Effects of bumetanide on changes of NKCC1 and KCC2 mRNA expression in hypothalamus and anxiety in adulthood induced by multiple sevoflurane exposure in neonatal rats / 中华行为医学与脑科学杂志

ABSTRACT

Objective:To investigate the effects of bumetanide on changes of NKCC1 and KCC2 mRNA expression in hypothalamus and anxiety in adulthood induced by multiple sevoflurane exposure in neonatal rats.Methods:Eighty-one healthy male Sprague-Dawley rats, at postnatal 5 days (P5), were randomly divided into 3 groups ( n=27 in each group) control group (group C), multiple sevoflurane group (group MS) and bumetanide group (group B). The rats were commonly reared in the cage and received no anesthesia in group C. Animals were exposed to 2.1% sevoflurane for 2 h on P5, P7, P9 in group MS and group B. In group B, animals received intraperitoneal injection of 1.82 mg/kg bumetanide(Na + -K + -2Cl - cotransporter 1 blocker, NKCC1 blocker)at 30 min before every anesthesia.The animals in group C and group MS received the same dose of dimethyl sulfoxide subcutaneously at the same time as group B. The rats were observed for 30 minutes after recovery from anesthesia, and then breastfed normally.On the 9th day after birth, six rats were taken from each group immediately at the end of anesthesia and the blood was collected by left ventricular puncture for blood gas analysis.At 30 min after anesthesia, 6 animals in each group were decapitated and the hypothalamus part of brain tissue was collected.Then the expression level of IL-6 mRNA, NKCC1 mRNA and KCC2 mRNA were detected by RT-PCR.The other rats in each group were raised to 60 days for the elevated plus maze (EPM) test. Results:Compared with group C, the expression of IL-6 mRNA and NKCC1 mRNA in hypothalamus of MS group was up-regulated (IL-6 (1.000±0.207) vs (1.782±0.231); t=6.899, P<0.01; NKCC1 (1.000±0.255) vs (1.639±0.290); t=3.518, P<0.01), the KCC2 mRNA expression was down-regulated ((1.000±0.140) vs (0.733±0.115); t=3.017, P<0.001) and the NKCC1/KCC2 mRNA ratio increased ((1.000±0.276) vs (2.054±0.521); t=5.078, P<0.001) and the differences were statistically significant.Compared with MS group, the expression of IL-6 mRNA and NKCC1 mRNA in hypothalamus of group B was down-regulated (IL-6 (1.147±0.140); t=5.635, P<0.01; NKCC1 (1.038±0.385); t=3.310, P=0.01), KCC2 mRNA expression was up-regulated((0.988±0.194); t=2.880, P<0.05), NKCC1 / KCC2 mRNA ratio was decreased((1.027±0.200); t=4.950, P<0.001), and the differences were statistically significant.EPM behavioral test showed that compared with group C, the open arm residence time in MS group was significantly shorter than that in group C ((18.4±10.1)s vs (4.3±3.1)s; P<0.01); compared with group MS, the open arm residence time in group B was significantly prolonged((16.6±7.6)s, P<0.05). Conclusion:Bumetanide can reduce the up-regulation of NKCC1 level and the down-regulation of KCC2 level in neonatal rats after sevoflurane anesthesia, and alleviate the anxiety state of adult rats.