Arginase attenuates inhibitory nonadrenergic noncholinergic nerve-induced nitric oxide generation and airway smooth muscle relaxation

Abstract

Background: Recent evidence suggests that endogenous arginase activity potentiates airway responsiveness to methacholine by attenuation of agonist-induced nitric oxide (NO) production, presumably by competition with epithelial constitutive NO synthase for the common substrate, L-arginine. Using guinea pig tracheal open-ring preparations, we now investigated the involvement of arginase in the modulation of neuronal nitric oxide synthase (nNOS)-mediated relaxation induced by inhibitory nonadrenergic noncholinergic (iNANC) nerve stimulation.

Methods: Electrical field stimulation (EFS; 150 mA, 4 ms, 4 s, 0.5 - 16 Hz)-induced relaxation was measured in tracheal preparations precontracted to 30% with histamine, in the presence of 1 microM atropine and 3 microM indomethacin. The contribution of NO to the EFS-induced relaxation was assessed by the nonselective NOS inhibitor L-NNA (0.1 mM), while the involvement of arginase activity in the regulation of EFS-induced NO production and relaxation was investigated by the effect of the specific arginase inhibitor nor-NOHA (10 microM). Furthermore, the role of substrate availability to nNOS in EFS-induced relaxation was measured in the presence of various concentrations of exogenous L-arginine.

Results: EFS induced a frequency-dependent relaxation, ranging from 6.6 +/- 0.8% at 0.5 Hz to 74.6 +/- 1.2% at 16 Hz, which was inhibited with the NOS inhibitor L-NNA by 78.0 +/- 10.5% at 0.5 Hz to 26.7 +/- 7.7% at 8 Hz (P < 0.01 all). In contrast, the arginase inhibitor nor-NOHA increased EFS-induced relaxation by 3.3 +/- 1.2-fold at 0.5 Hz to 1.2 +/- 0.1-fold at 4 Hz (P < 0.05 all), which was reversed by L-NNA to the level of control airways in the presence of L-NNA (P < 0.01 all). Similar to nor-NOHA, exogenous L-arginine increased EFS-induced airway relaxation (P < 0.05 all).

Conclusion: The results indicate that endogenous arginase activity attenuates iNANC nerve-mediated airway relaxation by inhibition of NO generation, presumably by limiting L-arginine availability to nNOS.

Figure 1

Role of NO and arginase in iNANC nerve-induced relaxation of guinea pig tracheal smooth muscle. Electrical field stimulation-induced relaxation of precontracted guinea pig tracheal open-ring preparations in the absence and presence of the NOS inhibitor L-NNA (100 μM), the arginase inhibitor nor-NOHA (10 μM) or a combination of both inhibitors. Results are means ± s.e.m. of 8 experiments. *P < 0.05 and **P < 0.01 compared to control, ^†P < 0.05 and ^‡P < 0.01 compared to nor-NOHA-treated.

Figure 2

Role of L-arginine availability and arginase in iNANC nerve-induced relaxation of guinea pig tracheal smooth muscle. Electrical field stimulation-induced relaxation of precontracted guinea pig tracheal open-ring preparations in the absence and presence of exogenous L-arginine (5.0 mM) or the arginase inhibitor nor-NOHA (10 μM). Results are means ± s.e.m. of 5–13 experiments. *P < 0.05 and **P < 0.01 compared to control.