Comparison of two soluble guanylyl cyclase inhibitors, methylene blue and ODQ, on sodium nitroprusside-induced relaxation in guinea-pig trachea

Abstract

1. To clarify further the role of cyclic GMP in mediating the relaxant response in guinea-pig trachea induced by sodium nitroprusside (SNP), the effects of soluble guanylyl cyclase inhibitors, methylene blue and 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ) on SNP-induced muscle relaxation and cyclic GMP accumulation were determined.

2. SNP (0.3–100 μM) evoked a concentration-dependent relaxation of guinea-pig isolated tracheas precontracted with 0.3 μM carbachol. Preincubation of the preparations with methylene blue (10, 30 and 100 μM) resulted in a slight but concentration-dependent prevention of the relaxant response to SNP. In contrast, the relaxation to SNP was extensively prevented by 3 μM ODQ and almost abolished by 10 μM ODQ.

3. SNP (30 μM) induced a significant elevation of cyclic GMP accumulation (from 1.34±0.14 to 5.39±0.28 pmol mg⁻¹ protein, n=5; P<0.001), which was partially attenuated by 100 μM methylene blue (3.11±0.51 pmol mg⁻¹ protein, n=5; P<0.05), whereas completely abolished by 10 μM ODQ (1.31±0.28 pmol mg⁻¹ protein, n=5; P<0.001).

4. Methylene blue, but not ODQ and N^ω-nitro-L-arginine methyl ester (L-NAME), caused a concentration-dependent contraction in the tracheal preparation. The tension produced by 100 μM methylene blue was 41.8±4.3% (0.3 μM carbachol as 100%; n=12). Moreover, the non-selective muscarinic receptor antagonist atropine and the M₃-selective antagonist 4-diphenylacetoxy-N-methylpiperidine methiodine greatly inhibited the contractile effect evoked by methylene blue (100 μM).

5. In conclusion, this study provides substantial evidence that SNP-induced muscle relaxation in guinea-pig trachea is completely via a cyclic GMP-dependent mechanism. Furthermore, ODQ, but not methylene blue, will likely become an important tool in differentiating between cyclic GMP-dependent and -independent effects of nitric oxide.

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