Skaggs School of Pharmacy and Pharmaceutical Sciences , University of California, San Diego , La Jolla , California 92093 , United States.
Department of Neurosciences and Department of Pharmacology, School of Medicine , University of California, San Diego , La Jolla , California 92093 , United States.
ACS Chem Neurosci. 2019 Mar 20;10(3):1369-1379. doi: 10.1021/acschemneuro.8b00432. Epub 2019 Jan 23.
The classical small molecule neurotransmitters are essential for cell-cell signaling in the nervous system for regulation of behaviors and physiological functions. Metabolomics approaches are ideal for quantitative analyses of neurotransmitter profiles but have not yet been achieved for the repertoire of 14 classical neurotransmitters. Therefore, this study developed targeted metabolomics analyses by full scan gas chromatography/time-of-flight mass spectrometry (GC-TOF) and hydrophilic interaction chromatography-QTRAP mass spectrometry (HILIC-MS/MS) operated in positive ionization mode for identification and quantitation of 14 neurotransmitters consisting of acetylcholine, adenosine, anandamide, aspartate, dopamine, epinephrine, GABA, glutamate, glycine, histamine, melatonin, norepinephrine, serine, and serotonin. GC-TOF represents a new metabolomics method for neurotransmitter analyses. Sensitive measurements of 11 neurotransmitters were achieved by GC-TOF, and three neurotransmitters were analyzed by LC-MS/MS (acetylcholine, anandamide, and melatonin). The limits of detection (LOD) and limits of quantitation (LOQ) were assessed for linearity for GC-TOF and LC-MS/MS protocols. In neurotransmitter-containing dense core secretory vesicles of adrenal medulla, known as chromaffin granules (CG), metabolomics measured the concentrations of 9 neurotransmitters consisting of the catecholamines dopamine, norepinephrine, and epinephrine, combined with glutamate, serotonin, adenosine, aspartate, glycine, and serine. The CG neurotransmitters were constitutively secreted from sympathoadrenal chromaffin cells in culture. Nicotine- and KCl-stimulated release of the catecholamines and adenosine. Lithium, a drug used for the treatment of bipolar disorder, decreased the constitutive secretion of dopamine and norepinephrine and decreased nicotine-stimulated secretion of epinephrine. Lithium had no effect on other secreted neurotransmitters. Overall, the newly developed GC-TOF with LC-MS/MS metabolomics methods for analyses of 14 neurotransmitters will benefit investigations of neurotransmitter regulation in biological systems and in human disease conditions related to drug treatments.
经典的小分子神经递质对于神经系统中的细胞间信号传递对于调节行为和生理功能至关重要。代谢组学方法是定量分析神经递质谱的理想方法，但尚未实现 14 种经典神经递质的全部神经递质谱。因此，本研究通过全扫描气相色谱/飞行时间质谱（GC-TOF）和亲水相互作用色谱-三重四极杆质谱（HILIC-MS/MS）在正离子模式下开发了靶向代谢组学分析，用于鉴定和定量分析由乙酰胆碱、腺苷、花生四烯酸、天冬氨酸、多巴胺、肾上腺素、γ-氨基丁酸、谷氨酸、甘氨酸、组胺、褪黑素、去甲肾上腺素、丝氨酸和 5-羟色胺组成的 14 种神经递质。GC-TOF 代表了一种新的神经递质分析代谢组学方法。通过 GC-TOF 实现了 11 种神经递质的敏感测量，通过 LC-MS/MS 分析了 3 种神经递质（乙酰胆碱、花生四烯酸和褪黑素）。评估了 GC-TOF 和 LC-MS/MS 方案的线性检测限（LOD）和定量限（LOQ）。在肾上腺髓质的称为嗜铬粒（CG）的致密核心分泌小泡中，代谢组学测量了由儿茶酚胺多巴胺、去甲肾上腺素和肾上腺素组成的 9 种神经递质的浓度，结合谷氨酸、5-羟色胺、腺苷、天冬氨酸、甘氨酸和丝氨酸。CG 神经递质由培养中的交感肾上腺嗜铬细胞组成性分泌。尼古丁和 KCl 刺激儿茶酚胺和腺苷的释放。用于治疗双相情感障碍的药物锂可减少多巴胺和去甲肾上腺素的组成性分泌，并减少尼古丁刺激的肾上腺素分泌。锂对其他分泌的神经递质没有影响。总之，新开发的 GC-TOF 与 LC-MS/MS 代谢组学方法用于分析 14 种神经递质将有助于研究生物系统中的神经递质调节以及与药物治疗相关的人类疾病状况中的神经递质调节。
