Low Utilization of Synchronous Telemedicine in Pediatric Critical Care Interfacility Transport: Barriers and Lessons.

OBJECTIVE: The use of telemedicine has increased and may enhance the care of children during medical transport. We aimed to evaluate the feasibility of synchronous telemedicine connectivity before interfacility transport of critically ill children by a pediatric transport team. METHODS: We performed a prospective, observational feasibility study of the introduction of synchronous telemedicine into an established pediatric transport team from 2019 to 2020. The outcomes examined included connectivity, physician workload, transport team satisfaction, and patient care outcomes. RESULTS: Among 118 eligible transports, telemedicine was considered in 23 transports (19%), including 11 transports in which an attempt to connect was sought and 12 in which telemedicine activation was offered but not attempted. The median connection time was 2.9 minutes (interquartile range, 1.7-4.4 minutes), and clinical care was altered in 1 case. Connection failed in 2 cases (18.2%). In 50% of cases, concurrent medical control physician workload prevented activation. There were no perceived benefits in 41.7% of cases. Team members indicated the desire for future telemedicine use in only 54.6% of cases. CONCLUSIONS: We found low utilization of synchronous telemedicine in interfacility pediatric transport. The identified barriers included reliable connectivity, physician workload, and low perceived benefit. Lessons learned and future research suggestions are presented to mitigate these barriers.

Adrenergic Receptors in Individual Ventricular Myocytes: The Beta-1 and Alpha-1B Are in All Cells, the Alpha-1A Is in a Subpopulation, and the Beta-2 and Beta-3 Are Mostly Absent.

RATIONALE: It is unknown whether every ventricular myocyte expresses all 5 of the cardiac adrenergic receptors (ARs), ß1, ß2, ß3, α1A, and α1B. The ß1 and ß2 are thought to be the dominant myocyte ARs. OBJECTIVE: Quantify the 5 cardiac ARs in individual ventricular myocytes. METHODS AND RESULTS: We studied ventricular myocytes from wild-type mice, mice with α1A and α1B knockin reporters, and ß1 and ß2 knockout mice. Using individual isolated cells, we measured knockin reporters, mRNAs, signaling (phosphorylation of extracellular signal-regulated kinase and phospholamban), and contraction. We found that the ß1 and α1B were present in all myocytes. The α1A was present in 60%, with high levels in 20%. The ß2 and ß3 were detected in only ≈5% of myocytes, mostly in different cells. In intact heart, 30% of total ß-ARs were ß2 and 20% were ß3, both mainly in nonmyocytes. CONCLUSION: The dominant ventricular myocyte ARs present in all cells are the ß1 and α1B. The ß2 and ß3 are mostly absent in myocytes but are abundant in nonmyocytes. The α1A is in just over half of cells, but only 20% have high levels. Four distinct myocyte AR phenotypes are defined: 30% of cells with ß1 and α1B only; 60% that also have the α1A; and 5% each that also have the ß2 or ß3. The results raise cautions in experimental design, such as receptor overexpression in myocytes that do not express the AR normally. The data suggest new paradigms in cardiac adrenergic signaling mechanisms.

An Alpha-1A Adrenergic Receptor Agonist Prevents Acute Doxorubicin Cardiomyopathy in Male Mice.

Alpha-1 adrenergic receptors mediate adaptive effects in the heart and cardiac myocytes, and a myocyte survival pathway involving the alpha-1A receptor subtype and ERK activation exists in vitro. However, data in vivo are limited. Here we tested A61603 (N-[5-(4,5-dihydro-1H-imidazol-2-yl)-2-hydroxy-5,6,7,8-tetrahydronaphthalen-1-yl]methanesulfonamide), a selective imidazoline agonist for the alpha-1A. A61603 was the most potent alpha-1-agonist in activating ERK in neonatal rat ventricular myocytes. A61603 activated ERK in adult mouse ventricular myocytes and protected the cells from death caused by the anthracycline doxorubicin. A low dose of A61603 (10 ng/kg/d) activated ERK in the mouse heart in vivo, but did not change blood pressure. In male mice, concurrent subcutaneous A61603 infusion at 10 ng/kg/d for 7 days after a single intraperitoneal dose of doxorubicin (25 mg/kg) increased survival, improved cardiac function, heart rate, and cardiac output by echocardiography, and reduced cardiac cell necrosis and apoptosis and myocardial fibrosis. All protective effects were lost in alpha-1A-knockout mice. In female mice, doxorubicin at doses higher than in males (35-40 mg/kg) caused less cardiac toxicity than in males. We conclude that the alpha-1A-selective agonist A61603, via the alpha-1A adrenergic receptor, prevents doxorubicin cardiomyopathy in male mice, supporting the theory that alpha-1A adrenergic receptor agonists have potential as novel heart failure therapies.

A Myocardial Slice Culture Model Reveals Alpha-1A-Adrenergic Receptor Signaling in the Human Heart.

BACKGROUND: Translation of preclinical findings could benefit from a simple, reproducible, high throughput human model to study myocardial signaling. Alpha-1A-adrenergic receptors (ARs) are expressed at very low levels in the human heart, and it is unknown if they function. OBJECTIVES: To develop a high throughput human myocardial slice culture model, and to test the hypothesis that alpha-1A- ARs are functional in the human heart. METHODS: Cores of LV free wall 8 mm diameter were taken from 52 hearts (18 failing and 34 nonfailing). Slices 250 µm thick were cut with a Krumdieck apparatus and cultured using a rotating incubation unit. RESULTS: About 60 slices were cut from each LV core, and a typical study could use 96 slices. Myocyte morphology was maintained, and diffusion into the slice center was rapid. Slice viability was stable for at least 3 days in culture by ATP and MTT assays. The beta-AR agonist isoproterenol stimulated phospholamban phosphorylation, and the alpha-1A-AR agonist A61603 stimulated ERK phosphorylation, with nanomolar EC50 values in slices from both failing and nonfailing hearts. Strips cut from the slices were used to quantify activation of contraction by isoproterenol, A61603, and phenylephrine. The slices supported transduction by adenovirus. CONCLUSIONS: We have developed a simple, high throughput LV myocardial slice culture model to study signaling in the human heart. This model can be useful for translational studies, and we show for the first time that the alpha-1A-AR is functional in signaling and contraction in the human heart.

The alpha-1A adrenergic receptor agonist A61603 reduces cardiac polyunsaturated fatty acid and endocannabinoid metabolites associated with inflammation in vivo.

INTRODUCTION: Alpha-1-adrenergic receptors (α1-ARs) are G-protein coupled receptors (GPCRs) with three highly homologous subtypes (α1A, α1B, and α1D). Of these three subtypes, only the α1A and α1B are expressed in the heart. Multiple pre-clinical models of heart injury demonstrate cardioprotective roles for the α1A. Non-selective α1-AR activation promotes glycolysis in the heart, but the functional α1-AR subtype and broader metabolic effects have not been studied. OBJECTIVES: Given the high metabolic demands of the heart and previous evidence indicating benefit from α1A activation, we chose to investigate the effects of α1A activation on the cardiac metabolome in vivo. METHODS: Mice were treated for one week with a low, subpressor dose of A61603, a highly selective and potent α1A agonist. Cardiac tissue and serum were analyzed using a non-targeted metabolomics approach. RESULTS: We identified previously unrecognized metabolic responses to α1A activation, most notably broad reduction in the abundance of polyunsaturated fatty acids (PUFAs) and endocannabinoids (ECs). CONCLUSION: Given the well characterized roles of PUFAs and ECs in inflammatory pathways, these findings suggest a possible role for cardiac α1A-ARs in the regulation of inflammation and may offer novel insight into the mechanisms underlying the cardioprotective benefit of selective pharmacologic α1A activation.

ß-myosin heavy chain is induced by pressure overload in a minor subpopulation of smaller mouse cardiac myocytes.

RATIONALE: Induction of the fetal hypertrophic marker gene ß-myosin heavy chain (ß-MyHC) is a signature feature of pressure overload hypertrophy in rodents. ß-MyHC is assumed present in all or most enlarged myocytes. OBJECTIVE: To quantify the number and size of myocytes expressing endogenous ß-MyHC by a flow cytometry approach. METHODS AND RESULTS: Myocytes were isolated from the left ventricle of male C57BL/6J mice after transverse aortic constriction (TAC), and the fraction of cells expressing endogenous ß-MyHC was quantified by flow cytometry on 10,000 to 20,000 myocytes with use of a validated ß-MyHC antibody. Side scatter by flow cytometry in the same cells was validated as an index of myocyte size. ß-MyHC-positive myocytes constituted 3 ± 1% of myocytes in control hearts (n=12), increasing to 25 ± 10% at 3 days to 6 weeks after TAC (n=24, P<0.01). ß-MyHC-positive myocytes did not enlarge with TAC and were smaller at all times than myocytes without ß-MyHC (≈70% as large, P<0.001). ß-MyHC-positive myocytes arose by addition of ß-MyHC to α-MyHC and had more total MyHC after TAC than did the hypertrophied myocytes that had α-MyHC only. Myocytes positive for ß-MyHC were found in discrete regions of the left ventricle in 3 patterns: perivascular, in areas with fibrosis, and in apparently normal myocardium. CONCLUSIONS: ß-MyHC protein is induced by pressure overload in a minor subpopulation of smaller cardiac myocytes. The hypertrophied myocytes after TAC have α-MyHC only. These data challenge the current paradigm of the fetal hypertrophic gene program and identify a new subpopulation of smaller working ventricular myocytes with more myosin.

Functional alpha-1B adrenergic receptors on human epicardial coronary artery endothelial cells.

Alpha-1-adrenergic receptors (α1-ARs) regulate coronary arterial blood flow by binding catecholamines, norepinephrine (NE), and epinephrine (EPI), causing vasoconstriction when the endothelium is disrupted. Among the three α1-AR subtypes (α1A, α1B, and α1D), the α1D subtype predominates in human epicardial coronary arteries and is functional in human coronary smooth muscle cells (SMCs). However, the presence or function of α1-ARs on human coronary endothelial cells (ECs) is unknown. Here we tested the hypothesis that human epicardial coronary ECs express functional α1-ARs. Cultured human epicardial coronary artery ECs were studied using quantitative real-time reverse transcription polymerase chain reaction, radioligand binding, immunoblot, and (3)H-thymidine incorporation. The α1B-subtype messenger ribonucleic acid (mRNA) was predominant in cultured human epicardial coronary ECs (90-95% of total α1-AR mRNA), and total α1-AR binding density in ECs was twice that in coronary SMCs. Functionally, NE and EPI through the α1B subtype activated extracellular signal-regulated kinase (ERK) in ECs, stimulated phosphorylation of EC endothelial nitric oxide synthase (eNOS), and increased deoxyribonucleic acid (DNA) synthesis. These results are the first to demonstrate α1-ARs on human coronary ECs and indicate that the α1B subtype is predominant. Our findings provide another potential mechanism for adverse cardiac effects of drug antagonists that nonselectively inhibit all three α1-AR subtypes.

Pharmacological properties of the active metabolites of the antidepressants desipramine and citalopram.

Although major metabolites of some antidepressant drugs are known to be active, their pharmacological effects are poorly characterized. Two of the most selective antidepressants, desipramine (selectively inhibits norepinephrine reuptake) and citalopram (selectively inhibits serotonin reuptake) are frequently used in animal studies of antidepressant action, as well as being useful therapeutically. The primary aim of this study was to determine the affinity of desmethyldesipramine, an active metabolite of desipramine, for the rat norepinephrine and serotonin transporters, as well as for the rat alpha(2)-adrenoceptor. The pharmacological characteristics of desmethyldesipramine and desmethylcitalopram, an active metabolite of citalopram, were also determined for various human transporters and neurotransmitter receptors. Competition binding studies using [(3)H]nisoxetine and [(3)H]citalopram showed desipramine to be 25 times more selective for the rat norepinephrine as compared to serotonin transporter (6.2 nM vs. 158 nM) whereas desmethyldesipramine is 12 times more selective for the serotonin over the norepinephrine transporter (12.8 nM vs. 153 nM). Interestingly, the affinity of desmethyldesipramine for the serotonin transporter is similar to the affinity of desipramine for the norepinephrine transporter. Desipramine and desmethyldesipramine were found to have a lower affinity for the rat alpha(2A(D))-adrenoceptor than the transporters, suggesting that this receptor is not a major site of action for either compound. Thus, the pharmacological effects of desipramine in rats may be attributed not only to the inhibition of the norepinephrine transporter by desipramine but also to the inhibition of serotonin transporter by the active metabolite desmethyldesipramine.