(732.10) SPARC: Development of a TRPA1 Reporter Mouse Model

Poster Board Number: E255

Parmvir Bahia (University of South Florida), Mayur Patil (University of South Florida), Seol-Hee Kim (University of South Florida), Jana Harsányiová (University of South Florida), Marian Kollarik (University of South Florida), Thomas Taylor-Clark (University of South Florida)

Transient receptor potential ankyrin 1 (TRPA1) is a cation channel activated by electrophilic irritants and oxidative stress. It is expressed in nociceptive sensory neurons and plays a role in pain and reflex signaling. Its expression in other cell types is controversial.

We generated a TRPA1-Flp knock-in mouse by incorporating a 2A-FlpO after the last endogenous TRPA1 exon. This TRPA1-Flp mouse was crossed with the Ai65f reporter strain which expresses the fluorescent protein tdTomato in a flp recombinase-dependent manner. From these animals, we collected tissue to visualize tdTomato, as an indicator of TRPA1 expression.

37% of vagal neurons expressed tdTomato. Of these 74% expressed TRPV1 (capsaicin receptor), and 88% expressed the neuropeptide CGRP. In fura 2AM studies of dissociated neurons, the TRPA1 agonist AITC activated gt;90% of tdTomato+ neurons and only 24% of tdTomato- neurons. tdTomato expression was limited in the dorsal root ganglion, with only 10% of neurons expressing tdTomato. Of these 61% expressed TRPV1, and 81% expressed CGRP. AITC activated 80% of tdTomato+ neurons and 26% of tdTomato- neurons.

We found tdTomato in fibers projecting to the nucleus of the solitary tract and to superficial lamina of the spinal cord dorsal horn. Very little tdTomato expression was found elsewhere in the CNS. In the periphery, we found tdTomato+ fibers innervating the lungs and GI tract. We also found tdTomato expression in some stomach and colonic cells, although none were enterochromaffin cells.

In summary, we have created a knock-in FlpO mouse model that can be used to identify TRPA1 expression.

This study is funded by National Institutes of Health Common Fund SPARC OT2 (2016-2019): "Functional mapping of peripheral and central circuits for airway protection and breathing"