Endogenous TRPV4 Expression of a Hybrid Neuronal Cell Line N18D3 and Its Utilization to Find a Novel Synthetic Ligand

Sungjae Yoo, Seung-In Choi, Seul Lee, Jiho Song, Chungmi Yang, Sangsu Bang, Seung Up Kim, Kyung Hoon Min, Sun Wook Hwang

Primary sensory afferent neurons detect environmental and painful stimuli at their peripheral termini. A group of transient receptor potential ion channels (TRPs) are expressed in these neurons and constitute sensor molecules for the stimuli such as thermal, mechanical, and chemical insults. We examined whether a mouse sensory neuronal line, N18D3, shows the sensory TRP expressions and their functionality. In Ca(2+) imaging and electrophysiology with these cells, putative TRPV4-mediated responses were observed. TRPV4-specific sensory modalities including sensitivity to a specific agonist, hypotonicity, or an elevated temperature were reproduced in N18D3 cells. Electrophysiological and pharmacological profiles conformed to those from native TRPV4 of primarily cultured neurons. The TRPV4 expression in N18D3 was also confirmed by RT-PCR and Western blot analyses. Thus, N18D3 cells may represent TRPV4-expressing sensory neurons. Further, using this cell lines, we discovered a novel synthetic TRPV4-specific agonist, MLV-0901. These results suggest that N18D3 is a reliable cell line for functional and pharmacological TRPV4 assays. The chemical information from the novel agonist will contribute to TRPV4-targeting drug design.