Effect Of Maternal Deprivation And Chronic Mild Stress On The Depressive Behavior And Methylation Mechanism Of Prostate Apoptosis Response-4 Expression In Adult Rats

Objective:To study the effect of early life stress and chronic stress on the depressive behavior and the expression of prostate apoptosis response-4 (par-4) in adult rats'Striatum and to explore whether the DNA methylation was involved in the regulation mechanism of par-4 expression.Methods:49 newborn rats were randomly divided into four groups, maternal deprivation group (MD, n=17), chronic mild stress group (CMS, n=17), dual stress group (DS, n=10) and control group (C, n=17). MD rats received maternal deprivation. CMS rats received chronic mild stress. DS rats received both maternal deprivation and chronic mild stress. Control rats received no experimental handle. Behavior tests including sucrose consumption test, forced swimming test and open field test were carried out to evaluate depressive and anxious level of the rats. Par-4 and DRD2 mRNA expression in striatum were detected by Real-Time PCR; par-4 protein expression in striatum was detected by Western Blot; and DNA methylation level of par-4 was investigated by bisulfite-sequencing PCR (BSP). Before sequencing, the target DNA fragment was implanted into T-vehicle and cloned.Result:1. the result of behavior tests showed that the MD group rats displayed depressive-like behaviors including reduction of sucrose preference, craw distance and vertical movement (P<0.05), extend time of float (P<0.05), but the rate of central area and number of stool were not significantly different compared with control group rats (P>0.05). The CMS group rats showed a reduction of sucrose preference and vertical movement(P<0.05), extend time of float(P<0.05), but the craw distance, central area rate and stool number were not significantly different compared with control group rats(P>0.05). The float time of dual stress rats was significantly longer and vertical movement was significantly less than the control group(P> 0.05), and the sucrose preference, craw distance, rate of central area, stool number were not significant different compared with the control group rats (P>0.05).2. The mRNA level of par-4 and DRD2 in MD, CMS, DS group rats were significantly lower than the control group rats. The par-4 expression of MD, CMS and DS group rats compared to control group rats were 0.53,0.33 and 0.61 separately, and the DRD2 expression of MD, CMS and DS group rats compared to control group rats were 0.31,0.33 and 0.54 separately. There was a significant relationship between par-4 mRNA level and sucrose preference rate, float time, craw distance, stool number, vertical movement (r were 0.408,0.537,0.504,0.478,0.464 separately, P<0.05), and there was no significant relationship between par-4 mRNA level and the rate of central area (r=0.155, P>0.05). There was a significant relationship between DRD2 mRNA level and float time, craw distance, vertical movement (r were 0.90,0.389,0.400 separately, P<0.05), and there was no significant relationship between DRD2 mRNA level and sucrose preference rate, the rate of central area, stool number (r were 0.133,0.070,0.094 separately, P>0.05). Also, the difference between groups in par-4 protein level was significant (F=14.597, P=0.000), the expression of par-4 protein was significantly lower in MD, CMS, DS group rats than that in control group (P<0.05), and the CMS and DS group rats had a significant lower expression of par-4 protein than the MD group rats (P<0.05).3. The difference between groups in total methylation level was not significantly different (F=1.079, P=0.359). But there were significant difference in four CG site when counted according to CG site (F= 3.103, 3.605,6,862,5.196 separately, P<0.05). CMS rats had 3 CG site hypermethylated and 1 CG site hypomethylated compared to the control rats (P<0.05).Conclusion:1. MD, CMS and DS would cause depressive-like behavior in rats, and CMS may also cause anxiety behavior in rats.2. MD, CMS and DS would cause par-4 and DRD2 mRNA expression decreased in rats'striatum, and there was a relationship between par-4 or DRD2 mRNA expression and depressive-like behavior. MD, CMS and DS also would cause par-4 protein expression decreased in rats'striatum.3. DNA methylation may not be the main mechanism of down regulation of par-4.