Molecular identification and functional analysis of a caffeine receptor on the plasma membrane.

咖啡因具有多种药理作用，如通过兰尼定受体的兴奋和磷酸二酯酶的抑制。然而，最近有报道称，咖啡因作用于STC-1小鼠胃肠内分泌细胞后，细胞内钙离子迅速增加，并被磷脂酶C抑制。这提示了咖啡因的一种新的药理作用。我们观察到咖啡因诱导Fura-2负载的STC-1细胞钙离子浓度升高依赖于细胞外钙离子，并被Trp(瞬时受体电位)通道阻滞剂所抑制。此外，钙离子成像和电生理分析表明，在HEK293T细胞和非洲爪哇卵母细胞等异源表达系统中，小鼠TRPA1通道被毫米级浓度的咖啡因激活。这些对咖啡因的反应在野生型(WT)小鼠急性分离的背根节感觉神经元中得到证实，已知表达TRPA1，但在TRPA1基因敲除(KO)小鼠的神经元中未见。免疫组织化学方法检测TRPA1蛋白在小鼠舌内神经纤维和神经束中的表达。此外，在两瓶偏好测试中，WT小鼠，而不是KO小鼠，对含咖啡因的水表现出显著的厌恶。最后，我们观察到人的TRPA1通道的活性被咖啡因抑制，这与小鼠的通道形成了鲜明的对比。我们的发现表明，咖啡因激活了小鼠钙离子通透性的TRPA1通道，但抑制了人类同源基因的活性。我们还发现，在感觉神经元中表达的小鼠TRPA1通道会引起对饮用含咖啡因的水的厌恶，这表明它们介导了对咖啡因的感知。

Caffeine has various well-characterized pharmacological effects mediated e.g. via ryanodine receptor stimulation and phosphodiesterase inhibition. However, it was recently reported that application of caffeine to STC-1 mouse gastrointestinal endocrine cells induces a rapid increase in intracellular Ca^<2+> (Ca^<2+>; ) that is suppressed by inhibition of phospholipase C. This suggests a novel pharmacological action for caffeine. We observed that caffeine-induced increases in Ca^<2+>i in fura-2-loaded STC-1 cells were dependent on extracellular Ca^<2+> and were suppressed by TRP (Transient Receptor Potential) channel blockers. In addition, Ca^<2+>i imaging and electrophysiological analyses showed that mouse TRPA1 channels were activated by mM concentrations of caffeine in heterologous expression systems such as HEK293T cells and Xenopus oocytes. These responses to caffeine were confirmed in acutely dissociated dorsal root ganglion sensory neurons from wild-type (WT) mice, which are known to express TRPA1, but were not seen in neurons from TRPA1 knock-out (KO) mice. Expression of TRPA1 protein was detected immunohistochemically in nerve fibers and bundles in the mouse tongue. Moreover, WT mice, but not KO mice, showed a remarkable aversion to caffeine-containing water in two-bottle preference tests. Finally, we observed that the activity of human TRPA1 channels is suppressed by caffeine, in clear contrast to the mouse channels. Our findings demonstrated caffeine activates a Ca^<2+>-permeable TRPA1 channel in mouse but suppresses the activity of the human orthologue. We also showed that mouse TRPA1 channels expressed in sensory neurons cause an aversion to drinking caffeine-containing water, suggesting they mediate the perception of caffeine.