The effect of okadaic acid on non-adrenergic non-cholinergic contraction in guinea-pig isolated bronchus

We have investigated the role of phosphatases in modulating contractile responses to electrical field stimulation (EFS), methacholine, substance P and capsaicin in guinea-pig isolated main bronchus by use of the phosphatase 1 and 2A inhibitor okadaic acid. Non-adrenergic non-cholinergic (eNANC) contractile responses were elicited by EFS (3 Hz, 20 s, 0.5 ms max. voltage) in the guinea-pig isolated main bronchus in the presence of the non-selective muscarinic antagonist, atropine (1 μM), the non-selective β-adrenoceptor antagonist, propranolol (1 μM), the neutral endopeptidase inhibitor thiorphan (10 μM) and the cyclo-oxygenase inhibitor, indomethacin (5 μM). Okadaic acid significantly attenuated eNANC contractile responses (% inhibition) elicited by EFS (0.01 μM, 15.2±26.9%; 0.03 μM, 30.4±13.9%; 0.01 μM, 39.8±5.1%; 0.3 μM, 59.5±8.7%; 1 μM 77.8±7.8%; P 0.05, n=4). Cholinergic contractile responses were elicited by EFS (1–30 Hz, 10 s, 0.5 ms max. voltage) in guinea-pig isolated bronchus in the presence of the nitric oxide synthase inhibitor, Nω-nitro-L-arginine methyl ester (L-NAME, 30 μM). Okadaic acid failed to attenuate significantly the contractile (% methacholine Emax) response elicited by EFS at all frequencies tested compared with the control (1 Hz, control, 22±7.9% vs okadaic acid, 18±7.7%; 3 Hz, control, 26±6.9% vs okadaic acid, 27±9.1%; 10 Hz, control, 36±7.6% vs okadaic acid, 33±8.9%; 30 Hz, control, 50±7.6% vs okadaic acid, 42±14%; P>0.05, n=4). Okadaic acid (0.3 μM) failed to alter significantly the contractile potency (pD2) to capsaicin (okadaic acid, 9.0±0.5, vs DMSO, 9.2±0.4; P>0.05 n=6), substance P (okadaic acid, 7.6±0.3 vs DMSO, 8.2±0.2; P>0.05 n=7) or methacholine (okadaic acid, 6.4±0.2 vs DMSO, 6.4±0.3; P>0.05 n=4). Okadaic acid (0.01–1 μM) did not appear to reverse substance P-induced tone. The maximal relaxant response (% reversal of substance P-induced tone) mediated by okadaic acid (1 μM) was 33±11.7% (n=4), this was not significantly different from the DMSO (0.8%) or a time-dependent fall in tone of 34.3±23.1% (n=4) and 33±15.8% (n=4), respectively. Okadaic acid (0.3 μM) failed to augment isoprenaline-induced relaxation repsonses in substance P contracted bronchus (okadaic acid, 6.5±0.4 vs DMSO, 5.9±0.3; P>0.05, n=9). These results indicate that protein phosphatases appear to regulate the release of sensory neuropeptides from airway sensory nerves in response to electrical field stimulation. British Journal of Pharmacology (1997) 121, 181–186; doi:10.1038/sj.bjp.0701114