Educational Forum with Clinical Studies Current Science and Research

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J Dermatol. 2004 Nov;31(11):87883.
Oxidative stressmediated
skin damage in an experimental mobile phone model
can be prevented by melatonin.
Ayata A, Mollaoglu H, Yilmaz HR, Akturk O, Ozguner F, Altuntas I.
Department of Pediatrics, Faculty of Medicine, Suleyman Demirel University, Isparta, Turkey.
Most mobile phones emit 900 MHz of radiation that is mainly absorbed by the external organs. The
effects of 900 MHz of radiation on fibrosis, lipid peroxidation, and antioxidant
enzymes and the
ameliorating effects of melatonin (Mel) were evaluated in rat skin. Thirty WistarAlbino
rats were
used in the study. The experimental groups were the control group, the irradiated group (IR), and
the irradiated+Mel treated group (IR+Mel). A dose of 900 MHz, 2 W radiation was applied to the
IR group every day for 10 days (30 min/day). The IR+Mel group received 10 mg/kg/day melatonin
in tap water for 10 days before the irradiation. At the end of the 10th day, a skin specimen was
excised from the thoracoabdominal area. The levels of malondialdehyde (MDA) and
hydroxypyroline and the activities of superoxide dismutase (SOD), glutathione peroxidase (GSHPx),
and catalase (CAT) were studied in the skin samples. MDA and hydroxyproline levels and
activities of CAT and GSHPx
were increased significantly in the IR group compared to the control
group (p<0.05) and decreased significantly in the IR+Mel group (p<0.05). SOD activity was decreased significantly in the IR group and this decrease was not prevented by the Mel treatment. These results suggest that rats irradiated with 900 MHz suffer from increased fibrosis and lipid peroxidation (LPO). Mel treatment can reduce the fibrosis and LPO caused by radiation. PMID: 15729859 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=9816579&query_hl=35&itool=pubmed _docsum Med Pr. 1998;49(4):3339. [Platelets oxygen metabolism in women and men in different age groups] Article in Polish] Buczynski A, Kocur J, Stopczyk D, DziedziczakBuczynska M, Kowalski W. Zakladu Medycyny Zapobiegawczej, Instytutu OOZW Wojskowej Akademii Medycznej, Lodzi. The aim of the study as to assess the influence of the physiological aging process on the platelet cell metabolism in middleaged people. A group of 17 healthy women (aged 4759 years), and a group of healthy men (aged 4560 years) were examined. The control group was composed of healthy women aged 1925 years, and healthy men aged 1927 years. The activity of hyperoxide dismutase, catalase, glutathione peroxidase and the concentration of malonyldialdehyde were determined in platelets. In comparison to the control group, a significant decrease in the activity of hyperoxide dismutase and glutathione peroxidase as well as an enhanced concentration of malonyldialdehyde were observed in the group studied. Moreover, a diminished catalase activity was noted in platelets of men, while in women there were no significant changes. The study indicated that disorders in the function of thrombocytes, an excessive generation of oxygen free radicals, and impaired mechanisms of cellular antioxidative defence accelerate atherosclerosis and aging process. Therefore, it is necessary to cover middleaged people, particularly those occupationally exposed to factors affecting defence mechanisms, with adequate preventive programmes. PMID: 9816579 [PubMed indexed for MEDLINE] DunXian Tan, Lucien C. Manchester, Maria P. Terron, Luis J. Flores and Russel J. Reiter. « MINI REVIEW One molecule, many derivatives : A neverending interaction of melatonin with reactive oxygen and nitrogen species ? ». J. Pineal Res. 2006 Doi:10.1111/j.1600079X. 2006.00407.x Full study in pdf format. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=11989597&query_hl=19&itool=pubme d_docsum Tumori. 2001 NovDec; 87(6):41722. Ginkgo biloba extract (EGb 761) modulates bleomycininduced acute lung injury in rats. ElKhatib AS, Moustafa AM, AbdelAziz AA, AlShabanah OA, ElKashef HA. Department of Pharmacology, College of Pharmacy, King Saud University, Riyadh, Kingdom of Saudi Arabia. The effect of Ginkgo biloba extract (EGb 761) on bleomycin (BLM)induced acute lung injury was studied in rats. The responsiveness of isolated pulmonary arterial rings to 5hydroxytryptamine (5HT) as well as the levels of some relevant biochemical markers in the lung tissue were taken as evidence for the acute lung injury. BLM was given intraperitoneally at a dose of 15 mg/kg/day for five consecutive days. It was found that BLM treatment attenuated the vasoconstrictor effect of 5HT on the isolated pulmonary arteries. In lung tissues BLM also elevated the level of lipid peroxides and enhanced the activity of glutathione peroxidase. On the other hand, the level of glutathione and the activity of alkaline phosphatase were reduced. Body weight, lung weight and tissue glutathioneStransferase activity were, however, not altered. Oral administration of EGb 761 at a dose of 100 mg/kg/day for five consecutive days did not alter any of the chosen biochemical parameters in the lung tissue except for a slight reduction in alkaline phosphatase activity. However, treatment with EGb 761 reduced the responsiveness of the pulmonary artery to 5HT. Administration of EGb 761 (100 mg/kg/day; po) two hours prior to BLM (15 mg/kg/day; ip), for five consecutive days blunted the occurrence of further reduction in the vasoconstrictor response of the pulmonary artery to 5HT. Furthermore, EGb 761 tended to normalize BLMinduced alterations in the measured biochemical markers in the lung tissue. The apparent modulatory influence of EGb 761 on BLMinduced acute lung injury stems, at least in part, from its beneficial free radical scavenging properties that provide the extract with antioxidant activity. PMID: 11989597 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=16715528&dopt=Abstract Bioelectromagnetics. 2006 Sep;27(6):48793. Oxidative stress effects on the central nervous system of rats after acute exposure to ultra high frequency electromagnetic fields. Ferreira AR, Bonatto F, de Bittencourt Pasquali MA, Polydoro M, DalPizzol F, Fernandez C, de Salles AA, Moreira JC. Departamento de Bioquimica, Centro de Estudos em Estresse Oxidativo, ICBS, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brazil. arferreira@yahoo.com Mobile telephones and their base stations are an important source of ultra high frequency electromagnetic fields (UHFEMFs; 8001800 MHz) and their utilization is increasing all over the world. Epidemiological studies have suggested that low energy UHFEMFs may have biological effects, such as changes in oxidative metabolism after exposure. Therefore, we have investigated the effect of acute UHFEMF exposure on nonenzymatic antioxidant defense and lipid and protein oxidative damage in the rat frontal cortex and hippocampus. We have used malondialdehyde (MDA) and carbonyl assays to assess lipid and protein oxidative damages, respectively. No changes in lipid and protein damage, and also in nonenzymatic defense were found in frontal cortex or hippocampus. These results suggest that acute UHFEMF exposure is not able to produce detectable oxidative stress in rats from any age tested. However, more tests using a longer period of exposure and evaluating other tissues are necessary to ensure that there is no health risk associated with the use of mobile phones. PMID: 16715528 [PubMed in process] Aim of study (according to author) To study the effect of acute ultrahigh frequency electromagnetic field exposure on nonenzymatic antioxidant defense and lipid and protein oxidative damage in the rat frontal cortex and hippocampus. Background/further details: A total of 31 rats were divided by age into three groups (30, 80, and 210 days) and exposed. Endpoint  effects on the neurological system: oxidative metabolism in the brain Exposure General: analog mobile phone Field characteristics Parameters 834 MHz continuous wave (CW) exposure duration: repeated daily exposure, 7.5 h per day for 6 days electric field strength: 23 V/m effective value (to 35.7 V/ m) magnetic field strength: 65 mA/m (to 90 mA/m) power flux density: 0.15 mW/cm² mean value (to 0.35 mW/cm²) SAR: 0.41 W/kg (to 0.98 W/kg; for 30 day old rats) SAR: 0.36 W/kg (to 0.86 W/kg; for 70 day old rats) http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=16037959&query_hl=24&itool=pubme d_docsum Bioelectromagnetics. 2005 Oct;26(7):58994. Effects of a 50 Hz electric field on plasma lipid peroxide level and antioxidant activity in rats. Harakawa S, Inoue N, Hori T, Tochio K, Kariya T, Takahashi K, Doge F, Suzuki H, Nagasawa H. National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Japan. harakawa@bd5.sonet. ne.jp The effects of exposure to extremely low frequency electric fields (ELF EFs) on plasma lipid peroxide levels and antioxidant activity (AOA) in SpragueDawley rats were studied. The test was based on comparisons among rats treated with a combination of the oxidizing agent, 2,2'azobis( 2aminopropane) dihydrochloride (AAPH) and 50 Hz EF of 17.5 kV/m intensity for 15 min per day for 7 days, AAPH alone, EF alone or no treatment. EF significantly decreased the plasma peroxide level in rats treated with AAPH, similar to treatment by ascorbic acid or the superoxide dismutase. Ascorbic acid increased AOA; however, EF and superoxide dismutase did not change AOA compared with sham exposure in stressed rats. No influence on the lipid peroxide level and AOA in unstressed rats was observed with EF exposure alone. Although the administration of AAPH decreased AOA, this decrease did not change when EF was added. These data indicate that the ELF EF used in this study influenced the lipid peroxide level in an oxidatively stressed rat. (c) 2005 WileyLiss, Inc. PMID: 16037959 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=14734207&dopt=Abstract Clin Chim Acta. 2004 Feb;340(12): 15362. Ginkgo biloba prevents mobile phoneinduced oxidative stress in rat brain. Ilhan A, Gurel A, Armutcu F, Kamisli S, Iraz M, Akyol O, Ozen S. Department of Neurology, Inonu University, Turgut Ozal Medical Center, 44069 Malatya, Turkey. ailhan@inonu.edu.tr BACKGROUND: The widespread use of mobile phones (MP) in recent years has raised the research activities in many countries to determine the consequences of exposure to the lowintensity electromagnetic radiation (EMR) of mobile phones. Since several experimental studies suggest a role of reactive oxygen species (ROS) in EMRinduced oxidative damage in tissues, in this study, we investigated the effect of Ginkgo biloba (Gb) on MPinduced oxidative damage in brain tissue of rats. METHODS: Rats (EMR+) were exposed to 900 MHz EMR from MP for 7 days (1 h/day). In the EMR+Gb groups, rats were exposed to EMR and pretreated with Gb. Control and Gbadministrated groups were produced by turning off the mobile phone while the animals were in the same exposure conditions. Subsequently, oxidative stress markers and pathological changes in brain tissue were examined for each groups. RESULTS: Oxidative damage was evident by the: (i) increase in malondialdehyde (MDA) and nitric oxide (NO) levels in brain tissue, (ii) decrease in brain superoxide dismutase (SOD) and glutathione peroxidase (GSHPx) activities and (iii) increase in brain xanthine oxidase (XO) and adenosine deaminase (ADA) activities. These alterations were prevented by Gb treatment. Furthermore, Gb prevented the MPinduced cellular injury in brain tissue histopathologically. CONCLUSION: Reactive oxygen species may play a role in the mechanism that has been proposed to explain the biological side effects of MP, and Gb prevents the MPinduced oxidative stress to preserve antioxidant enzymes activity in brain tissue. PMID: 14734207 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=12415560&dopt=Abstract Cell Biochem Funct. 2002 Dec;20(4):27983. Effects of electromagnetic radiation from a cellular telephone on the oxidant and antioxidant levels in rabbits. Irmak MK, Fadillioglu E, Gulec M, Erdogan H, Yagmurca M, Akyol O. Department of Biochemistry, Faculty of Medicine, Inonu University, 44069 Malatya, Turkey. mkirmak@yahoo.com The number of reports on the effects induced by electromagnetic radiation (EMR) in various cellular systems is still increasing. Until now no satisfactory mechanism has been proposed to explain the biological effects of this radiation. Oxygen free radicals may play a role in mechanisms of adverse effects of EMR. This study was undertaken to investigate the influence of electromagnetic radiation of a digital GSM mobile telephone (900 MHz) on oxidant and antioxidant levels in rabbits. Adenosine deaminase, xanthine oxidase, catalase, myeloperoxidase, superoxide dismutase (SOD) and glutathione peroxidase activities as well as nitric oxide (NO) and malondialdehyde levels were measured in sera and brains of EMRexposed and shamexposed rabbits. Serum SOD activity increased, and serum NO levels decreased in EMRexposed animals compared to the sham group. Other parameters were not changed in either group. This finding may indicate the possible role of increased oxidative stress in the pathophysiology of adverse effect of EMR. Decreased NO levels may also suggest a probable role of NO in the adverse effect. Copyright 2002 John Wiley & Sons, Ltd. PMID: 12415560 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=10437415&query_hl=7&itool=pubmed _docsum Vojnosanit Pregl. 1999 MarApr; 56(2):1137. Oxidative stress in the thalamus of Wistar rats treated with 1methyl4phenyl1,2,3,6tetrahydropyridine. Jovanovic MD, Ninkovic M, Malicevic Z, Mihajlovic R, Dukic M, Vasiljevic I, Jelenkovic A, Jovicic A. Military Medical Academy, Institute for Medical Research, Belgrade. Experimental parkinsonism was induced in adult Wistar rats by selective nigrostriatal neurotoxine, 1methyl4phenyl1,2,3,6tetrahydropyridine (MPTP) in a single dose of 0.09 g/kg, by unilateral intrastriatal application using stereotaxic instrument. Control group included rats treated with 0.9% saline solution in the same manner. Animals were sacrificed by decapitation seven days after the treatment. Total glutathione was measured in the crude mitochondrial fraction of thalamus and striatum. Total glutathione content, as a measure of reduced cell atmosphere, was mutually decreased in the thalamus and striatum of MPTPtreated animals, compared to controls: thalamus ipsi= 24.8 +/3.11, contralateral = 26.81 +/5.31; striatum ipsi= 19.96 +/4.13, contralateral = 17.3 +/4.09 nmol/mg prot. Mutually depleted glutathione content in the thalamus and contralateral striatum, the structures distant from ipsilateral treated striatum, could indicate on spatial propagation of oxidative stress, not only in the selective vulnerable dopaminergic nigrostriatal neurons, but in the structures included in the motor and cognitive loops of basal ganglia. PMID: 10437415 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=16898263&query_hl=2&itool=pubmed _docsum Toxicol Ind Health. 2006 Jun;22(5):2116. Melatonin modulates 900 Mhz microwaveinduced lipid peroxidation changes in rat brain. Koylu H, Mollaoglu H, Ozguner F, Nazyroglu M, Delibab N. Department of Physiology, Faculty of Medicine, Suleyman Demirel University, 32260, Isparta, Turkey. Microwaves (MW) from cellular phones may affect biological systems by increasing free radicals, which may enhance lipid peroxidation levels of the brain, thus leading to oxidative damage. Melatonin is synthesized in and secreted by the pineal gland at night and exhibits antioxidant properties. Several studies suggest that supplementation with antioxidant can influence MWinduced brain damage. The present study was designed to determine the effects of MW on the brain lipid peroxidation system, and the possible protective effects of melatonin on brain degeneration induced by MW. Twentyeight SpragueDawley male rats were randomly divided into three groups as follows: (1) shamoperated control group (N = 8); (2) study 900MHz MWexposed group (N = 8); and (3) 900MHz MWexposed+ melatonin (100 microg/kg sc before daily MW exposure treated group) (N = 10). Cortex brain and hippocampus tissues were removed to study the levels of lipid peroxidation as malonyl dialdehyde. The levels of lipid peroxidation in the brain cortex and hippocampus increased in the MW group compared with the control group, although the levels in the hippocampus were decreased by MW+melatonin administration. The brain cortex lipid peroxidation levels were unaffected by melatonin treatment. We conclude that melatonin may prevent MWinduced oxidative changes in the hippocampus by strengthening the antioxidant defense system, by reducing oxidative stress products. PMID: 16898263 [PubMed in process] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=14732250&dopt=Abstract J Photochem Photobiol B. 2004 Jan 23;73(12): 438. Effects of extremely low frequency magnetic field on the antioxidant defense system in mouse brain: a chemiluminescence study. Lee BC, Johng HM, Lim JK, Jeong JH, Baik KY, Nam TJ, Lee JH, Kim J, Sohn UD, Yoon G, Shin S, Soh KS. School of Physics, College of Natural Sciences, Seoul National University, San 561, Shillimdong, Kwanakgu, Seoul 151742, South Korea. donlee@phya.snu.ac.kr Among the putative mechanisms, by which extremely low frequency (ELF) magnetic field (MF) may affect biological systems is that of increasing free radical life span in organisms. To test this hypothesis, we investigated whether ELF (60 Hz) MF can modulate antioxidant system in mouse brain by detecting chemiluminescence and measuring superoxide dismutase (SOD) activity in homogenates of the organ. Compared to sham exposed control group, lucigenininitiated chemiluminescence in exposed group was not significantly increased. However, lucigeninamplified tbutyl hydroperoxide (TBHP)initiated brain homogenates chemiluminescence, was significantly increased in mouse exposed to 60 Hz, MF, 12 G for 3 h compared to sham exposed group. We also measured SOD activity, that plays a critical role of the antioxidant defensive system in brain. In the group exposed to 60 Hz, MF, 12 G for 3 h, brain SOD activity was significantly increased. These results suggest that 60 Hz, MF could deteriorate antioxidant defensive system by reactive oxygen species (ROS), other than superoxide radicals. Further studies are needed to identify the kind of ROS generated by the exposure to 60 Hz, MF and elucidate how MF can affect biological system in connection with oxidative stress. PMID: 14732250 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=11516912&query_hl=19&itool=pubme d_docsum J Pharm Biomed Anal. 2001 Nov;26(4):6058. Effects of acute exposure to the radiofrequency fields of cellular phones on plasma lipid peroxide and antioxidase activities in human erythrocytes. Moustafa YM, Moustafa RM, Belacy A, AbouElEla SH, Ali FM. Department of Pharmacology and Toxicology, Faculty of Pharmacy, Suez Canal University, Ismailia 41522, Egypt. Radiofrequency fields of cellular phones may affect biological systems by increasing free radicals, which appear mainly to enhance lipid peroxidation, and by changing the antioxidase activities of human blood thus leading to oxidative stress. To test this, we have investigated the effect of acute exposure to radiofrequency fields of commercially available cellular phones on some parameters indicative of oxidative stress in 12 healthy adult male volunteers. Each volunteer put the phone in his pocket in standby position with the keypad facing the body. The parameters measured were lipid peroxide and the activities of superoxide dismutase (SOD), total glutathione peroxidase (GSHPx) and catalase. The results obtained showed that the plasma level of lipid peroxide was significantly increased after 1, 2 and 4 h of exposure to radiofrequency fields of the cellular phone in standby position. Moreover, the activities of SOD and GSHPx in human erythrocytes showed significant reduction while the activity of catalase in human erythrocytes did not decrease significantly. These results indicate that acute exposure to radiofrequency fields of commercially available cellular phones may modulate the oxidative stress of free radicals by enhancing lipid peroxidation and reducing the activation of SOD and GSHPx, which are free radical scavengers. Therefore, these results support the interaction of radiofrequency fields of cellular phones with biological systems. PMID: 11516912 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=16318001&query_hl=1 Vopr Kurortol Fizioter Lech Fiz Kult. 2005 SepOct;( 5):1720. [Influence of (460 MHz) electromagnetic fields on the induced lipid peroxidation in the structures of visual analyzer and hypothalamus in experimental animals] [Article in Russian] Musaev AV, Ismailova LF, Gadzhiev AM. Changes in the intensity of ascorbateand NADPN2dependent induced lipid peroxidation (LPO) were studied in exposure of the visual analyzer and hypothalamus of 3and 12monthold rats to radiation with microwaves of high and low intensity. The exposure to microwaves of high intensity stimulated basal LPO but suppressed activity of LPOinducing systems. This suggests disturbances in the activity of different sources of active oxygen forms. Microwaves of low intensity activated systems of induced LPO. This is accompanied with synchronous activity of the antioxidant defense system maintaining a normal oxidationreduction balance of the cell. The conclusion is that, depending on their intensity, microwaves can be either beneficial to health or be a factor of oxidative stress. PMID: 16318001 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=11039304&query_hl=7&itool=pubmed _docsum Vojnosanit Pregl. 2000 MayJun; 57(3):25763. Effects of nerve growth factor on antioxidative system in the thalamus of MPTP treated Wistar rats. Ninkovic MB, Jovanovic MD, Malicevic Z, Dukic M, Jelenkovic A, Mihajlovic R, Vasiljevic I, Jovicic A. Military Medical Academy, Institute for Medical Research, Belgrade. 1methyl4phenyl1,2,3,6tetrahydropyridine (MPTP)induced parkinsonism is one of the most useful models for the study of that disease. It has been suggested that MPTPinduced neurotoxicity may involve the production of reactive oxygen species. MPTP was applied intracerebrally, unilaterally, in the striatum in single dose of 0.09 g/kg b.w. The second group was treated both with MPTP and nerve growth factor (NGF) in dose of 7 ng/ml. NGF was applied immediately after the neurotoxin. Control group was treated with 0.9% saline solution in the same manner. Animals were decapitated 7 days after the treatment. In the group treated with MPTP, the activity of superoxide dismutase (SOD) and glutathione peroxidase (GSHPx) was decreased in ipsilateral thalamus, compared to control values as well as to the contralateral thalamus. In the same structures superoxide anion production was increased, compared to controls. Following the application of both MPTP and NGF, the activity of SOD and GSHPx remained on control values, while the superoxide anion content was decreased, compared to controls. These results indicate a temporal and spatial propagation of oxidative stress and spread protective effects of NGF on the thalamus, the structure that is distant, but very tightly connected with striatum, the place of direct neurotoxic damage. PMID: 11039304 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=17191663&query_hl=7&itool=pubmed _docsum Acta Physiol Hung. 2006 Dec;93(4):31523. Oxidative stress in the rats brain capillaries in sepsisthe influence of 7nitroindazole. Ninkovic M, Malicevic I, Jelenkovic A, Jovanovic DM, Dukic M, Vasiljevic I. Institute of Medical Research, Military Medical Academy, Crnotvarska 17, 11002 Belgrade, Serbia vmaimi@Eunet.yu As a part of bloodbrain barrier, brain capillaries participate in pathophysiological events during systemic inflammation. We investigated the effects of 7nitroindazole (7NI), selective neuronal nitric oxide synthase (NOS) inhibitor, to oxidative status (OS) of brain capillaries. Adult Wistar rats were randomized at groups: control group (CG) (sham operated), sepsis group (GS) (cecal ligation and perforation with inoculation of Escherichia coli (ATCC 25922), 7NI group (G7NI), (30 mg/kg b/w i.p.) and 7NI + sepsis group (G7NIS), (7NI was applied 30 minutes before operation). Lipid peroxidation index (LPI), nitrite concentration, superoxide dismutase (SOD) activity and superoxide anion (O2*) content were determined 3, 6, 24 and 48 hour in each group. Cerebral capillaries were separated from nonvascular brain tissue using sucrose gradient. Compared to controls, LPI, nitrite and O2*increased at SG. In the G7NIS, LPI reached control values at the 24th and 48th hour, while nitrite were decreased at the 3rd and 24th hour, compared to controls. In the same group, O2*decreased at the 3rd, 6th and 24th hour, although SOD showed variable activity. The systematic nNOS inhibition with 7NI forces OS on early terms of sepsis, but lately it contributes to the normalization of OS in cerebral capillaries. PMID: 17191663 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=15950073&dopt=Abstract Arch Med Res. 2005 JulAug; 36(4):3505. Oxidative damage in the kidney induced by 900MHzemitted mobile phone: protection by melatonin. Oktem F, Ozguner F, Mollaoglu H, Koyu A, Uz E. Department of Pediatric Nephrology, School of Medicine, Suleyman Demirel University, Isparta, Turkey. BACKGROUND: The mobile phones emitting 900MHz electromagnetic radiation (EMR) may be mainly absorbed by kidneys because they are often carried in belts. Melatonin, the chief secretory product of the pineal gland, was recently found to be a potent free radical scavenger and antioxidant. The aim of this study was to examine 900MHz mobile phoneinduced oxidative stress that promotes production of reactive oxygen species (ROS) on renal tubular damage and the role of melatonin on kidney tissue against possible oxidative damage in rats. METHODS: The animals were randomly grouped as follows: 1) shamoperated control group and 2) study groups: i) 900MHz EMR exposed (30 min/day for 10 days) group and ii) 900MHz EMR exposed+melatonin (100 microg kg(1) s.c. before the daily EMR exposure) treated group. Malondialdehyde (MDA), an index of lipid peroxidation), and urine Nacetylbetadglucosaminidase (NAG), a marker of renal tubular damage were used as markers of oxidative stressinduced renal impairment. Superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSHPx) activities were studied to evaluate the changes of antioxidant status. RESULTS: In the EMRexposed group, while tissue MDA and urine NAG levels increased, SOD, CAT, and GSHPx activities were reduced. Melatonin treatment reversed these effects as well. In this study, the increase in MDA levels of renal tissue and in urine NAG and also the decrease in renal SOD, CAT, GSHPx activities demonstrated the role of oxidative mechanism induced by 900MHz mobile phone exposure, and melatonin, via its free radical scavenging and antioxidant properties, ameliorated oxidative tissue injury in rat kidney. CONCLUSIONS: These results show that melatonin may exhibit a protective effect on mobile phoneinduced renal impairment in rats. PMID: 15950073 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=16132682&dopt=Abstract Mol Cell Biochem. 2005 Aug;276(12): 317. Comparative analysis of the protective effects of melatonin and caffeic acid phenethyl ester (CAPE) on mobile phoneinduced renal impairment in rat. Ozguner F, Oktem F, Armagan A, Yilmaz R, Koyu A, Demirel R, Vural H, Uz E. Department of Physiology, School of Medicine, Suleyman Demirel University, P. K. 13 32100 Isparta, Turkey. drmfehmi@yahoo.com Melatonin and caffeic acid phenethyl ester (CAPE), a component of honeybee propolis, were recently found to be potent free radical scavengers and antioxidants. There are a number of reports on the effects induced by electromagnetic radiation (EMR) in various cellular systems. Mechanisms of adverse effects of EMR indicate that reactive oxygen species may play a role in the biological effects of this radiation. The present study was carried out to compare the protective effects of melatonin and CAPE against 900 MHz EMR emitted mobile phoneinduced renal tubular injury. Melatonin was administered whereas CAPE was given for 10 days before the exposure. Urinary NacetylbetaDglucosaminidase (NAG, a marker of renal tubular injury) and malondialdehyde (MDA, an index of lipid peroxidation), were used as markers of oxidative stressinduced renal impairment in rats exposed to EMR. Superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSHPx) activities were studied to evaluate the changes of antioxidant status in renal tissue. Urinary NAG and renal MDA were increased in EMR exposed rats while both melatonin and CAPE caused a significant reduction in the levels of these parameters. Likewise, renal SOD and GSHPx activities were decreased in EMR exposed animals while melatonin caused a significant increase in the activities of these antioxidant enzymes but CAPE did not. Melatonin caused a significant decrease in urinary NAG activity and MDA levels which were increased because of EMR exposure. CAPE also reduced elevated MDA levels in EMR exposed renal tissue, but the effect of melatonin was more potent than that of CAPE. Furthermore, treatment of EMR exposed rats with melatonin increased activities of SOD and GSHPx to higher levels than those of control rats. In conclusion, melatonin and CAPE prevent renal tubular injury by reducing oxidative stress and protect the kidney from oxidative damage induced by 900 MHz mobile phone. Nevertheless, melatonin seems to be a more potent antioxidant compared with CAPE in kidney. (Mol Cell Biochem 276: 3137, 2005). PMID: 16132682 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=16132717&dopt=Abstract Mol Cell Biochem. 2005 Sep;277(12): 7380. A novel antioxidant agent caffeic acid phenethyl ester prevents longterm mobile phone exposureinduced renal impairment in rat. Prognostic value of malondialdehyde, NacetylbetaDglucosaminidase and nitric oxide determination. Ozguner F, Oktem F, Ayata A, Koyu A, Yilmaz HR. Department of Physiology, School of Medicine, Suleyman Demirel University, P. K. 13, Isparta, 32100, Turkey. drmfehmi@yahoo.com Caffeic acid phenethyl ester (CAPE), a flavonoid like compound, is one of the major components of honeybee propolis. It has been used in folk medicine for many years in Middle East countries. It was found to be a potent free radical scavenger and antioxidant recently. The aim of this study was to examine longterm applied 900 MHz emitting mobile phoneinduced oxidative stress that promotes production of reactive oxygen species (ROS) and, was to investigate the role of CAPE on kidney tissue against the possible electromagnetic radiation (EMR)induced renal impairment in rats. In particular, the ROS such as superoxide and nitric oxide (NO) may contribute to the pathophysiology of EMRinduced renal impairment. Malondialdehyde (MDA, an index of lipid peroxidation) levels, urinary NacetylbetaDglucosaminidase (NAG, a marker of renal tubular injury) and nitric oxide (NO, an oxidant product) levels were used as markers of oxidative stressinduced renal impairment and the success of CAPE treatment. The activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSHPx) in renal tissue were determined to evaluate the changes of antioxidant status. The rats used in the study were randomly grouped (10 each) as follows: i) Control group (without stress and EMR), ii) Shamoperated rats stayed without exposure to EMR (exposure device off), iii) Rats exposed to 900 MHz EMR (EMR group), and iv) A 900 MHz EMR exposed + CAPE treated group (EMR + CAPE group). In the EMR exposed group, while tissue MDA, NO levels and urinary NAG levels increased (p < 0.0001), the activities of SOD, CAT, and GSHPx in renal tissue were reduced (p < 0.001). CAPE treatment reversed these effects as well (p < 0.0001, p < 0.001 respectively). In conclusion, the increase in NO and MDA levels of renal tissue, and in urinary NAG with the decrease in renal SOD, CAT, GSHPx activities demonstrate the role of oxidative mechanisms in 900 MHz mobile phoneinduced renal tissue damage, and CAPE, via its free radical scavenging and antioxidant properties, ameliorates oxidative renal damage. These results strongly suggest that CAPE exhibits a protective effect on mobile phoneinduced and free radical mediated oxidative renal impairment in rats. PMID: 16132717 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=16342473&dopt=Abstract Toxicol Ind Health. 2005 Oct;21(9):22330. Mobile phoneinduced myocardial oxidative stress: protection by a novel antioxidant agent caffeic acid phenethyl ester. Ozguner F, Altinbas A, Ozaydin M, Dogan A, Vural H, Kisioglu AN, Cesur G, Yildirim NG. Department of Physiology, School of Medicine, Suleyman Demirel University, Isparta, Turkey. drmfehmi@yahoo.com Electromagnetic radiation (EMR) or radiofrequency fields of cellular mobile phones may affect biological systems by increasing free radicals, which appear mainly to enhance lipid peroxidation, and by changing the antioxidant defense systems of human tissues, thus leading to oxidative stress. Mobile phones are used in close proximity to the heart, therefore 900 MHz EMR emitting mobile phones may be absorbed by the heart. Caffeic acid phenethyl ester (CAPE), one of the major components of honeybee propolis, was recently found to be a potent free radical scavenger and antioxidant, and is used in folk medicine. The aim of this study was to examine 900 MHz mobile phoneinduced oxidative stress that promotes production of reactive oxygen species (ROS) and the role of CAPE on myocardial tissue against possible oxidative damage in rats. Thirty rats were used in the study. Animals were randomly grouped as follows: shamoperated control group (N: 10) and experimental groups: (a) group II: 900 MHz EMR exposed group (N: 10); and (b) group III: 900 MHz EMR exposed+CAPEtreated group (N: 10). A 900 MHz EMR radiation was applied to groups II and III 30 min/day, for 10 days using an experimental exposure device. Malondialdehyde (MDA, an index of lipid peroxidation), and nitric oxide (NO, a marker of oxidative stress) were used as markers of oxidative stressinduced heart impairment. Superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSHPx) activities were studied to evaluate the changes of antioxidant status. In the EMR exposed group, while tissue MDA and NO levels increased, SOD, CAT and GSHPx activities were reduced. CAPE treatment in group III reversed these effects. In this study, the increased levels of MDA and NO and the decreased levels of myocardial SOD, CAT and GSHPx activities demonstrate the role of oxidative mechanisms in 900 MHz mobile phoneinduced heart tissue damage, and CAPE, via its free radical scavenging and antioxidant properties, ameliorates oxidative heart injury. These results show that CAPE exhibits a protective effect on mobile phoneinduced and free radical mediated oxidative heart impairment in rats. PMID: 16342473 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=16317515&query_hl=1 Mol Cell Biochem. 2006 Jan;282(12): 838. Protective effects of melatonin and caffeic acid phenethyl ester against retinal oxidative stress in longterm use of mobile phone: a comparative study. Ozguner F, Bardak Y, Comlekci S. Department of Physiology, School of Medicine, Suleyman Demirel University, P. K. 13, 32100 Isparta, Turkey. drmfehmi@yahoo.com There are numerous reports on the effects of electromagnetic radiation (EMR) in various cellular systems. Melatonin and caffeic acid phenethyl ester (CAPE), a component of honeybee propolis, were recently found to be potent free radical scavengers and antioxidants. Mechanisms of adverse effects of EMR indicate that reactive oxygen species may play a role in the biological effects of this radiation. The present study was carried out to compare the efficacy of the protective effects of melatonin and CAPE against retinal oxidative stress due to longterm exposure to 900 MHz EMR emitting mobile phones. Melatonin and CAPE were administered daily for 60 days to the rats prior to their EMR exposure during our study. Nitric oxide (NO, an oxidant product) levels and malondialdehyde (MDA, an index of lipid peroxidation), were used as markers of retinal oxidative stress in rats following to use of EMR. Superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSHPx) activities were studied to evaluate the changes of antioxidant status in retinal tissue. Retinal levels of NO and MDA increased in EMR exposed rats while both melatonin and CAPE caused a significant reduction in the levels of NO and MDA. Likewise, retinal SOD, GSHPx and CAT activities decreased in EMR exposed animals while melatonin and CAPE caused a significant increase in the activities of these antioxidant enzymes. Treatment of EMR exposed rats with melatonin or CAPE increased the activities of SOD, GSHPx and CAT to higher levels than those of control rats. In conclusion, melatonin and CAPE reduce retinal oxidative stress after longterm exposure to 900 MHz emitting mobile phone. Nevertheless, there was no statistically significant difference between the efficacies of these two antioxidants against to EMR induced oxidative stress in rat retina. The difference was in only GSHPx activity in rat retina. Melatonin stimulated the retinal GSHPx activity more efficiently than CAPE did. PMID: 16317515 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=PubMed&list_uids=12006087&dopt=Abstract Biochem J. 2002 Aug 15;366(Pt 1):2039. Selenium deficiency increases the expression of inducible nitric oxide synthase in RAW 264.7 macrophages: role of nuclear factorkappaB in upregulation. Prabhu KS, ZamamiriDavis F, Stewart JB, Thompson JT, Sordillo LM, Reddy CC. Department of Veterinary Science and The Center for Molecular Toxicology and Carcinogenesis, 115 Henning, The Pennsylvania State University, University Park, PA 16802, U.S.A. The inducible isoform of nitric oxide synthase (iNOS) is implicated in atherosclerosis, malignancy, rheumatoid arthritis, tissue and reperfusion injuries. A key determinant of the prooxidant versus protective effects of NO is the underlying redox status of the tissue. Selenoproteins, such as glutathione peroxidases (GPxs) and thioredoxin reductases, are key components of cellular defence and promote optimal antioxidant/oxidant balance. In this study, we have investigated the relationship between Se status, iNOS expression and NO production in Sedeficient and Sesupplemented RAW 264.7 macrophage cell lines. The cellular GPx activity, a measure of Se status, was 17fold lower in Sedeficient RAW 264.7 cells and the total cellular oxidative tone, as assessed by flow cytometry with 2',7'dichlorodihydrofluorescein diacetate, was higher in the Sedeficient cells than the Sesupplemented cells. Upon lipopolysaccharide (LPS) stimulation of these cells in culture, we found significantly higher iNOS transcript and protein expression levels with an increase in NO production in Sedeficient RAW 264.7 cells than the Sesupplemented cells. Electrophoretic mobilityshift assays, nuclear factorkappaB (NFkappaB) luciferase reporter assays and Western blot analyses indicate that the increased expression of iNOS in Se deficiency could be due to an increased activation and consequent nuclear localization of the redoxsensitive transcription factor NFkappaB. These results suggest an inverse relationship between cellular Se status and iNOS expression in LPSstimulated RAW 264.7 cells and provide evidence for the beneficial effects of dietary Se supplementation in the prevention and/or treatment of oxidativestressmediated inflammatory diseases. PMID: 12006087 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=12474410&query_hl=35&itool=pubme d_docsum Med Pr. 2002;53(4):3114. [Effect of electromagnetic field produced by mobile phones on the activity of superoxide dismutase (SOD1) and the level of malonyldialdehyde (MDA) in vitro study] [Article in Polish] Stopczyk D, Gnitecki W, Buczynski A, Markuszewski L, Buczynski J. Zakladu Medycyny Zapobiegawczej i Promocji Zdrowia, Wojskowej Akademii Medycznej w Lodzi. darstop@poczta.onet.pl The aim of the study was to assess in vitro the effect of electromagnetic field produced by mobile phones on the activity of superoxide dismutase (SOD1) and the level of malonyldialdehyde (MDA) in human blood platelets. The suspension of blood platelets was exposed to the electromagnetic field with the frequency of 900 MHz for 1, 3, 5, and 7 min. Our studies demonstrated that microwaves produced by mobile phones significantly depleted SOD1 activity after 1, 5, and 7 min of exposure and increased after 3 min in comparison with the control test. There was a significant increase in the concentration of MDA after 1, 5, and 7 min and decrease after 3 min of exposure as compared with the control test. On the grounds of our results we conclude that oxidative stress after exposure to microwaves may be the reason for many adverse changes in cells and may cause a number of systemic disturbances in the human body. PMID: 12474410 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=16602439&itool=pubmed_DocSum Ann Acad Med Stetin. 2005;51 Suppl 1:1258. [Effect of electromagnetic field produced by mobile phones on the activity of superoxide dismutase (SOD1) in vitro researches] [Article in Polish] Stopczyk D, Gnitecki W, Buczynski A, Kowalski W, Buczynska M, Kroc A. Zaklad Medycyny Zapobiegawczej i Promocji Zdrowia Wojskowej Akademii Medycznej ul. Zeligowskiego 7/9, 90643 lodz. The aim of the paper was to estimate in vitro the effect of electromagnetic field produced by mobile phones on the activity of superoxide dismutase (SOD1) in human blood platelets. Suspension of blood platelets exposed to the electromagnetic field of 900 MHz frequency for 1, 3, 5, 7 minutes. Our studies demonstrated that microwaves produced by mobiles significally depleted the activity of SOD1 after 1, 5, 7 min. of exposition and increased after 3 min. nn comparison wit control test. On the ground of our results we conclude that oxidative stress after exposition to microwaves can be the reason of many disadvantageous changes in cells and may cause many systemic consequences in human organism. PMID: 16602439 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=12552762&query_hl=28&itool=pubme d_docsum Sheng Wu Yi Xue Gong Cheng Xue Za Zhi. 1999 Sep;16(3):35961. [The influence of pulsed magnetic fields on SOD's activity and MDA value in metabolism of mice] [Article in Chinese] Wu D, Wang D, Xi X, Li X, Mo J. Laboratory for Free Radical Research, Fourth Military Medical University, Xi'an 710032. This experiment aimed to investigate the influence of pulsed extremelylowfrequencymagnetic fields on free radical metabolism of mice. Thirtytwo mice were randomly divided into four groups and were exposed to 0 T, 0.25 T, 0.34 T and 0.64 T intensity pulsed magnetic fields of 20 Hz for 40 min. The 0 T exposed group was the control group. The free radical metabolism, SOD's activity and MDA, of mice were measured respectively. The result showed the SOD's activity and MDA of the 0.34 T exposed group were both significantly lower (P < 0.01, P < 0.05) than that of the control group, while the 0.25 T and 0.64 T exposed groups were not significantly different from the control group. This demonstrates that the biological effect of pulsed extremelylowfrequency magnetic fields on free radical metabolism of mice varies and depends on the intensity of the fields. The "window" effect may exist. PMID: 12552762 [PubMed indexed for MEDLINE] http://www.ncbi.nlm.nih.gov/entrez/query.fcgi? db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=16954120&query_hl=3&itool=pubmed _docsum Electromagn Biol Med. 2006;25(3):17788. GSM base station electromagnetic radiation and oxidative stress in rats. Yurekli AI, Ozkan M, Kalkan T, Saybasili H, Tuncel H, Atukeren P, Gumustas K, Seker S. TubitakUekae, EMC TEMPEST Test Center, GebzeKocaeli, Turkey. yurekli@uekae.tubitak.gov.tr The ever increasing use of cellular phones and the increasing number of associated base stations are becoming a widespread source of nonionizing electromagnetic radiation. Some biological effects are likely to occur even at lowlevel EM fields. In this study, a gigahertz transverse electromagnetic (GTEM) cell was used as an exposure environment for plane wave conditions of farfield free space EM field propagation at the GSM base transceiver station (BTS) frequency of 945 MHz, and effects on oxidative stress in rats were investigated. When EM fields at a power density of 3.67 W/m2 (specific absorption rate = 11.3 mW/kg), which is well below current exposure limits, were applied, MDA (malondialdehyde) level was found to increase and GSH (reduced glutathione) concentration was found to decrease significantly (p < 0.0001). Additionally, there was a less significant (p = 0.0190) increase in SOD (superoxide dismutase) activity under EM exposure. PMID: 16954120 [PubMed in process] http://www.buergerwelle.de/pdf/microwave_and_oxidative_stress_studies.htm L'étude complète par lien n'est pas complète J Physiol Pharmacol. 2005 Dec;56 Suppl 6:1018. Effect of extremely low frequency of electromagnetic fields on cell proliferation, antioxidative enzyme activities and lipid peroxidation in 3T3L1 preadipocytes an in vitro study. ZwirskaKorczala K, Jochem J , AdamczykSowa M , Sowa P , Polaniak R , Birkner E , Latocha M , Pilc K , Suchanek R . Department of Physiology, Zabrze, Medical University of Silesia, Katowice, Poland. The exposure to extremely low frequency electromagnetic field (ELFMF, frequencies less than 200300 Hz) can alter the transcription and translation of genes, influence the cell proliferation rate and affect enzyme activities. Moreover, the hypothesis that ELFMF increases free oxygen metabolites generation has been proposed. Since recent in vivo studies suggest that electric and magnetic fields are able to affect adipose cells metabolism. The aim of the study was to examine the effects of ELFMF (frequency of basic impulse 180195 Hz, induction 120 muT) on cell proliferation, antioxidative enzyme activities and malondialdehyde (MDA) concentration in 3T3L1 preadipocyte cell culture. We found that ELFMF application lasting 36 minutes daily failed to influence cell count after 24h and 48 h of incubation. After 24 h, in the ELFMF treated group, manganeseand copperzinccontaining superoxide dismutase (MnSOD and Cu/ZnSOD) isoenzymes media activities were decreased, catalase activity was increased, whereas there were no significant differences in glutathione peroxidase (GSHPx) and glutathione reductase (GSSGRd) activities in comparison to the control. After 48 h of incubation, all enzyme activities were reduced, except for GSSGRd, in which no changes were noticed. MDA concentration at 24 h after incubation with the exposure to ELFMF was significantly higher in comparison to the control, without ELFMF. After 48 h of incubation, MDA levels were significantly lower in both groups with no differences between the groups without and with ELFMF. We conclude that ELFMF influences antioxidative enzyme activities and increases lipid peroxidation in 3T3L1 preadipocyte cultures.

Acupuncture: neuropeptide release
produced by electrical stimulation of
different frequencies

Ji-Sheng Han

Neuroscience Research Institute, Peking University, 38 Xue Yuan Road, Beijing 100083, China

Brain functions are regulated by chemical messengers
that include neurotransmitters and neuropeptides.
Recent studies have shown that acupuncture or electrical
stimulation in specific frequencies applied to certain
body sites can facilitate the release of specific neuropeptides
in the CNS, eliciting profound physiological
effects and even activating self-healing mechanisms.
Investigation of the conditions controlling this neurobiological
reaction could have theoretical and clinical
implications
Neuropeptides play important roles in various aspects of
brain function (e.g. opioid peptides in pain control [1] and
neuropeptide Y (NPY) in appetite modulation [2], among
others). This review discusses evidence that neuropeptides
could be mobilized by peripheral electric stimulation to
benefit human health.
It has been shown that physiological and pathological
conditions can induce release of neuropeptides. Two wellknown
examples are a severe painful stimulus inducing
the release of opioid peptides to ease pain, and sucking of
the nipples promoting the secretion of milk. Oxytocinergic
neurons fire at a very low rate, of ,1 Hz (0.1–2.6 Hz) in
basal conditions, but prolonged sucking by ten or more
pups can bring the firing rate up to 16–50 Hz, followed by
strong milk ejection within 10–12 seconds [3]. This finding
suggests that neuropeptide release could be modulated by
external stimulation.
Clinically, intracranial [4] or intra-spinal [5] electrical
stimulation has been used through neurosurgical procedures
to provide relief for patients suffering from chronic
pain, with a success rate of 50–80% after one year of
treatment. This pain-relief effect could involve the release
of neuropeptides [6], raising the attractive possibility that
non-invasive methods might be used to modulate neuropeptide
release for therapeutic intervention. The question
is, would such an approach be effective and practical?
Frequency-dependent neuropeptide release in vitro
In isolated rat neurohypophyses, field electrical stimulation
induces the release of arginine vasopressin (AVP)
and oxytocin (OT) into the incubation medium. Stimulation
at a frequency such as 15–30 Hz was much more
effective than a lower frequency such as 2–3 Hz in
triggering peptide release [7], and burst stimulation was
more effective than constant-frequency stimulation [8].
Furthermore, in superfused rat spinal cord slices, the
release of the neuropeptide substance P (SP) per pulse of
electrical stimulation was increased by frequencies in the
range of 20–50 Hz, whereas release of the small-molecule
neurotransmitter 5-hydroxytryptamine (5-HT) per pulse
remained constant [9]. Hokfelt proposed that peptide
release requires bursting or high-frequency activities,
whereas individual action potentials firing at a low
frequency will not induce the release of peptides [10,11].
The facilitation of peptide release by high-frequency
stimulation was considered to be due to the lengthening
of the action potential duration, together with the increase
in frequency, leading to an increase in Ca2þ entry and in
the amount of secretion per unit of action potential [12].
This concept has been supported by more recent reports
[13]. However, frequency requirement can vary for
different neuropeptides. In a similar experimental setting,
thyrotropin-releasing hormone (TRH) could be released by
electrical stimulation at a frequency as low as 0.5 and 3 Hz
[14].
Frequency-dependent release of CNS opioid peptides by
peripheral electrical stimulation
Peripheral electrical stimulation can be provided via
electrodes placed on the skin (transcutaneous electrical
nerve stimulation, TENS) or via a probe inserted through
skin into the tissue (percutaneous electrical nerve stimulation,
PENS). If the point of stimulation is selected
according to traditional acupuncture therapy, the process
is usually called electroacupuncture (EA). In fact, the
difference between PENS and EA is more hypothetical
than practical. One study compared the analgesic potency
and the underlying neurobiological mechanisms of EA and
TENS, with the acupuncture needles or the skin electrodes
placed at the same ‘acupoints’, and concluded that they
operate through very similar, if not identical, mechanisms
[15]. Thus, the mechanisms of the aforementioned
methods of peripheral stimulation are discussed under
the same heading.
To facilitate the release of opioid peptides in the CNS,
one can use manual acupuncture [16] or EA [17]
stimulation. The parameters of the latter can be precisely
Corresponding author: Ji-Sheng Han (hanjisheng@263.net). characterized. It was interesting to note that analgesia
Opinion TRENDS in Neurosciences Vol.26 No.1 January 2003 17
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induced by low-frequency (4 Hz) stimulation, but not that
induced by high-frequency (200 Hz) stimulation, can be
reversed by low doses of the opioid antagonist naloxone
[17], suggesting that low-frequency stimulation can
increase the release of opioid peptides in the CNS. By
changing the dose of naloxone or using various opioid
receptor subtype-specific antagonists, we were able to
show that analgesia induced by either low- or highfrequency
stimulation are both mediated by opioid peptides
[18,19]. The difference was that the former was
mediated by m and/or d opioid receptors, whereas the latter
was mediated by k opioid receptors [20]. These results
suggest that different kinds of opioid peptides are released
under these different conditions.
Direct evidence comes from our study using radioimmunoassay
of spinal perfusates from the rat [21],
showing that 2 Hz peripheral stimulation produces a
significant increase in the content of enkephalin-like
immunoreactivity (IR) but not in that of dynorphin IR,
whereas 100 Hz increases dynorphin IR but not enkephalin
IR. In a follow-up double-blind study, in collaboration
with Lars Terenius of the Karolinska Institute (Stockholm,
Sweden), the results obtained in rats were fully confirmed
in humans [22]. These studies suggest that (1) the
principle proposed by Hokfelt in 1991 [11] might have to
be revised, and (2) to support our hypothesis, more
evidence, obtained using different approaches, is needed.
To test whether analgesia induced by stimulation at 2
and 100 Hz are mediated differentially in the spinal cord
by enkephalin and dynorphin, respectively, we performed
an antibody microinjection study. Our idea was that
binding of an opioid peptide molecule to its antibody to
form a large protein complex would hinder its approach to
the receptor, resulting in a loss of its biological function.
Indeed, intrathecal injection of enkephalin antiserum
resulted in a dramatic decrease in the efficacy of 2 Hz
EA analgesia. This effect of antiserum diminished as the
EA frequency was increased to 128 Hz. By contrast,
dynorphin antiserum produced an equally dramatic
decrease in the analgesic effect produced by 128 Hz
EA, but this effect diminished gradually with
decreasing frequency, reaching zero at 4 Hz [23]
(Fig. 1). A similar approach was used to study the
possible effect of b-endorphin in mediating EA
analgesia. Injection of b-endorphin antiserum into
rat periaqueductal grey matter resulted in an 88%
decrease of analgesia at 2 Hz EA and a 61% decrease in
analgesia at 15 Hz EA, with no blockade of the analgesic
effect of 100 Hz EA [24].
Another example is endomorphin, a small peptide
composed of only four amino acid residues that has been
recognized as an endogenous opioid peptide with highly
selective affinity for the m-opioid receptors [25]. Antibodies
against endomorphin injected into the cerebral ventricle
[26] or the spinal subarachnoid space [27] dose-dependently
reduced the analgesia induced by 2 Hz EA stimulation, but
not that induced by 100 Hz EA stimulation. This result is
very similar to that obtained with the other two agonists
of m and d receptor already mentioned, enkephalin and
b-endorphin. Taken together, these studies support the
proposition that, although high-frequency stimulation is
preferable for the release of many CNS peptides, it should
not be taken as a gold standard in determining the
parameters of electrical stimulation for activating a
specific neuropeptide for either experimental or therapeutic
purposes.
Putative neural pathways mediating low- and
high-frequency electroacupuncture-induced analgesia
The afferent impulses induced by acupuncture have been
characterized to be mainly transmitted by Ab and Ad fibres
[28]. Wang and colleagues have conducted a series of
experiments to analyze the possible neural pathways
responsible for the frequency-specific release of different
kinds of opioid peptides in rat CNS [29] (Fig. 2). Lesion of
the arcuate nuclei of the hypothalamus abolished analgesia
induced by low-frequency EA but not that induced by
high-frequency EA, whereas selective lesion of the parabrachial
nuclei of the brainstem attenuated the effects of
high-frequency EA but not those of low-frequency EA. The
periaqueductal grey matter is a common element for both
of the descending pain inhibitory systems. These findings
have been partially supported by subsequent morphological
studies using fos gene expression as marker of brain
activation in the rat [30], and functional magnetic
resonance imaging (fMRI) study in human volunteers
(W.T. Zhang, et al., unpublished).
Optimization of peripheral electrical stimulation for
maximal release of central opioid peptides
From the research already mentioned, stimulation at a
single frequency, whether low or high, would not be
sufficient to trigger the full release of all four kinds of
opioid peptide together. To elicit the maximal release of all
four, two models have been considered. Model A involves
stimulation at low (2 Hz) and high (100 Hz) frequencies
alternately (referred to as ‘2/100’), optimally spaced so that
Fig. 1. Antibody-microinjection study investigating the roles played by spinal metenkephalin
and dynorphin A in mediating the analgesic effects that are induced by
electroacupuncture (EA) of different frequencies. Rats were given intrathecal injection
of normal rabbit serum (NRS) or antisera against either met-enkephalin (Enk
AS, red) or dynorphin A (Dyn AS, blue), 30 minutes before the administration of
EA. The analgesic effect was measured by the percentage change of tail-flick
latency (data were normalized with NRS control group as 100%). The analgesic
effects of low- or high-frequency EA were blocked by Enk AS or Dyn AS, respectively.
Modified, with permission, from Ref. [23].
TRENDS in Neurosciences
Dyn AS
Enk AS
n = 13 or 14 rats
Analgesic effect of EA (%)
100
50
0
2 4 8 16 32 64 128
NRS
Frequency of EA (Hz)
18 Opinion TRENDS in Neurosciences Vol.26 No.1 January 2003
http://tins.trends.com
the residual effect produced by the low frequency
stimulation could overlap with that produced by the
high frequency and, therefore, elicit an synergistic effect
[31]. Model B involves stimulation at 2 and 100 Hz
simultaneously (referred to as ‘2 þ 100’) applied at
different parts of the body, in which case all four kinds of
opioid peptide might be released simultaneously (Fig. 3).
Model A has been tested carefully [32], showing that
automatic shifting between low- and high-frequency
stimulation for three seconds each (i.e. 2/100 stimulation)
did, indeed, produce a simultaneous activation of the
enkephalin and dynorphin systems, inducing a much more
potent analgesic effect than that induced by a constant
frequency stimulation.
Formodel B (2 þ 100), two possibilities exist. One (B1) is
that the brain is capable of clearly distinguishing two
different frequencies of stimulation (2 Hz versus 100 Hz)
and induces the two efferent systems to work simultaneously.
The other (B2) is that two different signals
(2 and 100 Hz), coming from two different sites, merge in
the reticular formation of the brainstem so that they are
received as a stimulation of 102 Hz, which is indistinguishable
from a stimulation of 100 Hz.Model B2 is supported by
at least three observations [33]. First, an increase of the
content of dynorphin IR in the spinal fluid (representing an
increase in release of the dynorphin peptide) was observed
in both the 2/100 and 2 þ 100 modes, yet an increase of
the release of endomorphin IR was observed only in rats
treated with 2/100 mode. Second, intrathecal injection of k
opioid-receptor antagonist norbinaltorphimide (Nor-BNI)
suppressed the analgesic effect of both the 2/100 and
2 þ 100 modes, whereas the m opioid-receptor antagonist
D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr amide (CTOP) produced
a selective blockade of the analgesia only in the 2/100
mode. Third, these results have been validated by the
antibody microinjection experiment. Taken together, the 2/
100 mode seems to activate both the m/d and k opioid
systems to induce a synergistic analgesic effect,whereas the
2 þ 100 mode activates only the k opioid system. In
accordance with this hypothesis, the analgesic effect
induced by 2/100 Hz was significantly stronger than that
induced by 2 þ 100 Hz [33]. A recent study using molecular
biology has supported the concept that endogenously
released dynorphin does indeed possesses a strong antinociceptive
effect in the spinal cord [34].
Clinical verification of laboratory findings
The findings obtained in experimental animals have since
been confirmed in humans in clinical practice. White et al.
Fig. 3. Possible mechanisms for the analgesic effects of acupuncture. (a) Opioid
peptides and opioid receptors involved in analgesia elicited by electroacupuncture
of different frequencies. Opioids and receptors involved at 2 Hz are in red, those
involved at 100 Hz, in blue. At 15 Hz, there is a partial involvement of components
involved at both of the other two frequencies (purple). Abbreviations: Dyn, dynorphin
A; b-End, b-endorphin; Em, endomorphin; Enk, enkephalins. Simultaneous
activation of all three types of opioid receptor elicits a synergistic analgesic effect.
Note that simultaneous receptor activation does not necessarily mean that the
opioids are released simultaneously – it could be that the residual presence of one
opioid overlaps with newly induced release of another. (b) Model for the synergistic
analgesic effect produced by alternating low and high frequency stimulation
(referred to as model A in the text). Stimulation at 2 Hz facilitates the release of
enkephalin (red); that at 100 Hz stimulates the release of dynorphin (blue). The
overlapping areas (purple) indicate the synergistic interaction between the two
peptides. Modified, with permission, from Ref. [32].
TRENDS in Neurosciences
Frequency of electrical
stimulation (Hz)
Opioid peptides
Opioid receptors
Interaction
Physiological effects
Em, Enk, β-End
μ
2 15 100
δ
Analgesia
Synergism
κ
Dyn
Peptide released in CNS
Enk Dyn Enk + Dyn
2 Hz 100 Hz 100 Hz
0 3 6 9 12 15
2 Hz
Time (s)
(a)
(b)
Fig. 2. Neural pathways mediating the analgesic effect elicited by low-frequency
(2 Hz, red) or high-frequency (100 Hz, blue) electroacupuncture stimulation.
Abbreviations: DHN, dorsal horn neuron of the spinal cord; Dyn, Dynorphin A;
b-End, b-endorphin; Enk, enkephalin; PAG, periaqueductal grey matter. Modified,
with permission, from Ref. [23].
TRENDS in Neurosciences
β-End
Enk Dyn
Parabrachial nucleus
Arcuate nucleus of
hypothalamus
PAG
2 Hz
Medulla
DHN
100 Hz
Opinion TRENDS in Neurosciences Vol.26 No.1 January 2003 19
http://tins.trends.com
at the University of Texas Southwestern Medical Center
(TX, USA) performed a series of studies to determine
whether peripheral electrical stimulation of the alternating-
frequency mode would produce a significantly stronger
analgesic effect than that produced by stimulation of fixed
frequency in various clinical settings. Observations on
the post-operative requirement of opioid analgesics [35]
revealed that the alternating-mode stimulation reduced
morphine requirement by 53%, whereas a constant low
(2 Hz) or constant high (100 Hz) frequency produced only a
32 or 35% decrease, respectively. Ghoname et al. [36] made
similar observations in patients with chronic lower-back
pain and found that the alternating mode of stimulation
was the most effective in decreasing pain, increasing
physical activity and improving the quality of sleep (when
compared with the pure low- and pure high-frequency
stimulation). Because the alternating mode produced a
more potent analgesic effect, it was used as a standard
mode of stimulation for further studies searching for the
optimal intensity [37] and optimal stimulation duration
[38]. Thus, controlled clinical studies performed in the past
six years using peripheral electrical stimulation for
the control of various forms of acute [35,37] and chronic
[36,38,39] pain have elegantly replicated what we have
found in animal studies over the past two decades.
Results obtained in EA-induced analgesia have been
applied to the treatment of heroin addiction with considerable
success. The withdrawal syndrome observed in
rats dependent on morphine can be effectively suppressed
by 100 Hz EA, which accelerates the release of dynorphin
in the spinal cord [40,41]. By contrast, morphine-induced
conditioned-place preference (CPP), an experimental
model simulating the craving of heroin addicts, can be
successfully suppressed by 2 Hz EA but not 100 Hz EA
[42,43]. This effect can be blocked by a small dose of
naloxone, indicating the involvement of endogenous opioid
peptides interacting with m and d opioid receptors [42,43].
As would thus be expected, in clinical practice the
alternating mode of stimulation has shown strong therapeutic
effects for both physical and psychological dependence
in heroin addicts [44,45].
Responses of other neuropeptides to peripheral
electrical stimulation
Orphanin FQ (OFQ, also known as nociceptin) [46,47] is
another opiate-related neuropeptide that modulates nociception.
Recent studies describe apparent paradoxical
effects of OFQ on pain modulation – analgesia in the
spinal cord and pronociception (an increase in pain
sensitivity) in the brain [48–52]. Analgesia induced by
100 Hz EA can be potentiated by antibodies to OFQ
injected into the cerebral lateral ventricle and suppressed
by the same antibodies injected into the spinal arachnoid
space [53], suggesting that endogenous OFQ released by
100 Hz EA plays opposite roles in brain and spinal cord.
Cholecystokinin octapeptide (CCK-8) has been recognized
as an anti-opioid peptide in the CNS [54]. The most
effective method for stimulating the release of CCK-8 in
the spinal cord with peripheral stimulation is to use
higher frequencies (15 or 100 Hz), whereas 2 Hz is only
marginally effective [55]. Liu et al. [56] measured the
amount of CCK-8 in rat spinal perfusate as an indicator of
CCK-8 release and found that those rats showing a
significant increase in CCK release during 100 Hz EA
stimulation were low responders (i.e exhibited weak EA
analgesia), whereas rats showing little increase in CCK
release were high responders (i.e. exhibited strong EA
analgesia). Moreover, the speed of response also plays an
important role. It seems that the effect of EA analgesia is
determined by, among other things, the magnitude and the
rapidity of CCK release in the spinal cord in response to
peripheral stimulation. This has been confirmed by the
finding that a rat that is not responsive to 100 Hz EA can
be transformed into a responder by injection of antisense
oligonucleotides to CCK mRNA into the cerebral ventricles,
which suppresses the expression of CCK in the
brain [57]. Furthermore, a responder rat can be changed
into a non-responder by inducing overexpression of CCK in
the brain [58].
Substance P mediates nociception at the first synapse in
the spinal cord. In vivo study revealed that peripheral
stimulation in the 8–100 Hz range elevated the content of
SP in rat spinal perfusate, with maximal effect at 15 Hz
[59]. Similar results were obtained in cats (maximal
release at 20 Hz) [60]. By contrast, 2 Hz peripheral
stimulation produced a 50% decrease in the SP content
of the spinal perfusate [59], possibly owing to the release of
enkepahlin [21], which in turn suppressed the release of
SP [61].
Angiotensin II (AII) is another neuropeptide with antiopioid
activity [62]. The release profile is unique, with a
significant decrease (þ62%, P , 0.01) at 15 Hz and a
significant increase (þ60%, P , 0.05) at 100 Hz [63]. The
decrease of AII release can be reversed by the m-preferring
opioid antagonist naloxone, which changed the 62%
decrease into a 125% increase. These results suggest
that opioid peptides are important modulators affecting
the release of other neuropeptides: 2 Hz EA releases
enkephalin, which activates AII and, thus, a negative
feedback control [63]; 100 Hz EA releases dynorphin,
which activates CCK-8 and, thus, another feedback control
[64]. These can be considered as examples of the finetuning
that is achieved by interactions among peptides.
Last, but not least, is the finding that brain-derived
neurotrophic factor (BDNF) can be released by peripheral
stimulation of 100 Hz bursts, but not by pure low- (1 Hz) or
pure high- (constant 100 Hz) frequency stimulation [65].
This has been verified in primary cultures of hippocampal
neurons, in which high-frequency bursts of stimuli evoke
instantaneous secretion of BDNF together with the
induction of long-term potentiation (LTP) [66]. The ability
of peripheral stimulation to accelerate the release of nerve
growth factors has obvious clinical implications.
Concluding remarks
It has long been a dream to cure diseases by non-invasive
measures that activate self-healing mechanisms, without
using drugs or surgical operations. One recent effort along
these lines was the use of repetitive transcranial magnetic
stimulation(rTMS) to stimulate certainareas of the cerebral
cortex; this has achieved limited success in the treatment
of depression [67]. Evidence presented in the present
20 Opinion TRENDS in Neurosciences Vol.26 No.1 January 2003
http://tins.trends.com
review demonstrates that it is possible to facilitate the
release of certain neuropeptides in the CNS by means of
peripheral electrical stimulation. In contrast to magnetic
stimulation, which stimulates the superficial areas of the
brain (i.e. the cortex) [67], peripheral stimulation of the
skin or deeper structures activates various brain structures
and/or the spinal cord via specific neural pathways
(Fig. 2). Any predictions made at this stage should not be
overly optimistic. But the clinical efficacy demonstrated
using frequency-specific parameters to ease post-operative
pain [35,37], lower-back pain [36,38] and diabetic neuropathic
pain [39], and the successful application of 100 Hz
(but not 2 Hz) stimulation for treating muscle spastic pain
of spinal origin [68], certainly hold exciting promise for the
future.
Acknowledgements
I wish to thank Tomas Hokfelt of the Karolinska Institute and Richard
Morris of the University of Edinburgh for their encouragement in
preparing this article. Special thanks go to many of my colleagues and
friends, at home and abroad, who provided helpful suggestions and
editorial comments. This work was supported by the National Basic
Research Programme (G1999054000), the National Natural Science
Foundation of China (39830160) and a grant from the NIDA/NIH of the
USA (DA 03983).
References
1 Hughes, J. et al. (1975) Identification of two related pentapeptides from
the brain with potent opiate agonist activity. Nature 258, 577–580
2 Zarjevski, N. et al. (1993) Chronic intracerebroventricular neuropeptide
Y administration to normal rats mimics hormonal and metabolic
changes of obesity. Endocrinology 133, 1753–1758
3 Summerlee, A.J. and Lincohn, D.W. (1981) Electrophysiological
recordings from oxytocinergic neurons during suckling in the
unanesthetized lactating rat. J. Endocrinol. 90, 255–265
4 Kumar, K. et al. (1997) Deep brain stimulation for intractable pain: a
15-year experience. Neurosurgery 40, 736–747
5 Burchiel, K.J. et al. (1996) Prospective, multicenter study of spinal
cord stimulation for relief of chronic back and extremity pain. Spine 21,
278–294
6 Wang, Q. et al. (1990) Lumbar intrathecal administration of naloxone
antagonizes analgesia produced by electrical stimulation of the
hypothalamic arcuate nucleus in pentobarbital anesthetized rats.
Neuropharmacology 29, 1123–1129
7 Racke, K. et al. (1989) Frequency-dependent effects of activation and
inhibition of protein kinase C on neurohypophysial release of oxytocin
and vasopressin. Naunyn Schmiedebergs Arch. Pharmacol. 339,
617–624
8 Cazalis, M. et al. (1985) The role of patterned burst and interburst
interval on the excitation-coupling mechanisms in the isolated rat
neural lobe. J. Physiol. (Lond.) 369, 45–60
9 Frank, J. et al. (1993) Differential release of endogenous 5-hydroxytryptamine,
substance P, and neurokinin A from rat ventral spinal cord
in response to electrical stimulation. J. Neurochem. 61, 704–711
10 Lundberg, J.M. and Hokfelt, T. (1983) Coexistence of peptides and
classical neurotransmitters. Trends Neurosci. 6, 325–333
11 Hokfelt, T. (1991) Neuropeptides in perspective: the last ten years.
Neuron 7, 867–879
12 Bondy, C.A. et al. (1987) Effects of stimulus frequency and potassium
channel blockade on the secretion of vasopressin and oxytocin from the
neurohypophysis. Neuroendocrinology 46, 258–267
13 Marvizon, J.C. et al. (1997) Neurokinin 1 receptor internalization in
spinal cord slices induced by dorsal root stimulation is mediated by
NMDA receptors. J. Neurosci. 17, 8129–8136
14 Iverfeldt, K. et al. (1989) Differential release of coexisting neurotransmitters:
frequency dependence of the efflux of substance P,
thyrotropin releasing hormone and [3H]serotonin from tissue slice of
rat ventral cord. Acta Physiol. Scand. 137, 63–71
15 Wang, J.Q. et al. (1992) Antinociceptive effects induced by
electroacupuncture and transcutaneous electrical nerve stimulation
in the rat. Int. J. Neurosci. 65, 117–129
16 Mayer, D.J. et al. (1977) Antagonism of acupuncture analgesia in man
by the narcotic antagonist naloxone. Brain Res. 121, 368–372
17 Cheng, R.S. and Pomeranz, B. (1979) Electroacupuncture analgesia
could be mediated by at least two pain-relieving mechanisms:
endorphin and non-endorphin systems. Life Sci. 25, 1957–1962
18 Han, J.S. et al. (1986) Frequency as the cardinal determinant for
electroacupuncture analgesia to be reversed by opioid antagonists.
Acta Physiol. Sinica 38, 475–482
19 Chen, X.H. and Han, J.S. (1992) Analgesia induced by electroacupuncture
of different frequencies is mediated by different types of
opioid receptors: another cross-tolerance study. Behav. Brain Res. 47,
143–149
20 Chen, X.H. and Han, J.S. (1992) All three types of opioid receptors in
the spinal cord are important for 2/15 Hz electroacupuncture
analgesia. Eur. J. Pharmacol. 211, 203–210
21 Fei, H. et al. (1987) Low and high frequency electroacupuncture
stimulation release [Met5]enkephalin and dynorphin A in rat spinal
cord. Sci. Bull. China 32, 1496–1501
22 Han, J.S. et al. (1991) Effect of low- and high-frequency TENS on metenkephalin-
Arg-Phe and dynorphin A immunoreactivity in human
lumbar cerebrospinal fluid. Pain 47, 295–298
23 Han, J.S. (1993) Acupuncture and stimulation-produced analgesia. In
Handbook of Experimental Pharmacology In Handbook of Experimental
Pharmacology (Vol 104/II, Opioids II) (Herz, A., ed.), pp. 105–125,
Springer
24 He, C.M. and Han, J.S. (1990) Attenuation of low- rather highfrequency
electro-acupuncture analgesia followingmicroinjection of bendorphin
antiserum into the periaqueductal gray in rats. Acupunct.
Sci. Int. J. 1, 94–99
25 Zadina, J.E. et al. (1997) A potent and selective endogenous agonist for
m-opiate receptor. Nature 386, 499–502
26 Huang, C. et al. (2000) Endomorphin and m-opioid receptors in mouse
brain mediate the analgesic effect induced by 2 Hz – but not 100 Hz –
electroacupuncture stimulation. Neurosc. Lett. 294, 159–162
27 Han, Z. et al. (1999) Endomorphin-1 mediates 2 Hz – but not 100 Hz –
electroacupuncture analgesia in the rat. Neurosc. Lett. 274, 75–78
28 Lu, G.W. (1983) Characteristics of afferent fiber innervation on
acupoint zusanli. Am. J. Physiol. 245, R606–R612
29 Han, J.S. and Wang, Q. (1992) Mobilization of specific neuropeptides
by peripheral stimulation of identified frequencies. News Physiol. Sci.
7, 176–180
30 Guo, H.F. et al. (1996) Brain substrates activated by electroacupuncture
of different frequencies (II): role of Fos/Jun proteins in the EAinduced
transcription of preproenkephalin and preprodynorphin
genes. Mol. Brain Res. 43, 167–173
31 Huang, L. et al. (1987) Mutual potentiation of the analgesic effects of
met-enkephalin, dynorphin-A-(1-13) and morphine in the spinal cord
of the rat. Acta Physiol. Sinica 39, 454–461
32 Chen, X.H. et al. (1994) Optimal conditions for eliciting maximal
electroacupuncture analgesia with dense and disperse mode stimulation.
Am. J. Acupunct. 22, 47–53
33 Wang, Y. et al. (2002) New evidence for synergistic analgesia produced
by endomorphin and dynorphin. Chin. J. Pain Med. 8, 118–119
34 Cheng, H.Y.M. et al. (2002) DREAM is a critical transcriptional
repressor for pain modulation. Cell 108, 31–43
35 Hamza, M.A. et al. (1999) Effect of the frequency of transcutaneous
electrical nerve stimulation on the postoperative opioid analgesic
requirement and recovery profile. Anesthesiology 91, 1232–1238
36 Ghoname, E.A. et al. (1999) Percutaneous electrical nerve stimulation
for low back pain: a randomized crossover study. J. Am. Med. Assoc.
281, 818–823
37 Wang, B.G. et al. (1997) Effect of the intensity of transcutaneous
acupoint electrical stimulation on the postoperative analgesic requirement.
Anesth. Analg. 85, 406–413
38 Hamza, M.A. et al. (1999) Effect of the duration of electrical
stimulation on the analgesic response in patients with low back
pain. Anesthesiology 91, 1622–1627
39 Hamza, M.A. et al. (2000) Percutaneous electrical nerve stimulation: a
novel analgesic therapy for diabetic neuropathic pain. Diab. Care 23,
365–370
40 Wu, L.Z. et al. (1999) Suppression of morphine withdrawal by
Opinion TRENDS in Neurosciences Vol.26 No.1 January 2003 21
http://tins.trends.com
electroacupuncture in rats: dynorphin and k opioid receptor implicated.
Brain Res. 851, 290–296
41 Cui, C.L. et al. (2000) Spinal k-opioid system plays an important role in
suppressing morphine withdrawal syndrome in the rat. Neurosc. Lett.
295, 45–48
42 Wang, B. et al. (2000) Peripheral electrical stimulation inhibits
morphine-induced place preference. Neurosc. Lett. 11, 1017–1020
43 Wang, B. et al. (2000) Stress or drug-priming induces reinstatement of
extinguished conditioned place preference. Neurosc. Lett. 11,
2781–2784
44 Wu, L.Z. et al. (1999) 2/100 Hz transcutaneous electrical stimulation
for the treatment of heroine addiction. J. Beijing Med. Univ. 31,
239–242
45 Wu, L.Z. et al. (2000) Effect of 2/100 Hz transcutaneous electrical
stimulation on the sexual dysfunction of 33 heroine addicts as revealed
by behavioral questionale and serum testosterone and leutinizing
hormone examination. J. Chin. Tradit. West. Med. 20, 15–18
46 Reinscheid, R.K. et al. (1995) Orphanin FQ: a novel neuropeptide
which is a natural legend of an opioid-like G protein-coupled receptor.
Science 270, 792–794
47 Meunier, J.C. et al. (1995) Isolation and structure of the endogenous
agonist of opioid receptor-like ORL1 receptor. Nature 377, 532–535
48 Mogil, J.S. et al. (1996) Orphanin FQ is a functional anti-opioid
peptide. Neuroscience 75, 333–337
49 Stanfa, L.C. et al. (1996) Inhibitory action of nociceptin on spinal dorsal
horn of the rat, in vivo. Br. J. Pharmacol. 118, 1875–1877
50 Xu, X.J. et al. (1996) Nociceptin or anti-nociceptin: potent spinal
antinociceptive effect of orphanin FQ/nociceptin in the rat. Neuroreport
7, 2092–2094
51 Tian, J.H. et al. (1997) Bidirectional modulatory effect of orphanin FQ
on morphine-induced analgesia: antagonism in brain and potentiation
in spinal cord of the rat. Br. J. Pharmacol. 120, 676–680
52 Tian, J.H. et al. (1998) Endogenous orphanin FQ: evidence for a role in
the modulation of electroacupuncture analgesia and the development
of tolerance to analgesia produced by morphine and electroacupuncture.
Br. J. Pharmacol. 124, 21–26
53 Tian, J.H. and Han, J.S. (2000) Functional studies using antibodies
against orphanin FQ/nociceptin. Peptides 21, 1047–1050
54 Faris, P.L. et al. (1983) Evidence for the neuropeptide Cholecystokinin
as an antagonist of opiate analgesia. Science 219, 310–312
55 Zhou, Y. et al. (1993) Increased release of immunoreactive CCK-8 by
electroacupuncture and enhancement of electroacupuncture analgesia
by CCK-B antagonist in rat spinal cord. Neuropeptides 24, 139–144
56 Liu, S.X. et al. (1999) Relationship between the analgesic effect of
electroacupuncture and CCK-8 content in spinal purfusate in rats.
Chin. Sci. Bull. 44, 240–243
57 Tang, N.M. et al. (1997) Cholecystokinin anti-sense RNA increases
the analgesic effect induced by electroacupuncture and low dose
morphine: conversion of low responder rats into high responders.
Pain 71, 71–80
58 Zhang, L.X. et al. (1992) Lipofectin-facilitated transfer of Cholecystokinin
gene corrects behavioral abnormalities of rats with audiogenic
seizures. Neuroscience 77, 15–22
59 Shen, S. et al. (1996) Frequency dependence of substance P release by
electroacupuncture in rat spinal cord. Acta Physiol. Sin 48, 89–93
60 Go, V.L. and Yaksh, T.L. (1987) Release of substance P from the cat
spinal cord. J. Physiol. 391, 141–167
61 Hirota, N.Y. et al. (1985) Met-enkephalin and morphine but not
dynorphin inhibit noxious stimulation release of substance P from
rabbit dorsal horn in situ. Neuropharmacology 244, 567–570
62 Keneko, S. et al. (1985) Intracerebroventricular administration of
angiotensin II attenuates morphine-induced analgesia in mice.
Neuropharmacology 24, 1131–1134
63 Shen, S. et al. (1996) Angiotensin II release and anti-electroacupuncture
analgesia in spinal cord of rat. Acta Physiol. Sinica 48, 543–550
64 Chen, X.H. et al. (1994) CCK receptor antagonist L-365,260 potentiated
electroacupuncture analgesia in Wistar rats but not in
audiogenic epileptic rats. Chin. Med. J. 107, 113–118
65 Lever, I.J. et al. (2001) Brain-derived neurotrophic factor is released in
the dorsal horn by distinctive patterns of afferent fiber stimulation.
J. Neurosci. 21, 4469–4477
66 Gartner, A. and Staiger, V. (2002) Neurotrophin release from
hippocampal neurons evoked by long term potentiation-inducing
electrical stimulation patterns. Proc. Natl Acad. Sci. USA 99,
6386–6391
67 Melmuth, L. (2001) Boosting brain activity from the outside in. Science
292, 1284–1286
68 Wang, J.Z. et al. (2000) Post-traumatic spinal spasticity treated with
Han’s Acupoint Nerve Stimulator (HANS). Chin. J. Pain Med. 6,
217–224
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22 Opinion TRENDS in Neurosciences Vol.26 No.1 January 2003
http://tins.trends.com

Sabine Ivancsits Æ Elisabeth Diem Æ Oswald Jahn
Hugo W. Ru¨ diger

ORIGINAL ARTICLE
Intermittent extremely low frequency electromagnetic fields cause
DNA damage in a dose-dependent way

Received: 23 October 2002 / Accepted: 7 March 2003 / Published online: 12 June 2003

Springer-Verlag 2003

Abstract Objectives: Epidemiological studies have
reported an association between exposure to extremely
low frequency electromagnetic fields (ELF-EMFs) and
increased risk of cancerous diseases, albeit without
dose–effect relationships. The validity of such findings
can be corroborated only by demonstration of dosedependent
DNA-damaging effects of ELF-EMFs in cells
of human origin in vitro. Methods: Cultured human
diploid fibroblasts were exposed to intermittent ELF
electromagnetic fields. DNA damage was determined by
alkaline and neutral comet assay. Results: ELF-EMF
exposure (50 Hz, sinusoidal, 1–24 h, 20–1,000 lT, 5 min
on/10 min off) induced dose-dependent and timedependent
DNA single-strand and double-strand
breaks. Effects occurred at a magnetic flux density as
low as 35 lT, being well below proposed International
Commission of Non-Ionising Radiation Protection
(ICNIRP) guidelines. After termination of exposure the
induced comet tail factors returned to normal within
9 h. Conclusion: The induced DNA damage is not based
on thermal effects and arouses concern about environmental
threshold limit values for ELF exposure.
Keywords ELF-EMF Æ 50-Hz sinusoidal Æ Intermittent
exposure Æ Comet assay
Introduction
The issue of adverse health effects of extremely low
frequency electromagnetic fields (ELF-EMFs) is highly
controversial. Numerous contradictory results regarding
the carcinogenic potential of ELF-EMFs have been
reported in the literature. Some epidemiological studies
indicate that exposure to ELF-EMFs may lead to an
increased risk of certain types of adult and childhood
cancer, including leukaemia, cancer of the central nervous
system, and lymphoma (Wertheimer and Leeper
1979; Savitz et al. 1988; Feychting et al. 1997; Li et al.
1997), while others (Verkasalo et al. 1993; Tomenius
1986; Schreibner et al. 1993) did not find such an association.
Interpretation of these studies is complicated,
due to different and unreliable methods of exposure
assessment. Therefore, in vitro studies with defined
exposure conditions and with genotoxic effect markers
as endpoints could provide evidence for a carcinogenic
potential of ELF-EMFs.
Up to date, several comprehensive reviews regarding
in vivo and in vitro laboratory studies on ELF-EMFs
have been published (McCann et al. 1993, 1998; Murphy
et al. 1993; Moulder 1998). Conflicting results have been
reported, with genotoxic endpoints such as sister chromatid
exchange (SCE), micronuclei (MN), chromosome
aberrations (CA) and assessment of DNA strand breaks
at exposure levels ranging from 1 lT to 10 mT. The
majority of these studies, however, did not show any
EMF-related genotoxic effects. Several studies with
whole-body exposure of rodents to ELF-EMFs revealed
DNA single-strand and double-strand breaks in the
brain (Lai and Singh 1997; Singh and Lai 1998;
Svedenstal et al. 1999a, 1999b). These results, in the first
place, gave rise to the classification of ELF magnetic
fields as being possibly carcinogenic to humans
(group 2B) by the International Agency for Research on
Cancer (IARC 2002).
As previously reported (Ivancsits et al. 2002a) we
were able to corroborate these findings by demonstration
of an increase in DNA single-strand breaks (SSBs)
and double-strand breaks (DSBs) in cultured human
diploid fibroblasts upon intermittent exposure to a
50-Hz magnetic field, using comet assay under alkaline
(detection of SSBs + DSBs) and neutral conditions
Int Arch Occup Environ Health (2003) 76: 431–436
DOI 10.1007/s00420-003-0446-5
Sabine Ivancsits Æ Elisabeth Diem Æ Oswald Jahn
Hugo W. Ru¨ diger
S. Ivancsits (&)
Division of Occupational Medicine, University Hospital/AKH,
Waehringer Guertel 18–20, 1090 Vienna, Austria
E-mail: Sabine.Ivancsits@akh-wien.ac.at
Tel.: +43-1-404004022
Fax: +43-1-4088011
S. Ivancsits Æ E. Diem Æ O. Jahn Æ H. W. Ru¨ diger
Division of Occupational Medicine
University of Vienna, Vienna, Austria
(detection of DSBs). The extent of EMF-induced DNA
damage was variable in relation to the setting of on and
off times, and was highest at an intermittence of 5 min
on/10 min off. No effects were detected during continuous
exposure.
Here we report studies on the influence of exposure
time and of magnetic-flux densities on the induction of
DNA strand breaks with human fibroblast cultures of
three healthy donors.
Materials and methods
ELF-EMF exposure conditions and cell culture
Human diploid fibroblast strains of donors with different ages
(ES1, male, 6 years old; IH9, female, 28 years old; KE1, male, 43
years old) were initiated from skin biopsies from healthy donors
and maintained in culture as previously described (Ivancsits et al.
2002a). The cells were seeded into 35-mm Petri dishes at a density
of 5·104 cells/3 ml, 24 h prior to ELF-EMF exposure.
The exposure system was built and provided by the Foundation
for Information Technologies in Society (IT’IS foundation), Zurich,
Switzerland, http://ww.itis.ethz.ch). The set-up, which generated
a vertical EMF, consisted of two four-coil systems (two coils
with 56 windings, two coils with 50 windings), each of which was
placed inside a l-metal box. The currents in the bi-filar coils could
be switched parallel for field exposure or non-parallel for control
(sham-exposure). The residual magnetic field in the sham chamber
was at least 150 times (43 dB) lower than the applied field in the
exposure chamber. In addition, both chambers were not completely
insulated from the earth’s magnetic field, which remained at 20–
50 lT. Both systems were placed inside a commercial incubator
(BBD 6220, Kendro, Vienna, Austria) to ensure constant environmental
conditions (37C, 5% CO2, 95% humidity). Two fans
per l-metal box ensured atmospheric exchange of the chambers. A
PC controlled and continuously monitored the exposure set-up.
Data (temperature, current) were collected and stored in an encoded
file. The temperature was monitored at the location of the
dishes during exposure and was maintained at 36.5–37.5C. The
temperature difference between the chambers did not exceed 0.3C.
A current source based on four audio-amplifiers (Agilent Technologies,
Zurich, Switzerland) allowed magnetic fields up to
2.3 mT. The field could be varied in the frequency range from DC
to 1.5 kHz by a computer-controlled function generator. To enable
blind exposures, the computer randomly determined which of the
two chambers was exposed. This information was provided to the
investigator by the IT’IS foundation in Zurich via e-mail in exchange
with the transmission of comet assay results. All experiments
were performed at a frequency of 50 Hz sinusoidal at
intermittent exposure (5 min field on/10 min field off). Timedependent
effects were studied at a magnetic flux density of 1 mT;
for dose–response effects, the magnetic flux density was varied
between 20 and 1,000 lT (5 min field on/10 min field off) at a
constant exposure time of 15 h. After exposure the fibroblasts were
detached with trypsin and suspended in fresh culture medium. To
study repair kinetics, we post-incubated fibroblasts at 37C for 0.5–
9 h. Each exposure level was tested in duplicate.
Comet assay analysis
We followed the technique described by O¨ stling and Johanson
(1984) with minor modifications by Singh et al. (1988, 1991). ELFexposed
and sham-exposed cells (10,000–30,000) were mixed with
100 ll low-melting agarose (0.5%, 37C), and this cell suspension
was pipetted onto 1.5% normal-melting agarose pre-coated slides,
spread with a cover slip, and kept on a cold flat tray for approximately
10 min to solidify. After the cover slip had been removed, a
third layer of 0.5% low-melting agarose was added and allowed to
solidify. The slides were then immersed in freshly prepared cold
lysis solution (2.5 mol/l NaCl, 100 mmol/l Na2EDTA, 10 mmol/l
Tris, pH 10, 1% sodium sarcosinate, 1% Triton X-100, 10%
DMSO, pH 10) and lysed for 90 min at 4C. Subsequently, the
slides were drained and placed in a horizontal gel electrophoresis
tank, side by side and nearest the anode. The tank was filled with
fresh electrophoresis buffer (1 mmol/l Na2EDTA, 300 mmol/l
NaOH, pH>13 or pH 12.1 in the case of alkaline comet assay, and
100 mmol/l Tris, 300 mmol/l sodium acetate, 500 mmol/l sodium
chloride, pH 8.5 in the case of neutral comet assay) to a level
approximately 0.4 cm above the slides. For both alkaline and
neutral comet assay, the slides were left in the solution for 40 min
to allow equilibration and unwinding of the DNA before electrophoresis.
Electrophoresis conditions (25 V, 300 mA, 4C, 20 min,
field strength 0.8 V/cm) were the same for neutral and alkaline
comet assay. All steps were performed under dimmed light to
prevent the occurrence of additional DNA damage. After electrophoresis
the slides were washed three times with Tris buffer
(0.4 mol/l Tris, pH 7.5), to be neutralized, then air-dried and stored
until required for analysis. Comets were visualized by ethidium
bromide staining (20 lg/ml, 30 s) and examined at 400· magnification
with a fluorescence microscope (Axiophot, Zeiss, Germany).
One thousand DNA spots from each sample were classified into
five categories corresponding to the amount of DNA in the tail, in
accordance with Anderson et al. (1994). The proposed classification
system provides a fast and inexpensive method for genotoxic
monitoring. Due to the classification to different groups by eye, no
special imaging software is required. The technique becomes more
sensitive, because many cells can be scored in a short time (1,000
cells instead of 50–100 cells with image analysing). The subsequent
calculation of a ‘‘comet tail factor’’ allows DNA damage to be
quantified as a single figure, which makes it easier for results to be
compared. Due to the scoring of 1,000 cells in one experiment,
which are ten times the cells processed with image analysing,
standard deviations are very low. Reproducibility has been thoroughly
checked.
Results were expressed as ‘‘comet tail factors’’, calculated in
accordance with Diem, with modifications as previously described
(Diem et al. 2002; Ivancsits et al. 2002a, 2002b). The same investigator
performed all analyses. Figure 1 shows the five classification
groups, with the group averages, and the microphotograph.
Fig. 1 Comet assay classification groups and respective microscopic
appearance (cell line ES-1)
432
Tail factors were calculated according to the following formula:
tailfactor% ¼
A
FA þ B
FB þ C
FC þ D
FD þ E
FE
1000
where
A = the number of cells classified to group A and FA
= the average of group A (=2.5)
B = the number of cells classified to group B and FB
= the average of group B (=12.5)
C = the number of cells classified to group C and FC
= the average of group C (=30)
D = the number of cells classified to group D and FD
= the average of group D (=67.5)
E = the number of cells classified to group E and FE
= the average of group E (=97.5)
Statistical analysis
Statistical analysis was performed with STATISTICA V. 5.0
package (Statsoft, Tulsa, USA). All data are presented as mean ±
SD. The differences between exposed and sham-exposed, as well as
between different exposure conditions, were tested for significance
with an independent Student’s t-test. A difference at P< 0.01 was considered statistically significant. Results Fibroblast cultures of three healthy donors were exposed to ELF-EMFs (50 Hz sinusoidal, 1,000 lT, intermittent 5 min on/10 min off) for 1 to 24 h. Alkaline and neutral comet tail factors increased with exposure time, being largest at 15–19 h (Fig. 2). Comet assay levels declined thereafter, but did not return to basal levels. The different cell donors exhibited different basal levels, different maxima, and different end levels. When exposure was terminated after 15 h the comet factor returned to basal levels after a repair time of 7 to 9 h (Fig. 3a), which comprised a fast repair rate of DNA SSBs (<1 h) and a slow repair rate of DNA DSBs (>7 h). The marked comet peak value between 12 and
17 h and the following repair kinetics could also be detected
when ELF exposure was terminated after 12 h
(Fig. 3b). However, it disappeared when comet assay
was performed at pH 12.1 instead of pH>13, thereby
eliminating the cleavage of alkali labile sites in the DNA
(Fig. 4).
Fig. 2a, b Influence of exposure
time on formation of DNA
SSBs and DSBs in three human
fibroblast strains (ES-1, IH-9,
KE-1) determined with comet
assay (1 mT, 5 min on/10 min
off cycles). a Alkaline
conditions; b neutral conditions
433
When magnetic flux densities were varied between 20
and 1,000 lT (cell strain ES-1) we observed a dosedependent
increase of comet factor at alkaline and
neutral conditions, which had already become significant
at 35 lT (Fig. 5, Table 1).
Discussion
Exposure to thermal stress may result in alterations in
the integrity of DNA, comprising DNA strand breaks or
Fig. 3a, b Repair kinetics of
DNA SSBs and DSBs in human
fibroblasts after termination of
ELF-EMF exposure (cell strain
ES-1, 1 mT, 5 min on/10 min
off cycles) determined with
alkaline and neutral comet
assay. a Repair after 15-h ELFEMF
exposure; b repair after
12-h ELF-EMF exposure
Fig. 4 Comet assay of exposed
human fibroblasts was
performed at different pH
(1 mT, intermittent 5 min on/
10 min off)
434
apoptosis (Fairbairn et al. 1995). The induced DNA
damage depends on the extent and duration of the applied
heat stress. Taking these findings into account, we
consider it highly unlikely that, in our experiments, the
observed genotoxic damage is caused non-specifically by
spots of increased temperature within the cell layer as a
secondary effect of the electromagnetic field. If so, the
damage would increase with prolongation of the on time
during the intermittent exposure and would be largest at
continuous exposure. It has previously been shown,
however, that the largest effects are obtained at 5¢ on/10¢
off cycles, and that continuous exposure has no effect at
all (Ivancsits et al. 2002a). Therefore, we conclude that
the observed induction of DNA SSBs and DSBs is a
direct consequence of an intermittent exposure to ELFEMFs.
We observed an increase in DNA breaks up to 15 h
of exposure and then a decline to a ‘‘steady-state level’’
of approximately 1.5-times the base line. This unexpected
finding can be explained if the exposure activates
DNA repair processes and this activation takes a time of
10 to 12 h. After this time the DNA damage is repaired
at an enhanced rate, which leads to a reduction in DNA
breaks, albeit not to a normalisation. This explanation is
supported experimentally by the observation that the
single-strand DNA breaks (alkaline conditions) are repaired
after approximately 30 min, and double-strand
breaks 7 to 9 h after shut down of the exposure (Fig. 3).
The repair process itself also leads to a temporary increase
of alkali-sensitive sites in the DNA, which are
detected as a peak at hours 12 to 17 at comet assay
conditions of pH>13, but not of pH 12.1, the latter not
being able cleave the alkali-sensitive sites (Fig. 4).
It is well known that the repair of SSBs is a fast and
almost error-free process, while the repair of more
complex DNA damage (i.e. DNA DSBs) by homologous
recombination, single-strand annealing or nonhomologous
end joining requires more time and is error
prone in part (Van den Bosch et al. 2002). Therefore,
DNA DSBs may affect the integrity of the genome and
can lead to cell death, uncontrolled cell growth, or
cancer (Van Gent at al. 2001).
In addition, we demonstrate here an increase in DNA
SSBs and DSBs in relation to an increasing magnetic
flux density, which becomes significant at 35 lT at 15 h
of intermittent ELF-EMF exposure. This threshold is
well below the guidelines of the International Commission
of Non-Ionising Radiation Protection (ICNIRP
1998), which propose 500 lT per working day for
occupational exposures and 100 lT per 24 h for the
general population. However, no proposal with regard
to intermittent exposures has been made by the ICNIRP
as yet.
In conclusion, our findings strongly indicate a genotoxic
potential of intermittent ELF-EMFs. The induced
DNA damage was time-dependent and dose-dependent
and points to the need for consideration of environmental
and occupational threshold limit values for ELFEMFs,
in particular with regard to intermittent exposures.
Acknowledgements This study was funded by the European Union
under the programme ‘‘Quality of Life and Management of Living
Resources’’, Key Action 4 ‘‘Environment and Health’’: QLK4-CT-
01574.
References
Anderson D, Yu TW, Phillips BJ, Schmerzer P (1994) The effect of
various antioxidants and other modifying agents on oxygenradical-
generated DNA damage in human lymphocytes in the
comet assay. Mutat Res 307:261–271
Diem E, Ivancsits S, Ru¨ diger HW (2002) Basal levels of DNA
strand breaks in human leukocytes determined by comet assay.
J Toxicol Environ Health A 65:641–648
Fairbairn JJ, Khan MW, Ward KJ, Loveridge BW, Fairbairn DW,
O’Neill KL (1995) Induction of apoptotic cell DNA fragmentation
in human cells after treatment with hyperthermia. Cancer
Lett 89:183–188
Fig. 5 Dose-dependent formation of DNA SSBs and DSBs
determined with comet assay under alkaline and neutral conditions
with cell strain ES-1 (exposure time 15 h, 5 min on/10 min off
cycles)
Table 1 Alkaline and neutral comet assay levels of ELF-exposed
(20–1,000 lT, 15 h, 5 min on/10 min off) and non-exposed human
diploid fibroblasts (ES-1)
Magnetic
flux density
Alkaline comet assay Neutral comet assay
(lT) Comet tail
factor (%)
(± SD) Comet tail
factor (%)
(± SD)
0 4.105 0.028 3.797 0.011
20 4.100 0.092 3.906 0.027
35 5.807a 0.025 4.455a 0.127
50 8.985a 0.170 5.812a 0.018
70 12.450a 0.134 7.605a 0.103
100 14.494a 0.012 8.799a 0.004
1,000 16.213a 0.124 9.311a 0.063
aSignificant differences (P<0.01) between exposed and non-exposed 435 Feychting M, Forssen U, Floderus B (1997) Occupational and residential magnetic field exposure and leukemia and central nervous system tumors. Epidemiology 8:384–389 International Agency for Research on Cancer (2002) Non-ionizing radiation, part 1: static and extremely low-frequency (ELF) electric and magnetic fields. IARC Monogr Eval Carcinog Risks Hum 80:1–395 International Commission on Non-Ionizing Radiation Protection (1998) Guidelines for limiting exposure to time-varying, magnetic, and electromagnetic fields up to 300 GHz. Health Phys 74:494–522 Ivancsits S, Diem E, Pilger A, Ru¨ diger HW, Jahn O (2002a) Induction of DNA strand breaks by exposure to extremely-lowfrequency electromagnetic fields in human diploid fibroblasts. Mutat Res 519:1–13 Ivancsits S, Pilger A, Diem E, Schaffer A, Ru¨ diger HW (2002b) Vanadate induces DNA strand breaks in cultured human fibroblasts at doses relevant to occupational exposure. Mutat Res 519:25–35 Lai H, Singh N (1997) Acute exposure to a 60 Hz magnetic field increases DNA strand breaks in rat brain cells. Bioelectromagnetics 18:156–165 Li CY, Theriault G, Lin RS (1997) Residential exposure to 60 Hz magnetic fields and adult cancers in Taiwan. Epidemiology 8:25–30 McCann J, Dietrich F, Rafferty C, Martin AO (1993) A critical review of the genotoxic potential of electric and magnetic fields. Mutat Res 297:61–95 McCann J, Dietrich F, Rafferty C (1998) The genotoxic potential of electric and magnetic fields: an update. Mutat Res 411:45–86 Moulder JE (1998) Power-frequency fields and cancer. Crit Rev Biomed Eng 26:1–116 Murphy JC, Kaden DA, Warren J, Sivak A (1993) Power frequency electric and magnetic fields: a review of genetic toxicology. Mutat Res 296: 221–240 O¨ stling O, Johanson KJ (1984) Microelectrophoretic study of radiation-induced DNA damages in individual mammalian cells. Biochem Biophys Res Commun 123:291–298 Savitz DA, Wachtel H, Barnes FA, John EM, Tvrdik JG (1988) Case control study of childhood cancer and exposure to 60 Hz magnetic fields. Am J Epidemiol 128:21–38 Schreibner GH, Swaen GMH, Meijers JMM, Slangen JJM, Sturmans F (1993) Cancer mortality and residence near electricity transmission equipment: a retrospective cohort study. Int J Epidemiol 22:9-15 Singh N, Lai H (1998) 60 Hz magnetic field exposure induces DNA crosslinks in rat brain cells. Mutat Res 400:313–320 Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175:184–191 Singh NP, Tice RR, Stephens RE, Schneider EL (1991) A microgelelectrophoresis technique for direct quantitation of DNA damage and repair in individual fibroblasts cultured on microscope slides. Mutat Res 252:289–296 Svedenstal BM, Johanson KJ, Mild K H (1999a) DNA damage induced in brain cells of CBA mice exposed to magnetic fields. In Vivo 13:551–552 Svedenstal BM, Johanson KJ, Mattsson MO, Paulsson LE (1999b) DNA damage, cell kinetics and ODC activities studied in CBA mice exposed to electromagnetic fields generated by transmission lines. In Vivo 13:507–513 Tomenius L (1986) 50 Hz electromagnetic environment and the incidence of childhood tumors in Stockholm County. Bioelectromagnetics 7:191–207 Van den Bosch M, Lohman PHM, Pastink (2002) A DNA double strand break repair by homologous recombination. Rev Biol Chem 383:873–892 Van Gent DC, Hoeijmakers JH, Kanaar R (2001) Chromosomal stability and the DNA double-strand break connection. Nat Rev Genet 2:196–206 Verkasalo PK, Pukkala E, Hongisto MY, Valjus JE, Ja¨ rvinen PJ, Heikkila¨ KV, Koskenvuo M (1993) Risk of cancer in Finnish children living close to power lines. Br Med J 307:895–899 Wertheimer N, Leeper E (1979) Electrical wiring configurations and childhood cancer. Am J Epidemiol 109: 273–284 436

P.R.J. ZUIM, A.R. GARCIA, K.H.L. TURCIO, M.M. HAMATA

According to some authors, headache, earache, TMJ and muscle pain are consequence of muscle hyperactivity. Pain reduction may be achieved by physiotherapy, drugs, psychotherapy and/or occlusal therapy. Physiotherapy by microcurrent (MENS) and its effects on tissue repair process has been studied. However, it is lacking studies comparing microcurrent effect and others TMD therapeutic methods. Objectives: The purpose of this study was to determine if microcurrent therapy aids the masticatory muscle pain reduction in TMD patients.
Methods: Twenty patients divided in four groups were evaluated by means of sensibility to muscle palpation using a visual analogue scale (VAS). One group used occlusal splint therapy and MENS (I); other one, splint and placebo MENS (II); the third, only MENS application (III) and the last group, placebo MENS (IV). The results were submitted to analysis of variance (5% of significance). Results: It was verified that in group I (splint and MENS) there was a pain level reduction about 47.7%; group II (occlusal splint and placebo MENS), 66.7%; group III (MENS), 49.7%; group IV (placebo MENS), 16.5%. Conclusions: There was not statistical difference between MENS application and occlusal splint therapy regarding muscle pain level reduction in TMD patients.

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Xanya Sofra Weiss

JACK SIMONS;   2006

We overview our recent theoretical predictions and the innovative experimental findings that inspired us concerning the mechanisms by which very low-energy (0.1-2 eV) free electrons attach to DNA and cause strong (ca. 4 eV) covalent bonds to break causing so-called single-strand breaks. Our primary conclusions are that (i) attachment of electrons in the above energy range to base π orbitals is more likely than attachment elsewhere and (ii) attachment to base π* orbitals most likely results in cleavage of sugar-phosphate C-O σ bonds. Later experimental findings that confirmed our predictions about the nature of the electron attachment event and about which bonds break when strand breaks form are also discussed. The proposed mechanism of strand break formation by low-energy electrons involves an interesting through-bond electron-transfer process.

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Xanya Sofra Weiss