Abstract:Objective To investigate the therapeutic effect of Qinggongtang in regulating Glu/GABA metabolic balance and the mechanism of its anxiolytic effect on rat models of anxiety.Method Fifty-four rats were randomly divided into normal， model， diazepam （0.225 mg·kg-1）， and low-dose， medium-dose， and high-dose groups of Qinggongtang （5.085， 10.17， 20.34 g·kg-1）， with nine rats in each group. Except for the normal group， the other groups were subjected to indeterminate vacutainer stress and chronic restraint stress for 12 days to prepare the anxiety model. On the 3rd day of the stress， 10 days of corresponding drug intervention was started. At the end of the drug treatment， the anxiety level of rats in each group was evaluated by the elevated cross maze experiment （EPM） and the light and dark box experiment （LDB）， and the effect of Qinggongtang on the anxiety behavior of rats was preliminarily analyzed. The levels of Glu and GABA in the amygdala tissue of the rats were detected by enzyme linked immunosorbent assay （ELISA）， and the changes in the synaptic ultrastructure of the amygdala of the rats in each group were observed by electron microscopy. The mRNA expression of glutamic acid decarboxylase （GAD65 and GAD67）， glutamine synthetase （GS）， and glutamate transporter-1 （GLT-1） in the amygdala were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）， and their protein expression was detected by Western blot.Result Compared with those in the normal group， rats in the model group showed an obvious anxiety state and dull yellow and lusterless fur. They were irritable， easy to anger， and preferred to curl up in the corner. The number of times the EPM entered the open arm and the residence time in the open arm were significantly reduced （P<0.01）， and the residence time in the open box and the number of times the LDB went through the box were significantly reduced （P<0.01）. The content of Glu in the amygdala was increased （P<0.01）， and the content of GABA was reduced （P<0.01）. The value of Glu/GAB was elevated （P<0.01）， and the number of synaptic and pre-synaptic membrane vesicles in the amygdala was decreased. Sparse dense material in the post-synaptic membrane， increased synaptic gap， slightly disrupted internal structure， and decreased mRNA and protein expressions of GAD65， GAD67， GS， and GLT-1 in the amygdala were observed （P<0.01）. Compared with those in the model group， rats in the medium-dose and high-dose groups of Qinggongtang and the diazepam group had bright fur， sensitive reactions， and more active behavior. The number of times EPM entered the open arm and the residence time in the open arm increased significantly （P<0.01）， and the residence time in the open box and the number of times the LDB went through the box increased significantly （P<0.01）. The content of Glu in all-dose groups of Qinggongtang and the diazepam group decreased （P<0.05， P<0.01）， while GABA content increased （P<0.05， P<0.01）. The value of Glu/GABA decreased （P<0.01）， and the internal and external synaptic structure of each groups of Qinggongtang and the diazepam group was more complete. Synapses and vesicles were numerous， and the synaptic gap was more clearly defined. The efficacy of the high-dose group of Qinggongtang and the diazepam group was the best， and the mRNA and protein expressions of GAD65， GAD67， GS， and GLT-1 in the amygdala were increased in the high-dose group of Qinggongtang and diazepam group （P<0.05， P<0.01）.Conclusion Qinggongtang can improve synaptic plasticity and affect the expression of GAD65， GAD67， GS， and GLT-1 in the amygdala of rats to regulate Glu/GABA metabolic balance and thus exert anxiolytic effects.
