柴胡-黄芩药对配伍及用量比例对抑郁小鼠行为学及脑海马BDNF/TrkB信号通路的影响

Effects of Different Dosage Proportions of Chaihu(Bupleri Radix)-Huangqin(Scutellariae Radix) on Behavior Tests and BDNF/TrkB Signaling Pathway in Depression Mice

目的通过实验的方法探究柴胡-黄芩药对的抗抑郁功效及机制。方法采用慢性不可预见温和应激(CUMS)的方法造模,将造模成功的小鼠随机分为模型组、阳性药组、柴胡-黄芩不同用量比例3组,灌胃给予柴胡-黄芩不同配伍比例的水煎剂,观察其对CUMS小鼠糖水偏好、悬尾、开野等行为学实验的影响;测定小鼠海马中脑源性神经营养因子(BDNF)、脑源性神经营养因子酪氨酸激酶受体B(TrkB)及蛋白激酶B(Akt)的表达,并观察其对海马凋亡相关蛋白B淋巴细胞瘤-2(Bcl-2)以及Bcl相关X蛋白(Bax)的调控作用,探讨柴胡-黄芩药对的抗抑郁机制。结果与空白组小鼠相比,模型组小鼠糖水偏好率降低、悬尾不动时间延长、开野穿格次数减低(P<0.01),柴胡-黄芩(1∶1)配伍及柴胡-黄芩(8∶3)配伍可增加糖水偏好率、缩短其悬尾不动时间、增加开野穿格次数(P<0.01,P<0.05),柴胡-黄芩(3∶8)配伍可缩短小鼠悬尾不动时间(P<0.05),柴胡-黄芩(1∶1)配伍在糖水偏好实验、悬尾不动实验及开野实验中均优于其他两组配伍组(P<0.01)。蛋白印迹结果显示,与空白组相比,模型组小鼠BDNF表达减少,TrkB、Akt磷酸化水平降低(P<0.01),柴胡-黄芩(1∶1)配伍及柴胡-黄芩(8∶3)配伍,可增加海马BDNF的表达、增加TrkB自磷酸化及Akt的磷酸化水平(P<0.01,P<0.05),其中柴胡-黄芩(1∶1)配伍组BDNF表达高于柴胡-黄芩(3∶8)配伍;TrkB及Akt磷酸化水平高于柴胡-黄芩(3∶8)配伍及柴胡-黄芩(8∶3)配伍。免疫组化结果显示,与空白组相比,模型组小鼠Bcl-2蛋白表达降低、Bax蛋白表达增加(P<0.01),柴胡-黄芩(1∶1)配伍及柴胡-黄芩(8∶3)配伍可上调Bcl-2的表达,下调Bax的表达,柴胡-黄芩(8∶3)配伍组可下调Bax的表达,其中柴胡-黄芩(1∶1)配伍优于柴胡-黄芩(3∶8)配伍(P<0.01)。结论柴胡-黄芩(1∶1)配伍及柴胡-黄芩(8∶3)配伍可改善CUMS小鼠抑郁样行为,发挥抗抑郁功效;其抗抑郁的作用机制可能与调控海马中BDNF/TrkB信号通路从而影响海马中凋亡蛋白的表达,保护海马神经元有关。

Objective To investigate the possible antidepressant mechanism of couplet medicine of Chaihu(Bulerui Radix) and Huangqi(Scutellariae Radix). Method Chronic unpredictable mild stress(CUMS) was taken for modeling. After three weeks of modeling, the successful modeling mice were randomly divided into five groups:model group, positive control group, Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(1∶1) group, Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(3∶8) group, Chaihu(Bulerui Radix)-Huangqi(Scutellariae Radix)(8∶3) group. The decoction of this couplet medicine was given by intragastric administration, and the antidepressant effect was observed by behavior tests. The expression of brain-derived neurotrophic factor(BDNF) and the phosphorylation levels of Tropomyosin receptor kinase B(TrkB) and proteinkinase B(Akt)were measured by Western bloting, and the expressions of Bcl-2 and Bax were observed by immunohistochemical method. Result Compared with the blank group, the sucrose preference rat and the number of crossing of the model group were significantly decreased(P<0.01) and the immobility time of tail suspension was significant increased. Compared with the model group, the sucrose preference rat and the crossing number of the Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(1∶1) group and the Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(8∶3) group were increased(P<0.01 or P<0.05). The immobility time of tail suspension was decreased(P<0.01 or P<0.05) and the number of crossing was also decreased in the Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(1∶1) group(P<0.01). Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(1∶1) group was superior to the other two groups. Besides, while comparing with the blank group, the expression of BDNF and the phosphorylation levels of TrkB and Akt were decreased in the model group(P<0.01). The results of immunochemistry showed that the expression of Bcl-2 in hippocampus was decreased(P<0.01), while the expression of Bax was increased(P<0.01). These decreases were reversed with the treatments of Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(1∶1) group and the Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(8∶3) group(P<0.01 or P<0.05). Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(1∶1) group was superior to the Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(3∶8) group in the effect of increasing the expression of BDNF, and superior to the other two groups in increasing the phosphorylation levels of TrkB and Akt. The expression of Bcl-2 was also increased and the expression of Bax was decreased in these two groups(P<0.01 or P<0.05). Conclusion The compatibility of Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix) can ameliorate depressive-like behavior, and the Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(8∶3) compatibility is superior to the Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix)(1∶1) compatibility. The couplet medicine of Chaihu(Bulerui Radix)-Huangqin(Scutellariae Radix) can regulate the BDNF and TrkB signaling pathway, protect neurons and ameliorate depressive-like behaviors.