Daniel C. Sigg

Molecular Architecture of the Human Sinus Node Insights Into the Function of the Cardiac Pacemaker

Background— Although we know much about the molecular makeup of the sinus node (SN) in small mammals, little is known about it in humans. The aims of the present study were to investigate the expression of ion channels in the human SN and to use the data to predict electrical activity. Methods and Results— Quantitative polymerase chain reaction, in situ hybridization, and immunofluorescence were used to analyze 6 human tissue samples. Messenger RNA (mRNA) for 120 ion channels (and some related proteins) was measured in the SN, a novel paranodal area, and the right atrium (RA). The results showed, for example, that in the SN compared with the RA, there was a lower expression of Nav1.5, Kv4.3, Kv1.5, ERG, Kir2.1, Kir6.2, RyR2, SERCA2a, Cx40, and Cx43 mRNAs but a higher expression of Cav1.3, Cav3.1, HCN1, and HCN4 mRNAs. The expression pattern of many ion channels in the paranodal area was intermediate between that of the SN and RA; however, compared with the SN and RA, the paranodal area showed greater expression of Kv4.2, Kir6.1, TASK1, SK2, and MiRP2. Expression of ion channel proteins was in agreement with expression of the corresponding mRNAs. The levels of mRNA in the SN, as a percentage of those in the RA, were used to estimate conductances of key ionic currents as a percentage of those in a mathematical model of human atrial action potential. The resulting SN model successfully produced pacemaking. Conclusions— Ion channels show a complex and heterogeneous pattern of expression in the SN, paranodal area, and RA in humans, and the expression pattern is appropriate to explain pacemaking.

Facial warming increases the threshold for shivering.

A decrease of 1-2 degrees C core temperature provides protection against cerebral ischemia. However, shivering usually prevents reduction in core temperature in unanesthetized patients. Therefore, it was tested whether facial and airway heating increases the shivering threshold and enables core cooling in unanesthetized patients. Nine trials were performed on seven healthy male volunteers. Each subject was positioned supine on a circulating-water mattress (8-15 degrees C) with a convective-air coverlet (15-18 degrees C) extending from the neck to the feet. A dynamic study protocol governed by individualized physiological responses was used. Focal facial (and airway) warming was employed to suppress involuntary motor activity (muscle tensing, shivering) and, thereby, enabling noninvasive cooling to lower the core temperature. The following parameters were monitored: 1) heart rate, 2) blood pressure, 3) core temperature (tympanic, axilla, and rectal), 4) cutaneous temperatures, and 5) a subjective shiver index (scale 1-10). In three, electromyograms and infrared thermographs were also obtained. Upon cooling without facial and airway warming, involuntary motor activity increased until it was widespread. This vigorous motor activity prevented any significant lowering of core temperature or caused it to slightly increase. Subsequently, in all subjects, within seconds after the application of facial focal warming, motor activity was suppressed almost completely, and within minutes core temperatures significantly decreased. Preliminary studies described here indicate that focal facial warming applied during active whole body cooling to initiate mild hypothermia might minimize the need to pharmacologically suppress involuntary motor activity. Such a procedure might be useful for initiating as soon as possible (such as during emergency transport), cerebral mild hypothermia in order to maximize protection and thus improve outcome in neurologically injured patients (head trauma, stroke).

Pericardial delivery of omega-3 fatty acid: a novel approach to reducing myocardial infarct sizes and arrhythmias

Basic and clinical evidence suggests that omega-3 (n-3) polyunsaturated fatty acids (PUFAs) decrease fatal arrhythmias and infarct sizes. This study investigated if pericardial delivery of n-3 PUFAs would protect the myocardium from ischemic damages and arrhythmias. Acute myocardial infarctions were induced in 23 pigs with either 45 min balloon inflations or clamp occlusions of the left anterior descending coronary arteries and 180 min reperfusion. Docosahexaenoic acid (C22:6n-3, DHA, 45 mg), one of the main n-3 PUFAs in fish oil, was infused within the pericardial space only during the 40-min stabilizing phase, 45 min ischemia and initial 5 min reperfusion. Hemodynamics and cardiac functions were very similar between the DHA-treated and control groups. However, DHA therapy significantly reduced infarct sizes from 56.8 +/- 4.9% for controls (n = 12) to 28.8 +/- 7.9% (P < 0.01) for DHA-treated animals (n = 11). Compared with controls, DHA-treated animals significantly decreased heart rates and reduced ventricular arrhythmia scores during ischemia. Furthermore, three (25%) control animals experienced eight episodes of ventricular fibrillation (VF), and two died subsequent to unsuccessful defibrillation. In contrast, only 1 (9%) of 11 DHA-treated pigs elicited one episode of VF that was successfully converted via defibrillation to normal rhythm; thus, mortality was reduced from 17% in the controls to 0% in the DHA-treated animals. These data demonstrate that pericardial infusion of n-3 PUFA DHA can significantly reduce both malignant arrhythmias and infarct sizes in a porcine infarct model. Pericardial administration of n-3 PUFAs could represent a novel approach to treating or preventing myocardial infarctions.

Opioid preconditioning: myocardial function and energy metabolism.

BACKGROUND Opioid receptor agonists are involved in ischemic preconditioning and natural hibernation. The aim of this study was to determine whether pretreatment with D-Ala2-Leu5-enkephalin or morphine confers cardioprotection in large mammalian hearts. We assessed myocardial functional recovery and global energy metabolism after ischemic cold storage. METHODS After pretreatment with D-Ala2-Leu5-enkephalin, morphine sulfate, or saline (n = 6 each), swine hearts were excised and stored for 75 minutes at 4 degrees C, then reperfused in a four-chamber isolated working heart apparatus. Serial myocardial biopsies were performed to assess cellular energy metabolism. RESULTS Improved systolic (cardiac output, contractility) and diastolic (tau) left ventricular functions were observed in hearts pretreated with D-Ala2-Leu5-enkephalin or morphine. These benefits were not correlated with changes in high-energy phosphate levels. Cardiac enzyme leakage (creatine kinase, troponin-I) was similar among treated and control groups. Lactate efflux increased significantly in controls, but not in opioid-pretreated hearts (p < 0.01) at 75 minutes of reperfusion. CONCLUSIONS D-Ala2-Leu5-enkephalin and morphine pretreatments improve postischemic function after cold storage of swine hearts. Postischemic lactate reduction, but not high-energy phosphate levels, may account for the observed cardioprotective effects.

Cardiac I-1c Overexpression With Reengineered AAV Improves Cardiac Function in Swine Ischemic Heart Failure

Cardiac gene therapy has emerged as a promising option to treat advanced heart failure (HF). Advances in molecular biology and gene targeting approaches are offering further novel options for genetic manipulation of the cardiovascular system. The aim of this study was to improve cardiac function in chronic HF by overexpressing constitutively active inhibitor-1 (I-1c) using a novel cardiotropic vector generated by capsid reengineering of adeno-associated virus (BNP116). One month after a large anterior myocardial infarction, 20 Yorkshire pigs randomly received intracoronary injection of either high-dose BNP116.I-1c (1.0 × 10(13) vector genomes (vg), n = 7), low-dose BNP116.I-1c (3.0 × 10(12) vg, n = 7), or saline (n = 6). Compared to baseline, mean left ventricular ejection fraction increased by 5.7% in the high-dose group, and by 5.2% in the low-dose group, whereas it decreased by 7% in the saline group. Additionally, preload-recruitable stroke work obtained from pressure-volume analysis demonstrated significantly higher cardiac performance in the high-dose group. Likewise, other hemodynamic parameters, including stroke volume and contractility index indicated improved cardiac function after the I-1c gene transfer. Furthermore, BNP116 showed a favorable gene expression pattern for targeting the heart. In summary, I-1c overexpression using BNP116 improves cardiac function in a clinically relevant model of ischemic HF.

Shiver Suppression Using Focal Hand Warming in Unanesthetized Normal Subjects

Background A decrease of 1 or 2°C in core temperature may provide protection against cerebral ischemia. However, during corporeal cooling of unanesthetized patients, the initiation of involuntary motor activity (shiver) prevents the reduction of core temperature. The authors’ laboratory previously showed that focal facial warming suppressed whole-body shiver. The aim of the current study was to determine whether the use of hand warming alone could suppress shiver in unanesthetized subjects and hence potentiate core cooling. Methods Subjects (n = 8; healthy men) were positioned supine on a circulating water mattress (8–15°C) with a convective-air coverlet (14°C) extending from their necks to their feet. A dynamic protocol was used in which focal hand warming was used to suppress involuntary motor activity, enabling noninvasive cooling to decrease core temperatures. The following parameters were monitored: (1) heart rate; (2) blood pressure; (3) core temperature (rectal, tympanic); (4) cutaneous temperature and heat flux; (5) subjective shiver level (SSL scale 0–10) and thermal comfort index (scale 0–10); (6) metabolic data (n = 6); and (7) electromyograms. Results During cooling without hand warming, involuntary motor activity increased until it was widespread. After subjects reported whole-body shiver (SSL ≥ 7), applied hand warming, in all cases, reduced shiver levels (SSL ≤ 3), decreased electromyographic root mean square amplitudes, and allowed core temperature to decrease from 37.0 ± 0.2 to 35.9 ± 0.5°C (measured rectally). Conclusions Focal hand warming seems to be valuable in minimizing or eliminating the need to suppress involuntary motor activity pharmacologically when it is desired to induce or maintain mild hypothermia; it may be used in conjunction with facial warming or in cases in which facial warming is contraindicated.

Glucagon-like peptide-1 enhances cardiac L-type Ca2+ currents via activation of the cAMP-dependent protein kinase A pathway

BackgroundGlucagon-like peptide-1 (GLP-1) is a hormone predominately synthesized and secreted by intestinal L-cells. GLP-1 modulates multiple cellular functions and its receptor agonists are now used clinically for diabetic treatment. Interestingly, preclinical and clinical evidence suggests that GLP-1 agonists produce beneficial effects on dysfunctional hearts via acting on myocardial GLP-1 receptors. As the effects of GLP-1 on myocyte electrophysiology are largely unknown, this study was to assess if GLP-1 could affect the cardiac voltage-gated L-type Ca2+ current (ICa).MethodsThe whole-cell patch clamp method was used to record ICa and action potentials in enzymatically isolated cardiomyocytes from adult canine left ventricles.ResultsExtracellular perfusion of GLP-1 (7-36 amide) at 5 nM increased ICa by 23 ± 8% (p < 0.05, n = 7). Simultaneous bath perfusion of 5 nM GLP-1 plus 100 nM Exendin (9-39), a GLP-1 receptor antagonist, was unable to block the GLP-1-induced increase in ICa; however, the increase in ICa was abolished if Exendin (9-39) was pre-applied 5 min prior to GLP-1 administration. Intracellular dialysis with a protein kinase A inhibitor also blocked the GLP-1-enhanced ICa. In addition, GLP-1 at 5 nM prolonged the durations of the action potentials by 128 ± 36 ms (p < 0.01) and 199 ± 76 ms (p < 0.05) at 50% and 90% repolarization (n = 6), respectively.ConclusionsOur data demonstrate that GLP-1 enhances ICa in canine cardiomyocytes. The enhancement of ICa is likely via the cAMP-dependent protein kinase A mechanism and may contribute, at least partially, to the prolongation of the action potential duration.

The potential for increased risk of infection due to the reuse of convective air-warming/cooling coverlets

Background: The use of convective air warming and/or cooling for the prevention of hypothermia or to induce hypothermia is growing rapidly. To date, there is no information available as to the potential risks for infection associated with either the postsurgical reuse or the repositioning of coverlets closer to the wound. We hypothesized that use of coverlets either intra‐ or postoperatively leads to increased contamination.

Hibernation induction trigger reduces hypoxic damage of swine skeletal muscle

A link between the cardioprotective benefits of pharmacological preconditioning and natural mammalian hibernation is considered to involve the cellular activation of opioid receptors and subsequent opening of KATP channels. In previous studies, we have demonstrated the protective effects of specific δ‐opioid agonists against porcine cardiac ischemia/reperfusion injury. We hypothesize here that preincubation with hibernation induction trigger (HIT) should confer a similar protection in skeletal muscles. Therefore, muscle bundles from swine were pretreated with plasma from hibernating woodchucks (HWP) for 30 min, then exposed to hypoxia for 90 min and reoxygenation for 120 min. Stimulated twitch forces were assessed. The functional effects of pretreatment with nonhibernation (summer) woodchuck plasma, a KATP blocker, or opioid antagonist were also studied. During the reoxygenation period, significantly greater force recoveries were observed only for bundles pretreated with HWP; this response was blocked by naloxone (P < 0.05). We conclude that HIT pretreatment could be used to confer protection against hypoxia/reperfusion injury of skeletal muscles of nonhibernators; it could potentially be utilized to prevent injury during surgical procedures requiring ischemia.

Images of the human coronary sinus ostium obtained from isolated working hearts

Due to increasing interest in using the coronary venous system for placement of intracardiac devices, the functional anatomy of the coronary sinus ostium is clinically important. Using Plegisol cardioplegia and stan-dard cardiac surgery procedures, six human hearts deemed not viable for transplant were explanted to an isolated heart apparatus. A modified Krebs-Henseleit buffer was used as a blood substitute to sustain the hearts, allowing for visualization of internal structures of the functioning hearts. Video footage of the coronary sinus ostia was obtained using a 6-mm diameter flexible videoscope inserted into the hearts through the superior vena cava or the right atrial appendage. A wide range of coronary sinus morphologies was observed including remnant Thebesian valves covering approximately 50% of the coronary sinus ostium and a large fenestrated Thebesian valve covering greater than 50% of the coronary sinus ostium. These images demonstrate why difficulties are sometimes encountered while cannulating the coronary sinus during surgical procedures. Figure 1 shows still images of the coronary sinus ostia of six human hearts. Each image represents a single frame captured from beta video recordings. In each image, the ostium of the coronary sinus is marked “O” and the Thebesian valve is marked “v.” Images in Figures 1A , 1B, and 1D show well-developed Thebesian valves covering large portions of the coronary sinus ostium. Images in Figures 1C, 1E, and 1F show remnant Thebesian valves. Figure 2 illustrates serial images showing the movement of Thebesian valves that cover the coronary sinus ostia in three hearts. Consecutive frames in images in Figures 2A and 2B are 0.033 seconds apart and frames in images in Figure 2C are 0.067 seconds apart. The image in Figure 2A is a different view of the Thebesian valve shown in Figure 1A. The Thebesian valves in images in Figures 2A and 2B completely cover the coronary sinus ostium during systole, except for the small fenestrations in both valves. The Thebesian valve image in Figure 2C, although large enough to cover the entire coronary sinus ostium, never actually completely covers the ostium.