Melatonin Suppression by Time-Varying and Time-Invariant

Downloaded by UNIV MASSACHUSETTS AMHERST on July 31, 2012 | http://pubs.acs.org Publication Date: May 5, 1995 | doi: 10.1021/ba-1995-0250.ch025

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Melatonin Suppression by Time-Varying and Time-Invariant Electromagnetic Fields Russel J. Reiter Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, TX 78284-7762

This chapter reviews the experimental evidence that shows that both time-varying and time-invariant electromagneticfieldexposures suppress the production and secretion of the pineal hormone mela tonin in experimental animals. In general, the evidence that magnetic fields alter the circadian production of melatonin is more substantial than the data illustrating the ability of electricfieldexposure to sup press melatonin. The chapter also discusses potential mechanisms by which thefieldscouple to the organism, and it considers the signifi cance of the melatonin changes with regard to the reported in creasedcancer risk in individuals exposed to an elevated electro magnetic environment.

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H E PINEAL G L A N D , a neuroendocrine organ located near the center of the human brain, produces and secretes an important hormone, melatonin. This gland is also present as a neuroectodermal outgrowth of the central nervous system of other mammals. The ability of the pineal gland to produce melatonin is directly related to the amount of visible electromagnetic radiation (i.e., light) detected by the eyes. Light exposure is always associated with minimal synthesis of melatonin in the pineal gland, whereas during darkness the production and secretion of the hormone increases. Because during the night blood melatonin

0065-2393/95/0250-0451$12.00/0 ©1995 American Chemical Society

In Electromagnetic Fields; Blank, M.; Advances in Chemistry; American Chemical Society: Washington, DC, 1995.


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levels are higher than they are during the day, melatonin has been referred to as the chemical expression of darkness (7). More than a decade ago both power-frequency electric (E) fields (2) and static magnetic (B) fields (3) were, like light, reported to inhibit the production of melatonin by the pineal gland. Subsequently, time-varying Β fields were also shown (4) to be effective in suppressing the synthesis and secretion of the pineal hormone melatonin. These initial reports set the stage for a flurry of studies; some confirmed the original observations, but others did not. In general, the melatonin rhythm is considered to be highly stable, and therefore the fact that it was reportedly altered by very low energy electromag netic fields was surprising. Likewise, the findings were important because the hormone melatonin has been functionally linked to a large number of important physiological events (5) including the suppression of cancer (6). Thus, very early a potential connection between the reported increased cancer risk (7) asso ciated with electric residential wire codes in epidemiology studies and the sup pression of melatonin was implied. This possibility has attracted even more at tention in recent years (8). The purpose of this brief review is to examine the published data related to the inhibition of melatonin production in the pineal gland of animals acutely or chronically exposed to Ε οτ Β fields and to consider the mechanisms by which the fields interact with the system in question. Additionally, the potential conse quences of a modification of the melatonin cycle will be summarized.

Pineal Melatonin Synthesis and Secretion Because the pineal gland responds to the prevailing light-dark cycle, and inas much as the gland in mammals is not directly light-sensitive, it was not surpris ing when an anatomic (and functional) connection was found between the eyes and the pineal gland. Thus, light detected by the retinas is transferred through the central and peripheral nervous systems to eventually end in the pineal gland. This circuitous pathway includes neuronal synapses in the suprachiasmatic nu clei (SCN or biological clock) of the hypothalamus and also involves fibers in the peripheral sympathetic nervous system (9). A s noted already, retinal activa tion by light results in minimal pineal production of melatonin, whereas at night synthesis and secretion of the hormone increases (7). These changes in the meta bolic activity of the pineal gland are reflected in blood levels of melatonin (Figure 1). Acute light exposure at night, like daytime light, quickly inhibits the production of melatonin in the pineal gland followed by a drop in circulating melatonin levels (10). In the pineal gland, melatonin is a product of the metabolism of the amino acid tryptophan. After its uptake by the endocrine cells of the pineal gland, the pinealocytes, tryptophan is quickly converted to serotonin (5-hydroxytryptamine), a monoamine that is in very high concentrations in the pineal (77). Serotonin is the common precursor for several potential secretory products of the gland, but the only documented hormone produced by this organ is melatonin.


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Figure 1. Twenty-four-hour rhythms in melatonin in the blood offour adult human males. Melatonin levels are always higher at night than during the day; however, the amplitude of the nighttime melatonin rise, as shown in this figure varies between individuals. Within an individual, the melatonin rhythm is high reproducible.

Serotonin is converted to melatonin by a two-step process initially involving the #-acetylation of serotonin by the enzyme iV-acetyltransferase (NAT) (Figure 2). The product of the action of N A T is #-acetylserotonin, which is O-methylated by the enzyme hydroxyindole-O-methyltransferase (HIOMT) to form melatonin (12). A t night, as the production of melatonin increases, the concentration of its precursor (i.e., serotonin) drops. Thus, the pineal concentrations of serotonin and melatonin are characteristically inversely related. Once melatonin is formed, it is quickly released from the pinealocytes into the blood presumably by simple diffusion, because it is a highly lipophilic molecule. A s a result, blood levels of melatonin are generally considered to be a good indicator of ongoing melatonin synthesis in the gland (7, 72). Besides its metabolic conversion to melatonin, serotonin in the pineal gland can be oxidatively deaminated via a pathway that is usually evaluated by measuring pineal levels of 5-hydroxyindole acetic acid (5-HIAA). Also, serotonin can be acted upon directly by H I O M T with the resultant formation of 5methoxytryptamine, a compound that has been proposed as a pineal hormone.


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TRYPTOPHAN TH 5 - HYDROXYTRYPTOPHAN AAAD SEROTONIN

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Figure 2. Diagrammatic representation of tryptophan metabolism in the pineal gland. The intermediate, serotonin, is metabolized via two pathways; that is., by monoamine oxidase (MAO) to 5-hydoxyindole acetic acid and by N-acetyltransferase (NAT) and hydroxylindole-O-methyltransferase (HIOMT) to melatonin. The most important pathway, and certainly the one that is most appli cable to the current report, is the conversion of serotonin to melatonin. Also rep resented is the reduction of melatonin in animals exposed to electromagnetic fields (see text). AAAD is aromatic amino acid decarboxylase; TH is tryptophan hydroxylase.

Time-Varying Ε Field Effects on Melatonin When Wilson and colleagues (2) described the inhibition of pineal melatonin concentrations in rats exposed to 60-Hz Ε fields, their evidence was the first to indicate that electromagnetic field wavelengths outside the visible range affected pineal metabolic activity. In this study, rats were exposed to a 39-kV (effective strength) field in a well-grounded exposure system in which the animals never experienced electrical shocks. The field was on 20 of each 24 h, and the expo sure was chronic (30 days). After this period of exposure, nighttime pineal levels of melatonin, as well as the activity of the rate-limiting enzyme in melatonin synthesis, N A T , were significantly depressed. The changes were not totally uniform, and to achieve statistically significant differences the data from two nighttime points had to be combined. The control animals were indeed shamexposed, having been placed in an exposure facility that was not energized. A l -



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though the study initially claimed an effective field strength of 39 kV/m, an erra tum appeared two years later that corrected this value to 1.7-1.9 kV/m (75). Wilson et al. (2) surmised that the responses of the pineal gland to sinu soidal Ε fields involved mechanisms similar to those related to the light sup pression of melatonin. This conclusion implied that the retinas were the likely sites of coupling of the fields to the organism although the authors never actually made this claim. The original publication was followed by another report by the same group in which they studied the time course of the change in pineal melatonin after rats were exposed to a 39-kV/m, 60-Hz Ε field (14). Rather than waiting 30 days to collect pineals, in this study the authors measured melatonin levels in the pineals of rats exposed for either 1, 2, 3, or 4 weeks. In this study they observed a gradual reduction in nocturnal melatonin over time with the suppression being significant after both three and four weeks of exposure. Whereas this study served as a confirmation of their original report, it also strongly suggested that the mechanism of the induced suppression by Ε fields was different from that by which light reduces the ability of the gland to convert serotonin to melatonin. In light-exposed animals, light exposure at night is followed by a rapid shutdown of pineal synthetic mechanisms with a similar quick reduction in melatonin (15). In the study of Wilson et al. (14) the animals had to experience 3 weeks of expo sure to Ε fields (20 h/day) before a significant drop in melatonin was recorded. With these two studies as a backdrop, another experiment was done on newborn animals exposed to sinusoidal Ε fields to determine whether exposing pregnant rats and, following their delivery, the young for 23 days, would com promise the development of the pineal melatonin cycle (16). Usually the circadian melatonin rhythm in 23-day-old rats is similar to that seen in adult ani mals (12). In this case three different field strengths were employed: 10, 65, or 130 kV/m. Control animals were sham-exposed as in the previous reports. This experiment showed that whereas all three experimental Ε fields (10, 65, and 130 kV/m) reduced nighttime pineal melatonin levels, the suppressions were very slight and occurred at only one time point during the night. This observation, although not directly contradictory to the original observations, raised some questions about the earlier findings. This doubt was further strengthened when the same group that had pub lished the 1981 (2) and 1986 (14) papers failed to confirm their own findings (17). In the presentation of the new data, the authors were at a loss to explain the different outcomes of the experiments. What was particularly perplexing was that all the studies seem to have been carefully done and the same exposure fa cility was used for all experiments. More recently, Grota et al. (18) completed a comprehensive study related to the interaction of 60-Hz Ε fields with pineal melatonin production in rats. This study was designed specifically to duplicate the experiments of Wilson and co-workers (2, 14) at another site. The duration of exposure was 30 days, and the field strength was 65 kV/m. Control animals were appropriately sham-exposed. A t the conclusion of the exposure, pineal and plasma melatonin levels were



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measured as were the two pineal enzymes, N A T and HIOMT, involved in the conversion of serotonin to melatonin. In this carefully conducted study, no ef fects of the sinusoidal field exposure in terms of nocturnal pineal melatonin, N A T activity, or H I O M T activity were apparent (18); a small drop in blood melatonin concentrations was recorded. On the basis of these results the authors concluded that no compelling evidence suggests that the time-varying Ε fields alter the circadian production of melatonin. Enthusiasm for the notion that purely sinusoidal Ε field exposure inter feres with the ability of the mammalian pineal gland to produce melatonin has waned. A s a consequence, few i f any follow-up experiments are currently being conducted.

Time-Invariant (Static) Β Field Effects on Melatonin Perturbed static magnetic fields have been frequently used (19, 20) as an ex perimental paradigm to alter the melatonin-forming ability of the pineal gland. The first of these studies dates back to the early 1980s when Semm (21) and Welker and co-workers (3) found that the mere inversion of the geomagnetic field induced a reduction of nocturnal melatonin in both the pineal and blood as well as a drop in the activity of the enzyme that iV-acetylates serotonin. These were acute studies with the exposure to the inverted field usually lasting for less than 120 min. A t least in the Welker et al. (3) study the pineal seemed to become refractory to the inverted field within about 120 min even when the geomagnetic field remained inverted. The circumstances under which perturbed geomagnetic fields impede the melatonin-forming ability of the pineal gland were further defined in a series of studies by Olcese and colleagues (22, 23). Nighttime pineal N A T activity and melatonin levels in rats were found to be suppressed after the exposure of the animals to a 50° rotation of the horizontal component of the geomagnetic field for 30 min, but this suppression did not occur in rats that had been optically enucleated. The inference of this finding, and certainly one espoused by the authors (22, 23), was that the eyes serve as magnetoreceptors that subsequently signal the reduction in melatonin production. In support of this concept, Reuss and Olcese (24) found that weak background red illumination was also a factor required for the perturbed geomagnetic fields to change pineal melatonin syn thesis. Although the importance of the eyes as a coupling organ for Β fields with the organisms remains unproven, we have proposed that, i f true, the isomerization of 11-cw-retinal to all-fraras-retinal in the rod photoreceptor could be the chemical event that initiates the processes that eventually reduce nocturnal pin eal melatonin formation (25). The isomerization of 11-cw-retinal requires very low energy input and leads to the activation of the photopigment rhodopsin; this activation is presumed to be the initial event in light-induced melatonin inhibi tion. Implicit in this proposal is that perturbed Β fields change the conversion of serotonin to melatonin by the same means that acute light exposure does at night. This assumption, however, remains unproven.



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Systemic factors seem to interfere with the ability of a changed geomag netic field to alter pineal metabolic activity in rodents. Thus, in the only study in which the Syrian hamster was used (26) in lieu of rats to test Β field effects on melatonin production, no change was reported; this result implies species speci ficity. Melatonin production in the pineal gland of the Syrian hamster is, like that in the rat, very sensitive to light inhibition (15). Cutaneous pigmentation has also been reported (26) to be a significant factor in determining the sensitivity of the Mongolian gerbil (Meriones unguiculatus) to time-invariant Β fields (26). When both albino and pigmented gerbils were exposed to a 60° rotation of the horizontal component of the Earth's mag netic field for 30 min, only in the albino animals was a drop in melatonin meas ured. Further confounding the findings in this report was the observation that only in the males, but not in the females, were pineal melatonin levels sup pressed (26); yet in both males and females pineal N A T activity was lowered as a consequence of the exposure. Such variable findings without seemingly a sound physiological explanation make generalizations concerning interactions of the static Β field with the pineal gland difficult. Using pineal metabolic parameters in addition to melatonin and N A T , Lerchl and co-workers (27) showed that the exposure of both rats and mice to intermittent changes in the Earth's static Β field caused changes consistent with a drop in melatonin synthesis. Thus, when the animals were exposed, at night, to a repeatedly inverted (at 5-min intervals) horizontal component of the geomag netic field (0.4 G or 40 μΤ) for 1 h, serotonin and 5-HIAA levels accumulated in the pineal gland. These findings signal a reduction in the conversion of serotonin to melatonin and thus lead to a buildup of serotonin and its subsequent metabo lism via an alternate pathway to 5-HIAA. This study provided additional support for the notion that time-invariant Β fields are effective, at least under some conditions, in curtailing the ability of the pineal gland to synthesize its most im portant hormone, melatonin. As a follow-up to this report, Lerchl et al. (28) examined the possible pa rameters of the Β fields that may be important in signaling the organisms of a change in the electromagnetic environment. They repeated their earlier studies with a couple of variations in the exposure parameters. In their initial studies and in half of the animals used in the second set of experiments, the geomagnetic Β field was "rapidly" inverted (dB/dt = 7.5 ms) intermittently for 1 h at night. The remaining half were also exposed to a repeatedly inverted Β field, but the inver sion occurred over a 1-s interval; these were referred to as "slowly" inverted fields. The significance of these different type inversions is as follows. When the Β field is rapidly inverted, weak electrical currents (eddy currents) are produced in the animals. Conversely, when the fields are slowly inverted, little or no eddy current is induced. The question being asked in this study was whether the in duced electric currents were operative in causing the pineal melatonin changes. The induced electric currents seemed to be operative in causing these changes in that the rapid dB/dt, which produced eddy currents, induced a reduc tion in pineal N A T and melatonin (Figure 3) and an elevation in pineal serotonin


ELECTROMAGNETIC FIELDS MELATONIN


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Figure 3. Pineal NAT and melatonin reductions following the exposure of rats to intermittent magnetic (B)fieldsat night. Thefieldswere either inverted "rapidly" (AA) or "slowly'* (MA). Only the rapidly invertedfieldsreduced pin eal serotonin metabolism. Also shown is the voltage required to invert the ge magnetic Βfieldas well as thefieldstrength. Thesefindingssuggest that the in duced electric current may be causative in the observed pineal changes. (Reproduced with permissionfromreference 19. Copyright 1992.)



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and 5-HIAA. None of these changes were seen when rats were exposed to re peatedly inverted fields in which the inversions occurred slowly. The implication of these findings is that induced electric currents may be the mechanism that couples Β fields to the organism (Figure 3). This report (28) is one of the first to be designed to specifically test for a coupling mechanism of perturbed Β fields, and if confirmed in similar studies in other laboratories, it could represent a very significant finding. Some of the variability in the response of the melatonin-generating system to Β fields may relate to the timing of exposure. According to Yaga et al. (29), the pineal gland at night becomes progressively more sensitive to Β field pertur bations as the night is prolonged. Thus, when rats were exposed to rapidly in verted Β fields either early in the night, at mid-dark, or late in the dark period, only at the mid- or late-dark times was melatonin significantly depressed; fur thermore, the suppression was greater at late dark than at mid-dark. Hereafter, studies designed to test the ability of Β fields to change pineal gland activity should take into account the time the exposure is done Many of these studies implied that the eyes may serve as the magnetoreceptors for the Β field effects on the pineal gland. However, isolated cultured pineal glands also responded to Β field changes with a reduced melatonin syn thesis (30). This finding does not prove, however, that in intact animals the eyes are not important coupling site for Β fields.

Time-Varying Β Field Effects on Melatonin Because most electromagnetic fields to which humans and animals are exposed are sinusoidal, interest in time-varying fields in reference to possible changes in physiology are of special importance. Surprisingly, however, rather few studies have used time-varying Β fields in the experimental setting to examine their ef fect on the pineal gland. According to Kato and colleagues (5i, 32), 50-Hz circularly polarized Β fields, but not 50-Hz horizontal or vertical Β fields, reduce plasma and pineal melatonin levels in rats. The study in which circularly polarized fields were used was very complete in the sense that a range of field strengths was used (0-120 μΤ) on a large number of rats. Also, both daytime and nighttime pineal and blood melatonin levels were examined. The animals were continually exposed to these fields for 6 weeks (except for twice weekly 2-h intervals). Examination of blood melatonin levels estimated by radioimmunoassay indicated that fields strengths of 1 μΤ or greater altered melatonin levels. Unexpectedly, daytime pineal levels of this hormone were actually slightly, albeit statistically signifi cantly, elevated. This unexpected finding detracts somewhat from a report that otherwise is of considerable interest. Although melatonin levels were lower in animals exposed to a variety of field strengths, no dose-response relationship was observed. In contrast to these 50-Hz rotating vector Β fields, as already mentioned, neither 50-Hz horizontal nor vertical field exposure influences melatonin in rats



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(52). These studies seem to have been well-controlled. Without specifying why these fields failed to depress melatonin whereas the 50-Hz circularized polarized fields did, the authors merely assumed that the outcomes differed because of differences in magnetic field characteristics. As part of a study related to the ability of 50-Hz Β fields to promote mammary carcinogenesis in rats, Mevissen et al. (33) also measured a significant reduction in the nocturnal serum concentrations of rats. The field strength in this experiment was 0.3-1 μΤ, and the duration of exposure was 91 days. Melatonin was measured in these studies because it is generally considered to be an oncostatic agent (6) and thus, i f lowered by the field exposure used, a high inci dence of tumorigenesis would be expected. Perhaps the most unique sinusoidal Β field exposure to reportedly curtail melatonin production is that used by Yellon (34). This worker used a 60-Hz, 1-G horizontal magnetic field during the day to test its effect on the subsequent night's melatonin rhythms. In this case the species employed was the Djungarian hamster (Phodopus sungorus), and the exposure duration was 15 min, beginning 2 h before darkness onset. The control animals were appropriately shamexposed. On the night following the daytime exposure, both pineal and serum melatonin levels were lower in the experimental compared to the control animals in the first study. When the experiment was replicated, similar results were ob tained, with the melatonin values again being depressed. A second replicate, however, failed to confirm the results of the previous two experiments. These perplexing observations have no obvious explanation, but presumably some ex posure parameter must have varied among the experiments or some technical failure must have occurred. Interestingly, the suppression of melatonin that Yellon (34) observed in the first two studies has been confirmed in an indepen dent laboratory (35). Although only a preliminary report, Graham et al. (36) claimed that in humans as well blood melatonin levels may be lowered by time-varying Β field exposure. The fields used were 60-Hz, 200-mG fields that were intermittently turned off and on during the night. The suppression of circulating melatonin was noted only in individuals who had an inherently dampened nighttime melatonin rise (Figure 1). Also, maximal suppression occurred later in the dark phase, similar to earlier observations made in rats (29). These important studies on hu mans require replication and extension.

Combined Time-Varying Ε and Β Field Effects on Melatonin A small number of studies have examined potential melatonin changes following the exposure of animals to a combined sinusoidal Ε and Β field. In a very thor ough and well-controlled study, 10 female Suffolk lambs were caged under a 500-V transmission line between the ages of 2 and 10 months (37). The respec tive mean Ε and Β fields were 6 V / m and 40 mG. Control lambs were housed in similar pens 229 m from the transmission lines, where the ambient Ε and Β fields were

Melatonin Suppression by Time-Varying and Time-Invariant

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