Effects Of Methyl Tert-Butyl Ether (MTBE) On Learning And Memory In Rats And Its Possible Mechanism

ObjectiveWidespread used as gasoline oxygenate additives, methyl t-butyl ether (MTBE) were added to gasoline to enhance the octane number, reduce carbon monoxide in exhausts and to substitute for tetraethyl lead as an anti-knock agent.starting in the winter of 1992, MTBE(up to 15wt.%)has been added to about 30% of all gasoline sold in the United States, at present, 31 petroleum refinery have MTBE manufacture equipment in our country, and 16 of them starte to produce. With the widespread used of no-lead gasoline, it is necessary that full scale operation and use in our country. due to leakage of gasoline, MTBE content in drinking water. Because of MTBE in atmosphere, environmental water and soil, it is very important and has far-reaching meaning that study toxity of MTBE no-lead gasoline to protection consumer healthy and to prevent the pollution of the environment.Expernment study reported that MTBE product higher affinity and damage to CNS. Neurological symptoms reported following human exposure to MTBE and reformulated gasoline include headache, dizziness, nausea, tinnitus, fatigue, memory fall-off, sleep even cognitive disorder by investigation. The mechanism of MTBE damage the CNS have not clearly, and have not better evaluation standard and method, so it is very importantly problems that research the biological effects and mechanis of action of MTBE to CNS need to solve.Learning and memory function is very complicated and subtile nervous activity, as importance contents of evaluation CNS function, from the effect of learning and memory to explore the damage and mechanism, objective evaluation toxicity efficacy and deep elucidated the toxicity mechanism, it is scientific and practically.MethodsTo establish the animal poisoning model by male Sprague-Dawley rats, searching reasonable and pragmatic method relatively from intragastric administration, inhalation and lateral ventricle injecting. At last we have established animal model of impact of MTBE for ability of learning and memory in rats by lateral ventricle injecting, detect the changes of learning and memory ability in rats by Morris water maze; observe the impact of MTBE for morphous and construction of hippocamp neuron by Nissl's staining; detect the transcriptional level change of GABAAα1 and NMDAR2A&2B in hippocamp by RT-PCR; detect the proteinum expression change of GABAAα1 and NMDAR2A&2B in hippocamp by immunohistochemical method; dete ct some proteinum molecule and its change of phosphorylation level in MAPK signal transduction pathway by Western blot; observe apoptosis of hippocamp neurone by TUNEL staining.Results1,The body weight was significantly decreased after MTBE intragastric administration the 6th days, and the body weight growth rate was gradually to lower with the time lasting. Morris water maze tests showed that the rat's ability of learning and memory were not obviously affected, Nissl's staining of brain slices showed that Nissl's body of CA1 region line disorder, cavities increased.2,MTBE and its metabolites have been determined in rats cerebrospinal fluid by Gas Chromatography after intragastric administration or inhalation subchronic exposed.3,After the MTBE was injected into the lateral ventricle of rats, the learning ability impaired, the memory time elongation, the ability of spatialization decreased, the reaction time of memory presentation lengthen.3.1 Nissl's staining of brain slices showed that Nissl's body of CA1 region line disorder, structural deformity, cavities significantly increased. The results of RT-PCR and immunohistochemistry showed that the level of GABAARα1 mRNA and protein was gradually increased and the level of NMDAR2A mRNA and protein was gradually decreased in the hippocampus than control group by different doses exposured of MTBE. The results of different exposure time of MTBE showed that the level of GABAAα1 mRNA gradually decreasing from 3h after, to 24h up to normal level, gradually increasing and keeping high transcriptional level. The level of NMDAR2A mRNA increasing in 6h, but decreasing after 24h and significantly decreased in 7d(P<0.05); the level of NMDAR2B mRNA decreaing postoperation, at 12h up to normal level; Morris water maze showed that the abilities of learning and memory of rat's significantly decreased by MTBE combine the GABAAR agonist muscimol, but the abilities of learning and memory increasing by MTBE combine the GABAAR antagonist bicuculline.3.2 TUNEL staining showed that the increasing of hippocampus neuron apoptosis induced by MTBE, Western blot showed that the level of p-ERK gradually increasing in 12h postoperation,then gradually decreasing and lower than control group in 5d and 7d, the high dose group decreased more than the low dose group. with the time lasted, the level of p-JNK gradually increasing and up to highest in 12h, then slow-moving decreasing, but still higher than the control group in7d; the level of pp38 lower than control group in 3h, then gradually increasing and up to highest in 5d, still higher than the control group in7d; the high dose group increased more than the low dose group.Conclusion1,MTBE can through the blood brain barrier and reach brain by subacute intragastric and inhalation administration;2,The abilities of learning and memory will be decreased by lateral ventricle injection MTBE;3,GABAAR and NMDAR may be as a role in the damage of MTBE on learning and memory in rats, the level of GABAAR mRNA and protein increasing and the level of NMDAR2A mRNA and protein decreasing; 4,MTBE can induce the hippocampus neuron apoptosis, the phosphorylation of JNK and p38 increasing and the phosphorylation of ERK decreasing, that changes may be as part of mechanisms of MTBE damage the abilities of learning and memory in rats.