Cardiac Innervation Research Articles

Chronotropic Response to Exercise is Decreased in Patients With Congenital Heart Disease Compared to Cardiomyopathy Following Pediatric Heart Transplantation.

Two common indications for pediatric heart transplantation are congenital heart disease and cardiomyopathy. Prior studies suggest differences in chronotropy on cardiopulmonary exercise testing outcomes depending on indication for heart transplantation. We aimed to determine whether the number of pretransplant sternotomies is associated with differences in heart rate response during exercise testing. A retrospective analysis of our institutional pediatric heart transplant data between 2004 and 2022 was performed. Patients were categorized by indication for transplantation into a cardiomyopathy (CM) group if they had a congenital or acquired cardiomyopathy or a congenital heart disease (CHD) group including all other forms of congenital cardiac anatomic abnormalities. CHD patients (n = 40) differed from CM patients (n = 53) by mean number of sternotomies prior to transplant (2.4 ± 1.8 vs. 0.5 ± 0.9, p < 0.001). There were no significant differences in echocardiographic function or catheterization hemodynamics. In cardiopulmonary exercise testing performance, the congenital heart disease group had a significantly higher resting heart rate (91.8 ± 11.2 vs. 86.4 ± 10.2 bpm, p = 0.019), lower percent predicted age-predicted maximal heart rate achieved (78.3 ± 8.5% vs. 83.2 ± 11.4%, p = 0.032), and lower heart rate reserve (68.6 ± 19.8 vs. 84.4 ± 24.0 bpm, p = 0.001) despite a similar age and average time from transplantation. Regression analysis confirmed number of pretransplant sternotomies as a main predictor of heart rate metrics. There is greater chronotropic incompetence in patients who underwent transplantation due to congenital heart disease compared to cardiomyopathy. The groups differ significantly by number of sternotomies, potentially supporting the hypothesis that prior surgical disruption of cardiac innervation may cause decreased chronotropic response to exercise following transplantation.

The hidden dangers: How obesity alters cardiac innervation and structure in Zucker rats

The hidden dangers: How obesity alters cardiac innervation and structure in Zucker rats

Cardiac sympathetic neurons are additional cells affected in genetically determined arrhythmogenic cardiomyopathy.

Arrhythmogenic cardiomyopathy (AC) is a familial cardiac disease, mainly caused by mutations in desmosomal genes, which accounts for most cases of stress-related arrhythmic sudden death, in young and athletes. AC hearts display fibro-fatty lesions that generate the arrhythmic substrate and cause contractile dysfunction. A correlation between physical/emotional stresses and arrhythmias supports the involvement of sympathetic neurons (SNs) in the disease, but this has not been confirmed previously. Here, we combined molecular, in vitro and ex vivo analyses to determine the role of AC-linked DSG2 downregulation on SN biology and assess cardiac sympathetic innervation in desmoglein-2mutant (Dsg2mut/mut) mice. Molecular assays showed that SNs express DSG2, implying that DSG2-mutation carriers would harbour the mutant protein in SNs. Confocal immunofluorescence of heart sections and 3-D reconstruction of SN network in clarified heart blocks revealed significant changes in the physiologialc SN topology, with massive hyperinnervation of the intact subepicardial layers and heterogeneous distribution of neurons in fibrotic areas. Cardiac SNs isolated from Dsg2mut/mut neonatal mice, prior to the establishment of cardiac innervation, show alterations in axonal sprouting, process development and distribution of varicosities. Consistently, virus-assisted DSG2 downregulation replicated, in PC12-derived SNs, the phenotypic alterations displayed by Dsg2mut/mut primary neurons, corroborating that AC-linked Dsg2 variants may affect SNs. Our results reveal that altered sympathetic innervation is an unrecognized feature of AC hearts, which may result from the combination of cell-autonomous and context-dependent factors implicated in myocardial remodelling. Our results favour the concept that AC is a disease of multiple cell types also hitting cardiac SNs. KEY POINTS: Arrhythmogenic cardiomyopathy is a genetically determined cardiac disease, which accounts for most cases of stress-related arrhythmic sudden death. Arrhythmogenic cardiomyopathy linked to mutations in desmoglein-2 (DSG2) is frequent and leads to a left-dominant form of the disease. Arrhythmogenic cardiomyopathy has been approached thus far as a disease of cardiomyocytes, but we here unveil that DSG2 is expressed, in addition to cardiomyocytes, by cardiac and extracardiac sympathetic neurons, although not organized into desmosomes. AC-linked DSG2 downregulation primarily affect sympathetic neurons, resulting in the significant increase in cardiac innervation density, accompanied by alterations in sympathetic neuron distribution. Our data supports the notion that AC develops with the contribution of several 'desmosomal protein-carrying' cell types and systems.

Comparison of cardiac autonomic innervation in post-mortem tissue from individuals with kidney failure and preserved kidney function.

Comparison of cardiac autonomic innervation in post-mortem tissue from individuals with kidney failure and preserved kidney function.

Importance of cardiac-synchronized vagus nerve stimulation parameters on the provoked chronotropic response for different levels of cardiac innervation.

The influence of vagus nerve stimulation (VNS) parameters on provoked cardiac effects in different levels of cardiac innervation is not well understood yet. This study examines the effects of VNS on heart rate (HR) modulation across a spectrum of cardiac innervation states, providing data for the potential optimization of VNS in cardiac therapies. Utilizing previously published data from VNS experiments on six sheep with intact innervation, and data of additional experiments in five rabbits post bilateral rostral vagotomy, and four isolated rabbit hearts with additionally removed sympathetic influences, the study explored the impact of diverse VNS parameters on HR. Significant differences in physiological threshold charges were identified across groups: 0.09 ± 0.06μC for intact, 0.20 ± 0.04μC for vagotomized, and 9.00 ± 0.75μC for isolated hearts. Charge was a key determinant of HR reduction across all innervation states, with diminishing correlations from intact (r = 0.7) to isolated hearts (r = 0.44). An inverse relationship was observed for the number of pulses, with its influence growing in conditions of reduced innervation (intact r = 0.11, isolated r = 0.37). Frequency and stimulation delay showed minimal correlations (r < 0.17) in all conditions. Our study highlights for the first time that VNS parameters, including stimulation intensity, pulse width, and pulse number, crucially modulate heart rate across different cardiac innervation states. Intensity and pulse width significantly influence heart rate in innervated states, while pulse number is key in denervated states. Frequency and delay have less impact impact across all innervation states. These findings suggest the importance of customizing VNS therapy based on innervation status, offering insights for optimizing cardiac neuromodulation.

Genetic markers of cardiac autonomic neuropathy in the Kazakh population

BackgroundCardiac autonomic neuropathy (CAN) is a complication of diabetes mellitus (DM) that increases the risk of morbidity and mortality by disrupting cardiac innervation. Recent evidence suggests that CAN may manifest even before the onset of DM, with prediabetes and metabolic syndrome potentially serving as precursors. This study aims to identify genetic markers associated with CAN development in the Kazakh population by investigating the SNPs of specific genes.Materials and methodsA case-control study involved 82 patients with CAN (cases) and 100 patients without CAN (controls). A total of 182 individuals of Kazakh nationality were enrolled from a hospital affiliated with the RSE “Medical Center Hospital of the President’s Affairs Administration of the Republic of Kazakhstan”. 7 SNPs of genes FTO, PPARG, SNCA, XRCC1, FLACC1/CASP8 were studied. Statistical analysis was performed using Chi-square methods, calculation of odds ratios (OR) with 95% confidence intervals (CI), and logistic regression in SPSS 26.0. Results: Among the SNCA gene polymorphisms, rs2737029 was significantly associated with CAN, almost doubling the risk of CAN (OR 2.03(1.09–3.77), p = 0.03). However, no statistically significant association with CAN was detected with the rs2736990 of the SNCA gene (OR 1.00 CI (0.63–1.59), p = 0.99). rs12149832 of the FTO gene increased the risk of CAN threefold (OR 3.22(1.04–9.95), p = 0.04), while rs1801282 of the PPARG gene and rs13016963 of the FLACC1 gene increased the risk twofold (OR 2.56(1.19–5.49), p = 0.02) and (OR 2.34(1.00-5.46), p = 0.05) respectively. rs1108775 and rs1799782 of the XRCC1 gene were associated with reduced chances of developing CAN both before and after adjustment (OR 0.24, CI (0.09–0.68), p = 0.007, and OR 0.43, CI (0.22–0.84), p = 0.02, respectively). Conclusion: The study suggests that rs2737029 (SNCA gene), rs12149832 (FTO gene), rs1801282 (PPARG gene), and rs13016963 (FLACC1 gene) may be predisposing factors for CAN development. Additionally, SNPs rs1108775 and rs1799782 (XRCC1 gene) may confer resistance to CAN. Only one polymorphism rs2736990 of the SNCA gene was not associated with CAN.

Angiotensin II-Induced Heart Failure Disrupts Cardiac Sympathetic Innervation

About 26 million people worldwide live with heart failure (HF), and 25% have hypertension (HTN) as the primary cause. These patients have a high risk for cardiac arrhythmias and sudden cardiac death. Autonomic dysfunction and sympathetic hyperactivity accompany these diseases, and sympathetic activation is crucial for triggering fatal arrhythmias. However, changes in cardiac sympathetic innervation in HF are not well understood. We hypothesized that cardiac sympathetic innervation is disrupted in a mouse model of Angiotensin II (AngII)-induced HF. To test this hypothesis, C57BL6 male (N=6, 10) and female (N=6, 10) mice 12-20 weeks old were implanted with minipumps releasing either AngII (712ng/min/Kg, N=6-10) or saline (N=6-10) for 4 weeks. HTN and cardiac function were measured with tail cuff quantification and echocardiography, respectively. Fibrosis was assessed using Trichrome staining. Cardiac sympathetic innervation was quantified using immunohistochemistry for Tyrosine Hydroxylase (TH). Cardiac norepinephrine (NE) was measured by High-Performance Liquid Chromatography with electrochemical detection. Values were compared with a Student’s unpaired t-test unless noted, data were represented as mean ± SD, and significance was defined as p&lt;0.05. AngII-treated mice had significantly increased mean arterial pressure (Saline 91.0±15.6 vs. AngII 152.9±22.3 mmHg, p&lt;0.05, N=6-7) 7 days after minipump implantation. Four weeks of AngII treatment led to HF, as indicated by reduced ejection fraction (Saline 54.8±3.8 vs. AngII 43.1±8.2 %, p&lt;0.05, N=10) and fractional shortening (Saline 14.3±2.6 vs. AngII 10.6±3.3 %, p&lt;0.05, N=10). Cardiac hypertrophy was also observed in AngII-treated mice and was reflected by increased heart weight to body weight ratio (Saline 5.2±0.2 vs. AngII 7.7±1.0, p&lt;0.05, N=6). Fibrosis was detected in the intraventricular septum (IVS) and left ventricle (LV) of AngII-treated mice. TH+ fibers in the LV and IVS were decreased significantly (LV: Saline 0.6±0.2 vs. AngII 0.4±0.1 %; IVS: Saline 0.3±0.1 vs. AngII 0.09±0.04 %, p&lt;0.05, N=6), while no differences were detected in the right ventricle (Saline 1.1±0.2 vs. AngII 0.8±0.3 %, P&gt;0.05, N=6). In addition, the subepicardium-subendocardium (EPI-ENDO) gradient was altered (EPI: Saline 0.9±0.2 vs. AngII 0.8±0.3 %, P&gt;0.05; ENDO: Saline 0.3±0.05 vs. AngII 0.1±0.1 %, p&lt;0.05; EPI:ENDO ratio: Saline 2.7±0.7 vs. AngII 15.7±16.8, p&lt;0.05, Mann-Whitney test, N=6). A significant decrease in NE content accompanied the loss of TH+ fibers in the LV (LV: Saline 3.9±0.4 vs. AngII 1.6±0.2 pmol/mg, p&lt;0.05; RV: Saline 4.4±1.0 vs. AngII 3.8±0.8 pmol/mg, P&gt;0.05, N=4-6). Our data demonstrate that AngII-induced HF disrupts cardiac sympathetic nerve density, distribution, and NE content, confirming our initial hypothesis. This evidence contrasts with sympathetic hyperactivity seen in cardiac diseases. Future studies will assess the overall cardiac innervation and, determine whether the loss of TH+ fibers is due to a reduced sympathetic innervation after cardiac remodeling or the cholinergic transdifferentiation of TH+ fibers. A better understanding of the cardiac sympathetic nervous system in HF is imperative to deciphering the mechanistic changes to cardiac innervation in disease, and may help to develop and implement novel and precise neuromodulatory therapies for patients with autonomic imbalance. Supported by NIH R01 HL146833 to Dr. Habecker. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Identification of Spinal Afferent Innervation in the Rat Heart: Atria and Ventricles: Anterograde Tracing

The spinal afferent innervation of the heart regulates cardiac functions by sending sensory information from the dorsal root ganglia (DRG) to the brain. However, the distribution and morphology of spinal afferents in the whole heart are not well characterized. The challenges here are: 1) diffculty in surgically accessing the upper thoracic DRG to label only cardiac spinal afferents. 2) handling flat-mounts of whole atria and ventricles (thickness ~850 μm) without using sections or partial tissues that could disrupt the continuity of axonal structures. To overcome these challenges, we injected anterograde tracer dextran biotin (DB) into the left DRG (C8-T3) of male SD rats (3-5 months) and allowed them to recover for 14 days for tracer transportation. Flat-mounts of the left and right atria and ventricles were prepared, and DAB (3,3′-Diaminobenzidine) stained, followed by imaging, tracing, and digitization using the Neurolucida system. Our findings are: In the atria, the DB-labeled axons entered the atria potentially through the left precaval vein (LPCV), with predominant innervation of the left side of the atrial wall and extended their projection towards the auricles, middle region of the atrium, and pulmonary veins. Most of the spinal afferent axons were present in the myocardial layer. Varicose DB-labeled axons were also observed on cardiac targets at different regions, and distinct morphological structures were found. In the cardiac muscle, DB-labeled axons ran mainly in the direction of muscle fibers. Also, varicose axons ran along the small vasculature on the atria. Spinal afferents formed varicose contacts with individual principal neurons within some intrinsic cardiac ganglia (ICG). In the ventricles, DB-labeled axons were present in the left and right ventricles, which ramified and expressed spherical-like varicosities on the cardiac muscle. In both the atria and ventricles, spinal afferent axons formed simple, branching, and complex terminal structures. Thus, our work, for the first time, successfully labeled, traced, and digitized the spinal cardiac afferents in the flat-mounts of whole atria and ventricles, which will provide a foundation for specific labeling of cardiac spinal afferent fibers. In addition, it will lead to the future study of topographical cardiac spinal afferent innervation, functional studies, and their remodeling in pathological conditions. This study was supported by NIH HEAL/SPARC U01 NS113867-01, NIH R15 R15HL137143-01A1, and UCF dissertation completion Fellowships. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The role of the cardiac vagus in maintaining cardiac function during exercise

There is a strong association between parasympathetic nervous system activity (vagal tone) and physical fitness. High vagal tone is associated with increased capacity to exercise and overall cardiovascular health. Evidence for vagal withdrawal during exercise comes from historic studies examining heart rate control and the use of cholinergic blockers. However, more recent studies have challenged this assumption. We hypothesized that cardiac vagal activity increases during exercise and maintains cardiac function via neurotransmitters other than acetylcholine. The left cardiac vagal branch was identified in anesthetized open-chested sheep. Cardiac innervation of the left cardiac vagal branch was confirmed with lipophilic tracer dyes (DiO). Custom-built coil electrodes were placed around the left cardiac vagal branch. 3-10 days after surgery, conscious, direct recordings of cardiac vagal nerve activity (CVNA), cardiac output, coronary artery blood flow and heart rate were recorded in adult female sheep during whole-body treadmill exercise. Sheep were exercised with pharmacological blockers of acetylcholine (atropine, 250 mg), vasoactive intestinal peptide (VIP) ([4Cl−D-Phe6,Leu17]-VIP 25 μg), or saline control, randomized on different days. In a subset of sheep, the left cardiac vagal branch was denervated. Neural innervation from the cardiac vagal branch is seen at major cardiac ganglionic plexi, and within the fat pads associated with the coronary arteries. Directly recorded CVNA increased during exercise. CVNA increases during the lowest exercise intensity and plateaus as exercise intensity increases. Left cardiac vagal branch, denervation attenuated the maximum changes in coronary artery blood flow (maximum exercise, control: 63.5 ± 5.9 ml/min, n = 8, cardiac vagal denervated: 32.7 ± 5.6 ml/min, n = 6, mean ± SEM. P = 2.5x10−7. Two-way ANOVA-interaction), cardiac output and heart rate during exercise. Atropine did not affect any cardiac parameters during exercise, but VIP antagonism significantly reduced coronary artery blood flow during exercise to a similar level to vagal denervation (maximum exercise, paired: control: 68.2 ± 6.0 ml/min, VIP antagonism: 44.9 ± 3.9 ml/min, n = 6, mean ± SEM. P = 4.8x10−3. Two-way ANOVA-interaction). Our study challenges the conventional view and demonstrates that cardiac vagal nerve activity increases and is crucial for maintaining cardiac function during exercise. Furthermore, our findings show that the dynamic modulation of coronary artery blood flow during exercise is mediated by VIP and that acetylcholine has no role in modulating cardiac function during exercise. This work is supported by the Health Research Council of New Zealand 23/119 (Fellowship. JS) and the New Zealand Heart Foundation (JS, RR). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

From Psychostasis to the Discovery of Cardiac Nerves: The Origins of the Modern Cardiac Neuromodulation Concept.

This review explores the historical development of cardiology knowledge, from ancient Egyptian psychostasis to the modern comprehension of cardiac neuromodulation. In ancient Egyptian religion, psychostasis was the ceremony in which the deceased was judged before gaining access to the afterlife. This ritual was also known as the "weighing of the heart" or "weighing of the soul". The Egyptians believed that the heart, not the brain, was the seat of human wisdom, emotions, and memory. They were the first to recognize the cardiocentric nature of the body, identifying the heart as the center of the circulatory system. Aristotle (fourth century BC) considered the importance of the heart in human physiology in his philosophical analyses. For Galen (third century AD), the heart muscle was the site of the vital spirit, which regulated body temperature. Cardiology knowledge advanced significantly in the 15th century, coinciding with Leonardo da Vinci and Vesalius's pioneering anatomical and physiological studies. It was William Harvey, in the 17th century, who introduced the concept of cardiac circulation. Servet's research and Marcello Malpighi's discovery of arterioles and capillaries provided a more detailed understanding of circulation. Richard Lower emerged as the foremost pioneer of experimental cardiology in the late 17th century. He demonstrated the heart's neural control by tying off the vagus nerve. In 1753, Albrecht von Haller, a professor at Göttingen, was the first to discover the heart's automaticity and the excitation of muscle fibers. Towards the end of the 18th century, Antonio Scarpa challenged the theories of Albrecht von Haller and Johann Bernhard Jacob Behrends, who maintained that the myocardium possessed its own "irritability", on which the heartbeat depended, and was independent of neuronal sensitivity. Instead, Scarpa argued that the heart required innervation to maintain life, refuting Galenic notions. In contemporary times, the study of cardiac innervation has regained prominence, particularly in understanding the post-acute sequelae of SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2) infection (PASC), which frequently involves cardiorespiratory symptoms and dysregulation of the intrinsic cardiac innervation. Recently, it has been recognized that post-acute sequelae of acute respiratory infections (ARIs) due to other pathogens can also be a cause of long-term vegetative and somatic symptoms. Understanding cardiac innervation and modulation can help to recognize and treat long COVID and long non-COVID-19 (coronavirus disease 2019) ARIs. This analysis explores the historical foundations of cardiac neuromodulation and its contemporary relevance. By focusing on this concept, we aim to bridge the gap between historical understanding and modern applications. This will illuminate the complex interplay between cardiac function, neural modulation, cardiovascular health, and disease management in the context of long-term cardiorespiratory symptoms and dysregulation of intrinsic cardiac innervations.

The autonomic nerves around the vein of Marshall: apostmortem study with clinical implications.

This study aims to analyze the vein of Marshall (VOM) in human autopsy hearts and its correlation with clinical data to elucidate the morphological substrates of atrial fibrillation (AF) and other cardiac diseases. Twenty-three adult autopsy hearts were studied, assessing autonomic nerves by immunohistochemistry with tyrosine hydroxylase (sympathetic nerves), choline acetyltransferase (parasympathetic nerves), growth-associated protein 43 (neural growth), and S100 (general neural marker) antibodies. Interstitial fibrosis was assessed by Masson trichrome staining. Measurements were conducted via morphometric software. The results were correlated with clinical data. Sympathetic innervation was abundant in all VOM-adjacent regions. Subjects with a history of AF, cardiovascular cause of death, and histologically verified myocardial infarction had increased sympathetic innervation and neural growth around the VOM at the mitral isthmus. Interstitial fibrosis increased with age and heart weight was associated with AF and cardiovascular cause of death. This study increases our understanding of the cardiac autonomic innervation in the VOM area in various diseases, offering implications for the development of new therapeutic approaches targeting the autonomic nervous system.

Sleep-induced limb vasodilation in individuals confined to bed for 24 h

The sleep/wake rhythm in limbs has been scarcely studied, especially due to the difficulty associated with continuous monitoring of arterial flow to the forearm and leg for a 24-h period. Addressing this constraint, we employed indium-gallium-in-silicone strain-gauge venous-occlusion plethysmography, an automated method facilitating the measurement of 24-h limb arterial flow in bed-confined subjects without disturbing their natural sleep. This article presents the state of the art in this field. Our examination of 60 healthy normotensive individuals revealed a distinctive sleep/wake rhythm in limb arterial flow, characterized by elevated values during sleep (32.7% in the forearm, P &lt; 0.0001; 39.1% in the leg, P &lt; 0.0001). Correspondingly, limb resistance mirrored the trend of flow (-32.7%, P &lt; 0.0001; -33.5%, P &lt; 0.0001), with these variations attributed to sleep-induced limb vasodilation. Sleep-associated vasodilation was also evident in 21 hypertensive individuals (leg resistance: -33.1%, P &lt; 0.0001) and 13 heart transplant recipients lacking vagal and sympathetic cardiac innervation (resistance: -33.6%, P &lt; 0.0001). On the contrary, among 11 subjects with an interrupted spinal cord, we observed forearm vasodilation (resistance: -36.6%, P &lt; 0.0001) but observed no leg vasodilation if the spinal lesion was under T2 (innervating the leg). Furthermore, a loss of sleep-induced vasodilation occurred in both the forearm and leg if the injury was above C7 (innervating both forearm and leg). Our conclusion posits the existence of sleep-induced limb vasodilation, a phenomenon attributed to signals traveling along the spinal cord, with the heart playing no discernible role in this rhythmic process, and arterial hypertension deemed irrelevant. Comprehensive further studies are imperative to elucidate the precise triggers of limb vasodilation during sleep.

Exploring cardiac innervation by 3D light sheet imaging in horses with atrial fibrillation

Exploring cardiac innervation by 3D light sheet imaging in horses with atrial fibrillation

Cardiac remodeling, amyloidosis, neurotrophic signaling and innervation impairment in the Tg2576 model of Alzheimer’s disease

AbstractBackgroundIn Alzheimer’s disease (AD), the characteristic cerebral amyloid β (Aβ) accumulation and tau pathology, are accompanied by a gradual loss of the two main neuromodulators, NGF and BDNF. This AD‐mediated derangement of the brain’s neuro‐signaling pathways may extend to the periphery and, along with potential peripheral amyloid deposition, induce peripheral nervous system impairment, thus affecting other organs, including the heart. However, whether and how AD pathology affects cardiac physiology, neurotrophins, innervation, and heart amyloidosis is still undefined.MethodHere, we describe for the first‐time that cardiac remodeling and neuro‐signaling dysregulation are present in the hearts of Tg2576 mice, a widely used model of AD and cerebral amyloidosis.ResultsEchocardiographic analysis showed significant deterioration of left ventricle function, as demonstrated by a decline of both ejection fraction and fraction shortening in 12‐month‐old Tg2576 mice compared to age‐matched WT littermates. The transgenic mice hearts presented an increase in interstitial fibrosis, with an accumulation of amyloid aggregates, including Aβ, associated with severe cardiac nervous system dysfunction. Tg2576 mice exhibited a depletion in cardiac nerve fiber density, both adrenergic (stained with tyrosine hydroxylase‐ TH) and regenerating nerve terminals (labeled with GAP‐43) compared to the WT mice. This myocardial denervation was accompanied by a decline in NGF and BDNF protein expression as well as GAP‐43 expression in both the brain and heart of Tg2576 mice. Likewise, both human neuroblastoma cells (SH‐SY5Y) and human cardiomyocytes (AC16) challenged with human Aβ‐40 or Aβ‐42 oligomers showed significant downregulation of both BDNF and GAP‐43 activity.ConclusionOverall, this study highlights the deleterious impact of cerebral amyloidosis on the cardiac nervous system and heart physiology, providing potential therapeutic targets, such as neurotrophic factors, to prevent or limit the AD‐mediated degenerative mechanisms on the cardiovascular system.

Vascular senescence controls the neuro-cardiac interface in the aging heart

Abstract Aging is a major risk factor for impaired cardiovascular function. The aging heart is characterized by vascular dysfunction, increased hypertrophy, fibrosis and electrophysiological alterations. Studies show impaired endothelial cell (EC) function associates with senescence in aging. Since vessels and nerves align, and this interplay is critical for tissue homeostasis, we studied whether an impairment of the neuro-vascular interface may contribute to age-associated pathologies in the heart. To study the innervation of the aging mouse heart, we histologically assessed the cardiac autonomic nervous system in 3- and 20-month old mice using the pan-neuronal marker Tuj1 and the sympathetic marker TH. Interestingly, sympathetic innervation of the left ventricle, especially around arteries, was reduced by 0.66±0.07-fold with aging (p&amp;lt;0.05) indicating that aging reduces innervation of the left ventricle. Similar findings were observed in premature aging models of TERT-KO and EC-transgenic progeria mice. Electrophysiological studies confirmed an increased incidence of ventricular arrhythmias and reduced heart rate variability in aged mice. To determine the potential contribution of neurovascular cells, we analyzed gene expression of isolated ECs in aging mouse hearts by bulk RNA sequencing. Aging significantly induced the expression of genes associated with GO-terms related to neuronal cell death and axon injury. Sema3a, an axon repelling factor, was significantly increased in ECs of aged mice hearts, which was confirmed by qPCR and by histology (p&amp;lt;0.05). Next we determined the mechanism of Sema3a regulation. We show that miR-145 acts as an up-stream regulator of Sema3a expression. miR-145-/- mice showed increased Sema3a expression and reduced cardiac innervation. We observed an increase in acidic β-gal activity as a marker for cellular senescence in the aging heart, particularly in the proximate arterial space. In addition, Sema3a expression was significantly increased in senescent ECs in vitro, thus suggesting a putative role for senescence to impair cardiac innervation. To test whether eliminating senescent cells may restore left ventricular axon density, we treated 18-month old mice with a combination of the two senolytic drugs dasatinib and quercetin, which are known to deplete senescent cells in vivo. After 2 months of treatment, cellular senescence was reduced by 0.4±0.1-fold (p&amp;lt;0.05) in aged hearts. Endothelial expression of Sema3a was also reduced. Importantly, senolytic treatment restored ventricular innervation, augmented heart rate variability, and normalized electrophysiological abnormalities (all p&amp;lt;0.05). In conclusion, we show that aging augments axon repelling signals in ECs and reduces left ventricular innervation. The depletion of senescent cells prevented age-induced impairment of innervation and normalized electrophysiological abnormalities suggesting a critical role of senescence-associated axon repulsion in the aging heart.

The mesencephalic astrocyte-derived neurotrophic factor (MANF) as a novel regulator of post-infarction cardiac sympathetic hyperinnervation

Abstract Background Excessive outgrowth of sympathetic nerve fibers (sympathetic hyperinnervation) around the infarcted myocardium is associated with ventricular arrhythmia and an increased risk of sudden cardiac death. Post-infarction, epicardial cells exit quiescence through epithelial-to-mesenchymal transition (EMT), resulting in the formation of epicardium-derived cells (EPDCs). EPDCs support cardiac repair through differentiation into various cardiac cell lineages, as well as secretion of paracrine factors that stimulate processes such as angiogenesis. Recently, we demonstrated that human EPDCs reinforce neurite outgrowth in vitro. Whether EPDCs regulate cardiac innervation through paracrine signaling and contribute to post-infarction sympathetic hyperinnervation is unknown. Purpose To identify epicardial paracrine factors post-infarction that are involved in nerve growth. Methods Single-cell RNA-sequencing (scRNA-seq) datasets capturing epicardial cells of the healthy and infarcted adult mouse heart were collected from published research articles and were integrated into one object using Seurat. A list of gene products of known involvement in biological processes related to nerve growth (675 unique entries in total) was compiled. Based on this list, differential gene expression analysis on data from the integrated scRNA-seq object was performed to uncover highly-expressed candidates in the epicardial cell population. Epicardial expression of a candidate paracrine factor was validated by immunohistochemistry in the infarcted mouse heart as well as in an adult human in vitro model of EPDC EMT by immunocytochemistry and transcriptomic analysis. Results In our integrated scRNA-seq object, we identified high expression of Manf in epicardial cells post-infarction. Consistent with this finding, immunohistochemistry showed that Manf was up-regulated in the infarcted area, borderzone area, and epicardial region situated close to the infarct, an area where also hyperinnervation is observed. In vitro, MANF transcript expression was significantly down-regulated (p&amp;lt;0.01) in human EPDCs that were stimulated with TGFβ3 to induce EMT compared to EPDCs in which EMT was inhibited. In line with this, immunocytochemistry staining of MANF indicated that MANF expression was down-regulated in human EPDCs stimulated with TGFβ3. Conclusion(s) This study is the first to show that Manf is highly expressed in the epicardial region close to the infarct. In vitro, MANF is down-regulated in human adult EPDCs post-EMT. Future experiments will be directed toward determining the effect of MANF knockdown in human EPDC on neurite outgrowth in co-cultured human sympathetic neurons.MANF expression in human EPDCs after EMTManf expression after infarction

Cardiac Autonomic Innervation Is Decreased in Postmortem Tissue from Individuals with ESKD Compared with Those Without

Cardiac Autonomic Innervation Is Decreased in Postmortem Tissue from Individuals with ESKD Compared with Those Without

Cardiac Vagal Nerve Activity Increases During Exercise to Enhance Coronary Blood Flow.

The phrase complete vagal withdrawal is often used when discussing autonomic control of the heart during exercise. However, more recent studies have challenged this assumption. We hypothesized that cardiac vagal activity increases during exercise and maintains cardiac function via transmitters other than acetylcholine. Chronic direct recordings of cardiac vagal nerve activity, cardiac output, coronary artery blood flow, and heart rate were recorded in conscious adult sheep during whole-body treadmill exercise. Cardiac innervation of the left cardiac vagal branch was confirmed with lipophilic tracer dyes (DiO). Sheep were exercised with pharmacological blockers of acetylcholine (atropine, 250 mg), VIP (vasoactive intestinal peptide; [4Cl-D-Phe6,Leu17]VIP 25 µg), or saline control, randomized on different days. In a subset of sheep, the left cardiac vagal branch was denervated. Neural innervation from the cardiac vagal branch is seen at major cardiac ganglionic plexi, and within the fat pads associated with the coronary arteries. Directly recorded cardiac vagal nerve activity increased during exercise. Left cardiac vagal branch denervation attenuated the maximum changes in coronary artery blood flow (maximum exercise, control: 63.5±5.9 mL/min, n=8; cardiac vagal denervated: 32.7±5.6 mL/min, n=6, P=2.5×10-7), cardiac output, and heart rate during exercise. Atropine did not affect any cardiac parameters during exercise, but VIP antagonism significantly reduced coronary artery blood flow during exercise to a similar level to vagal denervation. Our study demonstrates that cardiac vagal nerve activity actually increases and is crucial for maintaining cardiac function during exercise. Furthermore, our findings show the dynamic modulation of coronary artery blood flow during exercise is mediated by VIP.

Abstract P2013: Cardiac Remodeling, Amyloidosis, Neurotrophic Signaling, And Innervation Impairment In The Tg2576 Model Of Alzheimer's Disease

Background: In Alzheimer's disease (AD), the characteristic cerebral amyloid β (Aβ) deposition and tau pathology are accompanied by a progressive loss of the two main neuromodulators, NGF and BDNF. This AD-mediated derangement of the brain's neuro-signaling pathway may extend to the periphery and, along with potential peripheral amyloid deposition, induce peripheral nervous system impairment, thus affecting other organs, including the heart. However, whether and how AD pathology affects cardiac physiology, neurotrophins, innervation, and heart amyloidosis is still unclear. Method: Here, we describe for the first time that cardiac remodeling and neuro-signaling dysregulation are present in the hearts of Tg2576 mice, a widely used model of AD and cerebral amyloidosis. Results: Echocardiographic analysis showed significant deterioration of left ventricle function, as demonstrated by a decline of both ejection fraction and fraction shortening in 12-month-old Tg2576 mice compared to age-matched WT littermates. The transgenic mice hearts displayed an increase in interstitial fibrosis, with an accumulation of amyloid aggregates, including Aβ, associated with severe cardiac nervous system dysfunction. Tg2576 mice exhibited a depletion in cardiac nerve fiber density, both adrenergic (stained with tyrosine hydroxylase- TH) and regenerating nerve terminals (labeled with GAP-43), compared to the WT mice. This myocardial denervation was accompanied by a decline in NGF and BDNF protein expression as well as GAP-43 expression in both the brain and heart of Tg2576 mice. Likewise, human neuroblastoma cells (SH-SY5Y) and human cardiomyocytes (AC16) challenged with human Aβ-40 or, Aβ-42 oligomers showed significant downregulation of both BDNF and GAP-43 activity. Conclusion: Overall, this study highlights the harmful impact of cerebral amyloidosis on the cardiac nervous system and heart physiology, providing potential therapeutic targets, such as neurotrophic factors, to prevent or limit AD-mediated degenerative mechanisms in the cardiovascular system.

Quantification of Large Transmural Biopsies Reveals Heterogeneity in Innervation Patterns in Chronic Myocardial Infarction

BackgroundAbnormal cardiac innervation plays an important role in arrhythmogenicity after myocardial infarction (MI). Data regarding reperfusion models and innervation abnormalities in the medium to long term after MI are sparse. Histologic quantification of the small-sized cardiac nerves is challenging, and transmural analysis has not been performed. ObjectivesThis study sought to assess cardiac innervation patterns in transmural biopsy sections in a porcine reperfusion model of MI (MI-R) using a novel method for nerve quantification. MethodsTransmural biopsy sections from 4 swine (n = 83) at 3 months after MI-R and 3 controls (n = 38) were stained with picrosirius red (fibrosis) and beta-III-tubulin (autonomic nerves). Biopsy sections were classified as infarct core, border zone, or remote zone. Each biopsy section was analyzed with a custom software pipeline, allowing calculation of nerve density and classification into innervation types at the 1 × 1–mm resolution level. Relocation of the classified squares to the original biopsy position enabled transmural quantification and innervation heterogeneity assessment. ResultsCoexisting hyperinnervation, hypoinnervation, and denervation were present in all transmural MI-R biopsy sections. The innervation heterogeneity was greatest in the infarct core (median: 0.14; IQR: 0.12-0.15), followed by the border zone (median: 0.05; IQR: 0.04-0.07; P = 0.02) and remote zone (median: 0.02; IQR: 0.02-0.03; P < 0.0001). Only in the border zone was a positive linear relation between fibrosis and innervation heterogeneity observed (R = 0.79; P < 0.0001). ConclusionsThis novel method allows quantification of nerve density and heterogeneity in large transmural biopsy sections. In the chronic phase after MI-R, alternating innervation patterns were identified within the same biopsy section. Persistent innervation heterogeneity, in particular in the border zone biopsy sections, may contribute to late arrhythmogenicity.