The Impacts Of Mid-to Late Prenatal Exposure Of Morphine On The Vulnerability To Drug Addiction In Adult Offspring And The Express Of Netrin-1 And TH In The Nucleus Accumbens

Objective:Prenatal exposure to drug of abuse often leads to physiological and neurobiological abnormalities including decreased brain and body weight,cognitive deficits and behavioral alterations.A handful of studies showed increased vulnerability to drug abuse in prenatally drug-exposed offspring.Prenatal exposure to analgesic doses of morphine during gestation days12 to 18 increases opioid receptor density in the nucleus accumbens and central amygdala of adult male rats. Both the nucleus accumbens and central amygdala play important roles in modulating drug-induced reward via the mesolimbic dopaminergic system.So we come to the hypothesis that prenatal exposure to morphine can increase the vulnerability to drug abuse.We carried out the experiment through injecting the offspring rats morphine or saline.We used conditioned place preference test model to detect the vulnerability of the offspring.(1)To establish and perfect the model of mid-to late pregnancy morphine exposure rats.(2)To study the influence of mid-to late pregnancy morphine exposure to the weight and the mortality of rat offspring.(3)To study the influence of mid-to late prenatal morphine exposure to the vulnerability to drug abuse in adult offspring.(4)To detect the level of axon guidance factors(Netrin-1) and tyrosine hydroxylase(TH) in the nucleus accumbens of the mid-to late morphine adult offspring,then compare it to the saline exposure group.(5)To detect the level of axon guidance factors(Netrin-1) and serotonin(5-HT) in the nucleus accumbens of the adult offspring after conditioned place Tpreference(CPP),then compare it to the saline exposure group.To explore the neurochemical mechanisms of prenatal mid-to late morphine exposure leading to increased susceptibitity to addiction in adult offspring.Methods: Pregnant Spraguee-Dawley rats were randomly divided into Morphine group(M group) and Normal saline group(N group).Pregnant rats assigned to the morphine group were injected with morphine twice daily at 09:00 h and 17:00 h on gestation da-ys 11e18. Female rats assigned to the saline group received a saline injection at the sa-me time.And the dose of morphine progressively increased with 1 mg/kg until it was 5mg/kg from a beginning dose of 2mg/kg. All injections were administered subcutaneously(s.c.). Rats assigned to the saline group received a saline injection at the same tim-e. And the volume of saline injected to pregnant rats of N group was equal to the M group. Observed the weight of the off-spring at 4 weeks, 8 weeks and the day of birt-h and the mortality of the offspring 4 weeks, 8 weeks and 3 days after birth.(1)Three day before the start of the experiments,the animals were placed in the room for 2h for habituation.(2)On day 1-3, pre-drug place preference was tested through letting the rats in the room freely for 15 min.(3) CPP conditionings and drug injections were performed from day 4 to day 10[morphine(3mg/kg,i.p.)/saline(1mg/kg,i.p.)/saline], alternating morphine injection in the morning with saline injection in the afternoon.(4)Post-drug place preference was measured and recorded on day 11.(1)The offspring rats were raised to 8weeks old,then selected 8 rats from each group randomly and removed their nucleus accumbens tissue after anesthesia.(2)After CPP test the adult offspring were divided into two groups-morphine exposure(CPPM)and saline exposure(CPPN).Selected 8 rats from each group randomly and removed their nucleus accumbens tissue.(3)Western-blot assay the level of Netrin-1 and 5-HT in the nucleus accumbens.(4)Immuofluorescence ssay the level of Netrin-1 and 5-HT in the nucleus accumbens.Results:(1)The offspring,s birth weight of M group was less than N group(P <0.01); M group male and female offspring born four weeks and eight weeks weighing less than N group(P <0.01). The mortality of M group rat offspring that born three days, four weeks and eight weeks was significantly higher than N group(P <0.01).(2)After 7 days of continuous injection of morphine,the time of M group of offspring staying in the drug side was significantly longer(P<0.01),while the time of N group of offspring staying in the drug side was not significantly longer(P<0.01)(3)he level of Netrin-1 and 5-HT in the nucleus accunbens of M group was higher than N group.(2)The level of Netrin-1 and 5-HT in the nucleus accunbens of CPPM group was higher than CPPN group.Conclusions :(1)Mid-to late gestational morphine exposure can cause growth retardation and increase the mortality of offspring.(2)Mid-to late prenatal exposure to morphine can increase the vulnerability to drug abuse.(3)The increased susceptibility to drug addiction of mid-to late morphine exposure adult offspring may be associated with the increases of Netrin-1and TH in the nucleus accumbens.