Neuroscience Research, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, IL 60064, USA.
Eur J Pharmacol. 2010 May 10;633(1-3):15-23. doi: 10.1016/j.ejphar.2010.02.004. Epub 2010 Feb 11.
A-778317 (1-((R)-5-tert-butyl-indan-1-yl)-3-isoquinolin-5-yl-urea) is a potent antagonist of human and rat transient receptor potential vanilloid-1 (TRPV1) receptors. We have previously reported that [(3)H]A-778317 is an excellent radioligand to study the recombinant human TRPV1 receptor in a heterologous expression system. These studies were extended to determine the feasibility of using [(3)H]A-778317 to label native TRPV1 channels in rat tissues. Saturable high-affinity binding of [(3)H]A-778317 was detected in membrane preparations of rat dorsal root ganglia (DRG) and spinal cord that was inhibited by TRPV1 receptor agonists and antagonists. [(3)H]A-778317 labeled a single class of high-affinity binding sites in both rat DRG and spinal cord membranes (K(D)=10 and 8.4nM, respectively). The number of binding sites was 10-fold higher in rat DRG membranes than spinal cord membranes (B(max)=3.3 and 0.35pmol/mg protein, respectively). The pharmacology of the high-affinity binding sites was similar in rat DRG and spinal cord, but differed from the recombinant rat TRPV1 (rTRPV1) receptor expressed in transiently transfected HEK293-F cells. In particular, a large disparity in potency (>300-fold) was observed for the TRPV1 receptor agonist resiniferatoxin between native and recombinant rTRPV1 receptors. Our data indicate that the binding of [(3)H]A-778317 to native rTRPV1 channels is pharmacologically distinct, and perhaps more complex, than its binding to the recombinant rTRPV1 receptor.
A-778317（1-((R)-5-叔丁基-1-茚满基)-3-异喹啉-5-基-脲）是一种有效的人源和大鼠瞬时受体电位香草酸 1（TRPV1）受体拮抗剂。我们之前曾报道过，[(3)H]A-778317 是一种研究重组人 TRPV1 受体在异源表达系统中活性的优异放射性配体。这些研究进一步扩展到确定使用 [(3)H]A-778317 标记大鼠组织中天然 TRPV1 通道的可行性。在大鼠背根神经节（DRG）和脊髓的膜制剂中检测到 [(3)H]A-778317 的可饱和高亲和力结合，该结合可被 TRPV1 受体激动剂和拮抗剂抑制。[(3)H]A-778317 在大鼠 DRG 和脊髓膜中标记了单一类高亲和力结合位点（分别为 K(D)=10 和 8.4nM）。在大鼠 DRG 膜中的结合位点数量比脊髓膜高 10 倍（B(max)=3.3 和 0.35pmol/mg 蛋白，分别）。高亲和力结合位点的药理学在大鼠 DRG 和脊髓中相似，但与瞬时转染 HEK293-F 细胞中表达的重组大鼠 TRPV1（rTRPV1）受体不同。特别是，在天然和重组 rTRPV1 受体之间，TRPV1 受体激动剂树脂毒素的效力（>300 倍）存在很大差异。我们的数据表明，[(3)H]A-778317 与天然 rTRPV1 通道的结合在药理学上与与重组 rTRPV1 受体的结合不同，而且可能更复杂。
