Caffeine is one of the most used ergogenic aid for physical exercise and sports. However, its mechanism of action is still controversial. The adenosinergic hypothesis is promising due to the pharmacology of caffeine, a nonselective antagonist of adenosine A1 and A2A receptors. We now investigated A2AR as a possible ergogenic mechanism through pharmacological and genetic inactivation. Forty-two adult females (20.0 ± 0.2 g) and 40 male mice (23.9 ± 0.4 g) from a global and forebrain A2AR knockout (KO) colony ran an incremental exercise test with indirect calorimetry (V̇O2 and RER). We administered caffeine (15 mg/kg, i.p., nonselective) and SCH 58261 (1 mg/kg, i.p., selective A2AR antagonist) 15 min before the open field and exercise tests. We also evaluated the estrous cycle and infrared temperature immediately at the end of the exercise test. Caffeine and SCH 58621 were psychostimulant. Moreover, Caffeine and SCH 58621 were ergogenic, that is, they increased V̇O2max, running power, and critical power, showing that A2AR antagonism is ergogenic. Furthermore, the ergogenic effects of caffeine were abrogated in global and forebrain A2AR KO mice, showing that the antagonism of A2AR in forebrain neurons is responsible for the ergogenic action of caffeine. Furthermore, caffeine modified the exercising metabolism in an A2AR-dependent manner, and A2AR was paramount for exercise thermoregulation.
The natural plant alkaloid caffeine (1,3,7-trimethylxantine) is one of the most common ergogenic substances for physical activity practitioners and athletes1,2,3,4,5,6,7,8,9,10. Caffeine increases endurance1,8,9,10,11,12, intermittent7,13,14 and resistance4,15 exercise in humans. In rodents, its ergogenic effects are conserved because caffeine increases running time on the treadmill at constant16,17 and accelerated speeds18,19. Sports sciences promote nonselective phosphodiesterase (PDE) inhibition7,8 and increased calcium mobilization2,7,8 as mechanisms for these ergogenic effects. However, the primary pharmacological effect of caffeine is the nonselective antagonism of adenosine A1 and A2A receptors (A1R, A2AR)20,21,22,23.
Adenosine can act as an inhibitory modulator of the Central Nervous System (CNS) associated with tiredness and drowsiness24,25,26,27,28,29. During exercise, circulating ADP/AMP/adenosine levels increase due to ATP hydrolysis30,31. However, there is still no substantial evidence on the role of adenosine in exercise-induced fatigue. It is just known that the nonselective A1R and A2AR agonist 5′-(N-ethylcarboxamido)adenosine (NECA), injected into the rat brain, abolishes the ergogenic effects of caffeine16.
Since there is increasing evidence that the adenosine modulation system critically controls allostasis29 and A2AR have a crucial role in the ability of caffeine to normalize brain function30, we hypothesized that caffeine decreases fatigue during exercise through antagonism of A2AR in the CNS. We combined the use of pharmacology (SCH 58261 and caffeine) and transgenic mice with tissue-selective deletion of A2AR, to test this hypothesis in an incremental running test with indirect calorimetry (or ergospirometry). A2AR knockout (KO) mice allow assessing if the ergogenic effect of caffeine persists in the absence of A2AR; the use of SCH 58261, the current reference for A2AR antagonists32,33, allows directly assessing the ergogenic role of A2AR. SCH 58261 has excellent selectivity and affinity for A2AR32,33, and affords motor benefits in animal models of Parkinson’s disease as does caffeine, supporting the recent FDA approval of the A2AR antagonist Istradefylline for PD treatment33. Our goal is to assess the ergogenicity of A2AR using the pharmacological and genetic tools described above.
SCH 58261: pharmacological inactivation of A2AR is ergogenic
SCH 58261 was psychostimulant for males, but not for females, since SCH 58261 only increased male locomotion in the open field (F1,39 = 4.5, η2 = 0.1, β = 0.54, 95% CI 58.8–72.1, P < 0.05, Fig. 1A).
The running power of females (F7,77 = 221, P < 0.05, Fig. 1B) and males (F7,84 = 183, P < 0.05, Fig. 1B’) increased at each stage of the exercise test. Submaximal V̇O2 also increased to the maximum (V̇O2max, dotted line) of females (F8,77 = 168, P < 0.05, Fig. 1B) and males (F7,84 = 14.3, P < 0.05, Fig. 1B’). Female (F8,70 = 180, P < 0.05, Fig.S2A) and male (F8,70 = 164, P < 0.05, Fig.S2B) submaximal V̇CO2 kinetics was similar to V̇O2. SCH 58261 had no effect on these submaximal values.
We demonstrated for the first time that SCH 58261 is ergogenic since SCH 58261 increased V̇O2max (F1,36 = 27.7, η2 = 0.44, β = 0.99, 95% CI 0.16–0.2, P < 0.5, Fig. 1C) and running power (F1,35 = 55, η2 = 0.61, β = 1.0, 95% CI 1.0–1.3, P < 0.05, Fig. 1D) in both sexes.
SCH 58261 had no effect on increasing RER of females (F7,70 = 6.9, P < 0.5, η2 = 0.43, β = 0.99, Fig. 1E) and males (F7,84 = 9.4, η2 = 0.57, β = 0.99, P < 0.5, Fig. 1E’). Exercise test raised the animals’ core (F1,26 = 5.5, η2 = 0.17, β = 0.62, 95% CI 28.7–29.39, P < 0.05, Fig. 1F) and tail temperature (F1,22 = 81, η2 = 0.78, β = 0.99, 95% CI 24.2–25.6, P < 0.05, Fig. 1G), with no effect of SCH 58261. Figure 1 shows the heating of the mouse’s tail in post-exercise recovery (rec) in relation to rest. Three females at estrus (Fig.S3C) were excluded from temperature experiments due to large exercise-induced tail hyperthermia40. The previous results refer to females in diestrus (Fig.S3A), proestrus (Fig.S3B), and metestrus (Fig.S3D).
Neuronal A2AR antagonism is ergogenic
Caffeine increases exercise performance in rodents16,17,19,26,40 and humans1,4,8,9,10,11,12,13,14,15,24,28,51,52. Our results show the key role of A2AR in the ergogenic effects of caffeine using pharmacological and genetic tools. Thus, the potent and selective A2AR antagonist SCH 58261 displayed an ergogenic effect similar to that of caffeine, and the ergogenic effect of caffeine was abrogated in A2AR KO mice.
SCH 58261 and caffeine improved V̇O2max, running and critical power of wild type mice. These results are in line with the improved running time observed in caffeine-treated rats16,26,53 and mice19. Further evidence for the ergogenic effect of caffeine is based on its ability to increase muscle power and endurance output in rodents54,55,56,57,58. For the first time, we demonstrated that the selective antagonism of A2AR is ergogenic. Also, for the first time, we demonstrated that the genetic inactivation of A2AR impaired the ergogenic effects of caffeine. Tissue-specific A2AR KO selectively in forebrain neurons further allowed showing that these ergogenic effects of caffeine are due to the antagonism of A2AR in forebrain neurons. Thus, we suggest that caffeine decreases central fatigue during exercise. Moreover, caffeine decreased RER in the submaximal stages of the exercise test, an effect also abrogated in A2AR KO mice. However, exercise-induced core and tail hyperthermia were similar among animals treated with SCH 58261 or caffeine, except for A2AR KO mice, suggesting possible A1R-A2AR-mediated interactions56,57 in the temperature control51.
In summary, we have now demonstrated that A2AR antagonism is a mechanism of action for ergogenicity, as SCH 58261 was ergogenic. Furthermore, we showed that the antagonism of forebrain A2AR was the mechanism underlying the ergogenic effect of caffeine since caffeine was not ergogenic in fb-A2AR KO. The use of selective A2AR KO in forebrain neurons further reinforces the ergogenic role of caffeine in decreasing central fatigue, with possible involvement of decreased perceived exertion, pain, and mental fatigue in humans. Despite methodological limitations, our data further suggest that caffeine modified the exercising metabolism in an A2AR-dependent manner and that A2AR is essential for exercise thermoregulation.
Authors: Aderbal S.Aguiar Jr1,2*; Ana Elisa Speck1,2; Paula M. Canas1; RodrigoA. Cunha1,3
1 CNC‑Center for Neuroscience and Cell Biology, University of Coimbra, 3004‑504 Coimbra, Portugal.
2 Biology of Exercise Lab, Department of Health Sciences, UFSC-Federal University of Santa Catarina, Araranguá, SC 88905‑120, Brazil.
3 FMUC – Faculty of Medicine, University of Coimbra, 3004‑504 Coimbra, Portugal.
published in: Nature Scientific Reports 10, 13414 (2020).
Neuronal adenosine A2A receptors signal ergogenic effects of caffeine
Reductions in prefrontal oxygenation near maximal exertion may limit exercise performance by impairing executive functions that influence the decision to stop exercising; however, whether deoxygenation also occurs in motor regions that more directly affect central motor drive is unknown. Multichannel near-infrared spectroscopy was used to compare changes in prefrontal, premotor, and motor cortices during exhaustive exercise. Twenty-three subjects performed two sequential, incremental cycle tests (25 W/min ramp) during acute hypoxia [79 Torr inspired Po(2) (Pi(O(2)))] and normoxia (117 Torr Pi(O(2))) in an environmental chamber. Test order was balanced, and subjects were blinded to chamber pressure. In normoxia, bilateral prefrontal oxygenation was maintained during low- and moderate-intensity exercise but dropped 9.0 +/- 10.7% (mean +/- SD, P < 0.05) before exhaustion (maximal power = 305 +/- 52 W). The pattern and magnitude of deoxygenation were similar in prefrontal, premotor, and motor regions (R(2) > 0.94). In hypoxia, prefrontal oxygenation was reduced 11.1 +/- 14.3% at rest (P < 0.01) and fell another 26.5 +/- 19.5% (P < 0.01) at exhaustion (maximal power = 256 +/- 38 W, P < 0.01). Correlations between regions were high (R(2) > 0.61), but deoxygenation was greater in prefrontal than premotor and motor regions (P < 0.05). Prefrontal, premotor, and motor cortex deoxygenation during high-intensity exercise may contribute to an integrative decision to stop exercise. The accelerated rate of cortical deoxygenation in hypoxia may hasten this effect.
After approval from the Colorado Multiple Institutional Review Board, 25 active, healthy volunteers (23 men and 2 women) from the Denver, CO, metropolitan area (elevation 1,650 m) provided written, informed consent to participate in a larger study investigating the etiology of acute mountain sickness. Physical examinations, including blood and urine tests, were conducted to verify general health before participation. All study procedures followed ethical guidelines established by the Declaration of Helsinki.
Two incremental exercise tests were performed in an environmental chamber under ambient normobaric [normoxic (Norm), 610 Torr barometric pressure (Pb), 118 Torr inspired Po2 (PiO2)] and hypobaric [hypoxic (Hypox), 425 Torr Pb, 79 Torr PiO2] conditions to assess aerobic fitness for the larger study. Both tests were performed during a single chamber session to allow direct comparisons between Norm and Hypox without introduction of error from sensor placement/replacement. Tests were counterbalanced to control for order using a blinding strategy that varied chamber pressure to elicit similar sounds and changes in ear pressure during standardized 15-min ascent and descent periods. After arrival at the target Pb, 15 min were needed to adjust the cycle ergometer (Velotron Dynafit Pro, Racermate, Seattle, WA) and equip subjects with instrumental sensors (see below). Resting data were collected for 2 min before a 5-min warm-up at 50 W. Work rate was then incrementally increased using a 25 W/min ramp protocol to exhaustion. Subjects were blinded to elapsed time, power output, pedal revolutions per minute, and all physiological signals. Cool-down exercise was performed at 50 W for 5 min before chamber pressure was adjusted. After 15 min of ascent/decent and 15 min of rest at the second Pb, the protocol was repeated.
Twenty-three subjects (29 ± 8 yr of age, 73.7 ± 10.0 kg body wt, 181.7 ± 8.0 cm) completed both trials. Two subjects (with NIRS configuration 1) completed the Norm trial but were excluded from further study because of nausea and/or paresthesia during the subsequent depressurization period. Metabolic and power responses to both conditions were representative of physically fit, age-matched individuals (Table 1). Hypox reduced maximal V̇o2 and Wmax by 16 ± 6% and 21 ± 12%, respectively (P < 0.01). Order of trials did not affect the difference in maximal V̇o2 (P = 0.20) or Wmax (P = 0.95) between conditions, and 8 of 23 subjects were unable to retrospectively identify the order of testing.
Cortical deoxygenation during high-intensity exercise is not restricted to prefrontal regions of the brain. Deoxygenation in premotor and motor cortices may contribute to fatigue and/or decisions to stop exercising. Acute hypoxia exacerbates cortical deoxygenation and, thus, may hasten these effects. Future functional NIRS studies are needed to expand our understanding of the role of cerebral activity in exhaustive whole body exercise.
Authors: Andrew W. Subudhi, Brittany R. Miramon, Matthew E. Granger, and Robert C. Roach
University of Colorado Altitude Research Center, Denver and Colorado Springs Campuses, Colorado Springs
published in: Journal of Applied Physiology 106: 1153–1158, 2009
Frontal and motor cortex oxygenation during maximal exercise in normoxia and hypoxia (pdf)
Turbine engine blades are subjected to extreme conditions characterized by significant and simultaneous excursions in both stress and temperature. These conditions promote thermo-mechanical fatigue (TMF) crack growth which can significantly reduce component design life beyond that which would be predicted from isothermal/constant load amplitude results. A thorough understanding of the thermo-mechanical fatigue crack behavior in single crystal superalloys is crucial to accurately evaluate component life to ensure reliable operations without blade fracture through the use of “retirement for cause” (RFC). This research was conducted on PWA1484, a single crystal superalloy used by Pratt & Whitney for turbine blades.
Initially, an isothermal constant amplitude fatigue crack growth rate database was developed, filling a void that currently exists in published literature. Through additional experimental testing, fractography, and modeling, the effects of temperature interactions, load interactions, oxidation and secondary crystallographic orientation on the fatigue crack growth rate and the underlying mechanisms responsible were determined. As is typical in published literature, an R Ratio of 0.7 displays faster crack growth when compared to R = 0.1. The effect of temperature on crack growth rate becomes more pronounced as the crack driving force increases. In addition secondary orientation and R Ratio effects on crack growth rate were shown to increase with increasing temperature. Temperature interaction testing between 649°C and 982°C showed that for both R = 0.1 and 0.7, retardation is present at larger alternating cycle blocks and acceleration is present at smaller alternating cycle blocks. This transition from acceleration to retardation occurs between 10 and 20 alternating cycles for R = 0.1 and around 20 alternating cycles for R = 0.7. Load interaction testing showed that when the crack driving force is near KIC the overload size greatly influences whether acceleration or retardation will occur at 982°C. Semi-realistic spectrum testing demonstrated the extreme sensitivity that relative loading levels play on fatigue crack growth life while also calling into question the importance of dwell times. A crack trajectory modeling approach using blade primary and secondary orientations was used to determine whether crack propagation will occur on crystallographic planes or normal to the applied load.
Crack plane determination using a scanning electron microscope enabled verification of the crack trajectory modeling approach. The isothermal constant amplitude fatigue crack growth results fills a much needed void in currently available data. While the temperature and load interaction fatigue crack growth results reveal the acceleration and retardation that is present in cracks growing in single crystal turbine blade materials under TMF conditions. This research also provides a deeper understanding of the failure and deformation mechanisms responsible for crack growth during thermo-mechanical fatigue. The crack path trajectory modeling will help enable “Retirement for Cause” to be used for critical turbine engine components, a drastic improvement over the standard “safe-life” calculations while also reducing the risk of catastrophic failure due to “chunk liberation” as a function of time. Leveraging off this work there exists the possibility of developing a “local approach” to define a crack growth forcing function in single crystal superalloys.
Author: Benjamin Scott Adair, Dissertation; George W. Woodruff School of Mechanical Engineering; Georgia Institute of Technology; August, 2013
Model O2iM – Oxygen Deficiency Safety Monitor:
The Oxigraf state of the art Oxygen Deficiency Monitor, the Model O2iM, is a fast response, accurate and reliable safety monitor for oxygen displacement monitoring in Quantum Computer Laboratory, MRI, NMR, and liquid nitrogen and helium storage facilities. Our reliable solid state sensor does not require routine maintenance or factory calibration, and the O2iM is equipped with an automatic/programmable auto-calibration system. The system easily interfaces with alarm system, HVAC controls, and building management systems.
Oxigraf Case Study:
State-of-the-art helium (and other rare gases) recovery, purification and liquefaction systems are required for operation of Helium-3/Helium-4 milli Kelvin dilution refrigerators in modern Quantum Computer Laboratories, liquid helium superconducting magnets (such as NMRs, MRIs, etc.), MEG systems for medical applications, cryogenic measurement cryostats, various size helium and cryogenic vacuum facilities.
Reliable solutions for sampling gas from remote locations in a Helium Processing Facility are needed in order to monitor equipment and personnel safety. During their operations, helium processing facilities are dealing with the presence of cryogenic nitrogen and helium, which presents oxygen deficiency hazards. Oxygen deficiency in the workplace can lead to blackouts, cause falls, and present more serious health risks — some of which can be fatal. The Oxigraf expert’s team can be brought in to help eliminate the risk of oxygen depletion.
The Oxigraf Model O2iM, which has a high-flow pump option and allows for sampling from long distances. This sensor allows for continual monitoring of the clients’ facilities atmosphere from a safe location, and provides local alarms and interfaces with sophisticated safety features to prevent hazards such as cryogenic spills, which can lead to rapid displacement of breathing air.
Oxigraf’s top-of-the-line oxygen deficiency monitor is flexible and efficient, and provided the client with a reliable, immediate oxygen alarm for concentrations of less than 19.5%. It also eliminated the need for frequent recalibration or replacement of oxygen sensors, as well as the comprehensive, time-consuming maintenance often involved in sampling systems. The risk of false alarms and alarm failures can also be eliminated.
This unique sensor features a rapid response time of less than a second. The built-in pump draws gas remotely, allowing for these quick response times. In fact, we offer the best response speed/signal in the industry, and can add multiplexors (valving) in order to monitor four or more locations from over 100 feet away.The transit time of the gas sample through the sampling tube may be 1 second per meter of sampling tube with our standard pump or using our high flow option, a much faster response is possible on long tubing lengths. The high flow pump operates at a much faster rate and pre-fetches samples.
Additionally, this sensor is insensitive to movement, temperature and pressure changes, has auto-calibration for absolute accuracy, and includes options for multi-port and high-flow sampling. It also features a remote display and optional battery backup to allow for proper functioning during power interruptions. In addition, it can be fitted with a Z-Purge system, which allows the unit to be used in Class 1 Div 2 hazardous areas. The monitor includes a sampling pump, hydrophobic filter, and flow sensor, while the microprocessor controller maintains the flow at a constant value.
When comparing the Oxigraf O2iM sensor to other O2 sensing solutions, it can be determined that O2iM is “the champion,” allowing for reliable performance 24/7. Oxigraf customers are particularly impressed with the unique engineering of the “Pre-Fetch” high-flow pump option, which allows for the monitoring of distant sample locations while maintaining fast response times.
Typical O2iM Installation:
Oxigraf has over 20 years of experience producing laser gas sensors and instruments, and is the leading manufacturer of laser absorption spectroscopy sensors for oxygen gas measurement and analysis. Oxigraf O2iM Oxygen Safety Monitors have been widely adapted in hundreds of facilities since 2004, replacing a wide range of less reliable electrochemical sensors. Oxigraf O2 and CO2 sensors, in particular, have been widely adapted by OEM manufacturers of medical respiratory gas monitors in order to measure breath waveforms, end-tidal gas values, anaerobic thresholds, VO2 maxs, and non-invasive cardiac outputs. For more information on our sensors, or to speak with an expert about your specific monitoring needs, contact the team today.
Please download your Oxigraf Case Study: Oxygen Monitor for Quantum Computer Helium Laboratory
The accurate measurement of oxygen consumption (VO2) and energy expenditure (EE) may be helpful to optimize the treatment of critically ill patients. However, current techniques are limited in their ability to accurately quantify these end points in infants due to a low VO2, low tidal volume, and rapid respiratory rate. This study describes and validates a new device intended to perform in this size range.
We created a customized device that quantifies inspiratory volume using a pneumotachometer and concentrations of oxygen and carbon dioxide gas in the inspiratory and expiratory limbs. We created a customized algorithm to achieve precise time alignment of these measures, incorporating bias flow and compliance factors. The device was validated in 3 ways. First, we infused a certified gas mixture (50% oxygen/50% carbon dioxide) into an artificial lung circuit, comparing measured with simulated VO2 and carbon dioxide production (VCO2) within a matrix of varying tidal volume (4–20 mL), respiratory rate (20–80 bpm), and fraction of inspired oxygen (0.21–0.8). Second, VO2, VCO2, and EE were measured in Sprague Dawley rats under mechanical ventilation and were compared to simultaneous Douglas bag collections. Third, the device was studied on n = 14 intubated, spontaneously breathing neonates and infants, comparing measured values to Douglas measurements. In all cases, we assessed for difference between the device and reference standard by linear regression and Bland–Altman analysis.
In vitro, the mean ± standard deviation difference between the measured and reference standard VO2 was +0.04 ± 1.10 (95% limits of agreement, −2.11 to +2.20) mL/min and VCO2 was +0.26 ± 0.31 (−0.36 to +0.89) mL/min; differences were similar at each respiratory rate and tidal volume measured, but higher at fraction of inspired oxygen of 0.8 than at 0.7 or lower. In rodents, the mean difference was −0.20 ± 0.55 (−1.28 to +0.89) mL/min for VO2, +0.16 ± 0.25 (−0.32 to +0.65) mL/min for VCO2, and −0.84 ± 3.29 (−7.30 to +5.61) kcal/d for EE. In infants, the mean VO2 was 9.0 ± 2.5 mL/kg/min by Douglas method and was accurately measured by the device (bias, +0.22 ± 0.87 [−1.49 to +1.93] mL/kg/min). The average VCO2 was 8.1 ± 2.3 mL/kg/min, and the device exhibited a bias of +0.33 ± 0.82 (−1.27 to +1.94) mL/kg/min. Mean bias was +2.56% ± 11.60% of the reading for VO2 and +4.25% ± 11.20% of the reading for VCO2; among 56 replicates, 6 measurements fell outside of the 20% error range, and no patient had >1 of 4 replicates with a >20% error in either VO2 or VCO2.
This device can measure VO2, VCO2, and EE with sufficient accuracy for clinical decision-making within the neonatal and pediatric size range, including in the setting of tachypnea or hyperoxia.
Authors: Nachman, Einav BS*; Clemensen, Peter MS*,†; Santos, Katheryn BS*; Cole, Alexis R. BS*; Polizzotti, Brian D. PhD*,‡; Hofmann, Grace RRT§; Leeman, Kristen T. MD‡,∥; van den Bosch, Sarah J. MS*; Kheir, John N. MD*,‡
*Department of Cardiology, Boston Children’s Hospital, Boston, Massachusetts
†Department of Research and Development, InnoCC, Glamsbjerg, Denmark
‡Department of Pediatrics, Harvard Medical School, Boston, Massachusetts
Departments of §Respiratory Care
∥Newborn Medicine, Boston Children’s Hospital, Boston, Massachusetts