Positive Chronotropic Effects Research Articles

The pancreas does not contribute to the non-adrenergic-non-cholinergic stimulation of heart rate in digesting pythons

Vertebrates elevate heart rate when metabolism increases during digestion. Part of this tachycardia is due to a non-adrenergic-non-cholinergic (NANC) stimulation of the cardiac pacemaker, and it has been suggested these NANC factors are circulating hormones that are released from either gastrointestinal or endocrine glands. The NANC stimulation is particularly pronounced in species with large metabolic responses to digestion, such as reptiles. To investigate the possibility that the pancreas may release hormones that exert positive chronotropic effects on the digesting Burmese python heart, a species with very large postprandial changes in heart rate and oxygen uptake, we evaluate how pancreatectomy affects postprandial heart rate before and after autonomic blockade of the muscarinic and the beta-adrenergic receptors. We also measured the rates of oxygen consumption and evaluated the short-term control of the heart using the spectral analysis of heart rate variability and the baroreflex sequence method. Digestion caused the ubiquitous tachycardia, but the intrinsic heart rate (revealed after the combination of atropine and propranolol) was not affected by pancreatectomy and therefore hormones, such as glucagon and insulin, do not appear to contribute to the regulation of heart rate during digestion in Burmese pythons.

Direct modulation of TRPC ion channels by Gα proteins.

GPCR-Gi protein pathways are involved in the regulation of vagus muscarinic pathway under physiological conditions and are closely associated with the regulation of internal visceral organs. The muscarinic receptor-operated cationic channel is important in GPCR-Gi protein signal transduction as it decreases heart rate and increases GI rhythm frequency. In the SA node of the heart, acetylcholine binds to the M2 receptor and the released Gβγ activates GIRK (I(K,ACh)) channel, inducing a negative chronotropic action. In gastric smooth muscle, there are two muscarinic acetylcholine receptor (mAChR) subtypes, M2 and M3. M2 receptor activates the muscarinic receptor-operated nonselective cationic current (mIcat, NSCC(ACh)) and induces positive chronotropic effect. Meanwhile, M3 receptor induces hydrolysis of PIP2 and releases DAG and IP3. This IP3 increases intracellular Ca2+ and then leads to contraction of GI smooth muscles. The activation of mIcat is inhibited by anti-Gi/o protein antibodies in GI smooth muscle, indicating the involvement of Gαi/o protein in the activation of mIcat. TRPC4 channel is a molecular candidate for mIcat and can be directly activated by constitutively active Gαi QL proteins. TRPC4 and TRPC5 belong to the same subfamily and both are activated by Gi/o proteins. Initial studies suggested that the binding sites for G protein exist at the rib helix or the CIRB domain of TRPC4/5 channels. However, recent cryo-EM structure showed that IYY58-60 amino acids at ARD of TRPC5 binds with Gi3 protein. Considering the expression of TRPC4/5 in the brain, the direct G protein activation on TRPC4/5 is important in terms of neurophysiology. TRPC4/5 channels are also suggested as a coincidence detector for Gi and Gq pathway as Gq pathway increases intracellular Ca2+ and the increased Ca2+ facilitates the activation of TRPC4/5 channels. More complicated situation would occur when GIRK, KCNQ2/3 (IM) and TRPC4/5 channels are co-activated by stimulation of muscarinic receptors at the acetylcholine-releasing nerve terminals. This review highlights the effects of GPCR-Gi protein pathway, including dopamine, μ-opioid, serotonin, glutamate, GABA, on various oragns, and it emphasizes the importance of considering TRPC4/5 channels as crucial players in the field of neuroscience.

Abstract 14259: Contractile Effects of Glucagon in Atrial Preparations From Transgenic Mice and Men

Glucagon, a peptide composed of 29 amino acids, produced mainly in the pancreas can also stimulate cardiac function. Glucagon might contribute to hyperglycemia in diabetes mellitus type 2 and is thus of clinical relevance. There is controversy in the literature whether glucagon affects force and beating rate in the human atrium and which receptor is involved. Hence, we studied the effect of glucagon on atrial preparations of wild type mouse hearts (WT), from transgenic mice with cardiac overexpression of the human glucagon receptor (GR-TG) and for comparison in human right atrial preparations. We noted that glucagon exerted concentration- and time-dependent positive chronotropic effects in atrial preparations from WT and GR-TG starting at 0.1 nM glucagon and amounting to 25 % of pre-drug value at 3 nM glucagon (n=8, p<0.05). These effects were antagonized by the glucagon receptor antagonist SC203973 (1 μM), the inhibitor of cAMP dependent protein kinase H89 (1 μM), by the M-cholinoceptor agonist 1 μM carbachol, by the A1-adenosine receptor antagonist R-phenylisopropyladenosine (1 μM) and the inhibitors of hyperpolarization-activated cyclic nucleotide gated channels namely ivabradine (1 μM) or cesium chloride (2 mM) (n=5-6, p<0.05). In left atrial preparations from GR-TG but not WT, glucagon exerted starting at 1 nM a positive inotropic effect. Interestingly, in human atrial preparations glucagon at 1 μM in the additional presence of the phosphodiesterase III inhibitor cilostamide (1 μM) increased force of contraction by 34 % (n=5, p<0.05). We conclude that glucagon can exert contractile effects in mammalian hearts via GR, but they exhibit considerable regional and species differences. The physiological and therapeutic relevance of glucagon in the human heart needs to be fully elucidated.

Endothelium-Derived Dopamine and 6-Nitrodopamine in the Cardiovascular System.

The review deals with the release of endothelium-derived dopamine and 6-nitrodopamine (6-ND) and its effects on isolated vascular tissues and isolated hearts. Basal release of both dopamine and 6-ND is present in human isolated umbilical cord vessels, human popliteal vessels, nonhuman primate vessels, and reptilia aortas. The 6-ND basal release was significantly reduced when the tissues were treated with Nω-nitro-l-arginine methyl ester and virtually abolished when the endothelium was mechanically removed. 6-Nitrodopamine is a potent vasodilator, and the mechanism of action responsible for this effect is the antagonism of dopamine D2-like receptors. As a vasodilator, 6-ND constitutes a novel mechanism by which nitric oxide modulates vascular tone. The basal release of 6-ND was substantially decreased in endothelial nitric oxide synthase knockout (eNOS-/-) mice and not altered in neuronal nitric oxide synthase knockout (nNOS-/-) mice, indicating a nonneurogenic source for 6-ND in the heart. Indeed, in rat isolated right atrium, the release of 6-ND was not affected when the atria were treated with tetrodotoxin. In the rat isolated right atrium, 6-ND is the most potent endogenous positive chronotropic agent, and in Langendorff's heart preparation, it is the most potent endogenous positive inotropic agent. The positive chronotropic and inotropic effects of 6-ND are antagonized by β1-adrenoceptor antagonists at concentrations that do not affect the effects induced by noradrenaline, adrenaline, and dopamine, indicating that blockade of the 6-ND receptor is the major modulator of heart chronotropism and inotropism. The review proposes that endothelium-derived catecholamines may constitute a major mechanism for control of vascular tone and heart functions, in contrast to the overrated role attributed to the autonomic nervous system.

SAT579 Newly Diagnosed Graves' Disease With Complete Heart Block In Elderly Patient. A Case Report

Abstract Disclosure: M.M. Eid: None. T. Zahra: None. J. Vargas-Jerez: None. Introduction: Hypothyroidism is the most common thyroid disorder in elderly. The prevalence of hyperthyroidism in elderly is 0.5-3% and mainly caused by toxic thyroid nodules. Graves’ disease is an autoimmune disorder mediated by TSH receptor antibody (TSHR AB), presents commonly with hyperthyroidism in middle age female(1). Complete heart block in elderly can be due to conduction system fibrosis, Ischemic/structural heart diseases, medications, electrolyte disturbance and hypothyroidism(2). Case 72yo, Female with history of DM and HTN. Denied smoking, alcohol, drug and allergy. Irrelevant family history. Medications include amlodipine, metformin and atorvastatin. She was referred to endocrinology clinic for low TSH (was normal last year). Review of system was negative, denied tremor, palpitation, anxiety, weight loss, bowel movement/mood/sleep change, vision/swallowing problem. Physical examination: ABP150/80, Heart Rate(HR) 75/min. Mild lid lag, no exophthalmos. No tremor. Thyroid gland: diffusely enlarged non tender, no bruit. Normal cardiopulmonary and abdomen examination. Laboratory work up TSH less than 0.01 normal 0.2 to 4.20 uIU/ml , FT4 3.5 normal 0.9 to1.8 ng/dl, FT3 6 normal 2 to 4.4 pg/ml. Repeat lab confirmed TSH suppression and high FT4 ,FT3. TSHR antibody 7.66 normal less than 1.75 IU/L, Thyroid stimulating immunoglobulin 5.05 normal less than 0.5 IU/L ,HBA1C 6.6, Normal liver and kidney function tests and Complete blood count. She was diagnosed with Graves’ disease, kept on Methimazole 10mg/day. 2 weeks later, she presented to the ED after syncope at home. Initial EKG showed complete heart block HR 35/min and right bundle branch block (RBBB), ABP 110/65. Hours later EKG showed transient first degree heart block HR 75/min, RBBB and left axis deviation, then complete heart block with HR 30/min. Normal Troponin, potassium 4.2 mmol/L, sodium 141 mmol/L, calcium 9.2mg/dl, TSH 0.015 uIU/ml,FT4 3 ng/dl. Negative Anti-Lyme IgG and IgM. Normal Echocardiogram. She underwent uneventful Dual chamber pacemaker implantation with close endocrinology follow up. Conclusion: Graves’ Disease has variable presentations in elderly. Hyperthyroidism causes tachyarrhythmia due to the positive chronotropic effect of thyroid hormones. Bradyarrhythmia in our case is most likely due to conduction system fibrosis and could be related to the inhibitory effect of TSHR AB in elderly in contrary to the simulating effect in young patients. It is still rare to diagnose symptomatic bradyarrhythmia with uncontrolled hyperthyroidism(1).

FRI478 Pharmacometrics-based Computer Model Utilizing Heart Rate To Monitor Thyroid Function In Children With Graves' Disease

Abstract Disclosure: B. Steffens: None. G. Koch: None. F. Claude: None. F. Bachmann: None. J. Schropp: None. M. Janner: None. D. l'Allemand: None. D. Konrad: None. M. Pfister: None. G. Szinnai: None. Graves’ disease (GD) with onset in childhood or adolescence is a pediatric rare disease (ORPHA:525731) with a ten times lower incidence than in adults. Leading clinical signs of hyperthyroidism are sinus tachycardia, weight loss, tremor, and goiter. First-line treatment are anti-thyroid drugs in order to normalize thyroid function. However, dose finding in pediatric GD is complex due to a broad spectrum of disease severity at diagnosis, and highly variable disease activity during follow-up, especially during puberty. As thyroid hormones have a strong positive chronotropic effect, heart rate (HR) turns out to be a useful clinical marker to monitor thyroid activity under treatment. Our overall aim was to provide a practical pharmacometrics-based (PMX-based) computer model characterizing both, individual FT4 dynamics and the relation between FT4 and tachycardia in children with various disease severity of GD during the first 120 days of treatment. Development of the PMX computer model was based on the non-linear mixed effects approach (i) linking FT4 kinetics with HR dynamics, (ii) accounting for inter-individual variability, and (iii) incorporating individual patient characteristics. Retrospectively collected clinical (resting heart rate during consultation) and laboratory data (FT4) from 41 children and adolescents with GD at four pediatric hospitals in Switzerland (75% female, median age 11.2 [IQR 8.5, 13.5] years) with 187 FT4 measurements and 132 HR measurements, and 124 paired measurements, were available and used for model development. Pediatric patients showed median FT4 of 59.6 [IQR 44.5, 73.0] pmol/l and median HR of 112 [IQR 100, 128] bpm at diagnosis. GD severity groups were defined based on FT4 measurement at diagnosis, resulting in equal numbers of mild (13), moderate (14) and severe (14) GD. We observed a significant difference in HR at diagnosis (p < 0.01) between the three GD severity groups based on FT4 at diagnosis. The final PMX computer model accounted for inter-individual variability and clinically relevant covariate effects such as age, gender, and GD severity, and was able to accurately predict FT4 and HR dynamics for each individual patient during the first 120 days of treatment. A PMX-based computer model that leverages individual HR dynamics can be applied to facilitate personalized pharmacotherapy in pediatric GD and mitigate the risk for under- or overdosing of anti-thyroid drugs in these patients. Prospective randomized validation trials are warranted to further validate and fine-tune such computer-supported personalized dosing in children with Graves’ disease. Presentation: Friday, June 16, 2023

Environmental concentrations of tire rubber-derived 6PPD-quinone alter CNS function in zebrafish larvae

N-(1,3-Dimethylbutyl)-N′-phenyl-p-phenylenediamine quinone (6PPD-quinone) is a degradation product of 6PPD, an antioxidant widely used in rubber tires. 6PPD-quinone enters aquatic ecosystems through urban stormwater runoff and has been identified as the chemical behind the urban runoff mortality syndrome in coho salmon. However, the available data suggest that the acute effects of 6PPD-quinone are restricted to a few salmonid species and that the environmental levels of this chemical should be safe for most fish. In this study, larvae of a “tolerant” fish species, Danio rerio, were exposed to three environmental concentrations of 6PPD-quinone for only 24 h, and the effects on exploratory behavior, escape response, nonassociative learning (habituation), neurotransmitter profile, wake/sleep cycle, circadian rhythm, heart rate and oxygen consumption rate were analyzed. Exposure to the two lowest concentrations of 6PPD-quinone resulted in altered exploratory behavior and habituation, an effect consistent with some of the observed changes in the neurotransmitter profile, including increased levels of acetylcholine, norepinephrine, epinephrine and serotonin. Moreover, exposure to the highest concentration tested altered the wake/sleep cycle and the expression of per1a, per3 and cry3a, circadian clock genes involved in the negative feedback loop. Finally, a positive chronotropic effect of 6PPD-quinone was observed in the hearts of the exposed fish. The results of this study emphasize the need for further studies analyzing the effects of 6PPD-quinone in “tolerant” fish species.

Ergometrine stimulates histamine H2 receptors in the isolated human atrium

Ergometrine (6aR,9R)-N-((S)-1-hydroxypropan-2-yl)-7-methyl-4,6,6a,7,8,9-hexa-hydro-indolo-[4,3-fg]chinolin-9-carboxamide or lysergide acid β-ethanolamide or ergonovine) activates several types of serotonin and histamine receptors in the animal heart. We thus examined whether ergometrine can activate human serotonin 5-HT4 receptors (h5-HT4R) and/or human histamine H2 receptors (hH2R) in the heart of transgenic mice and/or in the human isolated atrium. Force of contraction or beating rates were studied in electrically stimulated left atrial or spontaneously beating right atrial preparations or spontaneously beating isolated retrogradely perfused hearts (Langendorff setup) of mice with cardiac specific overexpression of the h5-HT4R (5-HT4-TG) or of mice with cardiac specific overexpression of the hH2R (H2-TG) or in electrically stimulated human right atrial preparations obtained during cardiac surgery. Western blots to assess phospholamban (PLB) phosphorylation on serine 16 were performed. Ergometrine exerted concentration- and time-dependent positive inotropic effects and positive chronotropic effects in atrial preparations starting at 0.3 µM and reaching a plateau at 10 µM in H2-TGs (n = 7). This was accompanied by an increase in PLB phosphorylation at serine 16. Ergometrine up 10 µM failed to increase force of contraction in left atrial preparations from 5-HT4-TGs (n = 5). Ten micrometer ergometrine increased the force of contraction in isolated retrogradely perfused spontaneously beating heart preparations (Langendorff setup) from H2-TG but not 5-HT4-TG. In the presence of the phosphodiesterase inhibitor cilostamide (1 µM), ergometrine at 10 µM exerted positive inotropic effects in isolated electrically stimulated human right atrial preparations, obtained during cardiac surgery, and these effects were eliminated by 10 µM of the H2R antagonist cimetidine but not by 10 µM of the 5-HT4R antagonist tropisetron. Furthermore, ergometrine showed binding to human histamine H2 receptors (at 100 µM and 1 mM) using HEK cells in a recombinant expression system (pKi < 4.5, n = 3). In conclusion, we suggest that ergometrine is an agonist at cardiac human H2Rs.

The pro-inflammatory signature of lipopolysaccharide in spontaneous contracting embryoid bodies differentiated from mouse embryonic stem cells.

Embryonic stem (ES) cells differentiate towards all three germ layers, including cardiac cells and leukocytes, and may be therefore suitable to model inflammatory reactions in vitro. In the present study, embryoid bodies differentiated from mouse ES cells were treated with increasing doses of lipopolysaccharide (LPS) to mimic infection with gram-negative bacteria. LPS treatment dose-dependent increased contraction frequency of cardiac cell areas and calcium spikes and increased protein expression of α-actinin. LPS treatment increased the expression of the macrophage marker CD68 and CD69, which is upregulated after activation on T cells, B cells and NK cells. LPS dose-dependent increased protein expression of toll-like receptor 4 (TLR4). Moreover, upregulation of NLR family pyrin domain containing 3 (NLRP3), IL-1ß and cleaved caspase 1 was observed, indicating activation of inflammasome. In parallel, generation of reactive oxygen species (ROS), nitric oxide (NO), and expression of NOX1, NOX2, NOX4 and eNOS occurred. ROS generation, NOX2 expression and NO generation were downregulated by the TLR4 receptor antagonist TAK-242 which abolished the LPS-induced positive chronotropic effect of LPS. In conclusion, our data demonstrate that LPS induced a pro-inflammatory cellular immune response in tissues derived from ES cells, recommending the in vitro model of embryoid bodies for inflammation research.

Effects of whole-body cryotherapy and static stretching are maintained 4 weeks after treatment in most patients with chronic fatigue syndrome

In the previous study, whole-body cryotherapy (WBC)+static stretching (SS) has been shown to reduce the severity of some symptoms in Chronic Fatigue Syndrome (CFS) noted just after the therapy. Here we consider the effects of treatment and explore the sustainability of symptom improvements at four weeks (one-month) follow-up. Twenty-two CFS patients were assessed one month after WBC + SS programme. Parameters related to fatigue (Chalder Fatigue Questionnaire (CFQ), Fatigue Impact Scale (FIS), Fatigue Severity Scale (FSS)), cognitive function (Trial Making test part A and B (TMT A and TMT B and its difference (TMT B-A)), Coding) hemodynamic, aortic stiffness (aortic systolic blood pressure (sBP aortic)) and autonomic nervous system functioning were measured. TMT A, TMT B, TMT B-A and Coding improved at one month after the WBC + SS programme. WBC + SS had a significant effect on the increase in sympathetic nervous system activity in rest. WBC + SS had a significant, positive chronotropic effect on the cardiac muscle. Peripheral and aortic systolic blood pressure decreased one month after WBC + SS in comparison to before. Effects of WBC + SS on reduction of fatigue, indicators of aortic stiffness and symptoms severity related to autonomic nervous system disturbance and improvement in cognitive function were maintained at one month. However, improvement in all three fatigue scales (CFQ, FIS and FSS) was noted in 17 of 22 patients. In addition, ten patients were treated initially but they were not assessed at 4 weeks, and are thus not included in the 22 patients who were examined on follow-up. The overall effects of WBC + SS noted at one month post-treatment should be interpreted with caution.

C42 OR 1896 EXERTS POSITIVE INOTROPIC EFFECTS IN ISOLATED HUMAN RIGHT ATRIAL MUSCLE

Abstract Levosimendan is approved for use as an intravenous agent to treat temporarily patients with severe systolic heart failure. It is thought, but hitherto never tested in isolated human cardiac tissue, that the main metabolite of levosimendan, called OR 1896, might cause the positive inotropic effect of levosimendan in patients. For comparison, we studied mouse atrial preparations. OR alone (up to 10 µM) failed to increase force of contraction in mouse atrial preparations while in the presence of the phosphodiesterase (PDE) IV–inhibitor rolipram (0.1 µM) profound positive inotropic effects and positive chronotropic effects of OR 1895 were recorded in mouse atrial preparations. In contrast to mouse atrium, we noticed that OR 1896 alone (0.01 µM to 10 µM cumulatively applied) exerted time– and concentration–dependent positive inotropic effects 1 µM increase by 72 ± 14.7%, p&amp;lt;0.05, n=7) and shortened the time of relaxation (by 7 .3 %, p&amp;lt;0.05, n=7) in isolated electrically paced human right atrial muscle preparations (obtained during cardiac surgery), that started at 0.1 µM and plateaued at 1 µM OR 1896. The positive inotropic effect of OR 1896 was antagonized by 10 µM propranolol. The positive inotropic effect of OR 1896 in mouse atrial preparations and human right atrial preparations was less effective than that of 1 µM isoprenaline (human atrium by 154 %). The positive inotropic effect of OR 1896 was diminished by pre–incubation of the samples with the PDE III inhibitor cilostamide (10 µM). EMD 57033 (10 µM), a calcium sensitizer, increased force of contraction further in the additional presence of 1 µM OR 1896. Our data suggest that OR 1896 is indeed a positive inotropic agent in the human heart and that OR 1896 acts mainly as a PDE III – inhibitor because the effect of OR 1896 was attenuated by a β–adrenoceptor antagonist and thus OR 1896 is unlikely to act as a calcium sensitizer in the human heart.

Type 2 phosphodiesterase regulates cardiac pacemaker activity

Elevated heart rate (HR) and lower HR variability (HRV) predict cardiovascular morbi-mortality. HR is mainly controlled by the autonomic nervous system (ANS). During sympathetic stimulation, the activation of β-adrenergic receptors increases cAMP levels leading to positive chronotropic effect. In the heart, cAMP can be hydrolyzed by 6 different phosphodiesterases (PDEs). One of them is PDE2 which has recently been reported to be beneficial for the failing heart. However, whether this is due to a direct control sinoatrial node (SAN) function or an indirect effect via the ANS is unknown. To explore the role of PDE2 in regulating HR. The hydrolytic cAMP activity was measured with radioenzymatic assay. The in vitro pacemaker activity was assessed by measuring spontaneous Ca2+ transients in Fluo4-loaded-SAN tissue using confocal microscopy. ECG telemetry was recorded in WT and cardiac PDE2-overexpressing (PDE2TG) conscious mice, in basal conditions, after ANS inhibition with atropine (atro, 1 mg/kg, ip) + propranolol (propra, 2 mg/kg, ip), or after β-adrenergic stimulation with isoprenaline (Iso, 1.5 mg/kg, ip). If current was measured by patch-clamp (whole-cell). PDE2 is expressed in WT SAN and overexpressed by ∼4-fold in the PDE2TG SAN. Total cAMP-hydrolyzing activity is increased ∼3-fold in PDE2TG SAN. PDE2TG mice display lower basal HR, during day and night. This phenotype was maintained after atro + prop and after Iso. Basal beating rate of PDE2TG SAN tissue was lower. While no difference in Ca2+ transient amplitude time to peak and duration at half maximum were unchanged in PDE2TG SAN cells, the time constant of decay was increased in basal conditions. Ca2+ sparks analysis between WT and PDE2TG SAN did not show any modification on ryanodine receptor activity. In addition, If current density was decreased in PDE2TG SAN cells. PDE2 regulates HR in vivo by a mechanism independent from ANS which takes place at the level of SAN and involves the voltage clock.

6-Nitrodopamine potentiates contractions of rat isolated vas deferens induced by noradrenaline, adrenaline, dopamine and electric field stimulation.

6-Nitrodopamine (6-ND) is a novel endogenous catecholamine that is released from the rat isolated vas deferens, and has been characterized as a major modulator of the contractility of rat isolated epididymal vas deferens (RIEVD). Drugs such as tricyclic antidepressants, α1 and β1β2 adrenoceptor blockers, act as selective antagonists of the 6-ND receptor in the RIEVD. In the rat isolated atria, 6-ND has a potent positive chronotropic action and causes remarkable potentiation of the positive chronotropic effects induced by dopamine, noradrenaline, and adrenaline. Here, whether 6-ND interacts with the classical catecholamines in the rat isolated vas deferens was investigated. Incubation with 6-ND (0.1 and 1 nM; 30min) caused no contractions in the RIEVD but provoked significant leftward shifts in the concentration-response curves to noradrenaline, adrenaline, and dopamine. Pre-incubation of the RIEVD with 6-ND (1 nM), potentiated the contractions induced by electric-field stimulation (EFS), whereas pre-incubation with 1 nM of dopamine, noradrenaline or adrenaline, did not affect EFS-induced contractions. In tetrodotoxin (1 μM) pre-treated (30 min) RIEVD, pre-incubation with 6-ND (0.1 nM) did not cause leftward shifts in the concentration-dependent contractions induced by noradrenaline, adrenaline, or dopamine. Pre-incubation of the RIEVD with the α2A-adrenoceptor antagonist idazoxan (30 min, 10 nM) did not affect dopamine, noradrenaline, adrenaline, and EFS-induced contractions. However, when idazoxan (10 nM) and 6-ND (0.1 nM) were simultaneously pre-incubated (30 min), a significant potentiation of the EFS-induced contractions of the RIEVD was observed. 6-nitrodopamine causes remarkable potentiation of dopamine, noradrenaline, and adrenaline contractions on the RIEVD, due to activation of adrenergic terminals, possibly via pre-synaptic adrenoceptors.

Investigation on the positive chronotropic action of 6-nitrodopamine in the rat isolated atria.

6-Nitrodopamine (6-ND) is released from rat isolated atria being 100 times more potent than noradrenaline and adrenaline, and 10,000 times more potent than dopamine as a positive chronotropic agent. The present study aimed to investigate the interactions of 6-ND with the classical catecholamines, phosphodiesterase (PDE)-3 and PDE4, and the protein kinase A in rat isolated atria. Atrial incubation with 1 pM of dopamine, noradrenaline, or adrenaline had no effect on atrial frequency. Similar results were observed when the atria were incubated with 0.01 pM of 6-ND. However, co-incubation of 6-ND (0.01 pM) with dopamine, noradrenaline, or adrenaline (1 pM each) resulted in significant increases in atrial rate, which persisted over 30 min after washout of the agonists. The increased atrial frequency induced by co-incubation of 6-ND with the catecholamines was significantly reduced by the voltage-gated sodium channel blocker tetrodotoxin (1 µM, 30 min), indicating that the positive chronotropic effect of 6-ND is due in part to activation of nerve terminals. Pre-treatment of the animals with reserpine had no effect on the positive chronotropic effect induced by dopamine, noradrenaline, or adrenaline; however, reserpine markedly reduced the 6-ND (1 pM)-induced positive chronotropic effect. Incubation of the rat isolated atria with the protein kinase A inhibitor H-89 (1 µM, 30 min) abolished the increased atrial frequency induced by dopamine, noradrenaline, and adrenaline, but only attenuated the increases induced by 6-ND. 6-ND induces catecholamine release from adrenergic terminals and increases atrial frequency independently of PKA activation.

Both osmolality-dependent and independent mechanisms are associated with acute hyperglycemia-induced cardiovascular adverse reactions: Analysis of the mutual interactions leading to cardiovascular phenotypes in dogs.

Acute hyperglycemia causes various cardiovascular responses; however, the underlying pathophysiology in vivo is myriad and complex, of which mutual interactions remain poorly understood. We analyzed the cardiovascular effects of acute hyperglycemia in comparison with those of hyperosmolality alone. Three g/kg of D-glucose (n = 4) or D-mannitol (n = 4) was intravenously infused to isoflurane-anesthetized intact dogs. Glucose infusion increased plasma glucose level and osmolality, whereas mannitol infusion similarly changed osmolality to glucose infusion but decreased glucose level. Glucose infusion decreased total peripheral vascular resistance, but increased heart rate, left ventricular contraction, left ventricular preload and cardiac output without altering mean blood pressure. Mannitol infusion likewise changed them, but its positive chronotropic and inotropic effects were less potent than those of glucose infusion. Glucose infusion prolonged PR interval, QRS width and QTcV. Mannitol infusion similarly changed them, but its QTcV prolongation was smaller than that of glucose infusion. Glucose infusion-induced cardiovascular responses would be basically attributed to osmolality-dependent mechanisms, whereas its positive chronotropic and inotropic effects along with repolarization delay may be enhanced by osmolality-independent mechanisms, including hyperglycemia by itself and insulin release.

Mechanisms of Modulation of Adrenergic Regulation of Spontaneous Activity Rate and Atrial Myocardial Contractility in Early Postnatal Ontogeny in Rats

We studied combined effect of the β1,2-adrenoreceptor agonist isoproterenol and the Y1,5 receptor agonist [Leu31, Pro34]neuropeptide Y on the frequency of spontaneous activity and myocardial contractility in 21- and 100-day-old rats. Isoproterenol increased the frequency of spontaneous activity and reduced the main parameters of isometric contraction of the atrial myocardium. When [Leu31, Pro34]neuropeptide Y was added, the frequency of spontaneous activity and the negative inotropic and the positive chronotropic effects of isoproterenol were reduced in 100-day-old rats. In 21-day-olds rats, a tendency to a decrease in the effect of isoproterenol was observed.

769 EFFECT OF GLUCOSE LOWERING AGENTS ON CARDIAC AUTONOMIC FUNCTION AND STRUCTURE IN TYPE 2 DIABETES MELLITUS

Effects of novel glucose-lowering agents on cardiac autonomic function and structure in type 2 diabetes Ilaria Cavallari, Ernesto Maddaloni, Francesco Veneziano, Carla Indennidate, Maria Valeria Giaccari, Dario Tuccinardi, Silvia Pieralice, Nicola Napoli, Raffaella Buzzetti, Francesco Grigioni Background Novel glucose-lowering agents, such as sodium glucose co-transporter-2 (SGLT-2) inhibitors and glucagon-like peptide-1 (GLP-1) receptor agonists, have the potential to reduce major adverse cardiovascular events among people with type 2 diabetes. However, mechanisms behind their cardiovascular protection and the implications of positive chronotropic effects of GLP-1 receptor agonists are not fully understood. Purpose To elucidate and compare the effects of novel glucose-lowering agents on cardiac autonomic function and structure. Methods This prospective observational study included subjects with type 2 diabetes and either a history of established cardiovascular disease or multiple risk factors and early initiation (within 2 weeks) of a SGLT-2 inhibitor or a GLP-1 receptor agonist on top of standard of care. Patients were followed for a median of 2 months. As per study protocol, background medical therapy should have remained stable for the entire duration of the study. Cardiac autonomic reflex tests (heart rate response to a deep breathing and to standing) and echocardiography were performed in all patients at baseline and at the end of follow-up. Endpoints were changes from baseline of E/I ratio, 30:15 ratio and indices of cardiac structure and function. Results A total of 37 patients completed follow-up, of whom 20 were on SGLT-2 inhibitors and 17 on GLP-1 receptor agonists. Mean age was 69 years, mean disease duration was 14 years and mean HbA1c value was 7.5%. Cardiac autonomic dysfunction, defined as at least an abnormal test, was found in 19% of patients (n=7). At a median follow-up of 2 months, there were no differences from baseline in heart rate response to a deep breathing test (p=0.92) and to standing (p=0.86). No differences were found when patients were stratified according to drug class. Among echocardiographic parameters, significant reductions of left ventricular mass (300.5±73.3 vs 280.4±68.3 g/m2, p=0.048) and left atrial volume (61.3±29.4 vs 54.7±18.0 ml, p=0.034) were found with no interaction by drug class. Left ventricular end-diastolic and systolic volumes, ejection fraction, filling pressures and pulmonary artery pressure did not substantially change. Conclusions In a population of patients with type 2 diabetes, short-term treatment with novel glucose-lowering agents significantly reduced left ventricular mass and left atrial volume but did not affect cardiac autonomic function.

Levosimendan increases the phosphorylation state of phospholamban in the isolated human atrium

Levosimendan (up to 10 µM) given alone failed to increase force of contraction in isolated electrically stimulated (1 Hz) left atrial (LA) preparations from wild-type mice. Only in the additional presence of 0.1 µM rolipram, an inhibitor of the activity of phosphodiesterase IV, levosimendan increased force of contraction in LA and increased the phosphorylation state of phospholamban at amino acid serine 16. Levosimendan alone increased the beating rate in isolated spontaneously beating right atrial preparations from mice and this effect was potentiated by rolipram. The positive inotropic and the positive chronotropic effects of levosimendan in mouse atrial preparations were attenuated by 10 µM propranolol. Finally, we studied the contractile effects of levosimendan in isolated electrically stimulated (1 Hz) right atrial preparations from the human atrium (HAP), obtained during cardiac surgery. We detected concentration-dependent positive inotropic effects of levosimendan alone that reached plateau at 1 µM levosimendan in HAP (n = 11). Levosimendan shortened time of tension relaxation in HAP. Cilostamide (1 µM), an inhibitor of phosphodiesterase III, or propranolol (10 µM) blocked the positive inotropic effect of levosimendan in HAP. Levosimendan (1 µM) alone increased in HAP the phosphorylation state of phospholamban. In conclusion, we present evidence that levosimendan acts via phosphodiesterase III inhibition in the human atrium leading to phospholamban phosphorylation and thus explaining the positive inotropic effects of levosimendan in HAP.

Myocardial perfusion recovery induced by an α-calcitonin gene-related peptide analogue

BackgroundEndogenous calcitonin gene-related peptide (CGRP) induces cardioprotective effects through coronary vasodilation. However, the systemic administration of CGRP induces peripheral vasodilation and positive chronotropic and inotropic effects. This study aims to examine the net effect on coronary perfusion of the systemically administered α-calcitonin gene-related peptide analogue, SAX, in rats during myocardial infarction. MethodsForty Sprague-Dawley rats underwent myocardial infarction. Following left anterior descending artery occlusion, [99mTc]Tc-sestamibi was administered to determine the myocardial perfusion before treatment. Twenty minutes, 24 and 48 h after [99mTc]Tc-sestamibi injection, the rats were treated with either SAX or placebo. Final infarct size was determined three weeks later by [99mTc]Tc-sestamibi SPECT/CT scan. ResultsThirty-one rats survived the surgery and 20 completed the follow-up SPECT/CT scan (SAX n = 12; Placebo n = 8). At baseline, there was no difference in size of perfusion defect between the groups (P = .88), but at follow-up the SAX group had improved myocardial recovery compared to the placebo group (P = .04), corresponding to a relative perfusion recovery of 55% in SAX-treated rats. ConclusionThe CGRP analogue, SAX, has a cardioprotective effect in this rat model of myocardial infarction, improving myocardial perfusion recovery after chronic occlusion of the coronary artery.

Human macrophages directly modulate iPSC-derived cardiomyocytes at healthy state and congenital arrhythmia model in vitro.

The electrophysiological regulation of cardiomyocytes (CMs) by the cardiac macrophages (MΦs) has been recently described as an unconventional role of MΦs in the murine heart. Investigating the molecular and physiological modulation of CM by MΦ is critical to understand the novel mechanisms behind cardiac disorders from the systems perspective and to develop new therapeutic approaches. Here, we developed an in vitro direct coculture system to investigate the cellular and functional interaction between human-induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) and monocyte-derived MΦs both in healthy-state and congenital arrhythmia disease model associated with SCN5A ion channel mutations. Congenital arrhythmia patient-derived (P) and healthy individual-derived control (C) monocytes and derived MΦs exhibited distinct M1- and M2-like polarization-related gene expression pattern. The iPSC-CMs and MΦs formed direct membrane contacts in cocultures demonstrated by time-lapse imaging, scanning electron microscopy, and immunolabeling. The intracellular Ca2+ transients were observed in iPSC-CMs and MΦs when in contact with each other. Interestingly, the C-MΦs in direct contact with C-CMs significantly accelerated the contraction rates, demonstrating the positive chronotropic effect of MΦs on healthy cardiac cultures. Furthermore, the MΦs carrying the SCN5A gene mutation significantly enhanced the arrhythmic events in both C-CMs and P-CMs, implying that the sodium channel mutation in the MΦ is important for the CM function. Importantly, when C-MΦs were coupled to tachycardic P-CMs, the contraction frequency drastically decreased, and rhythmicity enhanced implicating the amelioration of the disease phenotype in vitro. Consequently, our results indicated the functional regulatory role of MΦs on human iPSC-CM contractility by membrane contacts in a physiologically relevant in vitro coculture model of both steady-state and arrhythmia. Our findings could serve as a valuable source for the development of effective immunoregulatory therapies for cardiac arrhythmia in the future.