| ObjectivesThere are many sensitative factors to Posttraumatic Stress Disorder, including sex, age, race, predisposition, PTSD from family and early life stress. Recently, early life stress,due to a greater risk or increased vulnerability for developing psychiatric illnesses, becomes more and more important and it takes our attention gradually. So we used suppression subtractive hybridization (SSH) technique to investigate gene expression profiles in Hippocampus CA1 tissue collected from 3 months of age between maternal separation (MS) and control rats after contextual fear conditioning, try to explain the the mechanism of the gene action, to identify the function of differentially expressed genes and bring experimental data for medication through genetic approach.MethodsMaternal separationMS mission was performed daily for six consecutive hours (from 13:00 to 19:00), during which rat pups were removed from their home cage and kept alone in temperature-controlled cages at 32±1 ?C, where bedding was renewed every day. This procedure was applied between postnatal days 1 and 21. . Groups of rats were weaned and sacrificed on PND22.Contextual Fear ConditioningThe rats were placed into the box (conditioned stimulus, CS) and after 180 s were given a 2s, 0.8 mA shock for three times with an interval of 60 s. Rats were left in the conditioning box for 60 s after termination of the procedure and returned to their home cage. After 1h of the contextual fear acquisition the rats were returned to the box for a 5 min period,On day 2 (24 h later), the rats were returned to the box for a 5 min period.Morrris water mazeWe employ a method in which each rat is given four trials per day for four consecutive days. For each trial, the rat is allowed to swim a maximum of 90 sec in order to find the hidden platform. When successful, the rat is allowed a 30 sec rest period on the platform. If unsuccessful within the allotted time period, the rat is given a score of 90 seconds and then physically placed on the platform and also allowed the 30 sec rest period. In either case the rat is immediately given the next trial (ITI = 30 sec) after the rest period. On Day 5, two trials are given in which the platform is removed from the pool to measure spatial bias. The important measure will be the percentage of the total time elapsed and distance swam in which the rat is in the boundaries of the previous target quadrant.ResultsBehavioralContextual Fear ConditioningAcquisition: Animals in all 4 groups exhibited postshock freezing, in MS groups displaying lower postshock freezing compared to controls, but there was no reliable main effect of either maternal separation or sex.During contextual testing 1h later, depicts mean percentage of contextual freezing. There was a significant interaction effect between maternal separation and sex (P < 0.01), with females (P < 0.01), but not males (P=0.673), in females MS group showing lower contextual freezing levels compared to controls. And during contextual testing next day (24h after fear conditioning acquisition), there was a significant interaction effect between maternal separation and sex (P < 0.01), with females (P < 0.01), but not males (P=0.093), in females MS group showing lower contextual freezing levels compared to controls.Morrris water mazeIn the acquisition phase of MWM task, time spent and distance swam to reach the platform position for both groups (C and MS) improved significantly over trials. Overall analysis (repeated measures ANOVA) showed no significant effect of MS in latency or distance swam in female rats and male rats to find the platform. And also in the retention phase there was no reliable main effect of maternal separation.SSH9 up-regulated and 1 down-regulated genes were found by SSH, among them one is new EST sequence. And the expression changes of these genes are possibly related to the mechanism of impaired contextual fear conditioning induced by MS.ConclusionsEarly life stress impairs fear conditioning retention in female rats,which may result from greater corticosterone levels and stonger negative feedback of the HPA axis in response to foot shock. And the expression changes of these genes are possibly related to the mechanism of impaired contextual fear conditioning induced by MS... |
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