Pushing the sensing capacity for ultrasmall biological entities could open a window to the vast mysterious world of life. Take brain analysis, for example. Underneath neurons, the fundamental unit in the central nervous system (CNS), there exist vesicles (∼50–200 nm in diameter)─the smallest cell organelles responsible for the highest mental functions─encoding the pivotal information on neurotransmitter storage that bridges its synthesis and release. Therefore, identifying an approach to count neurotransmitter molecules in a single vesicle would give us an understanding of the chemical basis of either normal or abnormal processes of the brain. Traditional approaches for vesicle content analysis utilize ultracentrifugation, lysis, and HPLC-electrochemical (EC) or HPLC-UV–vis analysis, ignoring the vesicle-to-vesicle variation. Benefiting from high sensitivity and excellent temporal resolution, electrochemical sensing strategies show great potential for single vesicle analysis. (1) By using an amperometric sensing method, the catecholamine content in a single isolated vesicle can be quantified by an undefined mechanism, (2) which may involve the electroporation of the vesicle’s bilipid layer on an electrode biased at a proper potential. (3) Research has shown that the neurotransmitter content in each isolated vesicle varies in a wide range. Considering that vesicle damage and leakage during the isolation process may bias its content determination; sophisticated sensing tools for counting neurotransmitters directly in single intracellular vesicles are highly desired. Electrochemical sensing can accomplish this task with conical nanotip electrodes that can penetrate the cell membrane with little damage and analyze vesicles in situ. (4) Intracellular vesicular neurotransmitter storage sensing by nanoamperometry has proven successful on cultured neuronal cells, promoting the identification of both endogenous and exogenous factors that affect vesicular neurotransmitter storage. While intracellular single-vesicle sensing has been increasingly used to measure electroactive neurotransmitters, mostly catecholamine, it remains a challenge to sense electrochemically inactive neurotransmitters, such as acetylcholine, γ-amino butyric acid, and neuropeptides in single vesicles. (5) Therefore, more research is needed to develop sensing systems that can detect various neurotransmitters in a single vesicle in the CNS. Meanwhile, there are many cell organelles that have similar structures to vesicles, such as exosomes, phagolysosomes, and lysosomes. Developing sensing methods to qualify and quantify the contents in these cell organelles would represent another interesting and important field in life science study. Further, applying intracellular single vesicle sensing strategies to identify these structures would offer a new avenue to cell chemistry discovery. One approach for developing this type of method could begin by transplanting the nanotip electrode from cultured cells to the intact living brain, affording a brain slice with a few layers of neuronal cells to mimic the real CNS environment. Not surprisingly, investigating the vesicular neurotransmitter storage in vivo would be the ultimate goal of intracellular single vesicle sensing. Coupling behavioral tests and drug treatment, in vivo intracellular single vesicle sensing could help understand the physiology and pathology of neuronal functions and drug discovery for neuronal disorders as well. Intracellular single vesicle sensing constitutes an effective methodology for analyzing small amounts of a target in an extremely small vesicle, which has initiated an era of subcellular entity analysis. It is without a doubt that electrochemical sensing will have an important role in this area with its future direction foreseeable: from electrochemically inactive neurotransmitter counting in single vesicles to a comprehensive exploration of cell organelle chemistry in the brain. This article references 5 other publications. This article has not yet been cited by other publications. This article references 5 other publications.

中文翻译：

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在单个囊泡内瞥见大脑

推动超小型生物实体的感知能力可以为广阔的神秘生命世界打开一扇窗户。以大脑分析为例。在中枢神经系统 (CNS) 的基本单位神经元下方，存在囊泡（直径约 50-200 nm）——负责最高心理功能的最小细胞器——编码神经递质储存的关键信息，连接其合成并释放。因此，确定一种计算单个囊泡中神经递质分子的方法将使我们了解大脑正常或异常过程的化学基础。囊泡含量分析的传统方法利用超速离心、裂解和 HPLC-电化学 (EC) 或 HPLC-UV-vis 分析，忽略了囊泡间的差异。得益于高灵敏度和出色的时间分辨率，电化学传感策略显示出单囊泡分析的巨大潜力。(1) 通过使用电流传感方法，单个分离囊泡中的儿茶酚胺含量可以通过未定义的机制进行量化，(2) 这可能涉及在偏压在适当电位的电极上对囊泡的双脂层进行电穿孔。(3) 研究表明，每个孤立囊泡中的神经递质含量变化很大。考虑到分离过程中囊泡损伤和渗漏可能会影响其含量测定；非常需要用于直接在单个细胞内囊泡中计数神经递质的复杂传感工具。原位. (4) 纳米电流法的细胞内囊泡神经递质储存传感已被证明在培养的神经元细胞上是成功的，促进了影响囊泡神经递质储存的内源性和外源性因素的识别。虽然细胞内单囊泡传感已越来越多地用于测量电活性神经递质，主要是儿茶酚胺，但在单囊泡中感知电化学非活性神经递质（如乙酰胆碱、γ-氨基丁酸和神经肽）仍然是一个挑战。(5) 因此，需要更多的研究来开发能够检测中枢神经系统单个囊泡中各种神经递质的传感系统。同时，有许多细胞器具有与囊泡相似的结构，例如外泌体、吞噬溶酶体和溶酶体。开发传感方法来鉴定和量化这些细胞器中的内容将代表生命科学研究中另一个有趣且重要的领域。此外，应用细胞内单囊泡传感策略来识别这些结构将为细胞化学发现提供新途径。开发这种方法的一种方法可以首先将纳米尖端电极从培养的细胞移植到完整的活脑中，提供具有几层神经元细胞的脑切片来模拟真实的中枢神经系统环境。毫不奇怪，研究水泡神经递质的储存 应用细胞内单囊泡传感策略来识别这些结构将为细胞化学发现提供新途径。开发这种方法的一种方法可以首先将纳米尖端电极从培养的细胞移植到完整的活脑中，提供具有几层神经元细胞的脑切片来模拟真实的中枢神经系统环境。毫不奇怪，研究水泡神经递质的储存 应用细胞内单囊泡传感策略来识别这些结构将为细胞化学发现提供新途径。开发这种方法的一种方法可以首先将纳米尖端电极从培养的细胞移植到完整的活脑中，提供具有几层神经元细胞的脑切片来模拟真实的中枢神经系统环境。毫不奇怪，研究水泡神经递质的储存体内将是细胞内单囊泡传感的最终目标。体内耦合行为测试和药物治疗细胞内单囊泡传感可以帮助了解神经元功能的生理学和病理学以及神经元疾病的药物发现。细胞内单囊泡传感构成了一种分析极小囊泡中少量目标的有效方法，开启了亚细胞实体分析的时代。毫无疑问，电化学传感将在这一领域发挥重要作用，其未来方向可预见：从单个囊泡中的电化学非活性神经递质计数到对大脑细胞器化学的全面探索。本文引用了其他 5 篇出版物。这篇文章尚未被其他出版物引用。本文引用了其他 5 篇出版物。