Garrett L. Peltonen

Greater muscle protein synthesis and mitochondrial biogenesis in males compared with females during sprint interval training

Improved endurance exercise performance in adult humans after sprint interval training (SIT) has been attributed to mitochondrial biogenesis. However, muscle protein synthesis (MPS) and mitochondrial biogenesis during SIT have not been measured, nor have sex‐specific differences. We hypothesized that males and females would have similar rates of MPS, mitochondrial biogenesis, and synthesis of individual proteins during SIT. Deuterium oxide (D2O) was orally administered to 21 adults [11 male, 10 female; mean age, 23±1 yr; body mass index (BMI), 22.8±0.6 kg/m2; mean± se] for 4 wk, to measure protein synthesis rates while completing 9 sessions of 4–8 bouts of 30 s duration on a cycle ergometer separated by 4 min of active recovery. Samples of the vastus lateralis were taken before and 48 h after SIT. SIT increased maximum oxygen uptake (VO2max, males 43.4±2.1–44.0±2.3; females 39.5±0.9–42.5±1.3 ml/kg/min; P=0.002). MPS was greater in the males than in the females in the mixed (~150%; P < 0.001), cytosolic (~135%; P=0.038), and mitochondrial (~135%; P=0.056) fractions. The corresponding ontological clusters of individual proteins were significantly greater in the males than in the females (all P<0.00001). For the first time, we document greater MPS and mitochondrial biogenesis during SIT in males than in females and describe the synthetic response of individual proteins in humans during exercise training.—Scalzo, R. L., Peltonen, G. L., Binns, S. E., Shankaran, M., Giordano, G. R., Hartley, D. A., Klochak, A. L., Lonac, M. C., Paris, H. L. R., Szallar, S. E., Wood, L. M., Peelor, F. F., III, Holmes, W. E., Hellerstein, M. K., Bell, C., Hamilton, K. L., Miller, B. F. Greater muscle protein synthesis and mitochondrial biogenesis in males than in females during sprint interval training. FASEB J. 28, 2705–2714 (2014). www.fasebj.org

Sympathetic inhibition attenuates hypoxia induced insulin resistance in healthy adult humans

•  In low‐oxygen environments, such as high‐altitude, control of blood sugar is disrupted. Further, the activity of the sympathetic nervous system is known to increase when the availability of oxygen is decreased. •  We have investigated the possibility that the increase in sympathetic activity is partially responsible for the disruption in blood sugar control. •  Using gasbags filled with low‐oxygen gas, together with a commonly used blood pressure medication (clonidine) that inhibits the sympathetic nervous system, we have shown that breathing low oxygen disrupts blood sugar control, and that this disruption is prevented when the nervous system is inhibited. •  This finding has important implications for people travelling to high altitudes, and for people who suffer from conditions characterized by low oxygen, such as sleep apnoea and lung diseases.

Regulators of Human White Adipose Browning: Evidence for Sympathetic Control and Sexual Dimorphic Responses to Sprint Interval Training

The conversion of white adipose to the highly thermogenic beige adipose tissue has been proposed as a potential strategy to counter the unfavorable consequences of obesity. Three regulators of this conversion have recently emerged but information regarding their control is limited, and contradictory. We present two studies examining the control of these regulators. Study 1: In 10 young men, the plasma concentrations of irisin and fibroblast growth factor 21 (FGF21) were determined prior to and during activation of the sympathetic nervous system via hypoxic gas breathing (FIO2 = 0.11). The measurements were performed twice, once with and once without prior/concurrent sympathetic inhibition via transdermal clonidine administration. FGF21 was unaffected by basal sympathetic inhibition (338±113 vs. 295±80 pg/mL; P = 0.43; mean±SE), but was increased during hypoxia mediated sympathetic activation (368±135); this response was abrogated (P = 0.035) with clonidine (269±93). Irisin was unaffected by sympathetic inhibition and/or hypoxia (P>0.21). Study 2: The plasma concentration of irisin and FGF21, and the skeletal muscle protein content of fibronectin type III domain containing 5 (FNDC5) was determined in 19 young adults prior to and following three weeks of sprint interval training (SIT). SIT decreased FGF21 (338±78 vs. 251±36; P = 0.046) but did not affect FNDC5 (P = 0.79). Irisin was decreased in males (127±18 vs. 90±23 ng/mL; P = 0.045) and increased in females (139±14 vs. 170±18). Collectively, these data suggest a potential regulatory role of acute sympathetic activation pertaining to the browning of white adipose; further, there appears to be a sexual dimorphic response of irisin to SIT.

Total daily energy expenditure is increased following a single bout of sprint interval training

Regular endurance exercise is an effective strategy for healthy weight maintenance, mediated via increased total daily energy expenditure (TDEE), and possibly an increase in resting metabolic rate (RMR: the single largest component of TDEE). Sprint interval training (SIT) is a low‐volume alternative to endurance exercise; however, the utility of SIT for healthy weight maintenance is less clear. In this regard, it is feasible that SIT may evoke a thermogenic response above and beyond the estimates required for prevention of weight gain (i.e., >200–600 kJ). The purpose of these studies was to investigate the hypotheses that a single bout of SIT would increase RMR and/or TDEE. Study 1: RMR (ventilated hood) was determined on four separate occasions in 15 healthy men. Measurements were performed over two pairs of consecutive mornings; each pair was separated by 7 days. Immediately following either the first or third RMR measurement (randomly assigned) subjects completed a single bout of SIT (cycle ergometer exercise). RMR was unaffected by a single bout of SIT (7195 ± 285 kJ/day vs. 7147 ± 222, 7149 ± 246 and 6987 ± 245 kJ/day (mean ± SE); P = 0.12). Study 2: TDEE (whole‐room calorimeter) was measured in 12 healthy men, on two consecutive days, one of which began with a single bout of SIT (random order). Sprint exercise increased TDEE in every research participant (9169 ± 243 vs. 10,111 ± 260 kJ/day; P < 0.0001); the magnitude of increase was 946 ± 62 kJ/day (~10%). These data provide support for SIT as a strategy for increasing TDEE, and may have implications for healthy body weight maintenance.

Cerebrovascular regulation in men and women: stimulus-specific role of cyclooxygenase.

Greater cerebral artery vasodilation mediated by cyclooxygenase (COX) in female animals is unexplored in humans. We hypothesized that young, healthy women would exhibit greater basal cerebral blood flow (CBF) and greater vasodilation during hypoxia or hypercapnia compared to men, mediated by a larger contribution of COX. We measured middle cerebral artery velocity (MCAv, transcranial Doppler ultrasound) in 42 adults (24 women, 18 men; 24 ± 1 years) during two visits, in a double‐blind, placebo‐controlled design (COX inhibition, 100 mg oral indomethacin, Indo). Women were studied early in the follicular phase of the menstrual cycle (days 1–5). Two levels of isocapnic hypoxia (SPO2 = 90% and 80%) were induced for 5‐min each. Separately, hypercapnia was induced by increasing end‐tidal carbon dioxide (PETCO2) 10 mmHg above baseline. A positive change in MCAv (ΔMCAv) reflected vasodilation. Basal MCAv was greater in women compared to men (P < 0.01) across all conditions. Indo decreased baseline MCAv (P < 0.01) similarly between sexes. Hypoxia increased MCAv (P < 0.01), but ΔMCAv was not different between sexes. Indo did not alter hypoxic vasodilation in either sex. Hypercapnia increased MCAv (P < 0.01), but ΔMCAv was not different between sexes. Indo elicited a large decrease in hypercapnic vasodilation (P < 0.01) that was similar between sexes. During the early follicular phase, women exhibit greater basal CBF than men, but similar vasodilatory responses to hypoxia and hypercapnia. Moreover, COX is not obligatory for hypoxic vasodilation, but plays a vital and similar role in the regulation of basal CBF (~30%) and hypercapnic response (~55%) between sexes.

β-Adrenergic-mediated vasodilation in young men and women: Cyclooxygenase restrains nitric oxide synthase

We tested the hypothesis that women exhibit greater vasodilator responses to β-adrenoceptor stimulation compared with men. We further hypothesized women exhibit a greater contribution of nitric oxide synthase and cyclooxygenase to β-adrenergic-mediated vasodilation compared with men. Forearm blood flow (Doppler ultrasound) was measured in young men (n = 29, 26 ± 1 yr) and women (n = 33, 25 ± 1 yr) during intra-arterial infusion of isoproterenol (β-adrenergic agonist). In subset of subjects, isoproterenol responses were examined before and after local inhibition of nitric oxide synthase [N(G)-monomethyl-l-arginine (l-NMMA); 6 male/10 female] and/or cyclooxygenase (ketorolac; 5 male/5 female). Vascular conductance (blood flow ÷ mean arterial pressure) was calculated to assess vasodilation. Vascular conductance increased with isoproterenol infusion (P < 0.01), and this effect was not different between men and women (P = 0.41). l-NMMA infusion had no effect on isoproterenol-mediated dilation in men (P > 0.99) or women (P = 0.21). In contrast, ketorolac infusion markedly increased isoproterenol-mediated responses in both men (P < 0.01) and women (P = 0.04) and this rise was lost with subsequent l-NMMA infusion (men, P < 0.01; women, P < 0.05). β-Adrenergic vasodilation is not different between men and women and sex differences in the independent contribution of nitric oxide synthase and cyclooxygenase to β-mediated vasodilation are not present. However, these data are the first to demonstrate β-adrenoceptor activation of cyclooxygenase suppresses nitric oxide synthase signaling in human forearm microcirculation and may have important implications for neurovascular control in both health and disease.

Greater Beta-Adrenergic Receptor Mediated Vasodilation in Women Using Oral Contraceptives

Background: β-adrenergic receptors play an important role in mitigating the pressor effects of sympathetic nervous system activity in young women. Based on recent data showing oral contraceptive use in women abolishes the relationship between muscle sympathetic nervous system activity and blood pressure, we hypothesized forearm blood flow responses to a β-adrenergic receptor agonist would be greater in young women currently using oral contraceptives (OC+, n = 13) when compared to those not using oral contraceptives (OC–, n = 10). Methods: Women (18–35 years) were studied during the early follicular phase of the menstrual cycle (days 1–5) or placebo phase of oral contraceptive use. Forearm blood flow (FBF, Doppler ultrasound) and mean arterial blood pressure (MAP, brachial arterial catheter) were measured at baseline and during graded brachial artery infusion of the β-adrenergic receptor agonist, Isoproterenol (ISO), as well as Acetylcholine (ACH, endothelium-dependent vasodilation) and Nitroprusside (NTP, endothelium-independent vasodilation). Forearm vascular conductance was calculated (FVC = FBF/MAP, ml/min/100 mmHg) and the rise in FVC from baseline during infusion quantified vasodilation (ΔFVC = FVCinfusion − FVCbaseline). Results: ISO increased FVC in both groups (p < 0.01) and ISO-mediated ΔFVC was greater in OC+ compared to OC– (Main effect of group, p = 0.02). Expressing data as FVC and FBF resulted in similar conclusions. FVC responses to both ACH and NTP were also greater in OC+ compared to OC–. Conclusions: These data are the first to demonstrate greater β-adrenergic receptor-mediated vasodilation in the forearm of women currently using oral contraceptives (placebo phase) when compared to those not using oral contraceptives (early follicular phase), and suggest oral contraceptive use influences neurovascular control.

Concurrent Beet Juice and Carbohydrate Ingestion: Influence on Glucose Tolerance in Obese and Nonobese Adults

Insulin resistance and obesity are characterized by low nitric oxide (NO) bioavailability. Insulin sensitivity is improved with stimulation of NO generating pathways. Consumption of dietary nitrate (NO3−) increases NO formation, via NO3− reduction to nitrite (NO2−) by oral bacteria. We hypothesized that acute dietary nitrate (beet juice) ingestion improves insulin sensitivity in obese but not in nonobese adults. 12 nonobese (body mass index: 26.3 ± 0.8 kg/m2 (mean ± SE)) and 10 obese adults (34.0 ± 0.8 kg/m2) ingested beet juice, supplemented with 25 g of glucose (carbohydrate load: 75 g), with and without prior use of antibacterial mouthwash to inhibit NO3− reduction to NO2−. Blood glucose concentrations after beet juice and glucose ingestion were greater in obese compared with nonobese adults at 60 and 90 minutes (P = 0.004). Insulin sensitivity, as represented by the Matsuda Index (where higher values reflect greater insulin sensitivity), was lower in obese compared with nonobese adults (P = 0.009). Antibacterial mouthwash rinsing decreased insulin sensitivity in obese (5.7 ± 0.7 versus 4.9 ± 0.6) but not in nonobese (8.1 ± 1.0 versus 8.9 ± 0.9) adults (P = 0.048). In conclusion, insulin sensitivity was improved in obese but not in nonobese adults following coingestion of beet juice and glucose when oral bacteria nitrate reduction was not inhibited. Obese adults may benefit from ingestion of healthy nitrate-rich foods during meals.

The Effects of Sympathetic Inhibition on Metabolic and Cardiopulmonary Responses to Exercise in Hypoxic Conditions

OBJECTIVE Pre-exertion skeletal muscle glycogen content is an important physiological determinant of endurance exercise performance: low glycogen stores contribute to premature fatigue. In low-oxygen environments (hypoxia), the important contribution of carbohydrates to endurance performance is further enhanced as glucose and glycogen dependence is increased; however, the insulin sensitivity of healthy adult humans is decreased. In light of this insulin resistance, maintaining skeletal muscle glycogen in hypoxia becomes difficult, and subsequent endurance performance is impaired. Sympathetic inhibition promotes insulin sensitivity in hypoxia but may impair hypoxic exercise performance, in part due to suppression of cardiac output. Accordingly, we tested the hypothesis that hypoxic exercise performance after intravenous glucose feeding in a low-oxygen environment will be attenuated when feeding occurs during sympathetic inhibition. METHODS On 2 separate occasions, while breathing a hypoxic gas mixture, 10 healthy men received 1 hour of parenteral carbohydrate infusion (20% glucose solution in saline; 75 g), after which they performed stationary cycle ergometer exercise (~65% maximal oxygen uptake) until exhaustion. Forty-eight hours before 1 visit, chosen randomly, sympathetic inhibition via transdermal clonidine (0.2 mg/d) was initiated. RESULTS The mean time to exhaustion after glucose feeding both with and without sympathetic inhibition was not different (22.7 ± 5.4 minutes vs 23.5 ± 5.1 minutes; P = .73). CONCLUSIONS Sympathetic inhibition protects against hypoxia-mediated insulin resistance without influencing subsequent hypoxic endurance performance.

ATP‐mediated vasodilatation: all thanks to potassium?

Emerging evidence suggests circulating adenosine triphosphate (ATP) plays an important role in local blood flow control. ATP is primarily thought to bind to P2Y receptors on the endothelium, and activate downstream vasodilator pathways. Specific mechanisms behind ATP-mediated vasodilatation are currently unknown in humans; however, research supports the contribution of nitric oxide (NO), prostaglandins (PGs) and endothelial-derived hyperpolarization (EDH) via one or several potassium (K+) channels. Despite consistent findings in vitro, the contribution of NO and PGs to ATP-mediated vasodilatation in vivo is contradictory. Specifically, Crecelius et al. (2011) demonstrated NO and PGs to contribute to only ∼20–30% of the vasodilatory response to exogenous ATP in healthy humans, suggesting other mechanisms must be largely responsible for the increase in blood flow. Using evidence from in vitro research identifying a role for K+ channel activation in ATP-mediated vasodilation, Crecelius et al. (2012) continued to advance our understanding of local blood flow control in a recent article published in The Journal of Physiology. K+ efflux from small and intermediate-conductance Ca2+-activated K+ channels increases myoendothelial K+ concentration. This increase in K+ concentration results in activation of Na+/K+-ATPase and KIR channels leading to hyperpolarization of vascular smooth muscle cells. In vitro research has shown that application of ATP evokes local and conducted vasodilatation via these pathways. Thus, Crecelius et al. thought outside the NO/PG box by testing the hypotheses that ATP-mediated vasodilatation in the forearm is largely independent of NO and PG synthesis and occurs via Na+/K+ -ATPase and KIR channel activation in humans. To test their hypotheses, the authors assessed forearm vascular conductance (FVC) to intra-arterial infusions using standard venous occlusion plethysmography (VOP) procedures in 33 young, healthy adults. To confirm that ATP-mediated vasodilatation is largely independent of NO and PGs, and to focus on the role of K+ channels, Crecelius et al. used three separate but related protocols. First, the group tested the effect of combined NO synthase (NOS) and cyclooxygenase (COX) inhibition (l-NMMA and ketorolac, respectively) on ATP-mediated vasodilatation (n = 8). Confirming previous findings (Crecelius et al. 2011), combined NOS and COX inhibition significantly reduced FVC at rest, but not during ATP infusion. Thus, ATP vasodilatation appears to be largely independent of NO and PGs. The authors also examined K+-mediated vasodilatation using KCl. As expected, KCl infusion increased FVC; yet surprisingly, the relative response of KCl infusion was greater after NOS–COX inhibition. The second protocol examined the contribution of alternative downstream vascular control mechanisms to ATP-mediated vasodilatation via Na+/K+-ATPase and KIR channel inhibition. ATP (n = 8) or KCl (n = 6) were infused with and without ouabain and BaCl2 (Na+/K+-ATPase and KIR channel inhibitors, respectively). Co-infusion abolished forearm vasodilatation to KCl, and, as the authors hypothesized, oubain and BaCl2 attenuated vasodilatation in response to ATP (56% reduction). To test the independent effect of KIR channels on ATP-mediated smooth muscle relaxation, BaCl2 was administered alone in six additional subjects. The authors observed a significant reduction (51%) in the vasodilatory response to ATP. Taken together, inhibition of KIR had relatively the same effect as combined Na+/K+-ATPase and KIR inhibition, indicating KIR channels are a key player in ATP-mediated vasodilatation in the resting forearm circulation. However, the authors chose to compare double blockade of Na+/K+-ATPase and KIR channels vs. single blockade of KIR channels in separate research cohorts. Future studies might aim to examine the effect of single and double blockade in the same subset of subjects in order to control for inter-subject variability in blood flow responses to different pharmacological interventions. Additionally, figures directly comparing the two trials were not provided. A helpful approach to compare data from multiple protocols might be providing figures comparing percentage inhibition across doses and between various drug conditions. Such data presentation could provide readers with key information regarding the efficacy of inhibition and might allow insight into the relative importance of various K+ channels. In summary, Crecelius and colleagues provided novel and exciting research regarding specific mechanisms of ATP-mediated dilatation. They verified previous results (Crecelius et al. 2011) that blood flow responses to ATP in the forearm are largely independent of NO and PG synthesis. In addition, this research group was the first to identify vascular hyperpolarization in the forearm via KIR channel activation as one of the primary pathways underlying the vasodilator mechanisms of intravascular ATP in humans. Interestingly, blockade of KIR, with or without Na+/K+-ATPase blockade, did not completely abolish ATP-induced vasodilatation. One possible explanation for these findings is that ATP might have the ability to promote dilatation via auxiliary down-stream pathways. For example, ATP has been shown to stimulate epoxyeicosatrienoic acid (EETs) release from erythrocytes (Jiang et al. 2007), BKCa channel hyperpolarization of the smooth muscle, and subsequent vasodilatation independent of KIR channels. Therefore, a protocol utilizing blockade of KIR channels with and without blockade of cytochrome P450 epoxygenase (using miconazole or sulfaphenazole) could be considered in follow-up studies. This protocol would demonstrate the relative contribution of BKCa channel vasodilatation in response to ATP. The results from the current study have potential to provide insight to other applications regarding ATP-mediated vasodilatation such as exercise and disease. One exciting area to consider is the role of ATP dilatation and its associated pathways during steady-state exercise. ATP is thought to be released from red blood cells in response to a decrease in blood oxygen content and mechanical deformation and, thus, can contribute to the blood flow response seen with exercise. Future studies could test whether KIR channels are also a primary player in ATP-mediated dilatation during exercise. This study holds potential to provide clinical insight into patient populations such as those with hypertension. Adults with hypertension have been shown to exhibit endothelial dysfunction due, in part, to impairments in NO bioavailability, but other mechanisms contributing to the endothelial dysfunction have yet to be fully explored. Research in animal models suggests KIR channel function is impaired in the cerebral and mesenteric circulation during hypertension (Haddy et al. 2006). It is interesting to consider the potential for KIR channel impairments in other vascular beds, which could be explored in hypertensive humans using a protocol similar to that of the Crecelius group. If KIR channel function in response to ATP is indeed impaired in hypertensive humans, it might be interesting to consider the idea that expression and/or activity of BKCa channels is enhanced in this population. This could be in order to compensate for a reduction in KATP, Kv and KIR channel function in an attempt to ultimately restrict increases in vascular tone (Sobey, 2001). In conclusion, novel studies by Crecelius et al. uncovered one of the major players (KIR) of ATP-mediated vasodilatation. Their research allows for new avenues to test alternative downstream vasodilatory mechanisms and novel interactions between NO, PGs, EETs and KIR channels in a variety of vascular beds and cardiovascular conditions.