Using a guinea pig model of acute allergic asthma, we recently established that a deficiency of nitric oxide (NO) contributes to airway hyperreactivity (AHR) after the early asthmatic reaction (EAR) and that restoration of NO activity may contribute to the (partial) reversal of AHR after the late asthmatic reaction (LAR). In the present study, we investigated the role of iNOS-derived NO in the regulation of AHR to histamine after the LAR. Inhalation of a selective dose of the specific iNOS inhibitor aminoguanidine (0.1 mM, 3 min) had no effect on basal airway reactivity to histamine in unchallenged, ovalbumin-sensitized animals and did not affect the allergen-induced AHR after the EAR. By contrast, this dose of aminoguanidine significantly potentiated the partially reduced AHR after the LAR to the level of AHR observed after the EAR, indicating that induction of iNOS during the LAR contributes to the reversal of AHR. Inhalation of a higher aminoguanidine concentration (2.5 mM) shortly before the onset of the LAR diminished the AHR after the LAR and reduced the number of neutrophils, lymphocytes, and ciliated epithelial cells in the bronchoalveolar lavage at this time point. The results indicate that iNOS-derived NO may have both beneficial and detrimental effects on allergen-induced AHR to histamine after the LAR by functional antagonism of histamine-induced bronchoconstriction, and by promoting airway inflammation and epithelial damage on the other hand, respectively.

1 Using a conscious, unrestrained guinea-pig model of allergic asthma, we investigated the role of endogenous nitric oxide (NO) in the regulation of airway (hyper)reactivity to histamine before and after the allergen-induced early and late asthmatic reactions, by examining the e ect of inhalation of the NO synthase inhibitor N o -nitro-L-arginine methyl ester (L-NAME, 12 mM, 15 min) on the histamine-induced airway obstruction of ovalbumin-sensitized guinea-pigs before, and at 5.5 h and 23.5 h after allergen challenge. 2 Before allergen challenge, inhaled L-NAME caused a signi®cant 2.02+0.25 fold increase (P50.01) in airway reactivity to histamine; this e ect was reversed within 2.5 to 6 h after administration. 3 After the allergen-induced early asthmatic reaction at 5 h after ovalbumin provocation, a signi®cant 3.73+0.67 fold increase (P50.01) of the airway reactivity to histamine was observed; subsequent inhalation of L-NAME at 5.5 h had no e ect on the airway hyperreactivity, reassessed at 6 h. 4 After the late asthmatic reaction, at 23 h after ovalbumin provocation, a reduced, but still signi®cant airway hyperreactivity to histamine (2.18+0.40 fold; P50.05) was observed. Subsequent inhalation of L-NAME now signi®cantly potentiated the partially reduced airway hyperreactivity 1.57+0.19 fold (P50.05) to the level observed after the early asthmatic reaction. 5 When administered 30 min before allergen exposure, L-NAME signi®cantly enhanced the allergeninduced early asthmatic reaction. However, when administered at 5.5 h after allergen provocation, L-NAME did not a ect the subsequent late asthmatic reaction. 6 These results indicate that endogenous NO is involved the regulation of histamine-and allergeninduced bronchoconstriction and that a de®ciency of cNOS-derived NO contributes to the allergeninduced airway hyperreactivity to histamine after the early asthmatic reaction, while a recovery of NO de®ciency may account for the partial reversal of the allergen-induced airway hyperreactivity after the late asthmatic reaction.

The superimposed twitch technique is frequently used to study the degree of motor unit activation during voluntary effort. This technique is one of the preferred methods to determine the activation deficit (AD) in normal, athletic, and patient populations. One of the limitations of the superimposed twitch technique is its variability under given contractile conditions. The objective of this research was to determine the source(s) of variability in the superimposed twitch force (STF) for repeat measurements. We hypothesized that the variability in the AD measurements may be caused by the timing of the twitch force relative to the onset of muscle activation, by force transients during the twitch application, by small variations in the actual force from the nominal target force, and by variations in the resting twitch force. Twenty-eight healthy subjects participated in this study. Sixteen of these subjects participated in a protocol involving contractions at 50% of their maximal voluntary contraction (MVC) effort, whereas the remaining 12 participated in a protocol involving contractions at 100% of their MVC. Doublet-twitch stimuli were superimposed onto the 50 and 100% effort knee extensor muscle contractions, and the resting twitch forces, voluntary knee extensor forces, and STFs were then measured. The mean resting twitch forces obtained before and after 8 s of 50% of MVC were the same. Similarly, the mean STFs determined at 1, 3, 5, and 7 s into the 50% MVC were the same. The variations in twitch force were significantly smaller after accounting for the actual force at twitch application than those calculated from the prescribed forces during the 50% MVC protocol (P < 0.05). Furthermore, the AD and the actual force showed statistically significant negative correlations for the 50% MVC tests. The interpolated twitch torque determined for the maximal effort contractions ranged from 1 to 70%. In contrast to the protocol at 50% of MVC, negative correlations were only observed in 5 of the 12 subjects during the 100% effort contractions. These results suggest that small variations in the actual force from the target force can account for the majority of the variations in the STFs for submaximal but not maximal effort contractions. For the maximal effort contractions, large variations in the STF exist due to undetermined causes.

It has been suggested that tachykinin NK1 receptor-mediated neurogenic inflammation, characterized by microvascular leakage, mucus secretion, and infiltration and activation of inflammatory cells in the airways, may be involved in allergic asthma. Therefore, in a guinea pig model of allergic asthma, we investigated the involvement of the NK1 receptor in allergen-induced early (EAR) and late (LAR) asthmatic reactions, airway hyperreactivity (AHR) after these reactions and airway inflammation, using the selective nonpeptide NK1 receptor antagonist SR140333. On two different occasions, separated by 1 wk interval, OA-sensitized guinea pigs inhaled either saline (3 min) or SR140333 (100 nM, 3 min) at 30 min before as well as at 5.5 h after OA provocation (between the EAR and LAR) in a random crossover design. A control group, receiving saline inhalations before and at 5.5 h after the two OA provocations, was included as well. SR140333 had no significant effect on either the EAR or the LAR compared with saline control inhalations. However, the NK1 receptor antagonist significantly reduced the OA-induced AHR to histamine, both after the EAR at 5 h after OA challenge (1.77 +/- 0.13-fold increase in histamine reactivity versus 2.50 +/- 0.25-fold increase in the control animals, p < 0.01) and after the LAR at 23 h after OA challenge (1.15 +/- 0.12-fold increase versus 1.98 +/- 0. 34-fold increase, respectively, p < 0.05). Moreover, bronchoalveolar lavage studies performed at 25 h after the second OA provocation indicated that SR140333 significantly inhibited the allergen-induced infiltration of eosinophils, neutrophils, and lymphocytes in the airways (p < 0.05 for all observations), whereas a tendency to reduced accumulation of ciliated epithelial cells in the airway lumen was observed (p = 0.10). These results indicate that the NK1 receptor is involved in the development of allergen-induced AHR to histamine, and that NK1 receptor-mediated infiltration of inflammatory cells in the airways may contribute to this AHR.

1 The effects of increased cellular cyclic AMP levels induced by isoprenaline, forskolin and 8-bromoadenosine 3':5'-cyclic monophosphate (8-Br-cyclic AMP) on phosphoinositide metabolism and changes in intracellular Ca2+ elicited by methacholine and histamine were examined in bovine isolated tracheal smooth muscle (BTSM) cells.2 Isoprenaline (pD2 (-logI0 EC50)=6.32+0.24) and forskolin (pD2= 5.6 + 0.05) enhanced cyclic AMP levels in a concentration-dependent fashion in these cells, while methacholine (pD2= 5.64+0.12) and histamine (pD2=4.90+0.04) caused a concentration-related increase in [3H]-inositol phosphates (IP) accumulation in the presence of 10 mM LiCl.3 Preincubation of the cells (5 min, 37°C) with isoprenaline (1 giM), forskolin (10 ,UM) and 8-Br-cyclic AMP (1 mM) did not affect the IP accumulation induced by methacholine, but significantly reduced the maximal IP production by histamine (1 mM). However, the effect of isoprenaline was small (15.0+0.6% inhibition) and insignificant at histamine concentrations between 0.1 and 100 gM. 4 Both methacholine and histamine induced a fast (max. in 0.5-2 s) and transient increase of intracellular Ca2+ concentration ([Ca2 ]1) followed by a sustained phase lasting several minutes. EGTA (5 mM) attenuated the sustained phase, indicating that this phase depends on extracellular Ca2+.5 Preincubation of the cells (5 min, 37°C) with isoprenaline (1 uM), forskolin (10,M) and 8-Br-cyclic AMP (1 gM) significantly attenuated both the Ca2+-transient and the sustained phase generated at equipotent IP producing concentrations of 1 gM methacholine and 100 gM histamine (approx. 40% of maximal methacholine-induced IP response), but did not affect changes in [Ca2+] induced by 100 gM methacholine (95.2+3.5% of maximal methacholine-induced IP response). 6 Significant correlations were found between the isoprenaline-induced inhibition of BTSM contraction and inhibition of Ca2+ mobilization or influx induced by methacholine and histamine, that were similar for each contractile agonist. 7 These data indicate that (a) cyclic AMP-dependent inhibition of Ca2+ mobilization in BTSM cells is not primarily caused by attenuation of IP production, suggesting that cyclic AMP induced protein kinase A (PKA) activation is effective at a different level in the [Ca2+]i homeostasis, (b) that attenuation of intracellular Ca2+ concentration plays a major role in P-adrenoceptor-mediated relaxation of methacholine-and histamine-induced airway smooth muscle contraction, and (c) that the relative resistance of the muscarinic agonist-induced contraction to fi-adrenoceptor agonists, especially at (supra) maximal contractile concentrations is largely determined by its higher potency in inducing intracellular Ca2+ changes.