The Effects Of Lead Exposure At Different Levels On Learning And Memory In Mice

OBJECTIVELead widely exists in nature and extensively uesed in the industry and people's daily life because of its flexibility and antisepsis. However, lead is a neurotoxic heavy metal and can pollute the environment by long-term accumulation and enter human body through food chain, water, soil and air. Children are susceptible to lead exposure. Lead can seriously harm children's growth, development and intelligence.Nitric oxide (NO) is a retrograde messenger, and plays an important role in learning and memory. Its biosynthesis is modulated by nitric oxide synthase (NOS). By measuring NOS activity, we can estimate the content of NO in the tissue. There are three types of NOS, neurone NOS (nNOS), inducible NOS (iNOS) and endothelial NOS (eNOS), among which, nNOS and eNOS are merged as constructional NOS (cNOS). However, the research outcomes on the effects of NO level and NOS activity are still controversy.The study was aimed to investigate the effects of different lead exposure on lead content, NO levels, NOS activity and learning ability in mice. The relationships between lead exposure level and brain damage as well as the potential correlation between lead exposure and NOS were explored. MATERIALS AND METHODS1 Experimental animalsThe subjects were weaned KM mice with the body weight at 18g to 22g. The total number of subjects was 120 and half for each gender. They were randomly assigned into non-exposure control group and three levels of lead exposure groups, thirty for each group. The lead exposure was carried out by adding lead acetate in drinking water. The concentrations of lead acetate were 0.3g/L, 1.0g/L and 3.0g/L respectively.2 Sample collectionThe first day of starting lead exposure was assigned as day Zero. On the 7^th, 14^th and 21^st days, blood was collected from the mice tail. Ten mice from each of the groups were killed by decapitation. The blood was collected and the serum was separated. The brain, spleen, kidneys, liver, lungs and femurs were dissected and stored at -20℃for future use.3 Observation on physical developmentThe mice body weight was recorded once a week.4 Determination of the lead concentration in blood and tissuesThe lead levels in serum, brain and other tissues were measured using Z-5000 graphite furnace atomic absorption spectrometer.5 Evaluation of learning and memory abilityOn day seven, fourteen and twenty-one, of lead exposure, ten mice from each groups were evaluated for their learning and memory function using modified Morris water maze test. After three times training, the duration of escape was recorded.6 Determination of NO content and NOS activity in serum and brainAt these three time points, after behavior test, the mice were killed. Their serum and brain tissue were analyzed for the NO content and NOS activity.7 Data analysisThe results were analysed using SPSS 12.0. One way ANOVA meathod and rank sum test (K-W test) were used. The significant level was set at 0.05 .RESULTS1 The effects of lead exposure on body weightThe mice in both the non-exposure and exposure groups gained body weight, but the later ones gained less and showed the downward weight gain trend (P<0.05); the weekly weight growth rate of the lead-exposued mice showed a downward trend, and all less than the control group's, there were significant differences between them (P<0.05).2 The effects of exposure on blood and tissue lead levelsThe lead levels in blood, brain, bone tissues were increased with the lead exposure time and dosage (P<0.05); liver, kidneys, and spleen tissues were increased with the lead exposure dosage (P<0.05); The lead level in lung tissue was not significantly different between the exposure groups and the control group (P>0.05).3 The effects of lead exposure on the learning and memory3.1 Morris water-maze testThe escape latency was significant longer in the lead-exposed groups, the longer and higher dosage exposure, the poor learning performance (P<0.05)3.2 The effects of lead exposure on NO content and NOS activity in the brain and the serum3.2.1 The effects of lead exposure on NO content and NOS activity in the brainBoth of the content of NO and total NOS activity were decreased in the lead-exposed mice and more severe declines were observed in the mice with higher dosage and longer time exposure (P<0.05). The changes of the activity of iNOS and cNOS in brain were not obviously.3.2.2 The effects of lead exposure on NO content and NOS activity in mice serum The content of NO in serum was significantly decreased along with thelead-exposure time (P<0.05).The overall effect of lead exposure on serum NOS and iNOS activity was decreased. The cNOS activity in serum was decreased in exposed mice (P<0.05).3.3 Correlation analysisThe serum cNOS activity was negatively associated with serum lead on the 7^th day (r=-0.316, P=0.047); and serum lead, brain lead as well as the escape latency on 14^th day and 21^st day (r=-0.365, P=0.020; r=-0.361, P=0.022; r=-0.458, P=0.003; r=-0.731, P=0.000; r=-0.512, P=0.001; r=-0.698, P=0.000 respectively).CONCLUSION1 The body weight gain in mice was effected by lead exposure. The higher dosage and the longer time exposure showed more serious effects.2 The learning and memory performance was impaired in lead exposure mice.3 The content of NO and the NOS activity in the brain and serum was decreased along with the lead-exposure time, and the serum cNOS activity may be relevant with learning and memory of mice.