To evaluate whether cervical spinal neurons can influence cardiac indices and myocyte viability in the acutely ischemic heart, the hearts of anesthetized rabbits subjected to 30. min of LAD coronary ...arterial occlusion (CAO) were studied 3. h after reperfusion. Control animals were compared to those exposed to pre-emptive high cervical cord stimulation (SCS; the dorsal aspect of the C1-C2 spinal cord was stimulated electrically at 50. Hz; 0.2. ms; 90% of motor threshold, starting 15. min prior to and continuing throughout CAO). Four groups of animals were so tested: 1) neuroaxis intact; 2) prior cervical vagotomy; 3) prior transection of the dorsal spinal columns at C6; and 4) following pharmacological treatment muscarinic (atropine) or adrenergic (atenolol, prazosin or yohimbine) receptor blockade. Infarct size (IS) was measured by tetrazolium, expressed as percentage of risk zone. C1-C2 SCS reduced acute ischemia induced IS by 43%, without changing the incidence of sudden cardiac death (SCD). While SCS-induced reduction in IS was unaffected by vagotomy, it was no longer evident following transection of C6 dorsal columns or atropinization. Beta-adrenoceptor blockade eliminated ischemia induced SCD, while alpha-receptor blockade doubled its incidence. During SCS, myocardial ischemia induced SCD was eliminated following vagotomy while remaining unaffected by atropinization. These data indicate that, in contrast to thoracic spinal neurons, i) cranial cervical spinal neurons affect both adrenergic and cholinergic motor outflows to the heart such that ii) their activation modifies ventricular infarct size and lethal arrhythmogenesis.
Background-High catecholamine concentrations are cytotoxic to cardiac myocytes. We hypothesized that myocardial interstitial catecholamine levels are greatly elevated immediately after long-duration ...ventricular fibrillation (VF), defibrillation, and reperfusion and that the short-acting β-antagonist esmolol administered at reperfusion would protect against this catecholamine surge and improve survival. Methods and Results-In part 1 of this study, catecholamines from myocardial interstitial fluid (ISF) and aortic and coronary sinus plasma were quantified by use of 3H-labeled radioenzymatic assay in 8 open-chest, anesthetized pigs. Eight minutes of electrically induced VF was followed by internal defibrillation and reperfusion. By 4 minutes of VF, ISF norepinephrine increased significantly, from 1.3±0.3 to 7.4±2.4 ng/mL. Epinephrine increased significantly, from 0.4±0.2 to 1.5±0.7 ng/mL. ISF norepinephrine and epinephrine peaked at 219.2±92.1 and 63.7±25.1 ng/mL after defibrillation and reperfusion and decreased significantly to 12.2±3.5 and 6.7±3.1 ng/mL 23 minutes after defibrillation. Transcardiac catecholamine changes were similar. In part 2, 8 minutes of VF was followed by external defibrillation in anesthetized, closed-chest pigs. Animals received 1.0 mg/kg esmolol (n=8) or saline (n=8) intravenously at the start of cardiopulmonary resuscitation (CPR). Advanced cardiac life support, including CPR and epinephrine, was delivered to both groups. Esmolol before reperfusion improved return of spontaneous circulation and 4-hour survival (7/8 versus 3/8 survivors, χ2 P<0.05). Conclusions-Transcardiac and ISF norepinephrine and epinephrine levels are briefly massively elevated after 8 minutes of VF, defibrillation, and reperfusion. A short-acting β-antagonist administered immediately after defibrillation improves return of spontaneous circulation and 4-hour survival after this prolonged VF.
Background Myocardial infarction (MI) induces remodeling in stellate ganglion neurons (SGNs). Objective We investigated whether infarct site has any impact on the laterality of morphologic changes or ...neuropeptide expression in stellate ganglia. Methods Yorkshire pigs underwent left circumflex coronary artery (LCX; n = 6) or right coronary artery (RCA; n = 6) occlusion to create left- and right-sided MI, respectively (control: n = 10). At 5 ± 1 weeks after MI, left and right stellate ganglia (LSG and RSG, respectively) were collected to determine neuronal size, as well as tyrosine hydroxylase (TH) and neuropeptide Y immunoreactivity. Results Compared with control, LCX and RCA MIs increased mean neuronal size in the LSG (451 ± 25 vs 650 ± 34 vs 577 ± 55 μm2, respectively; P =.0012) and RSG (433 ± 22 vs 646 ± 42 vs 530 ± 41 μm2, respectively; P =.002). TH immunoreactivity was present in the majority of SGNs. Both LCX and RCA MIs were associated with significant decreases in the percentage of TH-negative SGNs, from 2.58% ± 0.2% in controls to 1.26% ± 0.3% and 0.7% ± 0.3% in animals with LCX and RCA MI, respectively, for LSG (P =.001) and from 3.02% ± 0.4% in controls to 1.36% ± 0.3% and 0.68% ± 0.2% in LCX and RCA MI, respectively, for RSG (P =.002). Both TH-negative and TH-positive neurons increased in size after LCX and RCA MI. Neuropeptide Y immunoreactivity was also increased significantly by LCX and RCA MI in both ganglia. Conclusion Left- and right-sided MIs equally induced morphologic and neurochemical changes in LSG and RSG neurons, independent of infarct site. These data indicate that afferent signals transduced after MI result in bilateral changes and provide a rationale for bilateral interventions targeting the sympathetic chain for arrhythmia modulation.
Calcitonin gene-related peptide (CGRP) is a nonadrenergic/noncholinergic (NANC) peptide with vasodilatative/inotropic action that may benefit the failing heart. However, precise mechanisms for its in ...vivo inotropic action remain unclear. To assess this, dogs with normal or failing (sustained tachypacing) hearts were instrumented for pressure-dimension analysis. In control hearts, CGRP (20 pmol/kg per minute) enhanced cardiac contractility (eg, +33±4.2% in end-systolic elastance) and lowered afterload (-14.2±2% in systemic resistance, both P<0.001). The inotropic response was markedly blunted by heart failure (+6.5±2%; P<0.001 versus control), whereas arterial dilation remained unaltered (-19.3±5%). CGRP-positive inotropy was not attributable to reflex activation because similar changes were observed in the presence of a ganglionic blocker. However, it was fully prevented by the β-receptor antagonist (timolol), identifying a dominant role of sympatho-stimulatory signaling. In control hearts, myocardial interstitial norepinephrine assessed by microdialysis almost doubled in response to CGRP infusion, whereas systemic plasma levels were unchanged. In addition, CGRP receptors were not observed in ventricular myocardium but were prominent in coronary arteries and the stellate ganglia. Ventricular myocytes isolated from normal and failing hearts displayed no inotropic response to CGRP, further supporting indirect sympatho-stimulation as the primary in vivo mechanism. In contrast, the peripheral vasodilatative capacity of CGRP was similar in femoral vascular rings from normal and failing hearts in dogs. Thus, CGRP-mediated positive inotropy is load-independent but indirect and attributable to myocardial sympathetic activation rather than receptor-coupled stimulation in canine hearts. This mechanism is suppressed in heart failure, so that afterload reduction accounts for CGRP-enhanced function in this setting.
Background - This study tested the hypothesis that β1-adrenoreceptor blockade modulates the angiotensin II (Ang II)-evoked neural release of norepinephrine (NE) and epinephrine (Epi) into the cardiac ...interstitial fluid (ISF) space in experimentally induced mitral regurgitation (MR) in the dog. Methods and Results - Normal dogs (n=8) were compared with dogs with MR of 2 (n=8) and 4 (n=6) weeks' duration and with dogs with MR treated with β1-receptor blockade (RB; extended-release metoprolol succinate, 100 mg QD; MR+β1-RB) that was started 24 hours after MR induction for 2 (n=6) and 4 weeks (n=8). Left ventricular end-diastolic dimension increased 20% as plasma Ang II levels increased >5-fold in both MR and MR+β1-RB dogs at 2 and 4 weeks. Ang II infusion into the left atrium produced increases in ISF NE and Epi in normal dogs, which were further increased in 2- and 4-week MR dogs but were restored to normal in 4-week MR+β1-RB dogs. Ang II infusion produced 4-fold increases in circulating NE and Epi in 2- and 4-week MR dogs that returned to normal in 4-week+β1-RB dogs. Left ventricular angiotensin-converting enzyme activity and ISF Ang II were increased in 4-week MR dogs but were decreased in 4-week MR+β1-RB dogs. Conclusions - β1-RB decreases renin-angiotensin system sympathostimulation and activation by attenuating the Ang II-mediated NE and Epi release into the cardiac ISF and circulation and by decreasing left ventricular angiotensin-converting enzyme expression in the early phases of volume overload.
Mitral regurgitation (MR) is associated with increased neuronal release of norepinephrine (NE) and epinephrine (EP) into myocardial interstitial fluid (ISF) that may be necessary in sustaining left ...ventricular (LV) function via activation of cardiomyocyte β-adrenergic receptors (ARs). However, activation of neuronal β-ARs on cardiac neurons may lead to further catecholamine release, with an attendant risk of functional deterioration. We hypothesize that a beneficial effect of β-AR blockade may therefore mitigate excessive catecholamine release from cardiac adrenergic neurons in dogs with MR. We measured the effects of chronic β-receptor blockade (β-RB) on ISF NE and EP release using in vivo microdialysis in open-chest anesthetized dogs after 4 wk of MR with or without extended release of metoprolol succinate (100 mg/day) as well as in control dogs. Fractional shortening increased by 30% in both MR and MR + β-RB dogs after 4 wk of MR. In MR + β-RB dogs, stellate-stimulated heart rate change was attenuated compared with control and MR dogs, whereas peak change of LV pressure over time (+dP/dt) increased equally in all groups. Stellate-stimulated ISF NE increased fivefold over baseline in MR versus twofold in control dogs (<0.05), but the NE release was significantly attenuated in MR + β-RB dogs. In contrast, stellate-stimulated increases in ISF EP did not differ in control, MR, and MR + β-RB dogs. This study demonstrates that β-RB attenuates ISF NE release from cardiac neurons and that the LV functional response to MR is not dependent on an excess increase in ISF NE. Thus β1-RB may exert a beneficial effect by attenuating untoward effects of excessive sympathetic efferent neural NE release while sustaining early LV functional adaptation to MR.
BACKGROUND: Neuronal remodeling in human heart disease is not well understood. METHODS: Stellate ganglia from patients with cardiomyopathy (CMY) and refractory ventricular arrhythmias undergoing ...cardiac sympathetic denervation (n = 8), and from organ donors with normal hearts (n = 8) collected at the time of organ procurement were compared. Clinical data on all subjects were reviewed. Electron microscopy (EM), histologic, and immunohistochemical assessments of neurotransmitter profiles, glial activation and distribution, and lipofuscin deposition, a marker of oxidative stress, were quantified. RESULTS: In CMY specimens, lipofuscin deposits were larger, and present in more neurons (26.3% ± 6.3% vs. 16.7% ± 7.6%, P < 0.043), than age-matched controls. EM analysis revealed extensive mitochondrial degeneration in CMY specimens. T cell (CD3+) infiltration was identified in 60% of the CMY samples, with one case having large inflammatory nodules, while none were identified in controls. Myeloperoxidase-immunoreactive neutrophils were also identified at parenchymal sites distinct from inflammatory foci in CMY ganglia, but not in controls. The adrenergic phenotype of pathologic samples revealed a decrease in tyrosine hydroxylase staining intensity compared with controls. Evaluation of cholinergic phenotype by staining for the vesicular acetylcholine transporter revealed a low but comparable number of cholinergic neurons in ganglia from both groups and demonstrated that preganglionic cholinergic innervation was maintained in CMY ganglia. S100 staining (a glial cell marker) demonstrated no differences in glial distribution and relationship to neurons; however, glial activation demonstrated by glial fibrillary acidic protein (GFAP) staining was substantially increased in pathologic specimens compared with controls. CONCLUSIONS: Stellate ganglia from patients with CMY and arrhythmias demonstrate inflammation, neurochemical remodeling, oxidative stress, and satellite glial cell activation. These changes likely contribute to excessive and dysfunctional efferent sympathetic tone, and provide a rationale for sympathectomy as a treatment for arrhythmias in this population. FUNDING: This work was made possible by support from NIH grants HL125730 to OAA, GM107949 to DBH, and HL084261 and OT2OD023848 to KS.
We developed a spatially-tracked single neuron transcriptomics map of an intrinsic cardiac ganglion, the right atrial ganglionic plexus (RAGP) that is a critical mediator of sinoatrial node (SAN) ...activity. This 3D representation of RAGP used neuronal tracing to extensively map the spatial distribution of the subset of neurons that project to the SAN. RNA-seq of laser capture microdissected neurons revealed a distinct composition of RAGP neurons compared to the central nervous system and a surprising finding that cholinergic and catecholaminergic markers are coexpressed, suggesting multipotential phenotypes that can drive neuroplasticity within RAGP. High-throughput qPCR of hundreds of laser capture microdissected single neurons confirmed these findings and revealed a high dimensionality of neuromodulatory factors that contribute to dynamic control of the heart. Neuropeptide-receptor coexpression analysis revealed a combinatorial paracrine neuromodulatory network within RAGP informing follow-on studies on the vagal control of RAGP to regulate cardiac function in health and disease.
Background. Whether cardiac reinnervation occurs after transplantation remains controversial. If reinnervation does occur, how sympathetic and parasympathetic efferent neurons do this remains ...unknown. Methods. Power spectral analysis of heart rate variability was assessed for 1 year after cardiac autotransplantation in 9 dogs. After induction of anesthesia 13 months after transplantation, cardiac and intrinsic cardiac neuronal responses elicited by both electrical stimulation of parasympathetic or sympathetic efferent neurons and systemic or local coronary artery administration of nicotine (5 μg/kg), angiotensin II (0.75 μg/kg), and tyramine (1.2 μg/kg) were studied. The transmembrane electrical properties of intrinsic cardiac neurons were studied in vitro. Ventricular tissue catecholamine content, α-tubulin expression, and β-adrenergic receptor density and affinity were studied. The presence of axons crossing suture lines was sought histologically. Results. Nerves were identified crossing suture lines. Electrical or chemical (ie, nicotine or angiotensin II) activation of sympathetic efferent neurons enhanced cardiodynamics, as did tyramine. Stimulating vagal efferent preganglionic axons induced bradycardia in half of the dogs. Functional reinnervation did not correlate with specific power spectra derived from rate variability in the conscious state. Responding to nicotine and angiotensin II in situ, transplanted intrinsic cardiac neurons generated spontaneous activity. These neurons displayed nicotine-dependent synaptic inputs in vitro. Ventricular tissue had normal β-adrenergic receptor affinity and density but reduced catecholamine and α- tubulin contents. Conclusions. The intrinsic cardiac nervous system receives reduced input from extracardiac sympathetic efferent neurons after transplantation and inconsistent input from parasympathetic efferent preganglionic neurons. These heterogeneous neuronal inputs are not reflected in heart rate variability or ventricular β-adrenergic receptor function. Transplanted angiotensin II-sensitive intrinsic cardiac neurons exert greater cardiac control than do nicotine-sensitive ones. The intrinsic cardiac nervous system remodels itself after cardiac transplantation, and this indicates that direct assessment of extracardiac and intrinsic cardiac neuronal behavior is required to fully understand cardiac control after transplantation.
Surgical disruption of the small (approximately 0.7 x 1.0 cm) epicardial fat pad situated at the junction of the inferior vena cava (IVC) and inferior surface of the left atrium (ILA) interrupts both ...right and left vagal input to the atrioventricular nodal (AVN) region of the canine heart. This intervention eliminates AV block during supramaximal stimulation of both cervical vagi, without interfering with sinus bradycardia normally associated with sinoatrial nodal (SAN) suppression. Independent modulation of SAN and AVN activities by the parasympathetic system is thereby revealed. Histology of the excised IVC-ILA fat pad reveals multiple well organized autonomic ganglia. These ganglia range from 2 to 80 cells per cluster and are associated with numerous nerve trunks. Individual ganglia are commonly surrounded by fatty connective tissue closely adjacent to epicardial muscle. They have not been found imbedded within atrial muscle and have been been found in or close to endocardial muscle layers. Other ganglia, imbedded in the fat pad overlying the posterior surface of the left atrium or in the atrioventricular groove, do not directly modulate A-V conduction. Surgical dissection around the extreme left or middle segments of the great cardiac vein and the coronary sinus failed to interrupt either left or right vagal input to the AVN region. Parasympathetic, preganglionic pathways to these AVN synapses do not, therefore, course from left to right along the atrioventricular groove. However, dissection around the extreme right portion of the coronary sinus at its penetration of the inferior interatrial septum, did interrupt vagal influences upon A-V conduction. Thus, numerous autonomic ganglia have been localized in the canine heart which serve as synaptic stations mediating both right and left vagal regulation of A-V conduction.
