Simultaneous oral and inhalational intake of molecular hydrogen additively suppresses signaling pathways in rodents.

Molecular hydrogen (H2) is an agent with potential applications in oxidative stress-related and/or inflammatory disorders. H2 is usually administered by inhaling H2-containing air (HCA) or by oral intake of H2-rich water (HRW). Despite mounting evidence, the molecular mechanism underlying the therapeutic effects and the optimal method of H2 administration remain unclear. Here, we investigated whether H2 affects signaling pathways and gene expression in a dosage- or dose regimen-dependent manner. We first examined the H2 concentrations in blood and organs after its administration and found that oral intake of HRW rapidly but transiently increased H2 concentrations in the liver and atrial blood, while H2 concentrations in arterial blood and the kidney were one-tenth of those in the liver and atrial blood. In contrast, inhalation of HCA increased H2 equally in both atrial and arterial blood. We next examined whether H2 alters gene expression in normal mouse livers using DNA microarray analysis after administration of HCA and HRW. Ingenuity Pathway Analysis revealed that H2 suppressed the expression of nuclear factor-kappa B (NF-κB)-regulated genes. Western blot analysis showed that H2 attenuated ERK, p38 MAPK, and NF-κB signaling in mouse livers. Finally, we evaluated whether the changes in gene expression were influenced by the route of H2 administration and found that the combination of both HRW and HCA had the most potent effects on signaling pathways and gene expression in systemic organs, suggesting that H2 may act not only through a dose-dependent mechanism but also through a complex molecular network.

Maintenance of upright standing posture during trunk rotation elicited by rapid and asymmetrical movements of the arms.

Nine healthy subjects standing upright, initiated small, medium and large (S, M and L conditions, respectively) forward movements of their right (Rt) arm together with backward movements of their left (Lt) arm. They also performed medium-size movements while holding a 3 kg dumbbell in each hand (D condition). Movements started with the arm hanging alongside the body and ended when the shoulder reached a desired orientation. The arm and trunk movements were videotaped and recorded by accelerometers taped to the wrists, shoulders, and hips bilaterally. The torque around the vertical axis was measured using a force-plate on which the subjects stood. EMGs were recorded with surface electrodes bilaterally from the shoulder, trunk, and thigh muscles. Trajectories of the center of foot pressure were measured in additional experiments. In association with arm movements, there was a small counterclockwise (ccw: the Rt shoulder forward and the Lt backward) rotation of the trunk, followed by large alternate rotations of the trunk, first clockwise (cw) and subsequently ccw. The intervals from the hand acceleration to the shoulder and hip accelerations were, respectively, 0+/-15 ms (mean and S.D. for all subjects) and -17+/-15 ms. The force-plate showed initial cw and later ccw torques 63+/-41 ms after hand acceleration. The EMGs of the Rt hamstrings (Ham) and Lt rectus femoris (RFem) were followed by those of the Lt Ham and Rt RFem which, respectively preceded the alternate trunk rotations. The integrated EMGs and torques increased with increasing amplitude of arm movement and load. The integrated torques increased in the order of S, M, L, and D conditions. The integrated EMGs of the thigh muscles correlated with the integrated torques (medians: r=0.880, 0.696, 0.785, and 0.688, respectively, in the Rt Ham, Lt Ham, Rt RFem, and Lt RFem). The trajectories of the center of foot pressure showed variations, initially toward the Rt side and then the Lt side which, respectively coincided with the initial and later phases of the trunk rotations and the muscle activation. The trunk muscles were generally coactivated between the Rt and Lt muscles, and the integrated EMGs increased with increasing the integrated torques. Our results showed that alternate rotations of the upper trunk, produced by rapid arm movements, were transmitted to the hip in part due to cocontraction of trunk muscles, and each pair of hip joint muscles contributed to the maintenance of the standing posture by stabilizing the hip joints against alternating trunk rotations.

Effects of hypoxia and hypoxic training on 8-hydroxydeoxyguanosine and glutathione levels in the liver.

The effects of hypoxia and hypoxic training on 8-hydroxydeoxyguanosine (8-OHdG), reduced glutathione (GSH), and oxidized glutathione (GSSG) levels and on glutathione reductase (GR) activity in the liver of rats were evaluated. Rats were divided into 3 groups: a hypoxia and exercise (HE) group, a hypoxia and sedentary (HS) group, and a normoxia and sedentary (NS) group. The liver 8-OHdG levels were lower in the HE and HS groups compared with the NS group (P <.05). No significant difference between in the liver 8-OHdG levels in the HE and HS groups were found. However, the liver GSH level in the HS group was lower than that in the NS group (P <.05), and the HE group had significantly higher levels of liver GSH than the HS group (P <.01). The activity of liver GR in the HS group was lower than that of the NS group (P <.05). Moreover, the liver GR activity of the HE group was significantly higher than that of the HS group (P <.01). No significant difference in liver GR activity between the HE and NS groups was noted. In conclusion, the present study confirmed that moderate hypoxia and hypoxic training attenuated liver DNA damage and decreased liver GSH levels and GR activity. These results indicate that moderate hypoxia and hypoxic training result in decreased oxidative stress.

Coactivation in arm and shoulder muscles during voluntary fixation of a single joint.

The objective of this study was to determine the organization of coactivation in the arm and shoulder muscles. Normal human subjects made alternate movements of a joint in the horizontal plane, either adduction-abduction of the second finger and shoulder, ulnar-radial deviation of the wrist, or extension-flexion of the elbow, during which they fixed a focal joint while decreasing the movement amplitude and increasing the fixation strength. They varied the fixation strength at four different levels up to the maximum. The focal-joint angle, and surface electromyograms (EMGs) from the intrinsic hand, antebrachial, upper-arm, and shoulder muscles were recorded. EMGs in the phase of fixation were quantified by integration after rectification. The degree of coactivation among the muscles was evaluated by calculating correlation coefficients across the integrated EMGs. There were correlations in the integrated EMGs among focal-joint muscles (FJMs), and also between one of the FJMs and the muscles distal and/or proximal to the FJMs: in the finger fixation between the hand and antebrachial muscles, in the wrist fixation between the antebrachial and hand/upper-arm muscles, in the elbow fixation between the upper-arm and antebrachial/shoulder muscles, and in the shoulder fixation between the shoulder and upper-arm muscles. Moderate or slight correlations were seen in muscles more distant from FJMs. Our results indicate that the longitudinal distance from FJMs in the shoulder and arm muscles is an important factor in determining levels of coactivation. This is discussed in relation to the fact that neighboring muscles share joints with FJMs.

Effects of long-term hypoxia and hypoxic exercise on brain monoamine levels in rat.

This study clarified the effects of long-term hypoxia and hypoxic exercise on monoamines in the whole brain, and in four specific regions of the rat brain. The male Wistar rat progenitors (P1 group) were randomly assigned to three groups: hypoxia (16.0% oxygen) and exercise (MHE-P1), hypoxia and sedentary (MHS-P1), normoxia and sedentary (MNS-P1). The male children of P1 (the first generation of hypoxic rats; F1) were randomly divided into two groups: hypoxia and exercise (MHE-F1) and hypoxic sedentary (MHS-F1). The monoamines of whole brain were measured in P1 males, and monoamines of cerebellum, frontal lobe, striatum and hippocampus were measured in F1 males. The monoamine levels of MHE-P1 were significantly lower than those of MHS-P1 and MNS-P1. No significant difference was found in monoamine levels between MHS-P1 and MNS-P1. Epinephrine, norepinephrine, and dopamine levels of the MHE-F1 group significantly decreased in the frontal lobe, cerebellum and striatum, compare with the other groups (hypoxic and sedentary; normoxic and sedentary, respectively). These monoamines in the hippocampus were not influenced by the hypoxia or hypoxic exercise conditions. This study suggests that long-term hypoxic exercise decreased monoamine levels in whole brain, and that sensitivity to hypoxia and hypoxic exercise differed according to brain region.

Rapid acceleration of the lower arm correlates with agonist EMGs during the initial phase.

Fifty-eight healthy subjects made rapid elbow extensions to a target over 54 degrees. Angular acceleration was measured and surface electromyograms (EMGs) were recorded from the antagonistic muscles using monopolar rather than bipolar electrode configurations. Marked individual differences were found in the peak value of the first derivative of acceleration (dAcc/dt_Pk). The dAcc/dt_Pk correlated with both quantitative and qualitative properties of the agonist EMGs, but not with those of the antagonist EMG. The agonist EMGs, integrated until the moment of dAcc/dt_Pk, were positively correlated with dAcc/dt_Pk. The interval between EMG onset and EMG peak decreased with increasing dAcc/dt_Pk. The duration of the initial negative phase in the EMGs, which was considered to index the time required to recruit high-threshold MUs, decreased with increasing dAcc/dt_Pk. The results indicate that the ability to rapidly accelerate the lower arm varies across subjects, probably due in part to individual differences in the neural capacity to drive the agonists.

Nitric oxide concentrations in gas emanating from the tails of obese rats.

This study was undertaken to investigate the effects of oral L-arginine administration and exercising training on the NO concentration emanating from rat tail and NOx in plasma. Obese (fa/fa) Zucker rats (n = 22) were divided into four groups: (1) oral L-arginine administration (A) (n = 6), (2) exercise training (E), (3) exercise training + L-arginine administration (E + A) (n = 5), and (4) non-exercise training + non-L-arginine administration (N) (n = 6). The control (+/+) Zucker rats (n = 22) were also divided into the same four groups. The body weight of the E + A and the A groups was significantly lower than that of the N group. The NO concentration emitted from the tail was higher in the L-arginine (E + A and A) groups than in the non-L-arginine (E and N) groups in both obese and control rats. Exercise training did not affect the skin gas NO concentration in either obese or control rats. Plasma NOx concentrations in four obese rats were significantly higher than those observed in control rats. Exercise training did not influence the level of plasma NOx in obese or control rats. In conclusion, this study confirmed that L-arginine administration increases the skin gas NO concentration and obesity increases the plasma NOx level. The plasma NOx concentrations were not affected by L-arginine administration or exercise training in obese or control rats.

Acetone concentration in gas emanating from tails of diabetic rats.

This study investigated the effects of diabetic rats induced by streptozotocin (STZ) on acetone concentration emanating from the tail of a rat. Experiments were carried out with male Wistar rats (9 weeks of age, 220 - 250 g body weight). Glucose concentration in the blood was 10.8 ± 0.7 mmol/l for the control group and 39.6 ± 2.4 mmol/l for the diabetic group. ß-Hydroxybutyrate concentration in blood was 218 ± 52 µmol/l for the control group and 1439 ± 101 µmol/l for the diabetic group. Both glucose and ß-hydroxybutyrate concentrations in the blood of the diabetic group were significantly higher than those of the control group (p < 0.001). Skin gas acetone concentration emanated from rat tail was 124 ± 46 ppb for control and 1134 ± 417 ppb for diabetic. Skin gas acetone concentration emitted from the tail of a rat with diabetes was significantly higher than that from a rat in the control group (p < 0.001). The result indicates that skin acetone emanating from a rat tail is a useful parameter to use for insulin-dependent diabetes (type I).

Influence of cycle exercise on acetone in expired air and skin gas.

This study investigated the influence of cycle exercise on acetone concentration in expired air and skin gas. The subjects for this experiment were eight healthy males. Subjects performed a continuous graded exercise test on a cycle ergometer. The workloads were 360 (1.0 kg), 720 (2.0 kg), 990 (2.75 kg) kgm/min, and each stage was 5 min in duration. A pedaling frequency of 60 rpm was maintained. Acetone concentration was analyzed by gas chromatography. The acetone concentration in expired air and skin gas during exercise at 990 kgm/min intensity was significantly increased compared with the basal level. The skin-gas acetone concentration at 990 kgm/min significantly increased compared with the 360 kgm/min (P < 0.05). The acetone excretion of expired air at 720 kgm/min and 990 kgm/min significantly increased compared with the basal level (P < 0.05). Acetone concentration in expired air was 4-fold greater than skin gas at rest and 3-fold greater during exercise (P < 0.01). Skin gas acetone concentration significantly related with expired air (r = 0.752; P < 0.01). This study confirmed that the skin-gas acetone concentration reflected that of expired air.

The Relationship between Exercise Intensity and Lactate Concentration on the Skin Surface.

We examined the relationship between skin surface lactate concentration on working muscle and heart rate during continuous graded cycling exercise. Sixteen healthy male volunteers participated in this study. A plastic container with 100 µl 1% ethanol was put on the skin surface on the belly of rectus femoris muscle. The workloads were 300, 600, 900 and 1080 (or 990) kpm/min, and each stage was 5 min in duration. Sample collections were performed at rest, during exercise, and recovery. The lactate concentration during exercise significantly increased compared to the basal level (p<0.05 or p<0.001). Skin surface lactate concentration was found to correlate significantly with heart rate at the exercise intensity of 360 kpm/min (r=-0.52, p<0.05), 720 kpm/min (r=-0.74, p<0.01) and 900 kpm (r=-0.53, p<0.05). This study confirmed that 1) the increase in lactate concentration on the skin surface on working muscle is associated with increase in exercise intensity (heart rate), and 2) the skin surface lactate concentration on the working muscle can be used as a parameter of exercise intensity in each subject.

Influence of dynamic hand-grip exercise on acetone in gas emanating from human skin.

This study investigated the effects of dynamic hand-grip exercise on skin-gas acetone concentration. The subjects for this experiment were seven healthy males. In the first experiment, to ascertain the reproducibility of the results for the skin-gas acetone concentration test, the skin gas was collected four times from one subject. In the second experiment, all subjects performed three different types of exercise (Exercises I-III) for a duration of 60 s. Exercise I was performed at 10 kg with one contraction every 3 s. Exercise II was 30 kg with one contraction every 3 s. Exercise III was 10 kg with one contraction per second. Acetone concentration was analyzed by gas chromatography. In the first experiment, reasonable reproducibility was obtained in measurements of skin-gas acetone concentration during the hand-grip exercise. In the second experiment, acetone concentration in skin gas during hand-grip exercise II was significantly higher than the basal level. Although skin-gas acetone levels increased in all subjects during exercises I and III, a significant difference was not found. No significant difference was found in skin-gas acetone concentration during dynamic hand-grip exercise among exercises I, II, and III. This study confirmed that skin-gas acetone levels increase during dynamic hand-grip exercise.

Effects of Hypoxia on Nitric Oxide (NO) in Skin Gas and Exhaled Air.

This study confirmed the effects of hypoxia on nitric oxide (NO) concentrations in skin gas and exhaled air. NO concentrations in skin gas and exhaled air were measured by a chemiluminescence analyzer. Arterial oxygen saturation (SpO2) of the right forefinger was determined using an oxygen saturation monitor. The M ± SEM of NO concentrations in skin gas at 20.93% (control), 15.1% and 14.8% oxygen concentrations were 23.7 ± 3.6, 32.3 ± 4.7 and 36.2 ± 5.2 ppb, respectively. M ± SEM of NO concentrations in exhaled air at 20.93% (control), 15.1%, and 14.8% were 25.0 ± 5.1, 35.01 ± 5.6 and 44.9 ± 7.2 ppb, respectively. There was no significant difference in NO concentration at the absolute value of skin gas and exhaled air between normoxia and hypoxia. But significant increase was found at relative changes in skin gas at 15.1% (p<0.01) and 14.8% (p<0.01) oxygen content compared with control. Significant increase was also found at relative changes in exhaled air at 15.1% (p<0.01) and 14.8% (p<0.01) oxygen content compared with control. In conclusion, we confirmed that exposure to hypoxia elicits an increase in NO concentrations at relative changes of skin gas and exhaled air compared to normoxia.

Effect of hypoxia on norepinephrine of various tissues in rats.

OBJECTIVE: To determine the effects of hypoxia and hypoxic exercise (HE) on the norepinephrine levels of various tissues in rats. METHODS: Male Wistar rats were randomly assigned to 3 groups: an HE group (n = 6), a hypoxic-sedentary (HS) group (n = 6), and a normoxic-sedentary (NS) group (n = 6). The HE rats had access, ad lib, to an exercise wheel for 8 weeks. HE and HS rats were maintained in a normobaric hypoxic chamber with an FIO2 of 16%. Norepinephrine levels were measured and compared in liver, heart, diaphragm, soleus, and gastrocnemius tissues from the 3 groups. RESULTS: Liver norepinephrine levels in the HE and HS groups were significantly lower than the levels in the NS group (P < .05). No significant difference was found in liver norepinephrine levels between the HE and the HS groups. The heart norepinephrine levels in the NS group were significantly lower than the levels in the HE (P < .01) and HS groups (P < .01). In contrast, no significant differences were found in the norepinephrine levels for the diaphragm and soleus muscle among the 3 groups. The norepinephrine levels in the gastrocnemius white muscles were significantly higher in the HS group than in the HE (P < .05) and NS groups (P < .01). P < .01 represents a significant difference at the level of 1%. CONCLUSIONS: This study demonstrated that hypoxia and HE both elicit a decreased sympathetic response in the liver tissue of male Wistar rats but cause an increased response in heart tissue. These results suggest that the sympathetic responses to long-term hypoxia and HE training are different in various rat tissues.

Comparison of t-PA and u-PA levels in maximal treadmill and deep-water running.

BACKGROUND: The differences of urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen activator (t-PA) between maximal treadmill and deep-water running have not been reported. The purpose of this investigation was to compare u-PA and t-PA levels during maximal treadmill and deep-water running. METHODS: Six male subjects carried out two maximal exercises, one on a treadmill and the other running in deep water using a vest. The u-PA, t-PA, total plasminogen activator inhibitor (PAI-1), epinephrine, and norepinephrine in plasma and lactate and ammonia in blood concentrations were measured after maximal exercise. RESULTS: The blood lactate and ammonia concentrations were significantly higher in treadmill running than in deep-water running during recovery following exercise (P < 0.05). At 1 min after exercise, the plasma epinephrine, norepinephrine levels, and the u-PA and t-PA levels were higher in treadmill running compared with that in deep-water running (P < 0.05). No significant difference between the two runs was found in PAI-1 level. CONCLUSION: The maximal treadmill running induced a greater increase in u-PA and t-PA levels than maximal deep-water running.