rats: For oral administration, A-784168 in 10% dehydrated ethanol, 20% polysorbate 80, and 70% polyethylene glycol 400 (PEG) was administered 1 h before testing using a volume of 2 ml/kg. For intrathecal administration, compound was dissolved in 10% di-methyl-sulfoxide/90% hydroxyl-β-cyclodextrin, injected in a volume of 10 μl, and followed by a 10 μl saline flush

A-784168 was identified as TRPV1 antagonists in a capsaicin-induced calcium flux assay using a fluorometric imaging plate reader. Concentration-effect curves for capsaicin-induced, increased intracellular Ca2+ in the presence of increasing concentrations of A-784168 (0-200 nM). A-784168 potently and competitively inhibited capsaicin-induced calcium flux into 1321N1 astrocytoma cells expressing recombinant human TRPV1. The IC50 value for inhibiting TRPV1 activation by 50 nM capsaicin was 25.0 nM. A-784168 inhibited pH 5.5-induced activation of recombinant TRPV1 expressed in HEK293 cells with similar potency IC50 = 14.0 ± 2.3 as well as activation by 3 μM NADA (IC50 = 33.7 ± 9.6) and 10 μM anandamide (IC50 = 35.1 ± 10.6). It also blocked 1 μM capsaicin-induced currents in rat dorsal root ganglion neurons using electrophysiologic methods (IC50 = 10 nM)