Wang Jing, Ni Xin-Qiang, Li Li-Min, Liao Ying-Zhao, Chen Yao, Huang Chun-Ling
Shenzhen Children's Hospital Shenzhen 518038, China.
the Fourth Clinical Medical College of Guangzhou University of Chinese Medicine Shenzhen 518035, China.
Zhongguo Zhong Yao Za Zhi. 2022 Jul;47(13):3554-3561. doi: 10.19540/j.cnki.cjcmm.20220412.402.
The present study investigated the effect of Rehmanniae Radix Praeparata(RRP) on the energy metabolism of prefrontal cortex(PFC) of spontaneously hypertensive rats with attention deficit hyperactivity disorder(ADHD) based on the "static Yin and dynamic Yang" theory.Thirty spontaneously hypertensive male rats aged 3 weeks were randomly divided into a model group, a methylphenidate(MPH) group(2 mg·kg(-1)), and an RRP group(2.4 g·kg(-1)).Wistar-Kyoto(WKY) male rats of the same age were assigned to the normal group.Rats were treated with corresponding drugs twice per day, and those in the model group and the normal group received the same volume of 0.9% sodium carboxymethyl cellulose(CMC-Na) solution by gavage.The open-field test was performed to evaluate the spontaneous and impulsive behaviors of rats before treatment and on the 4(th) week after treatment.Four weeks after treatment, PFC was isolated and mitochondria were prepared.The content of adenosine triphosphate(ATP), adenosine diphosphate(ADP), and adenosine monophosphate(AMP) in the PFC was determined by high-performance liquid chromatography(HPLC), and energy charge(EC) was calculated.The parameters related to mitochondrial respiratory function were measured by the Clark oxygen electrode, specifically, state 3 respiration(ST3), state 4 respiration(ST4), and respiratory control rate(RCR).Enzymatic activities of succinate dehydrogenase(SDH), cytochrome C oxidase(COX), Na+-K+-ATPase, and Ca(2+)-Mg(2+)-ATPase were measured by chemical colorimetry.Mitochondrial permeability transition pore(mPTP) opening was measured by spectrophotometry.Protein expression of glucose transporter 1(GLUT1) and GLUT3 in PFC was tested by Western blot.Compared with the results in the model group, RRP could significantly reduce the total distance of movement, vertical times, and distance in the central area in the open field test(P<0.05 or P<0.01), increase the content of ATP and EC, decrease the content of AMP(P<0.05), elevate ST3 and RCR(P<0.05), potentiate activities of SDH, COX, Na+-K+-ATPase, and Ca(2+)-Mg(2+)-ATPase(P<0.05 or P<0.01), inhibit the opening of mPTP, and increase the expression levels of GLUT1 and GLUT3 proteins(P<0.05).It was inferred that RRP could inhibit hyperacti-vity and impulsivity by improving the energy metabolism disorder in PFC of ADHD rats, and its mechanism may be related to the improvement of mitochondrial respiratory function, potentiation of Na+-K+-ATPase, Ca(2+)-Mg~(2+)-ATPase, and mitochondrial respiratory enzymes, inhibition of the opening of mPTP, and up-regulation of the expression of glucose transporter proteins.This study initially reveals the biological connotation of the "static Yin and dynamic Yang" theory in ADHD.
本研究基于“阴静阳动”理论，探讨了熟地黄对注意力缺陷多动障碍（ADHD）自发性高血压大鼠前额叶皮质（PFC）能量代谢的影响。将30只3周龄的自发性高血压雄性大鼠随机分为模型组、哌甲酯（MPH）组（2 mg·kg⁻¹）和熟地黄组（2.4 g·kg⁻¹）。将同龄的Wistar-Kyoto（WKY）雄性大鼠作为正常组。大鼠每天接受相应药物治疗两次，模型组和正常组大鼠通过灌胃给予相同体积的0.9%羧甲基纤维素钠（CMC-Na）溶液。在治疗前及治疗后第4周进行旷场试验，以评估大鼠的自发和冲动行为。治疗4周后，分离PFC并制备线粒体。采用高效液相色谱法（HPLC）测定PFC中三磷酸腺苷（ATP）、二磷酸腺苷（ADP）和一磷酸腺苷（AMP）的含量，并计算能量电荷（EC）。用Clark氧电极测定与线粒体呼吸功能相关的参数，即状态3呼吸（ST3）、状态4呼吸（ST4）和呼吸控制率（RCR）。采用化学比色法测定琥珀酸脱氢酶（SDH）、细胞色素C氧化酶（COX）、Na⁺-K⁺-ATP酶和Ca²⁺-Mg²⁺-ATP酶的酶活性。用分光光度法测定线粒体通透性转换孔（mPTP）的开放情况。通过蛋白质免疫印迹法检测PFC中葡萄糖转运蛋白1（GLUT1）和GLUT3的蛋白表达。与模型组结果相比，熟地黄可显著减少旷场试验中的总运动距离、垂直次数和中央区域距离（P<0.05或P<0.01），增加ATP含量和EC，降低AMP含量（P<0.05），提高ST3和RCR（P<0.05），增强SDH、COX、Na⁺-K⁺-ATP酶和Ca²⁺-Mg²⁺-ATP酶的活性（P<0.05或P<0.01），抑制mPTP的开放，并增加GLUT1和GLUT3蛋白的表达水平（P<0.05）。推测熟地黄可能通过改善ADHD大鼠PFC的能量代谢紊乱来抑制多动和冲动，其机制可能与改善线粒体呼吸功能、增强Na⁺-K⁺-ATP酶、Ca²⁺-Mg²⁺-ATP酶和线粒体呼吸酶活性、抑制mPTP开放以及上调葡萄糖转运蛋白表达有关。本研究初步揭示了“阴静阳动”理论在ADHD中的生物学内涵。
