High-resolution functional labeling of vertebrate and invertebrate olfactory receptor neurons using agmatine, a channel-permeant cation.
High-resolution functional labeling of vertebrate and invertebrate olfactory receptor neurons using agmatine, a channel-permeant cation.
复制标题
使用胍丁胺（一种通道渗透性阳离子）对脊椎动物和无脊椎动物嗅觉受体神经元进行高分辨率功能标记。
DOI：
10.1016/s0165-0270(99)00077-1
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发表时间：
1999
影响因子：
3
通讯作者：
Derby,CD
中科院分区：
文献类型：
--
作者：
Michel,WC;Steullet,P;Cate,HS;Burns,CJ;Zhainazarov,AB;Derby,CD
描述了淡水斑马鱼斑马鱼和海洋龙虾的嗅觉受体神经元 (ORN) 的气味刺激标记方法。阳离子分子 1-氨基-4-胍基丁烷（=胍基丁胺，AGB）在气味刺激后通过离子通道渗透，并通过抗 AGB 抗体进行检测，从而可以标记气味刺激的 ORN。调整参数以优化活性依赖性标记，包括标记介质离子组成、刺激时间和 AGB 浓度。对于龙虾，在最佳条件下，7% 的 ORN 被复杂的气味、牡蛎混合物标记，即在钠和钙浓度降低的人造海水中用 20 mM AGB 刺激 5 秒，持续 60 分钟。 AGB 对龙虾来说是一种弱气味剂；在没有气味的情况下，用 AGB 刺激时，它仅引起 ORN 的轻微电生理反应，并标记 <1% 的 ORN。对于斑马鱼，用鱼林格氏溶液中的 5 mM AGB 加气味剂（L-谷氨酰胺）刺激 10 秒/分钟，持续 10 分钟是最佳标记条件，导致 17% 的嗅觉上皮区域被标记。大约 6% 的嗅觉上皮在仅使用 AGB 的对照刺激物的刺激过程中被标记。仅 AGB 的标记表明它对斑马鱼具有嗅觉刺激作用；电生理记录支持的结论。我们使用电生理测定和通道阻滞剂来检查每个物种的 AGB 进入 ORN 的潜在离子通道。这些结果表明，AGB 可用作生活在各种环境条件下的不同门的化学感受器神经元的活性依赖性标签。
Methods are described for odor-stimulated labeling of olfactory receptor neurons (ORNs) of the freshwater zebrafish Danio rerio and the marine spiny lobster Panulirus argus. Permeation of a cationic molecule, 1-amino-4-guanidobutane (=agmatine, AGB), through ion channels following odor stimulation, and its detection by an anti-AGB antibody, allow labeling of odor-stimulated ORNs. Parameters adjusted to optimize activity-dependent labeling included labeling medium ionic composition, stimulation times, and AGB concentration. For lobsters, 7% of ORNs were labeled by a complex odor, oyster mixture, under optimal conditions, which was stimulation for 5 s per min for 60 min with 20 mM AGB in artificial seawater with reduced sodium and calcium concentrations. AGB was a weak odorant for lobsters; it elicited only a small electrophysiological response from ORNs and labeled<1% of the ORNs during stimulation with AGB in the absence of odors. For the zebrafish, stimulation for 10 s per min for 10 min with 5 mM AGB plus odorant (l-glutamine) in fish Ringer’s solution was the optimal labeling condition, resulting in labeling of 17% of the olfactory epithelial area. Approximately 6% of the olfactory epithelium was labeled during stimulation with a control stimulus, AGB alone. This labeling by AGB alone suggests it is an olfactory stimulus for zebrafish; a conclusion supported by electrophysiological recordings. We used electrophysiological assays and channel blockers to examine, for each species, potential ion channels for entry of AGB into ORNs. These results show that AGB can be used as an activity-dependent label for chemoreceptor neurons of diverse phyla living in a range of environmental conditions.