Protease-activated receptors mediate apamin-sensitive relaxation of mouse and guinea pig gastrointestinal smooth muscle  Thomas M. Cocks, Vitina Sozzi, James D. Moffatt, Stavros Selemidis  Gastroenterology  Volume 116, Issue 3, Pages 586-592 (March 1999) DOI: 10.1016/S0016-5085(99)70180-0 Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 1 Digitized original traces showing responses to a single concentration (↓) of the serine proteases (A) thrombin (0.3 U/mL) and (B) trypsin (0.3 U/mL) in separate strips of mouse gastric fundus contracted with carbachol to 50% Fmax. After responses to the enzymes, isoprenaline (Iso) was added to obtain the maximum relaxation. Vertical (force) and horizontal (time) scale bars apply for both traces. ↱, Changes in time scale from 6 to 2 minutes; ●, points of addition of carbachol and isoprenaline. Gastroenterology 1999 116, 586-592DOI: (10.1016/S0016-5085(99)70180-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 2 Digitized original traces showing responses to cumulative, half-log (M) additions of the PAR-1 and PAR-2 ligands SLIGRL-NH2 (0.1–30 μmol/L) and SFLLRN-NH2 (0.1–30 μmol/L), respectively, in strips of mouse gastric fundus contracted with carbachol to 50% Fmax in the (A) absence and (B) presence of 0.1 μmol/L apamin. After responses to the PAR ligands, isoprenaline (Iso) was added to obtain the maximum relaxation. Vertical (force) and horizontal (time) scale bars apply for both traces. ↘, Changes in time scale from 6 to 2 minutes; ● points of addition of all compounds. No washout occurred between SLIGRL-NH2 and SFLLRN-NH2 additions. Gastroenterology 1999 116, 586-592DOI: (10.1016/S0016-5085(99)70180-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 3 Cumulative concentration-response curves to the PAR-1 and PAR-2 ligands (A, C, and E) SFLLRN-NH2 and (B, D, and F) SLIGRL-NH2, respectively, in strips of mouse gastric fundus contracted to 50% Fmax with carbachol in the absence (control) and presence of either 0.1 μmol/L apamin, 0.3 μmol/L nifedipine, or 10 μmol/L ryanodine. The experimental design was as described in Figure 2. Responses are expressed as a percent relaxation of the contraction to carbachol and represent the mean ± SEM from 4–5 experiments. *P < 0.05, significant difference between the effect of ryanodine or nifedipine compared with apamin on responses to SFLLRN-NH2. Negative values represent contractions. Gastroenterology 1999 116, 586-592DOI: (10.1016/S0016-5085(99)70180-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 4 Cumulative concentration-contraction curves to the PAR-1 ligand SFLLRN-NH2 in strips of mouse gastric fundus contracted to 30% Fmax with carbachol in the presence of either 0.1 μmol/L apamin alone or in combination with 10 μmol/L ryanodine. Responses are expressed as a percentage of the contraction to carbachol and represent the mean ± SEM from 6 experiments. *Contractions at all concentrations of SFLLRN-NH2 (except for 30 μmol/L) were significantly less (P < 0.05) in the presence of a combination of apamin and ryanodine than apamin alone. Gastroenterology 1999 116, 586-592DOI: (10.1016/S0016-5085(99)70180-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 5 Guinea pig taenia coli. (A) Responses to single concentrations (0.3 U/mL) of the serine proteases trypsin and thrombin. ■ and 2, Relaxation and contraction components of each response, respectively. (B) Cumulative and (C) single concentration-effect curves to the PAR-1 and PAR-2 ligands SFLLRN-NH2, and SLIGRL-NH2, respectively, in the absence (control) and presence of either 0.1 μmol/L apamin or 10 μmol/L ryanodine. All responses (expressed as a percent relaxation of the contraction) were obtained in strips of taenia coli contracted to 50% Fmax with carbachol and represent the mean ± SEM from 6–11 experiments. Negative values represent contractions. Gastroenterology 1999 116, 586-592DOI: (10.1016/S0016-5085(99)70180-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions