Optogenetic Activation of Colon Epithelium of the Mouse Produces High-Frequency Bursting in Extrinsic Colon Afferents and Engages Visceromotor Responses
Optogenetic Activation of Colon Epithelium of the Mouse Produces High-Frequency Bursting in Extrinsic Colon Afferents and Engages Visceromotor Responses
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DOI：
10.1523/jneurosci.0837-18.2018
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发表时间：
2018-06-20
影响因子：
5.3
通讯作者：
Davis, Brian M.
中科院分区：
文献类型：
--
作者：
Makadia, Payal A.;Najjar, Sarah A.;Davis, Brian M.
结肠的上皮细胞提供了内部环境（结肠腔）和结肠实质之间的重要界面。为了研究在这个接口的上皮神经元信号，我们分析了小鼠，其中通道视紫红质（ChR 2）是针对TRPV 1阳性传入或绒毛表达结肠上皮细胞。通过将Ai 32小鼠分别与TRPV 1-Cre或villin-Cre小鼠杂交，将ChR 2-EYFP融合蛋白的表达导向初级感觉神经元或结肠上皮细胞。结肠的离体制备用于盆神经的结肠感觉传入的单纤维分析。使用先前描述的标准，如粘膜，肌肉，肌肉-粘膜，或serenum的传入，然后测试蓝光诱导的激活。结肠上皮细胞的光激活产生了强烈的动作电位放电，类似于生理刺激引起的（例如，周向拉伸），在ChR 2表达纯合子小鼠的50.5%的结肠传入中。光诱导的活动可以减少或取消在大多数纤维使用的鸡尾酒嘌呤受体阻滞剂表明ATP释放上皮细胞产生的感觉神经元动作电位。使用内脏反应的肌电图记录，我们发现，光刺激的结肠上皮细胞诱发的行为反应，在Vil-ChR 2小鼠，这是类似于所看到的气球扩张的结肠。这些离体和体内数据表明，光刺激结肠上皮细胞单独，而不添加机械或化学刺激，可以直接激活结肠传入神经，并引起行为反应。
Epithelial cells of the colon provide a vital interface between the internal environment (lumen of the colon) and colon parenchyma. To examine epithelial-neuronal signaling at this interface, we analyzed mice in which channelrhodopsin (ChR2) was targeted to either TRPV1-positive afferents or to villin-expressing colon epithelial cells. Expression of a ChR2-EYFP fusion protein was directed to either primary sensory neurons or to colon epithelial cells by crossing Ai32 mice with TRPV1-Cre or villin-Cre mice, respectively. An ex vivo preparation of the colon was used for single-fiber analysis of colon sensory afferents of the pelvic nerve. Afferents were characterized using previously described criteria as mucosal, muscular, muscular-mucosal, or serosal and then tested for blue light-induced activation. Light activation of colon epithelial cells produced robust firing of action potentials, similar to that elicited by physiologic stimulation (e.g., circumferential stretch), in 50.5% of colon afferents of mice homozygous for ChR2 expression. Light-induced activity could be reduced or abolished in most fibers using a cocktail of purinergic receptor blockers suggesting ATP release by the epithelium contributed to generation of sensory neuron action potentials. Using electromyographic recording of visceromotor responses we found that light stimulation of the colon epithelium evoked behavioral responses in Vil-ChR2 mice that was similar to that seen with balloon distension of the colon. These ex vivo and in vivo data indicate that light stimulation of colon epithelial cells alone, without added mechanical or chemical stimuli, can directly activate colon afferents and elicit behavioral responses.