Effect Of Microglia Activation On Lead Induced Learning And Memory Deficits And Its Possible Mechanism

【Background】Lead is the main risk factor of environmental chemistry which seriously endangers children's health in our country. Characteristics of childhood lead poisoning are wide popularity, lasting and hidden toxicity, and children's intellectual development damaged mainly, etc. Now about 10 % to 15 % of urban children's blood lead exceed the standards, industrial and mining enterprises around more serious, and even more serious events of lead pollution and lead poisoning in children happened. It influenced stability of social and physical and mental health of child severely. Therefore, to study the cellular and molecular mechanisms of lead exposure on hippocampus, learning and memory, and to propose effective preventive measures, are urgently needed to solve the problems.Microglia is an important residence of immune cells in the central nervous system. Microglial cells are very sensitive to external environmental stimuli. When many factors of micro-environment change, microglial cells react quickly, they turn into activating with deformed and phagocytosis from resting state, and can significantly upregulate the levels of secretion of many proinflammatory cytokines such as tumor necrosis factorα(TNF-α), interleukin-lβ(IL-1β) and so on. Activated microglia can also upregulate some of inflammatory-related factors such as the expression of iNOS, these factors are considered play important role in mediating neuronal injury.Some studies found that activation of microglia stimulated by lipopolysaccharide can induce memory impairment, and our preliminary study found that lead exposure can induce the activation of microglia. Accordingly, we speculate that activation of microglia may play an important role in hippocampal neurons injury, learning and memory decline induced by lead.【Aim】To study the effect of lead exposure on hippocampal neurons activities, learning and memory; To investgate the effect and mechanism of microglia in neuronal injury induced by lead; To provide theoretical and experimental evidences for prevention and treatment of childhood lead poisoning.【Methods】(1) Established animal models of low lead exposure through drinking water, established Transwell model with microglia and hippocampal neurons co-cultured in vitro.(2) Recorded LTP through the patch-clamp technique, observed the ability of learning and memory through morris water maze through detecting hidden platform latency and target quadrant time. (3) Detected hippocampus injury with double labeled by TUNEL and NeuN, detected effect of lead exposion on activation of microglial with immunohistochemistry.(4) Detected the expression of microglia activation products, IL-1βand TNF-α, by ELISA. Detected the expression of iNOS, microglia activation products with immunohistochemistry.【Results】1. Learning and memory injury caused by lead exposure.Long-range blocked of rats induced by 75mg/L of lead exposure for 8 weeks, suggested that lead exposure could induce learning and memory impairment.2. Hippocampal neurons injury caused by lead exposure.Hippocampal neurons injury induced by 75mg/L of lead exposion for 8 weeks in vivo or 50μmol/L of lead exposion for 24 h in microglia and hippocampal neurons co-cultured in vitro. It suggested that learning and memory injury caused by lead which might because hippocampal neurons injury caused by lead exposure.3. Activation of microglia induced by lead exposion.Activation of microglia could be detected in model which treated with 0.1 % of lead exposure for 8 weeks in vivo or in model treated with 50μmol/L of lead exposure for 24 h in microglia and hippocampal neurons co-cultured in vitro. Model of Transwell established through microglia treated with 50μmol/L of lead for 24 h in vitro then co-cultured with hippocampal neurons in normal nutrient medium for 24 h, hippocampal neurons injury was detected. The results showed significant difference, suggested that lead caused hippocampal neurons injury could through activation of microglia induced.4. Inhibited the expression of iNOS could pretected hippocampal neurons from injury induced by lead exposure to a certain extent.Microglia and hippocampal neurons co-cultured in vitro treated with 300μmol/L SMT and lead for 24 h, results showed that expression of iNOS in microglia in lead + SMT group significantly lower than lead exposure group (33.4±1.08 fold compare to control group, P<0.01). After SMT blocking the expression of iNOS, extent of hippocampal neurons injuries significantly decreased compared with lead exposure group (P<0.01), suggested that expression of iNOS induced by lead exposure could cause hippocampal neurons injury to a certain extent.5. Inhibition of microglia activation could significantly protect the hippocampal neurons from the damage of lead exposure.Microglia and hippocampal neurons co-cultured in vitro treated with 45μmol/L of SMT and lead exposure for 24h, results showed that extent of microglia activation and their activation products in SMT + lead exposure group both significantly lower than lead exposure group (P<0.01). After SMT blocking the activation of microglia and their activation products, the extent of hippocampal neurons injuries significantly decreased (P<0.01). Results suggested that lead could cause hippocampal neurons injury through inducing activation of microglia and its activation products.【Conclusion】1. Lead exposure can cause hippocampal neuronal injury, learning and memory decline, suggesting that lead may induce hippocampal neurons damage then cause learning and memory decline.2. Lead exposure causes microglia activation and also induces hippocampal neurons injury, suggesting activation of microglia may be one of the reasons that hippocampal neurons injury induced by lead and then cause learning and memory decline.3. Antagonist activation of microglia and upregulation of expression of its its activation products, to a certain extent, can protect hippocampal neuronal from injury, and further confirm that the activation of microglia play an important role in the process of hippocampal neuronal injury induced by lead, which suggest that lead cause hippocampal neurons injury may through inducing the activation of microglia and increases the expression of their activation products, then result learning and memory decline.