The Influence Of Bisphenol A On Androgen-regulated Social Behaviour

BPA?Bisphenol A,BPA?is A kind of typical environmental Endocrine disruptors?Endocrine disrupting chemicals,EDCs?,because it can improve the plasticity of plastic products,transparency and flexibility,widely used medical apparatus and instruments,food packaging materials and other products.Its widespread use not only causes serious pollution to the environment such as water,air and soil,but also exposes human and wildlife to it.BPA can affect brain and behavioral development and gender differentiation,as well as synaptic plasticity and a variety of neural behaviors in the adult brain.Social behavior refers to the social communication and social relations between different individuals of the same species,including communication,hierarchy,aggression,courtship,mating,feeding,etc.,which are crucial to the survival and reproduction of animals.Previous studies in our laboratory found that adolescent BPA exposure decreased the social behavior,social inquiry and aggression of adult male rats and the opposite sex,and increased the affinity.BPA has been reported to inhibit the binding of DHT to androgen receptor?AR?and antagonize the transcriptional activity mediated by AR.Sex hormones regulate the regulation of social behavior by the neuropeptides vasopressin?AVP?and oxytocin?OT?,and are also involved in the regulation of the reward system of the midbrain-limbic DA and affect social motivation.In this study,a male mouse model with testicular removal of exogenous testosterone supplementation was used to investigate the mechanism of BPA's influence on social behavior by detecting the changes in social behavior after BPA exposure in adolescence,analyzing the AVP and receptor and OT receptor levels in related brain regions,and the expression levels of proteins related to the DA system.Research methodThe castration model was established by excision of testis in 3 weeks of clean grade ICR male mice after 1 week of acclimatization.There were three groups:sham surgery?sham?,testicular excision?GDX?,and testosterone propionate?TP?supplementation?GDX+TP?.Each large group was given BPA?0.4,4 mg/kg/day?exposure and a solvent control.One week after the operation,daily subcutaneous injection of TP?0.5 mg/kg/day?supplement and BPA contamination was started.BPA was continuously contaminated for 18 days,and TP supplementation was carried out until the end of the experiment.At week 8,adolescent play behavior was detected,and at week 12,adult social behavior was detected.The non-social odor olfactory function was detected by the feed burial experiment,social competence and novelty preference were detected by the three-chamber model,social interaction and social behavior were detected by the social interaction model,and aggression was detected by the residential invasion model.Meanwhile,serum and amygdala testosterone?T?and estradiol?17?-E₂?levels were detected by ELISA.Western blotting?WB?was used to detect the expression levels of androgen receptor?AR?,estrogen receptor?ER??,neuropeptide AVP receptor 1a?V1aR?and OT receptor?OTR?,DA transporter?DAT?and DA type I receptor?DR1?and type II receptor?DR2?in amygdala brain region.The level of AVP in amygdala was detected by immunofluorescence.Research results1 Levels of sex hormones and receptorsCompared with sham group,GDX significantly reduced T and 17?-E2 in amygdala and serum T levels?p<0.001;p<0.05;p<0.001?,all of them recovered after TP supplementation,while GDX and TP supplementation did not affect serum 17?-E₂level,suggesting that the model was successfully established.BPA exposure significantly reduced T levels in the amygdala of Sham and GDX mice?p<0.001,p<0.001?.p<0.05);The serum T levels of sham,GDX and GDX+TP mice were decreased?all p<0.001?,and the levels of 17?-E₂ in the amygdala and serum were significantly increased?p<0.05;p<0.001,p<0.001;P<0.001,p<0.01?.WB analysis found that compared with sham,GDX significantly reduced the AR level of amygdala in mice?p<0.001?,which was reversed after TP supplementation?p<0.001?.BPA exposure reduced AR expression in the amygdala of sham and GDX+TP mice?p<0.01;p<0.05?,but had no effect on the GDX mice.Meanwhile,BPA exposure reduced ER?expression levels in the amygdala of sham and GDX+TP mice?p<0.05,p<0.01;P<0.05?,and increased ER?expression in GDX mice?p<0.01?.These results showed that BPA exposure reduced T levels in the serum and amygdala regions of the mice,but increased E2 levels in the 17?-E2.The expression levels of AR and ER?in sham and GDX+TP mice were down-regulated,and the level of ER?in testicular excision mice was up-regulated.2 Behavior tests2.1 Adolescent social play behavior Compared with sham group,the play ability of GDX mice with the opposite sex was reduced?p<0.01?,and was recovered after TP supplementation?p<0.01?.BPA did not affect adolescent play in mice.2.2 Adult behavior?1?Three-chamber test BPA exposure significantly reduced the time ratio between sham and GDX mice to sniff unfamiliar mice/empty cage?p<0.05;P<0.05,p<0.05?,significantly reduced the sniffing time ratio of sham mice to unfamiliar/familiar mice?p<0.05,p<0.01?.The olfaction time ratio of unfamiliar mice/empty cage and unfamiliar mice/familiar mice in GDX+TP mice was slightly reduced,but no statistical difference was found.BPA was found to have no effect on olfactory function.This suggests that BPA exposure during adolescence impairs the social and novelty preferences of the mice,but does not affect general olfactory function.?2?Social interactions In social interaction with same-sex individuals,neither GDX nor BPA exposure affected the cross-climbing behavior of the mice,but BPA exposure increased the time spent sniffing the genitals?p<0.05;p<0.05,p<0.01;p<0.01,p<0.05?.This suggests that BPA may increase homosexual proximity in mice.In the social interaction with the opposite sex,GDX reduced the interaction time,the time of sniffing the genitals and the incidence of climbing?p<0.05,p<0.001,p<0.05?,increased the incubation period of climbing?p<0.05?,and was reversed after TP supplementation?p<0.001?.p<0.05;p<0.05;p<0.01).BPA exposure in sham and GDX+TP mice significantly reduced the time of interaction with the opposite sex,the time of sniffing the genitals,and the incidence of straddling?p<0.01,p<0.05?.p<0.01,p<0.001;p<0.05;p<0.05;p<0.05;p<0.05,p<0.01),extended the crawl span latency period?p<0.05;P<0.05,p<0.01?,but no effect on GDX mice.It is suggested that BPA exposure in adolescence can antagonize the promotion effect of TP on male and female social interaction and social behavior.?3?Resident-intruder test Compared with sham group,GDX significantly reduced the attack time of male rats against the invaders?p<0.001?,and recovered after TP supplementation?p<0.001?.BPA exposure reduced the attack time of sham and GDX+TP mice?p<0.01?.p<0.01,p<0.01);Increased affinity between males in each group.This suggests that BPA may enhance same-sex affability and reduce aggression in adult male rats by antagonizing TP.3 Amygdala vasopressin?AVP?and its receptor?V1aR?and oxytocin receptor?OTR?levelsCompared with sham group,GDX decreased the AVP level of amygdala?P<0.001?,and was reversed after TP supplementation?p<0.001?.BPA exposure significantly reduced amygdala AVP levels in sham and GDX+TP mice?p<0.001,p<0.001?.p<0.001),but had no effect on GDX mice;WB results showed that the change trend of amygdala V1aR was consistent with that of AVP level in mice.But OTR levels did not change.It was suggested that BPA exposure inhibited the promoting effect of testosterone on AVP system activity in MeA in male rats,but had no effect on OT system.4 Dopamine receptor?DR1,DR2?and transporter levels?DAT?in striatumCompared with sham group,GDX significantly upregulated DAT level in striatum of mice?p<0.01?,and TP supplementation had no effect.BPA exposure significantly increased DAT levels in sham and GDX+TP mice?p<0.01,p<0.01;p<0.05?;4 mg/kg/d BPA exposure increased the expression level of DR1 in sham group?p<0.05?,which had no effect on other groups.BPA exposure and testicular removal did not affect DR2 levels in the amygdala.This suggests that BPA exposure affects the activity of the DA transmitter system in the striatum,but probably not by interfering with androgen.ConclusionBPA exposure did not affect adolescent play ability in male mice during adolescence but affected social behavior in adulthood.BPA inhibited the effects of TP on the reduction of homosexual aggression and the reduction of heterosexual interaction and social behavior in castrated male mice.These inhibitory effects of BPA may be related to the regulatory effect of androgens on the neuropeptide AVP system in the brain region of MeA associated with social recognition by reducing the level of androgens and their receptors in the brain.