The effect of hydro-ethanolic extract of Achillea millefolium on muscarinic receptors of guinea pig tracheal smooth muscle

Abstract

Objective: To investigate one possible mechanism for the observed relaxant effect of A. millefolium (Achillea millefolium), in the present study the inhibitory effect of the extract of this plant on muscarinic receptors was examined.

Materials and methods: The effects of three concentrations of aqueous-ethanolic extract, 10 nM atropine, and saline on muscarinic receptors were tested in three conditions: In non incubated tracheal smooth muscle (group 1), tracheal chain incubated with propranolol and chlorpheniramine (group 2), and the one incubated with propranolol (group 3).

Results: The EC₅₀ obtained in the presence of all three concentrations of the extract were significantly higher compared to saline in groups 2 and 3 (P < 0.001 and P < 0.01 in group 2 and 3 respectively). The EC₅₀ obtained in the presence of all concentrations of the extract in group 2 were significantly improved compared to groups 1 and 3 (P < 0.05 to P < 0.001). The maximum responses to methacholine in presence of only the higher concentration of the extract (0.8 mg/ml) was significantly lower than that of saline in groups 1 (P < 0.05). There was neither significant difference between slopes of methacholine-response curves obtained in the presence of different concentrations of the extract and that of saline nor between the three groups. The values of (CR-1), obtained in the presence of all concentrations of the extract, were significantly lower compared to atropine in the first group but were not significantly different in other groups. The values of (CR-1) obtained in the presence of all concentrations of the extract were significantly improved in groups 2, compared to groups 1 and 3 (P < 0.05 to P < 0.001).

Conclusion: These results showed an inhibitory effect for the extract of A. millefolium on muscarinic receptors of tracheal smooth muscle. A histamine (H₁) receptor blockade was also suggested for the extract.

Keywords: Achillea millefolium; guinea pig; hydroalcoholic extract; muscarinic receptors; trachea.

Figure 1

Cumulative log concentration-response curves of metacholin induced contraction of guinea pig tracheal chains, in the presence of saline, three concentrations of aqueous-ethanolic extract and 10 nM atropine in (a) non incubated trachea (group 1, open symbols, n= 6), (b) incubated trachea with 1 μM chlorpheniramine and 1 μM propranolol (group 2, filled symbols, n=9) and (c) on tracheal chains incubated with propranolol (Group 3, open symbols with a dot, n=7)

Figure 2

EC₅₀ of metacholin obtained in the presence of three concentrations of aqueous-ethanolic extract from A.millefolium (0.2 ∇, 0.4 Δ, and 0.8 mg/mL, ), 10 nM atropine (□) and saline (O) in non incubated trachea (a) (open symbols, group 1, n = 6), incubated trachea with 1 μM chlorpheniramine and 1 μM propranolol (b) (filled symbols, group 2, n = 9) and trachea incubated with propranolol (c) (open symbols with a dot, group 3, n = 7). Statistical significance for differences in EC50 between saline and other solutions, NS: non-significant difference, ^**: p < 0.01, ^***: p < 0.001. Statistical significance for the difference in EC50 for group 1 and 3 vs group 2, ⁺: p < 0.05, ⁺⁺⁺: p < 0.001. Statistical significance for the difference in EC₅₀ between groups 1 and 3, #: p < 0.5

Figure 3

The values of (CR-1) obtained in the presence of three concentrations of aqueous-ethanolic extract from A.millefolium (0.2 ∇, 0.4 Δ, and 0.8 mg/mL, ) and 10 nM atropine (□) in (a) non incubated trachea (open symbols, group 1, n = 6), (b) incubated trachea with 1 μM chlorpheniramine and 1 μM propranolol (filled symbols, group 2, n = 9) and (c) tracheal chains incubated with propranolol (open symbols with a dot, group 3, n = 7). Statistical significance for the difference in (CR-1) between atropine and the extract, ^*: p < 0.05, ^**: p < 0.01, ^***: p < 0.001. Statistical significance for the difference in (CR-1) for group 1 and 3 vs group 2, ⁺: p < 0.05, ⁺⁺: p < 0.01, ⁺⁺⁺: p < 0.001. There was no significant difference in (CR-1) between groups 1 and 3

Figure 4

The Schild plot, log (CR-1) against log extract concentrations in (a) non incubated trachea (open symbols, group 1), (b) incubated trachea with 1 μM chlorpheniramine and 1 μM propranolol (filled symbols, group 2) and (c) tracheal chains incubated with propranolol (open symbols with a dot, group 3). The slopes of Schild plot for the extract was .-0.568, -0.837 and -0.761 for group 1, 2 and 3 respectively