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Figure S1. Effects of a PKC inhibitor Go6976 and a ROCK inhibitor Y-27632 on [Ca2+]i elevation due to carbachol or high K+. (A–C) Representative results of [Ca2+]i change after stimulation with carbachol (1 μmol L−1) and high K+ (65.4 mmol L−1) in the presence or absence of Go6976 (1 μmol L−1) or Y-27632 (10 μmol L−1). (D) The area under [Ca2+]i-time curve (AUC) after stimulation was normalized against the AUC seen with 1st high K+. Results are expressed as means ± SEM of four experiments.

Figure S2. Effects of PKC inhibitors and ROCK inhibitors on phosphorylation of CPI-17 and MYPT1 due to carbachol. (A) Representative CPI-17 phosphorylation results after stimulation with carbachol (1 μmol L−1, 0.5 or 5 min) in the presence or absence of bisindolyl-maleimide I (Bis) (10 μmol L−1), Go6976 (1 μmol L−1), rottlerin (10 μmol L−1), Y-27632 (10 μmol L−1) and H-1152 (1 μmol L−1). (B) Representative MYPT1 phosphorylation results after stimulation with carbachol (1 μmol L−1, 0.5 or 5 min) in the presence or absence of bisindolyl-maleimide I (Bis) (10 μmol L−1), Y-27632 (10 μmol L−1) and H-1152 (1 μmol L−1). Each inhibitor was added 20 min before stimulation. (C, D, and E) CPI-17 phosphorylation at Thr38, MYPT1 phosphorylation at Thr696, and MYPT1 phosphorylation at Thr853 with or without inhibitors (bisindolyl-maleimide I or H-1152). Phosphorylation levels were quantified as described in the Methods section. Results are expressed as means ± SEM of 4–8 experiments. *,†< 0.05, **,††< 0.01, significantly different from phosphorylation stimulated by carbachol (*; 0.5 min, #; 5 min) without pretreatment of each inhibitor.

Figure S3. Effects of a PKC inhibitor bisindolyl-maleimide I and ROCK inhibitor H-1152 on resting MYPT1 phosphorylation. Effects of bisindolyl-maleimide I (Bis) (10 μmol L−1) and H-1152 (1 μmol L−1) on resting MYPT1 phosphorylation. Results are expressed as means ± SEM of four experiments. *< 0.05, **< 0.01, significantly different from resting phosphorylation.

Figure S4. Effects of neurotransmitter inhibitors on contraction due to high K+. Effects of atropine, L-NAME and suramin on contraction of ileal circular smooth muscle stimulated with high K+. Atropine (1 μmol L−1), L-NAME (300 μmol L−1) and suramin (100 μmol L−1) were added 20 min before carbachol stimulation. Force levels were normalized against the peak contraction seen with high K+. Results are expressed as means ± SEM of four experiments. *< 0.05, significantly different from phosphorylation stimulated by high K+ without pretreatment of each inhibitor. ns; not significantly different.

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NMO_1799_sm_FigS1.pdf362KSupporting info item
NMO_1799_sm_FigS2.pdf2026KSupporting info item
NMO_1799_sm_FigS3.pdf390KSupporting info item
NMO_1799_sm_FigS4.pdf294KSupporting info item

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