The Effects Of Maternal Sleep Deprivation On Behavior Of Offspring Rats Associated With Neuroinflammatory Response

Women in pregnancy are at particular risk of sleep deprivation or restriction. The investigation revealed that more than two-thirds of pregnant women reported poor sleep quality, shortened sleep duration and increased number of awakenings. Although sleep complaints are very common among pregnant women, the potential adverse effects of sleep disturbance on the offspring are not well studied. In the present study, we investigated the effects of maternal sleep deprivation(MSD) on long-term behavioral and cognitive consequences in offspring and its underlying mechanism, the main results of this study are as follows:1. Sleep deprivation was induced using multiple-platform method on pregnant Wistar rats to detect the physiological indicators of mothers. According to the modified multiple-platform method built sleep deprived model, confirmed this model succeed and did not have abortion phenomenon. Pregnant rats for sleep deprivation were deprived 72 hours at different pregnant periods: early gestational time(Gestational day 4, GD 4), middle gestational time(Gestational day 9, GD 9) and late gestational time(Gestational day 18, GD 18). After sleep deprivation has been established, weight of pregnant rats was detected at GD 1, GD 7, GD 12 and GD 21. The results showed that weight of pregnant rats has no difference between different groups at different gestational times.2. The pregnant Wistar rats received prolonged sleep deprivation(72 h) on GD 4, 9, and 18, respectively. The prepuberty(post-natal day 21, PND21) offspring was used to explore the effect of cognition, behavior and neurogenesis. Hippocampus-dependent spatial learning and memory in the Morris Water Maze task, neurogenesis in hippocampus by immunohistochemistry, weight in PND 1 and PND 21, litter size, sex ratio and preference of sucrose were discussed in offspring. The PND 21 offspring showed impaired hippocampus-dependent spatial learning and memory and decreased hippocampus neurogenesis in prepuberty offspring, especially for the late MSD(GD 18) group. This revealed that late pregnant time is a sensitive period for sleep deprivation, and the next experiments were all chose this time to investigate the effect of MSD on offspring. Born weight and weight in PND 21 decreased, litter size reduced and anhedonia raised in sucrose preference experiment in late MSD offspring. Sex ratio was not changed in late MSD offspring.3. The effect of maternal sleep deprivation during late pregnancy on neuroinflammation and microglia was detected by real-time PCR(RT-PCR) and immunohistochemistry staining. Neuroinflammation was revealed by the expression of inflammatory cytokines in four stages: PND 1、PND 7、PND 14 and PND 21, the microglial activated status and phenotype were indicated by Iba1+ staining. This study demonstrated that the expression of pro-inflammatory cytokines: IL-1β、TNFα and IL-6 was increased in hippocampus of late MSD offspring on PND 1、PND 7、PND 14 and PND 21, whereas anti-inflammatory cytokine IL-10 reduced at the same time. Immunofluorescence found that the cells of activated microglia were higher in the brains of MSD offspring, resting microglia were lower in hippocampus of MSD rats, and the pro-inflammatory cytokine IL-6 released from activated microglia. This illustrated microglial activation in hippocampus of MSD offspring played a central role on cognition and neurogenesis.4. To explore the protective effect of inhibiting neuroinflammation on cognition and neurogenesis of MSD offspring. Microglial activated inhibitor-minocycline was chosen to discuss the specific mechanism on microglia. The prepuberty male offspring were given the intraperitoneal injection with or without minocycline for five days. After minocycline administration, the MSD offspring showed improvement in Morris water maze and increase in BrdU+/DCX+ cells by immunofluorescence. Minocycline can ameliorate the damage of hippocampus-dependent spatial learning and memory, and improve the hippocampus neurogenesis. This demonstrated microglia is the key moderator in cognitive function and neurogenesis.5. The underlying mechanism on the cognition and neurogenesis of MSD offspring improved by minocycline. Microglial activation is often divided into two phenotypic profiles: the classical activation(M1) and the alternative activation/deactivation state(M2). RT-PCR, enzyme-linked immunosorbent assay(ELISA) and immunofluorescence were used to examine the expression of M1 and M2 markers in MSD offspring. The classical microglial activation markers(M1 phenotype): IL-1β, IL-6, TNF-α, CD68 and iNOS, were increased, while the alternative microglial activation markers(M2 phenotype): Arg1, Ym1, IL-4, IL-10 and CD206, were reduced in hippocampus of MSD offspring. Minocycline reduced Iba1+ cells, suppressed the production of pro-inflammatory molecules, and reversed the reduction of M2 microglial markers and neurogenesis-related genes in the MSD prepuberty offspring. Minocycline exerts its neuroprotective effect by inhibiting the microglial polarization into M1, facilitating M2 activation. These results indicate that dysregulation in microglial pro- and anti-inflammatory activation is involved in MSD-induced inhibition of neurogenesis and impairment of spatial learning and memory.In conclusion, the impairment on cognition, behavior and neurogenesis is due to microglial activation-induced neuroinflammation. Minocycline plays the role of neuroprotection through inhibiting microglial activation and regulating the rebalance of M1 and M2 in MSD offspring.