Comparative effects of L-NOARG and L-NAME on basal blood flow and ACh-induced vasodilatation in rat diaphragmatic microcirculation

1 The effects of N-omega-nitro-L-arginine (L-NOARG) and N-omega-nitro-L-arginine methyl ester (L-NAME) on diaphragmatic microcirculation in male Sprague-Dawley rats were assessed under basal conditions and after acetylcholine (ACh) stimulation. In addition, L-arginine (L-arg) was used with the aim of preventing L-NOARG and L-NAME from inhibiting ACh-induced vasodilatation, in order to explore the possibility that L-NOARG is not only a nitric oxide (NO) synthase inhibitor but also a muscarinic receptor antagonist. 2 Male Sprague-Dawley rats were anaesthetized with urethane and mechanically ventilated. The left hemi-diaphragm of each rat was prepared and microvascular blood flow was recorded during continuous superfusion with bicarbonate-buffered prewarmed Ringer solution by using laser-Doppler flowmetry. The drugs were topically applied to the surface of the hemi-diaphragm. 3 Baseline microvascular blood flow was unaffected after 15 min superfusion with any one of the following agents: L-NOARG (0.1 mM), L-NAME (0.1 mM), L-arg (10 mM). 4 ACh (0.03 mM, 0.1 mM and 0.3 mM) elicited a significant increase of microvascular blood flow (171+/-16%, 214+/-55%, and 323+/-68% of baseline values, respectively), via interaction with the muscarinic receptor, for the vasodilator response was severely inhibited by 15 min superfusion with atropine (0.3 mM). 5 Following 15 min superfusion with either of the L-arg analogues (0.1 mM), the ACh-induced vasodilator response was significantly inhibited. Pretreatment with L-arg (10 mM) for 5 min, followed by co-administration of L-arg (10 mM) and L-NOARG (0.1 mM) for another 15 min significantly prevented the inhibitory effect of L-NOARG on ACh-induced vasodilatation. However, a similar pretreatment schedule with L-arg failed to prevent L-NAME from exerting its inhibitory effect. 6 Neither of the L-arg analogues potentiated sodium nitroprusside (10 mu M and 30 mu M)-induced vasodilatation. However, adenosine (0.1 mM)-induced vasodilatation was slightly but significantly attenuated by either L-NOARG (0.1 mM) or L-NAME (0.1 mM), an effect which was prevented by L-arg (10 mM). 7 In conclusion, an increase in endothelium-dependent blood flow stimulated by ACh may occur in diaphragmatic microcirculation of anaesthetized rats independently of low baseline NO activity. The results also suggest that L-NAME has muscarinic receptor antagonist action in addition to its ability to inhibit NO synthase. Thus, we suggest that L-NAME should not be used as a specific NO synthase inhibitor in the rat diaphragm in situations in which there is potential for muscarinic receptors to be stimulated.