Effects of the single exposure to EMFs in glucose, lipids, and redox pair-metabolites in fed and fasted rats. Animals were subjected to a 15 min-exposure of 60 Hz-EMFs (time zero), and euthanized immediately after obtaining blood samples. In fed animals, the EMFs did not affect significantly serum level of glucose or that of TG, but decreased largely FFA serum levels (Fig. 1A). Moreover, EMFs exerted an opposite effect in the redox-pair metabolites: lactate was decreased and pyruvate was drastically increased in the serum; in addition, serum levels of AcAc were also significantly decreased (Fig. 1A).
In overnight fasted animals, in which the metabolic environment was changed, decreasing serum level of glucose, increasing those of FFA, and lactate and ketone bodies, the exposure to EMFs had not significant effects on these parameters, except that EMFs also decreased levels of FFA (Fig. 1B). Then, depending on the metabolic scenario, a single exposure to EMFs exerted significant metabolic changes in rats.
Time-course of EMFs effects on glucose, lactate, and pyruvate in fed and fasted rats. Since we did not find higher levels of glucose in animals subjected to a 15 min-exposure of 60 Hz-EMFs, as previously reported (20), we looked for a time-course of EMFs effects on some parameters. In fed animals, serum levels of glucose increased at 15 min after exposure termination, returned to the control levels (30 min), and started again to increase 60 min after exposure (Fig. 2A). In these animals, lactate was early decreased, and this significant diminution was maintained, thereafter (3 h, Fig. 2B). On the contrary, EMFs promoted three peaks of augmented serum levels of pyruvate at 0, 30, and 180 min (Fig. 2C), where the lactate/pyruvate ratio was largely decreased (not shown).
Interestingly, in animals with an overnight fast, a hyperglycemic status was observed after exposure to 60 Hz-EMFs, reaching the same higher blood levels for glucose as in fed animals, at the end of the experiment (Fig. 2A). However, lactate levels (which were much higher in these animals) did not decrease, but rather showed two increasing peaks at 30 and 180 min after treatment (Fig. 2B). Also interesting, EMFs promoted two increments of serum pyruvate at 30 and 120 min (Fig. 2C). Therefore, increases of serum glucose seem to coincide with lower lactate/pyruvate ratios (more oxidized state) after EMFs treatment (Fig. 2).
Curve of tolerance to glucose and levels of insulin and glucagon in rats treated with EMFs. Another set of rats were fasted overnight, administered with a glucose load of 2 g • kg− 1, and subjected to a 15 min-exposure of 60 Hz-EMFs. Control (sham) animals depicted a “typical” absorption and further decay of blood levels of glucose assumed to be due to its utilization. In rats treated with EMFs, we noted that, during the first 90 min, glucose tolerance was similar to that of the control group, but, at 120 min after glucose administration, a robust hyperglycemia occurred, which could be considered as a “diabetic-like response” curve (Fig. 3A). In this context, control animals elicited two peaks of insulin secretion, the first at 15 min and the second 2 h after treatment. Although EMFs indeed increased the first peak of insulin when compared with control animals, the second insulin peak was practically abolished by the treatment with EMFs (Fig. 3B). Interestingly, in control and treated animals, serum levels of glucagon showed an opposite pattern of release when compared to that of insulin. However, during the first 30 min of the tolerance curve, animals treated with EMFs had the lowest serum levels of this hormone (Fig. 3C). Therefore, the insulin/glucagon ratio could be involved in the hyperglycemic effect of EMFs in these rats.
In vivo oxidation of U-(14C)-glucose and its incorporation into glycogen and glycerol-containing lipids in rats treated with EMFs. Another set of fed rats received 0.5 mg of glucose-containing 4 µCi (148 mBq) of (U-14C)-glucose, and was subjected to a 15 min-exposure of 60 Hz-EMFs. Animals exposed to EMFs showed an increased glucose oxidation, as reflected by the production of 14CO2 (Fig. 4A), which was accompanied by a tendency of decreasing blood level of 14C-glucose (Fig. 4B) 30 min after EMFs treatment. Interestingly, synthesis of liver glycogen, as assessed by incorporation of glucose into glycogen, was decreased at 120 min after exposure (Fig. 4C), coinciding with the abatement of the second peak of insulin secretion (Fig. 3B). As to muscular glycogen, treatment with EMFs induced two decrements in its synthesis, at 30 and 120 min, respectively (Fig. 4D); these effects also seemed to be related with the initial serum insulin (Fig. 3B), as well as with the augmented glucose oxidation (Fig. 4A). Moreover, whereas EMFs promoted lipogenesis (as assessed by FFA esterification to 14C-glycerol) in the liver (120 min; Fig. 4E), in the epididymal adipose tissue there was a transient decrease of the rate of lipogenesis 60 min after EMFs exposure (Fig. 4F).
Correlations among serum glucose levels, insulin, glucagon, and redox-pair metabolites. We looked for possible relationships between the hyperglycemic status induced by EMFs exposure and the serum levels pancreatic hormones, as well as with redox-pair metabolites (Fig. 5). The levels of glucose in control and treated rats correlated straightly with fluctuations in insulin levels (r = 0.687, p < 0.01; Fig. 5A), with those of glucagon (r = 0.403, p < 0.05; Fig. 5B), and much better with the insulin/glucagon ratio (r = 0.965, p < 0.001; Fig. 5C). As to the lactate and pyruvate, serum glucose did not correlate with those of lactate (r = -0.132, n.s.; Fig. 5D), but a very high significant relationship was found with pyruvate (r = 0.993, p < 0.001; Fig. 5E); indeed, glucose level also showed an inverse correlation with the lactate/pyruvate ratio (r = -0.806, p < 0.001; Fig. 5F), which indicates the relevance of fluctuations in pyruvate levels. In the same context, glucose did not significantly correlate with β-OH-but (r = 0.146, n.s.; Fig. 5G), but correlated highly and directly with those levels for AcAc (r = -0.963, p < 0.001; Fig. 5H). Moreover, glucose also showed an inverse correlation with the β-OH-but /AcAc ratio (r = -0.584, p < 0.01; Fig. 5I); therefore, glucose increase was highly correlated with an enhancement in oxidized metabolic products, namely pyruvate and AcAc (Fig. 5).
Effects of chronic exposure to EMF on rat blood glucose, lipids, and parameters indicative of redox state and energy status. Most of the reported effects of EMFs on cell proliferation, cell signaling, oxidant status, or metabolic changes have been obtained through chronic exposure to EMFs in animals or isolated cells (2). Therefore, we searched whether the acute response to EMFs remains even after exposing the animals for 14 consecutive days to EMFs. For this, a set of rats was exposed for 15 min to 60 Hz-EMFs per day for two weeks; then, rats were fasted overnight at the 15th day, and treated again for the same time. Table 1 shows that this chronically-treated group also elicited a significant hyperglycemia, but the decrease in FFA serum levels was not seen. The lactate/pyruvate ratio was decreased and, consequently, the NAD/NADH ratio was augmented in both groups, but through different mechanism; in the group of a single exposure, pyruvate levels were increased, whereas in the chronic group, the levels of lactate were diminished (Table 1). Total levels for ketone bodies were not significantly changed in both groups, but, in the chronic group, the β-OH-but /AcAc ratio was decreased, indicating a more oxidized NAD/NADH status, presumably reflecting liver mitochondria redox state (Table 1). Finally, when examining blood energy parameters (ATP/ADP ratio, energy charge, and total adenine nucleotides), we did not find any significant difference in these indicators in either the “acute” or the “chronic” groups (Table 1). Therefore, chronic application of EMFs does not induce refractory actions on the effects of a single exposure of 60 Hz-EMFs to rats.
Table 1
Effects of chronic exposure to EMF on rat blood glucose, lipids, and parameters indicative of redox state and energy status.
| Fasted (single) Fasted (chronic) |
Parameter | Controls | With EMF | Control | With EMF |
Glucose (mg • dL− 1) | 77.1 ± 5.4 | 118.5 ± 14.9* | 72.3 ± 6.7 | 109.6 ± 4.6* |
TG (mg • dL− 1) | 167 ± 14 | 203 ± 26 | 187 ± 19 | 162 ± 20 |
FFA (µmoles • dL− 1) | 5.2 ± 0.5 | 3.2 ± 0.3* | 6.5 ± 0.8 | 6.1 ± 0.6 |
Lactate (mM) | 1.89 ± 0.37 | 2.01 ± 0.30 | 1.61 ± 0.25 | 0.89 ± 0.14* |
Pyruvate (mM) | 0.08 ± 0.02 | 0.19 ± 0.03* | 0.07 ± 0.02 | 0.13 ± 0.03 |
Lactate/Pyruvate | 23.6 ± 4.9 | 10.6 ± 1.8* | 22.5 ± 4.0 | 6.7 ± 1.6* |
NAD+/NADH (Cyto) | 382 ± 79 | 852 ± 136* | 392 ± 60 | 1316 ± 261* |
β-OH-butyrate (mM) | 0.58 ± 0.12 | 0.49 ± 0.06 | 0.33 ± 0.07 | 0.27 ± 0.06 |
Acetoacetate (mM) | 0.18 ± 0.02 | 0.24 ± 0.03 | 0.14 ± 0.02 | 0.23 ± 0.03 |
β-OH-but/AcAc | 3.2 ± 0.5 | 2.1 ± 0.3 | 2.4 ± 0.4 | 1.2 ± 0.2* |
NAD+/NADH (Mito) | 7.6 ± 1.1 | 11.5 ± 1.7 | 10.1 ± 1.7 | 19.6 ± 3.1* |
ATP (mM) | 0.77 ± 0.05 | 0.72 ± 0.04 | 0.71 ± 0.05 | 0.58 ± 0.05 |
ADP (mM) | 0.10 ± 0.01 | 0.09 ± 0.01 | 0.17 ± 0.02 | 0.16 ± 0.03 |
AMP (mM) | 0.06 ± 0.01 | 0.04 ± 0.01 | 0.09 ± 0.01 | 0.08 ± 0.02 |
ATP/ADP | 7.7 ± 0.8 | 8.0 ± 0.6 | 4.7 ± 0.5 | 3.6 ± 0.7 |
Total Nucleotides | 0.93 ± 0.12 | 0.85 ± 0.11 | 0.97 ± 0.14 | 0.82 ± 0.11 |
Energy charge | 0.88 ± 0.03 | 0.89 ± 0.04 | 0.81 ± 0.04 | 0.80 ± 0.03 |
Results are the mean ± SE of four individual observations per experimental group, which consisted in rats exposed to 60-Hz EMFs during 15 min/day for 14 consecutive days before the single (acute) exposure at the day of the experiment, and blood samples were taken 30 min after ending exposure. Abbreviations: β-OH-but, β-hydroxybutyrate, AcAc, acetoacetate, Cyto, cytoplasm, and Mito, mitochondria. Statistical significance: *p < 0.01 vs. control group. |