Computational insight into energy control balance by Ca2+ and cAMP-PKA signaling in pacemaker cells.

Cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) signaling controls sinoatrial node cell (SANC) function by affecting the degree of coupling between Ca2+ and membrane clocks. PKA is known to phosphorylate ionic channels, Ca2+ pump and release from the sarcoplasmic reticulum, and enzymes controlling ATP production in the mitochondria. While the PKA cytosolic targets in SANC have been extensively explored, its mitochondrial targets and its ability to maintain SANC energetic balance remain to be elucidated. To investigate the role of PKA in SANC energetics, we tested three hypotheses: (i) PKA is an important regulator of the ATP supply-to-demand balance, (ii) Ca2+ regulation of energetics is important for maintenance of NADH level and (iii) abrupt reduction in ATP demand first reduces the AP firing rate and, after dropping below a certain threshold, leads to a reduction in ATP. To gain mechanistic insights into the ATP supply-to-demand matching regulators, a modified model of mitochondrial energy metabolism was integrated into our coupled-clock model that describes ATP demand. Experimentally, increased ATP demand was accompanied by maintained ATP and NADH levels. Ca2+ regulation of energetics was found by the model to be important in the maintenance of NADH and PKA regulation was found to be important in the maintenance of intracellular ATP and the increase in oxygen consumption. PKA inhibition led to a biphasic reduction in AP firing rate, with the first phase being rapid and ATP-independent, while the second phase was slow and ATP-dependent. Thus, SANC energy balance is maintained by both Ca2+ and PKA signaling.

cAMP-PKA signaling modulates the automaticity of human iPSC-derived cardiomyocytes.

Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have been used to screen and characterize drugs and to reveal mechanisms underlying cardiac diseases. However, before hiPSC-CMs can be used as a reliable experimental model, the physiological mechanisms underlying their normal function should be further explored. Accordingly, a major feature of hiPSC-CMs is automaticity, which is regulated by both Ca2+ and membrane clocks. To investigate the mechanisms coupling these clocks, we tested three hypotheses: (1) normal automaticity of spontaneously beating hiPSC-CMs is regulated by local Ca2+ releases (LCRs) and cAMP/PKA-dependent coupling of Ca2+ clock to M clock; (2) the LCR period indicates the level of crosstalk within the coupled-clock system; and (3) perturbing the activity of even one clock can lead to hiPSC-CM-altered automaticity due to diminished crosstalk within the coupled-clock system. By measuring the local and global Ca2+ transients, we found that the LCRs properties are correlated with the spontaneous beat interval. Changes in cAMP-dependent coupling of the Ca2+ and M clocks, caused by a pharmacological intervention that either activates the ß-adrenergic or cholinergic receptor or upregulates/downregulates PKA signaling, affected LCR properties, which in turn altered hiPSC-CMs automaticity. Clocks' uncoupling by attenuating the pacemaker current If or the sarcoplasmic reticulum Ca2+ kinetics, decreased hiPSC-CMs beating rate, and prolonged the LCR period. Finally, LCR characteristics of spontaneously beating (at comparable rates) hiPSC-CMs and rabbit SAN are similar. In conclusion, hiPSC-CM automaticity is controlled by the coupled-clock system whose function is mediated by Ca2+-cAMP-PKA signaling.

Increase in Ca2+-Activated cAMP/PKA Signaling Prevents Hydroxychloroquine-Induced Bradycardia of the Cardiac Pacemaker.

Bradycardia or tachycardia are known side effects of drugs that limit their clinical use. The heart pacemaker function which control the heart rate under normal conditions is determined by coupled clock system. Thus, interfering with specific clock mechanism will affect other clock mechanisms through changes in interconnected signaling and can lead to rhythm disturbance. However, upregulation of a different clock components can compensate for this change. We focus here on hydroxychloroquine (HCQ), which has been shown effective in treating COVID-19 patients, however its bradycardic side effect limits its clinical use. We aim to decipher the mechanisms underlying the effect of HCQ on pacemaker automaticity, to identify a potential drug that will eliminate the bradycardia. We used isolated rabbit sinoatrial node (SAN) cells, human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and mouse SAN cells residing in SAN tissue. Further, we employed SAN cell computational model to suggest mechanistic insights of the effect of HCQ on pacemaker function. HCQ increased mean spontaneous beat interval and variability in all three models in parallel to slower intracellular kinetics. The computational model suggested that HCQ affects the pacemaker (funny) current (If), L-type Ca2+ current (ICa,L), transient outward potassium (Ito) and due to changes in Ca2+ kinetics, the sodium-calcium exchanger current (INCX). Co-application of 3'-isobutylmethylxanthine (IBMX) and HCQ prevented the increase in beat interval and variability in all three experimental models. The HCQ-induced increase in rabbit and mice SAN cell and hiPSC-CM spontaneous beat interval, can be prevented by a phosphodiester inhibitor that restores automaticity due to slower intracellular Ca2+ kinetics.

The Cardiovascular Reserve Index-A Noninvasive Clinical Insight Into Heat Intolerance.

OBJECTIVE: To noninvasively explore the heat intolerance condition during exercise-heat stress by assessing cardiovascular (CV) performance. DESIGN: Prospective study of participants undergoing a standard heat-tolerance test (HTT). SETTING: Institutional study. PARTICIPANTS: Ninety-five young males: 16 heat-intolerant (HI) and 79 heat-tolerant (HT). INTERVENTIONS: Cardiovascular performance during an HTT was estimated by heart rate (HR) and blood pressure measurements. MAIN OUTCOME MEASURES: The sensitivity of the cardiovascular reserve index (CVRI) and the dynamic heart rate reserve (dHRR) index to predict heat intolerance was compared. RESULTS: A significant difference in the CV reserve during exercise-heat stress was exhibited between the HI and the HT groups. Starting at a similar level, the reduction in the CV reserve at HTT endpoint was much greater in the HI than the HT individuals (P < 0.0001), as depicted by both the CVRI and the dHRR. This result indicates a greater utilization of the CV reserve by HI individuals. The CVRI is likely to be better predictor of heat intolerance than the dHRR because the partial area under the curve in the high sensitivity (>90%) region of its receiver operating characteristic curve is higher (93.2 vs 76.8). CONCLUSIONS: More than being a predictor, the CVRI may provide a new clinical insight into heat intolerance because it noninvasively characterizes the efficiency of an individual's thermoregulatory mechanism and hints that an impaired CV reserve might underlie heat intolerance. The CVRI provides a noninvasive measurement of thermoregulation, which has been long awaited to enable on-field studies and dynamic monitoring of heat-exposed task forces.

Measuring core body temperature with a non-invasive sensor.

In various occupations, workers may be exposed to extreme environmental conditions and physical activities. Under these conditions the ability to follow the workers' body temperature may protect them from overheating that may lead to heat related injuries. The "Dräger" Double Sensor (DS) is a novel device for assessing body-core temperature (Tc). The purpose of this study was to evaluate the accuracy of the DS in measuring Tc under heat stress. Seventeen male participants performed a three stage protocol: 30min rest in a thermal comfort environment (20-22°C, 50% relative humidity), followed by an exposure to a hot environment of 40°C, 40% relative humidity -30min at rest and 60min of exercise (walking on a treadmill at 5km/h and 2% elevation). Simultaneously temperatures measured by the DS (TDS) and by rectal temperature (Tre) (YSI-401 thermistor) were recorded and then compared. During the three stages of the study the average temperature obtained by the DS was within±0.3°C of rectal measurement. The correlation between TDS and Tre was significantly better during the heat exposures phases than during resting under comfort conditions. These preliminary results are promising for potential use of the DS by workers under field conditions and especially under environmental heat stress or when dressed in protective garments. For this goal, further investigations are required to validate the accuracy of the DS under various levels of heat stress, clothing and working levels.