Effects of A213 on production of muscarinic acetylcholine receptor (M3R)‐specific cytokines in vitro. (a,b) Splenocytes from immunized M3R–/– mice obtained on day 20 after the first immunization were cultured without and with different concentrations of A213 [(a) 5 μM, (b) 0·04, 0·2, 1 or 5 μM] under stimulation with M3R peptide mixture (5 μg/ml each). After 72 h of culture, interferon (IFN)‐γ and interferon (IL)‐17 levels in culture supernatants were measured by enzyme‐linked immunosorbent assay (ELISA). Data are mean ± standard error of the mean (s.e.m.) of three mice per group (representative data of two independent experiments). *P < 0·05 [(a) Student's t‐test, (b) Kruskal–Wallis test)]. (c) Effects of A213 at different doses on co‐culture assay using CD4+ T cells and mitomycin C‐treated CD11c+ cells (at a ratio of 5 : 1) obtained from immunized M3R–/– mice and stimulated with M3R peptides. After 72 h of co‐culture, IFN‐γ and IL‐17 levels in culture supernatants were analysed by enzyme‐linked immunosorbent assay (ELISA). Data are mean ± s.e.m. of three mice per group (representative data of two independent experiments). *P < 0·05 (Kruskal–Wallis test).