The Effects Of ELF-EMF Exposure On Calcium Signals And Expression Of Glutamate Receptors In Rat Entorhinal Cortex

With the rapid development of science and technology. the electronic equipments gain its popularity in every walk of life. However. with the widely using of electronic equipments. the electromagnetic field(EMF). considered as a kind of pollutant source. has been flooding human’s living space. In terms of it’s the energy. the rates of EMF from high to low is particle radiation. X ray. ultraviolet. infrared radiation. high frequency and low frequency radiation respectively; while in terms of spectrum. it can be mainly divided into ionizing radiation. non ionizing radiation and extremely low frequency magnetic fields-Electromagnetic field(ELF-EMF). It is well known that electronic radiation with high energy and radio frequency(RF) exerts a harmful effect on human being. Recently. people realize that ELF-EMF also have the same harmful effect on health of human being. ELF-EMF is produced by cables for household and equipments for industrial power transmission. It has been proved that exposure to ELF-EMF will deteriorate the body of human and animals including cardiovascular. immune. nervous and productive system. Recent researches mainly focused on the connection between ultra low frequency radiation or radio frequency and some diseases. such as childhood leukemia. brain tumors. malformation. neurogenic diseases. decreased immune function. allergy. mammary cancer. abortion. angiocardiopathy. etc. As a vulnerable target organ for electromagnetic radiation,the nervous system is considered to be associated with lateral sclerosis,Alzheimer’s disease,insomnia. headache. energy loss. memory problem and some other diseases.Cognitive function is a complex psychological and physiological process. which needs activities of abundant neurons in multiple brain areas including hippocampus. entorhinal cortex(EC). prefrontal cortex. etc. In 19 th century. people had found that cables for electrical transmission could deteriorate cognitive function. which indicated that ELF-EMF does affect the learning and memory functions. The mechanisms underlying EMF-induced deficit of learning and memory remain elusive.As the interface of hippocampus,the entorhinal cortex(EC) connects senior cortex and hippocampus. The entorhinal cortex is divided into MEC. miainly participating in the processing space information. and LEC. which is relative to non-space informationprosessing. The anatomic structure of entorhinal cortex makes it an important part in hippocampus-entorhinal-neocortex system. and also the fundament of the formation and consolidation of episodic memory.Hence. to explore the mechnisms underlying EMF-induced cognitive dsfunction. this research focus on entorhinal cortex and investigate how EMF affects the animals’ spatial congnition behavior,the physiological activities of neurons and the protein molecule expression.The main results are as follows:1. The effects of ELF-EMF exposure on intracellular calcium in rat entorhinal cortexPrimary cultured neurons from entrohinal cortex of P0 rats were individed in to 2 groups. One group was for vehicle control. The other group was for ELF-EMF exposure(50Hz. 1 or 3 m T) for 24 hr. After exposure. the neurons were removed to a stage for whole cell recordings or calcium imaging.(1) Whole-cell currents in cultured neurons were tested after ELF-EMF exposure. The findings showed that there were no significant differences in amplitude and density of whole-cell currents in cultured neurons from EC between ELF-EMF and control group. The outward and inward currents were also being analyzed including transient outward currents and persistent outward currents. No obvious differences were dectected in these outward and inward currents in cultured neurons between two groups.(2) ELF-EMF exposure does not affect the amplitude and density of high voltage-activated calcium currents(HVA) in cultured EC neurons. The statistic data showed that there was no detected difference in HVA currents between ELF-EMF(1m T). ELF-EMF(3m T) and control group.(3) ELF-EMF exposure has no influence on the magnitude and density of low voltage-activated calcium currents(LVA) in cultured EC neurons. LVA is composed of transient inactivation currents. The peak of transient currents was analyzed by plotting I-V curve. The data demonstrated that calcium currents from LVA channels after ELF-EMF were voltage-dependent. Difference of LVA currents was not siginificant in three groups.The above data suggested that ELF-EMF has no significant influence on activation of voltage-gated calcium currents in EC cultured neurons.(4) Effects of ELF-EMF exposure on inactivation of voltage-gated calcium currents in these cultured neurons were further investigated. Inactivation potentials and slope factors were analyzed. The data demonstrated that neither HVA nor LVA inactivation would be changed after ELF-EMF exposure.(5) Calcium imaging experiments showed that ELF-EMF exposure has no influence on cconcentration of intracellular calcium. but attenuates high K+-evoked intracellular calcium elevation in cultured neurons. The maximum concentration of intracellular calcium induced by high K+ was decreased by ELF-EMF in a dose-dependent manner. which could be blocked by thapsigargin inhibition of calcium releasing from internal calcium store(6) In the primary cultured neurons of the entorhinal cortex. the effects of the magnetic field exposure(1m T or 3 m T. 24 h) on the high potassium(10 m M) stimulation-induced changes in intracellular calcium dynamics were completely blocked after depletion the inner intracellular calcium stores by 5 u M thapsigargin(TG). an inhibitor for the Ca2+-ATP enzyme.2. The effects of ELF-EMF exposure on ionotropic glutamate receptor expressionIt is widely accepted that changes in number of glutamate receptors is an indicator of synaptic plasticity. which is closely related to learning and memory function. The different subunits of AMAP and NMDA receptors were examined in EC. hippocampus and prefrontal cortex. including Glu A1. Glu A2. Glu A3. Glu N1. Glu N2 A and Glu N2 B.(1) 14 days’ exposure(4 hours per day. 0. 5 m T) did not alter the subunits expression of AMPA receptors(Glu A1. Glu A2 and Glu A3) in EC. Compared to EC. the expression of AMPA subunit receptors showed no change in hippocampus after ELF-EMF exposure. While decrease of Glu A3 but not Glu A1 and Glu A2 expression was detected in prefrontal cortex(n=5. P<0. 05).(2) After 14 days’ exposure. the expression levels of Glu N1 and Glu N2 A but not Glu N2 B increased in EC(n=5. P<0. 05). The same changes of NMDA subunits were also detected in prefrontal cortex. In hippocampus. the expression of Glu N2A(n = 5,P < 0. 05) increased. but Glu N1 expressions remained unchanged(n = 5,P > 0. 05).(3) In addition. we conducted 28 days’(4 hours per day) magnetic field exposure of 0. 5m T. and checked the number of protein isoforms in in entorhinal cortex. We found that. in entorhinal cortex. the number in Glu A2 expression decreased(n = 6,P < 0. 05). Compared to EC. exposure of 0. 5 m T did not alter AMPA subunits expression in hippocampus and prefrontal cortex. The data indicated that Glu A2 in EC were more sensitive to ELF-EMF exposure.(4) Glu N2 A expression in EC decreased after 28days’ exposure to ELF-EMF. While the amount of Glu N2 B proteins increased and Glu N1 did not show any changes. In hippocampus. expression of Glu N2 B but not Glu N1 increased after ELF-EMF exposure. Compared to EC. changes of Glu N2 A expression were not detected in hippocampus and prefrontal cortex. Glu N1 but not Glu N2 B expression in prefrontal cortex increased after ELF-EMF exposure.(5) Finally. T-maze task was used to test the effect of ELF-EMF on rat’s spatial working memory. The correct rate increased gradually with the argument of training days. 14 days’ and 28 days’ exposure to ELF-EMF did not affect learning and memory in rats. which suggested that the changes of proteins in EC were insufficent to alter spatial working memory.Overall. entorhinal cortex is prone to be damaged by ELF-EMF exposure. which decreased intracellular calcium in EC in a dose-dependent manner. Exposure to EMF for long term. AMPA and NMDA receptor. related tosynaptic transmission and plasticity. in EC and other brain regions will show significant changes. While in our experimental conditions. the changes of these receptors expression did not alter spatial working memory in rats.