| Objective: To investigate whether repeated high dose propofol exposure impaire the developing hippocampus of rats and the spatial learning/ memory ability in later adulthood; whether fasudil hydrochloride had a effect against propofol-induced apoptosis in hippocampus.Methods:1 Animals and model preparation65 healthy postnatal day 7(P7) Sprague-Dawley rats(12–16 g), half male and half female were used in this study.They were randomly assigned into five groups(n=13): group B received no treatment,group S were dosed intraperitoneally with fat emulsion(7.5ml/kg) and group P with propofol(75mg/kg); group PF were injected with fasudil(10mg/kg) and propofol(75mg/kg); group F were dosed intraperitoneally with fasudil(10mg/kg). All treatmen groups were given a 10ml/kg dose by using fat emulsion once a day at regular 24 h intervals for 7 days. Throughout the anesthesia procedure,the pups were placed in a incubator with continuous oxygen supply(2L/min) until they could successfully perform the rightingreflex. After rightingreflex were recovery, the pups were turned back to their maternal side(maternal separation time were not more than 5 hours).2 Materials and method2.1 Blood glucose and oxygen saturation15min after the last injection, selected 5 rats randomly from eath group to detect the blood glucose and oxygen saturation.2.2 Western blot24h after the last injection, 5 rats were anesthetized with 1% sodium pentobarbital(0.003ml/g), decapitated, isolated the hippocampus in ice, and then added the hippocampus in a mixture of lysis buffer and protein phosphatase inhibitors. The hippocampus was homogenized and centrifuged at 4℃, 10000 rpm for 5min. Collect the supernatant to detect the expression of Bcl-2 by western blot.2.3 Detect the caspase3 activity in hippocampus by caspase3 activity assay kitThe method of protein mention was same to 2.2 The reaction substrate was added to the protein to detect the expression of caspase3 enzyme through the absorbance value of the product.2.4 Morris water mazeAt postnatal day 60, morris water maze test was performed to detect brain development for long-term learning/memory ability. Each rat received 2 trials per day for 5 consecutive days. The time spent(escape latency) to reach the platform was recorded. On the sixth day, the rats were placed in the quadrant for the 120-s probe trial where the platform was removed. The number of crossing the platform was recorded.Results:1 The level of blood glucose and oxygen saturation in each group was within normal range and had no significant difference between each two groups(P>0.05).2 Western blot results showed that: Compared with group B, the expression of Bcl-2 was higher then other groups(P<0.05). Compared with group S and group PF,the expression of group P was significantly decreased.(P<0.05). Group PF was decreased compared with the group F(P<0.05).3 Compared with the expression of caspase3 in group B, each group expressed higher(P<0.05). Group P was significantly higher than the other groups(P<0.01). Compared with group F, group PF was higher. There was significant differences between them(P<0.05).4 In five days training, the escape latency of group P and PF were longer than other groups on d3, d4 and d5(P<0.01). Compared with group P, the escape latency in group PF was less on d3, d4 and d5. The two groups were significantly different(P<0.05) and there were no significant difference on the first two days. In the probe trials period, the change trend of the platform crossing number was reversed to the escape latency.Either scape latency nor the number of crossing the platform had no statistically significant among group B, group S and group F.Conclusion: In this study, repeated propofol exposure can induce apoptosis in hippocampal neurons and impair the learning/memory ability in adulthood. Fasudil can reduce the harmful effects of propofol.In addition, repeated injection can also impair the development of the brain in pups, but cause little influence to long-term spatial learning/ memory ability. |
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