| Objective: After establishing conditioned fear stress model, the indicators of stress-reaction and brain damage of the rats were tested, analyzed and evaluated to screen the indexes of anti-fear drug's efficacy, and to provide the material for evaluation system of anti-fear drug's efficacy.Methods: 64 male Wistar rats were divided into 2 groups at random: fear-stress group and control group. In each group, there are 32 rats. After 10 days, the fear-stress group was given 2s tone +3s inescapable electric foot-shock for 30 times at 1mA. Retention training was conducted 24 hours after the first foot shock by placing the animals back into the same chamber, so is repeated on 13th day. On the 14th day, there was only tone for 5 min and no foot-shock, then took a video of rat's behavior change and recorded the freezing time in 6～30 min. The freezing time can help to evaluate success of the conditioned fear stress model. Three observers scored the rat's behavior after seeing the video in the blind way. In the 1st, 2nd, 3rd, 4th week(on 18th, 25th, 32nd, 39th day), the rats were observed and measured for their general statuses, stress behavioral reactions (freezing, spontaneous activities and so on), physiological reactions (serum corticosterone), and brain damages (neon-specific enolase and morphology of hippocampus).Results: 1. In fear stress group rats, score of ethology had discrepancy significantly compared with control group. Score of ethology in fear stress group rats was higher than control group (P <0.01), suggesting the conditioned fear stress model was successfully established. 2. Compared with control group, there were languid bearing, matte disheveled hair, reduced diet amount and loose stool appeared in fear stress group rats. The weights of two group rats were increased, the lower weight gain had been observed in fear stress group rats. There were discrepancy significantly between fear stress group and control group at week2 (P <0.05), week3 (P <0.05) and week4 (P <0.01). 3. Compared with control group, there were increased irritability, increased fighting behavior, less action and idleness appeared in fear stress group. For the rest time in spontaneous activities, there were discrepancy significantly between fear stress group and control group at week1 (P <0.05), week2 (P <0.01), week3 (P <0.05) and week4 (P <0.05). 4. For the level of serum corticosterone, there were discrepancy significantly between fear stress group and control group at week1(P <0.05),week2(P <0.01),week3(P <0.05) and week4(P <0.05). Fear stress could increase the level of serum corticosterone. 5. For the level of serum NSE, there were discrepancy significantly between fear stress group and control group at week1(P <0.05). Fear stress could increase the level of serum NSE at week1. There was no discrepancy significantly between fear stress group and control group at week2, week3 and week4. To observe the pathological variation of hippocampus tissue by H-E staining: compared with control group, there were different degrees degeneration and cell loss in hippocampal neurons in fear stress group. In the right side's hippocampus region of control group rats, the cells were in good arrangement and the form was complete. At week1 and week2 of fear stress group, neurons in the regions of CA1, CA3 and DG showed a remarkable edema, lightly stained and sparse. At week3, neuronal atrophy and necrosis in hippocampal CA1, CA3 and DG regions occurred in the fear stress rats. At week4, neurons presented irregular shape, a large number of neurons were necrosis, nuclei pyknosis and fragmentation.Conclusion: The conditioned fear stress model was established successfully through modified Takeuchi's method. Conditioned fear stress could result in acute and chronic fear stress, affected physiological responses, behavioral reactions and brain damages in rats. The results provide the important information of fear stress influencing the changes of stress reaction and brain functions, and the material for evaluation system of anti-fear drug's efficacy. |
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