Experimental strategy and transgene expression. A, Strategy for IRNA. The Tg mice expressing IR84a/IR8a genes in specific cell types are treated with exogenous ligands in the brain regions, resulting in the activation of target neurons. B, Structure of the gene cassette encoding GFP-IR84a-2A-IR8a with a signal peptide (SP) downstream of the TH gene promoter. B, BamHI, E, EcoRI, H, HindIII, S, SalI; pA, polyadenylation signal. C, Transgene expression in the LC revealed by GFP immunohistochemistry. 4V, Fourth ventricle. D, Confocal microscopic images of LC sections obtained from double immunohistochemistry for TH/GFP and NET/IR8a. Scale bars: C, 500 μm; D, 20 μm.