by the left and right coronary subplexuses. Clusters of intrinsic cardiac neurons were concentrated at the heart base at the roots of pulmonary veins with some positioned on the infundibulum. The mean number of intrinsic neurons in the rabbit heart is not significantly affected by aging: 2200 ± 262 (range 1517-2788; aged) vs. 2118 ± 108 (range 1513-2822; juvenile). In conclusion, despite anatomic differences in the distribution of intrinsic cardiac neurons and the presence of well-developed nerve plexus within the heart hilum, the topography of all seven subplexuses of the intrinsic nerve plexus in rabbit heart corresponds rather well to other mammalian species, including humans.

The Intrinsic Cardiac Nervous System: From Pathophysiology to Therapeutic Implications.

The cardiac autonomic nervous system (CANS) plays a pivotal role in cardiac homeostasis as well as in cardiac pathology. The first level of cardiac autonomic control, the intrinsic cardiac nervous system (ICNS), is located within the epicardial fat pads and is physically organized in ganglionated plexi (GPs). The ICNS system does not only contain parasympathetic cardiac efferent neurons, as long believed, but also afferent neurons and local circuit neurons. Thanks to its high degree of connectivity, combined with neuronal plasticity and memory capacity, the ICNS allows for a beat-to-beat control of all cardiac functions and responses as well as integration with extracardiac and higher centers for longer-term cardiovascular reflexes. The present review provides a detailed overview of the current knowledge of the bidirectional connection between the ICNS and the most studied cardiac pathologies/conditions (myocardial infarction, heart failure, arrhythmias and heart transplant) and the potential therapeutic implications. Indeed, GP modulation with efferent activity inhibition, differently achieved, has been studied for atrial fibrillation and functional bradyarrhythmias, while GP modulation with efferent activity stimulation has been evaluated for myocardial infarction, heart failure and ventricular arrhythmias. Electrical therapy has the unique potential to allow for both kinds of ICNS modulation while preserving the anatomical integrity of the system.

Neural control hierarchy of the heart has not evolved to deal with myocardial ischemia.

The consequences of myocardial ischemia are examined from the standpoint of the neural control system of the heart, a hierarchy of three neuronal centers residing in central command, intrathoracic ganglia, and intrinsic cardiac ganglia. The basis of the investigation is the premise that while this hierarchical control system has evolved to deal with "normal" physiological circumstances, its response in the event of myocardial ischemia is unpredictable because the singular circumstances of this event are as yet not part of its evolutionary repertoire. The results indicate that the harmonious relationship between the three levels of control breaks down, because of a conflict between the priorities that they have evolved to deal with. Essentially, while the main priority in central command is blood demand, the priority at the intrathoracic and cardiac levels is heart rate. As a result of this breakdown, heart rate becomes less predictable and therefore less reliable as a diagnostic guide as to the traumatic state of the heart, which it is commonly used as such following an ischemic event. On the basis of these results it is proposed that under the singular conditions of myocardial ischemia a determination of neural control indexes in addition to cardiovascular indexes has the potential of enhancing clinical outcome.

Acute Modulation of Left Ventricular Control by Selective Intracardiac Sympathetic Denervation.

BACKGROUND: The sympathetic nervous system plays an integral role in cardiac physiology. Nerve fibers innervating the left ventricle are amenable to transvenous catheter stimulation along the coronary sinus (CS). OBJECTIVES: The aim of the present study was to modulate left ventricular control by selective intracardiac sympathetic denervation. METHODS: First, the impact of epicardial CS ablation on cardiac electrophysiology was studied in a Langendorff model of decentralized murine hearts (n = 10 each, ablation and control groups). Second, the impact of transvenous, anatomically driven axotomy by catheter-based radiofrequency ablation via the CS was evaluated in healthy sheep (n = 8) before and during stellate ganglion stimulation. RESULTS: CS ablation prolonged epicardial ventricular refractory period without (41.8 ± 8.4 ms vs 53.0 ± 13.5 ms; P = 0.049) and with ß1-2-adrenergic receptor blockade (47.8 ± 7.8 ms vs 73.1 ± 13.2 ms; P < 0.001) in mice. Supported by neuromorphological studies illustrating a circumferential CS neural network, intracardiac axotomy by catheter ablation via the CS in healthy sheep diminished the blood pressure increase during stellate ganglion stimulation (Δ systolic blood pressure 21.9 ± 10.9 mm Hg vs 10.5 ± 12.0 mm Hg; P = 0.023; Δ diastolic blood pressure 9.0 ± 5.5 mm Hg vs 3.0 ± 3.5 mm Hg; P = 0.039). CONCLUSIONS: Transvenous, anatomically driven axotomy targeting nerve fibers along the CS enables acute modulation of left ventricular control by selective intracardiac sympathetic denervation.

Catecholaminergic axon innervation and morphology in flat-mounts of atria and ventricles of mice.

Sympathetic efferent axons regulate cardiac functions. However, the topographical distribution and morphology of cardiac sympathetic efferent axons remain insufficiently characterized due to the technical challenges involved in immunohistochemical labeling of the thick walls of the whole heart. In this study, flat-mounts of the left and right atria and ventricles of FVB mice were immunolabeled for tyrosine hydroxylase (TH), a marker of sympathetic nerves. Atrial and ventricular flat-mounts were scanned using a confocal microscope to construct montages. We found (1) In the atria: A few large TH-immunoreactive (IR) axon bundles entered both atria, branched into small bundles and then single axons that eventually formed very dense terminal networks in the epicardium, myocardium and inlet regions of great vessels to the atria. Varicose TH-IR axons formed close contact with cardiomyocytes, vessels, and adipocytes. Multiple intrinsic cardiac ganglia (ICG) were identified in the epicardium of both atria, and a subpopulation of the neurons in the ICG were TH-IR. Most TH-IR axons in bundles traveled through ICG before forming dense varicose terminal networks in cardiomyocytes. We did not observe varicose TH-IR terminals encircling ICG neurons. (2) In the left and right ventricles and interventricular septum: TH-IR axons formed dense terminal networks in the epicardium, myocardium, and vasculature. Collectively, TH labeling is achievable in flat-mounts of thick cardiac walls, enabling detailed mapping of catecholaminergic axons and terminal structures in the whole heart at single-cell/axon/varicosity scale. This approach provides a foundation for future quantification of the topographical organization of the cardiac sympathetic innervation in different pathological conditions.

Recording plasticity in neuronal activity in the rodent intrinsic cardiac nervous system using calcium imaging techniques.

The intrinsic cardiac nervous system (ICNS) is composed of interconnected clusters of neurons called ganglionated plexi (GP) which play a major role in controlling heart rate and rhythm. The function of these neurons is particularly important due to their involvement in cardiac arrhythmias such as atrial fibrillation (AF), and previous work has shown that plasticity in GP neural networks could underpin aberrant activity patterns that drive AF. As research in this field increases, developing new techniques to visualize the complex interactions and plasticity in this GP network is essential. In this study we have developed a calcium imaging method enabling the simultaneous recording of plasticity in neuronal activity from multiple neurons in intact atrial GP networks. Calcium imaging was performed with Cal-520 AM labeling in aged spontaneously hypertensive rats (SHRs), which display both spontaneous and induced AF, and age-matched Wistar Kyoto (WKY) controls to determine the relationship between chronic hypertension, arrhythmia and GP calcium dynamics. Our data show that SHR GPs have significantly larger calcium responses to cholinergic stimulation compared to WKY controls, as determined by both higher amplitude and longer duration calcium responses. Responses were significantly but not fully blocked by hexamethonium, indicating multiple cholinergic receptor subtypes are involved in the calcium response. Given that SHRs are susceptible to cardiac arrhythmias, our data provide evidence for a potential link between arrhythmia and plasticity in calcium dynamics that occur not only in cardiomyocytes but also in the GP neurons of the heart.

Closed-loop modeling of central and intrinsic cardiac nervous system circuits underlying cardiovascular control.

The baroreflex is a multi-input, multi-output control physiological system that regulates blood pressure by modulating nerve activity between the brainstem and the heart. Existing computational models of the baroreflex do not explictly incorporate the intrinsic cardiac nervous system (ICN), which mediates central control of the heart function. We developed a computational model of closed-loop cardiovascular control by integrating a network representation of the ICN within central control reflex circuits. We examined central and local contributions to the control of heart rate, ventricular functions, and respiratory sinus arrhythmia (RSA). Our simulations match the experimentally observed relationship between RSA and lung tidal volume. Our simulations predicted the relative contributions of the sensory and the motor neuron pathways to the experimentally observed changes in the heart rate. Our closed-loop cardiovascular control model is primed for evaluating bioelectronic interventions to treat heart failure and renormalize cardiovascular physiology.

Human apoCIII transgenic mice with epicardial adipose tissue inflammation and PRESERVATION of the cardiac plexus.

Hypertriglyceridemia is a result of the increase in the serum levels of lipoproteins, which are responsible for the transport of triglycerides and can be caused by genetic and/or metabolic factors. Animal models which either express or lack genes related to changes in the lipoproteins profile are useful to understand lipid metabolism. Apolipoprotein CIII (apoCIII) is an important modulator of hepatic production and peripheral removal of triglycerides. Mice that overexpress the apoCIII gene become hypertriglyceridemic, showing high concentrations of free fatty acids in the blood. Since hypertriglyceridemia is related to atherosclerosis, and the latter refers to cardiac alterations, this study aimed at evaluating the morphological, morphometric and quantitative profiles of the cardiac plexus, as well as the morphometric and histopathological aspects of the epicardial adipose tissue in human apoCIII transgenic mice. Therefore, 8-12-month-old male C57BL/6 mice that overexpressed human apoCIII (CIII) and their respective controls were used. Our results showed that overexpression of human apoCIII did not modify morphological or quantitative parameters of cardiac plexus neurons; however, age increased both, the area and the number of such cells. Furthermore, there was a direct correlation of this dyslipidemia to the thickening of periganglionar type 1 collagens. On the other hand, this overexpression caused epicardial adipose tissue inflammation and an increase in the area of the adipocytes, thus, favoring the recruitment of inflammatory cells in this tissue. In conclusion, this overexpression is harmful since it is related to an increase in cardiac adiposity, as well as to a predisposition to an inflammatory environment in the epicardial fat and to the incidence of cardiovascular diseases.

Ivabradine modulates the autonomic nervous system by affecting the "little brain" of the heart: A hypothesis.

Ivabradine decreases heart rate by selective inhibition of the If current in the sinoatrial node. Ivabradine is declared to have no direct effect on the autonomic nervous system (ANS). However, there are some data suggesting an (at least indirect) effect of ivabradine on the ANS. The pathomechanism behind is unclear. Based on the complex of plexuses and ganglia in the heart, the existence of the intrinsic cardiac nervous system (ICNS), also known as the "little brain" of the heart, has been suggested. The ICNS is supposed to process information on the cardiac milieu and provide the central nervous system with these data. We put forward a hypothesis that part of ivabradine's protective effects might reside in the modulation of the ANS by affecting the ICNS. Setting a new autonomic balance by ivabradine might be of benefit in the treatment of autonomic dysfunction-related pathologies.

Influence of cardiac nerve status on cardiovascular regulation and cardioprotection.

Neural elements of the intrinsic cardiac nervous system transduce sensory inputs from the heart, blood vessels and other organs to ensure adequate cardiac function on a beat-to-beat basis. This inter-organ crosstalk is critical for normal function of the heart and other organs; derangements within the nervous system hierarchy contribute to pathogenesis of organ dysfunction. The role of intact cardiac nerves in development of, as well as protection against, ischemic injury is of current interest since it may involve recruitment of intrinsic cardiac ganglia. For instance, ischemic conditioning, a novel protection strategy against organ injury, and in particular remote conditioning, is likely mediated by activation of neural pathways or by endogenous cytoprotective blood-borne substances that stimulate different signalling pathways. This discovery reinforces the concept that inter-organ communication, and maintenance thereof, is key. As such, greater understanding of mechanisms and elucidation of treatment strategies is imperative to improve clinical outcomes particularly in patients with comorbidities. For instance, autonomic imbalance between sympathetic and parasympathetic nervous system regulation can initiate cardiovascular autonomic neuropathy that compromises cardiac stability and function. Neuromodulation therapies that directly target the intrinsic cardiac nervous system or other elements of the nervous system hierarchy are currently being investigated for treatment of different maladies in animal and human studies.

Characterization of the intrinsic cardiac nervous system.

Heart disease is the number one cause of mortality in the developed world and it is well recognised that neural mechanisms are important in pathology. As well as peripheral autonomic nerves, there is a rich intrinsic innervation of the heart that includes cardiac ganglia, collectively termed ganglionic plexuses (GP). Understanding the role that the intrinsic cardiac nervous system (ICNS) play in controlling cardiac function and how it interacts with information between central command centers and its integration with sensory information from the myocardium could prove crucial for prophylactic and corrective treatments of heart disease. This article in the timely and important special issue on central and peripheral nervous control of the heart in Autonomic Neuroscience; Basic and Clinical will focus on the anatomical and physiological characteristics that define the ICNS.

Pathological effects of chronic myocardial infarction on peripheral neurons mediating cardiac neurotransmission.

OBJECTIVE: To determine whether chronic myocardial infarction (MI) induces structural and neurochemical changes in neurons within afferent and efferent ganglia mediating cardiac neurotransmission. METHODS: Neuronal somata in i) right atrial (RAGP) and ii) ventral interventricular ganglionated plexi (VIVGP), iii) stellate ganglia (SG) and iv) T1-2 dorsal root ganglia (DRG) bilaterally derived from normal (n=8) vs. chronic MI (n=8) porcine subjects were studied. We examined whether the morphology and neuronal nitric oxide synthase (nNOS) expression in soma of RAGP, VIVGP, DRG and SG neurons were altered as a consequence of chronic MI. In DRG, we also examined immunoreactivity of calcitonin gene related peptide (CGRP), a marker of afferent neurons. Chronic MI increased neuronal size and nNOS immunoreactivity in VIVGP (but not RAGP), as well as in the SG bilaterally. Across these ganglia, the increase in neuronal size was more pronounced in nNOS immunoreactive neurons. In the DRG, chronic MI also caused neuronal enlargement, and increased CGRP immunoreactivity. Further, DRG neurons expressing both nNOS and CGRP were increased in MI animals compared to controls, and represented a shift from double negative neurons. CONCLUSIONS: Chronic MI impacts diverse elements within the peripheral cardiac neuraxis. That chronic MI imposes such widespread, diverse remodeling of the peripheral cardiac neuraxis must be taken into consideration when contemplating neuronal regulation of the ischemic heart.

Intrinsic and extrinsic innervation of the heart in zebrafish (Danio rerio).

In the vertebrate heart the intracardiac nervous system is the final common pathway for autonomic control of cardiac output, but the neuroanatomy of this system is not well understood. In this study we investigated the innervation of the heart in a model vertebrate, the zebrafish. We used antibodies against acetylated tubulin, human neuronal protein C/D, choline acetyltransferase, tyrosine hydroxylase, neuronal nitric oxide synthase, and vasoactive intestinal polypeptide to visualize neural elements and their neurotransmitter content. Most neurons were located at the venous pole in a plexus around the sinoatrial valve; mean total number of cells was 197 ± 23, and 92% were choline acetyltransferase positive, implying a cholinergic role. The plexus contained cholinergic, adrenergic, and nitrergic axons and vasoactive intestinal polypeptide-positive terminals, some innervating somata. Putative pacemaker cells near the plexus showed immunoreactivity for hyperpolarization-activated cyclic nucleotide-gated channel 4 (HCN4) and the transcription factor Islet-1 (Isl1). The neurotracer neurobiotin showed that extrinsic axons from the left and right vagosympathetic trunks innervated the sinoatrial plexus proximal to their entry into the heart; some extrinsic axons from each trunk also projected into the medial dorsal plexus region. Extrinsic axons also innervated the atrial and ventricular walls. An extracardiac nerve trunk innervated the bulbus arteriosus and entered the arterial pole of the heart to innervate the proximal ventricle. We have shown that the intracardiac nervous system in the zebrafish is anatomically and neurochemically complex, providing a substrate for autonomic control of cardiac effectors in all chambers.

Dynamic remodeling of the guinea pig intrinsic cardiac plexus induced by chronic myocardial infarction.

Myocardial infarction (MI) is associated with remodeling of the heart and neurohumoral control systems. The objective of this study was to define time-dependent changes in intrinsic cardiac (IC) neuronal excitability, synaptic efficacy, and neurochemical modulation following MI. MI was produced in guinea pigs by ligation of the coronary artery and associated vein on the dorsal surface of the heart. Animals were recovered for 4, 7, 14, or 50 days. Intracellular voltage recordings were obtained in whole mounts of the cardiac neuronal plexus to determine passive and active neuronal properties of IC neurons. Immunohistochemical analysis demonstrated an immediate and persistent increase in the percentage of IC neurons immunoreactive for neuronal nitric oxide synthase. Examination of individual neuronal properties demonstrated that after hyperpolarizing potentials were significantly decreased in both amplitude and time course of recovery at 7 days post-MI. These parameters returned to control values by 50 days post-MI. Synaptic efficacy, as determined by the stimulation of axonal inputs, was enhanced at 7 days post-MI only. Neuronal excitability in absence of agonist challenge was unchanged following MI. Norepinephrine increased IC excitability to intracellular current injections, a response that was augmented post-MI. Angiotensin II potentiation of norepinephrine and bethanechol-induced excitability, evident in controls, was abolished post-MI. This study demonstrates that MI induces both persistent and transient changes in IC neuronal functions immediately following injury. Alterations in the IC neuronal network, which persist for weeks after the initial insult, may lead to alterations in autonomic signaling and cardiac control.