Gut–brain axis (GBA) communication relies on serotonin (5-HT) signaling between the gut epithelium and the peripheral nervous system, where 5-HT release patterns from the basolateral (i.e., bottom) side of the epithelium activate nerve afferents. There have been few quantitative studies of this gut-neuron signaling due to a lack of real-time measurement tools that can access the basolateral gut epithelium. In vitro platforms allow quantitative studies of cultured gut tissue, but they mainly employ offline and endpoint assays that cannot resolve dynamic molecular-release patterns. Here, we present the modification of a microporous cell culture membrane with carbon nanotube-coated gold (Au-CNT) electrodes capable of continuous, label-free, and direct detection of 5-HT at physiological concentrations. Electrochemical characterization of single-walled carbon nanotube (SWCNT)-coated Au electrodes shows increased electroactive surface area, 5-HT specificity, sensitivity, and saturation time, which are correlated with the CNT film drop-cast volume. Two microliters of CNT films, with a 10-min saturation time, 0.6 μA/μM 5-HT sensitivity, and reliable detection within a linear range of 500 nM–10 μM 5-HT, can be targeted for high-concentration, high-time-resolution 5-HT monitoring. CNT films (12.5 μL) with a 2-h saturation time, 4.5 μA/μM 5-HT sensitivity, and quantitative detection in the linear range of 100 nM–1 μM can target low concentrations with low time resolution. These electrodes achieved continuous detection of dynamic diffusion across the porous membrane, mimicking basolateral 5-HT release from cells, and detection of cell-released 5-HT from separately cultured RIN14B cell supernatant. Electrode-integrated cell culture systems such as this can improve in vitro molecular detection mechanisms and aid in quantitative GBA signaling studies.

肠-脑轴 (GBA) 通讯依赖于肠上皮和周围神经系统之间的血清素 (5-HT) 信号传导，其中 5-HT 从上皮的基底外侧（即底部）释放模式激活神经传入。由于缺乏可以进入基底外侧肠道上皮的实时测量工具，对这种肠道神经元信号传导的定量研究很少。体外平台允许对培养的肠道组织进行定量研究，但它们主要采用无法解析动态分子释放模式的离线和终点测定。在这里，我们提出了用碳纳米管涂层金 (Au-CNT) 电极对微孔细胞培养膜进行修饰，能够连续、无标记、直接检测生理浓度的 5-HT。单壁碳纳米管 (SWCNT) 涂层金电极的电化学表征显示电活性表面积、5-HT 特异性、灵敏度和饱和时间增加，这些与 CNT 薄膜滴铸体积相关。2 微升 CNT 薄膜，具有 10 分钟饱和时间、0.6 μA/μM 5-HT 灵敏度以及在 500 nM–10 μM 5-HT 线性范围内可靠检测，可针对高浓度、高浓度时间分辨率 5-HT 监测。CNT 膜 (12.5 μL) 具有 2 小时饱和时间、4.5 μA/μM 5-HT 灵敏度以及 100 nM–1 μM 线性范围内的定量检测，可以以低时间分辨率瞄准低浓度。这些电极实现了对穿过多孔膜的动态扩散的连续检测，模拟细胞的基底外侧 5-HT 释放，以及从单独培养的 RIN14B 细胞上清液中检测细胞释放的 5-HT。诸如此类的电极集成细胞培养系统可以改善体外分子检测机制并有助于定量 GBA 信号研究。