Jeonnam Bioindustry Foundation, Jeonnam Institute of Natural Resources Research (JINR), 288, Woodland-gil, Anyang-myeon, Jangheung-gun, Jeollanamdo 59338, Republic of Korea.
Department of Biomedical Science, College of Natural Science, Chosun University, 309, pilmun-daero, Dong-gu, Gwangju 61452, Republic of Korea; Department of Integrative Biological Sciences & BK21 FOUR Educational Research Group for Age-associated Disorder Control Technology, Chosun University, 309, pilmun-daero, Dong-gu, Gwangju 61452, Republic of Korea.
Phytomedicine. 2023 Jul 25;116:154871. doi: 10.1016/j.phymed.2023.154871. Epub 2023 May 10.
Vaccinium bracteatum Thunb. leaves (VBL) are used in traditional herbal medicines to treat various biological diseases. p-coumaric acid (CA), the main active component of VBL, has neuroprotective effects against corticosterone-induced damage in vitro. However, the effects of CA on immobility induced by chronic restraint stress (CRS) in a mouse model and 5-HT receptor activity have not been investigated.
HYPOTHESIS/PURPOSE: We investigated the antagonistic effects of VBL, NET-D1602, and the three components of Gαs protein-coupled 5-HT receptors. Additionally, we identified the effects and mechanism of action of CA, the active component of NET-D1602, in the CRS-exposed model.
For in vitro analyses, we used 1321N1 cells stably expressing human 5-HT receptors and CHO-K1 expressing human 5-HT or 5-HT receptors cell lines to study the mechanism of action. For in vivo analyses, CRS-exposed mice were orally administered CA (10, 50, or 100 mg/kg) daily for 21 consecutive days. The effects of CA were analyzed by assessing behavioral changes using a forced swim test (FST), measuring levels of hypothalamic-pituitary-adrenal (HPA) axis-related hormones in ntial therapeutic effects as 5-HT6 receptor antagonists for neurodegenerative diseases and depressioserum, and acetylcholinesterase (AChE), monoamines, including 5-HT, dopamine, and norepinephrine, using enzyme-linked immunosorbent assay kits. The underlying molecular mechanisms of the serotonin transporter (SERT), monoamine oxidase A (MAO-A), and extracellular signal-regulated kinase (ERK)/protein kinase B (Akt)/mTORC1 signaling were detected using western blotting.
CA was confirmed to be an active component in the antagonistic effects of NET-D1602 on 5-HT receptor activity through decreases in cAMP and ERK1/2 phosphorylation. Moreover, CRS-exposed mice treated with CA showed a significantly reduced immobility time in the FST. CA also significantly decreased corticosterone, corticotropin-releasing hormone (CRH), and adrenocorticotropic hormone (ACTH) levels. CA enhanced 5-HT, dopamine, and norepinephrine levels in the hippocampus (HC) and prefrontal cortex (PFC) but decreased MAO-A and SERT protein levels. Similarly, CA significantly upregulated the ERK, Ca/calmodulin-dependent protein kinase II (CaMKII), Akt/mTOR/p70S6K/S6 signaling pathways in both HC and the PFC.
CA contained in NET-D1602 may play the antidepressant effects against CRS-induced depression-like mechanism and the selective antagonist effect of 5-HT receptor.
蔓越莓叶（VBL）被用于传统草药医学中，以治疗各种生物疾病。对香豆酸（CA）是 VBL 的主要活性成分，具有体外对抗皮质酮诱导损伤的神经保护作用。然而，CA 对慢性束缚应激（CRS）诱导的小鼠模型中不动性的影响以及 5-HT 受体活性尚未得到研究。
假设/目的：我们研究了 VBL、NET-D1602 和 Gαs 蛋白偶联 5-HT 受体的三个组成部分的拮抗作用。此外，我们确定了 CA（NET-D1602 的活性成分）在 CRS 暴露模型中的作用机制和作用。
对于体外分析，我们使用稳定表达人 5-HT 受体的 1321N1 细胞和表达人 5-HT 或 5-HT 受体的 CHO-K1 细胞系来研究作用机制。对于体内分析，CRS 暴露的小鼠每日口服给予 CA（10、50 或 100mg/kg）连续 21 天。通过强迫游泳试验（FST）评估行为变化来分析 CA 的作用，测量下丘脑-垂体-肾上腺（HPA）轴相关激素在血清中的水平，使用酶联免疫吸附试验试剂盒测量乙酰胆碱酯酶（AChE）、单胺类物质，包括 5-HT、多巴胺和去甲肾上腺素。使用 Western blot 检测 5-HT 转运体（SERT）、单胺氧化酶 A（MAO-A）和细胞外信号调节激酶（ERK）/蛋白激酶 B（Akt）/哺乳动物雷帕霉素靶蛋白（mTORC1）信号转导的下游分子机制。
通过降低 cAMP 和 ERK1/2 磷酸化，CA 被证实是 NET-D1602 对 5-HT 受体活性的拮抗作用中的一种活性成分。此外，CA 处理的 CRS 暴露小鼠在 FST 中的不动时间明显减少。CA 还显著降低了皮质酮、促肾上腺皮质激素释放激素（CRH）和促肾上腺皮质激素（ACTH）水平。CA 增加了海马（HC）和前额叶皮层（PFC）中的 5-HT、多巴胺和去甲肾上腺素水平，但降低了 MAO-A 和 SERT 蛋白水平。同样，CA 显著上调了 HC 和 PFC 中 ERK、钙/钙调蛋白依赖性蛋白激酶 II（CaMKII）、Akt/mTOR/p70S6K/S6 信号通路。
NET-D1602 中含有的 CA 可能发挥抗 CRS 诱导的抑郁样机制和 5-HT 受体选择性拮抗作用的抗抑郁作用。
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