Mechanisms of organophosphate insecticide-induced airway hyperreactivity

It has been suggested that pesticide exposure may be a contributing factor underlying the increased incidence of asthma in the United States and other industrialized nations. To test this hypothesis, airway hyperreactivity was measured in guinea pigs exposed to chlorpyrifos, a widely used organophosphate pesticide. Electrical stimulation of the vagus nerves caused frequency-dependent bronchoconstriction that was significantly potentiated in animals 24 h or 7 days after a single subcutaneous injection of either 390 mg/kg or 70 mg/kg of chlorpyrifos, respectively. Mechanisms by which chlorpyrifos may cause airway hyperreactivity include inhibition of acetylcholinesterase (AChE) or dysfunction of M3 muscarinic receptors on airway smooth muscle or of autoinhibitory M2 muscarinic receptors on parasympathetic nerves in the lung. AChE activity in the lung was significantly inhibited 24 h after treatment with 390 mg/kg of chlorpyrifos, but not 7 days after injection of 70 mg/kg of chlorpyrifos. Acute exposure to eserine (250 microg/ml) also significantly inhibited lung AChE but did not potentiate vagally induced bronchoconstriction. Neuronal M2 receptor function was tested using the M2 agonist pilocarpine, which inhibits vagally induced bronchoconstriction in control animals. In chlorpyrifos-treated animals, pilocarpine dose-response curves were shifted significantly to the right, demonstrating decreased responsiveness of neuronal M2 receptors. In contrast, chlorpyrifos treatment did not alter methacholine-induced bronchoconstriction, suggesting that chlorpyrifos does not alter M3 muscarinic receptor function on airway smooth muscle. These data demonstrate that organophosphate insecticides can cause airway hyperreactivity in the absence of AChE inhibition by decreasing neuronal M2 receptor function.     

Effects of inflammatory cells on neuronal M2 muscarinic receptor function in the lung

In the lungs, acetylcholine released from the parasympathetic nerves stimulates M3 muscarinic receptors on airway smooth muscle inducing contraction and bronchoconstriction. The amount of acetylcholine released from these nerves is limited locally by neuronal M2 muscarinic receptors. These neuronal receptors are dysfunctional in asthma and in animal models of asthma. Decreased M2 muscarinic receptor function results in increased release of acetylcholine and in airway hyperreactivity. Inflammation has long been associated with hyperreactivity and the role of inflammatory cells in loss of neuronal M2 receptor function has been examined. There are several different mechanisms for loss of neuronal M2 receptor function. These include blockade by endogenous antagonists such as eosinophil major basic protein, decreased expression of M2 receptors following infection with viruses or exposure to pro inflammatory cytokines such as gamma interferon. Finally, the affinity of acetylcholine for these receptors can be decreased by exposure to neuraminidase.     

Substance P-induced airway hyperreactivity is mediated by neuronal M(2) receptor dysfunction

Neuronal muscarinic (M(2)) receptors inhibit release of acetylcholine from the vagus nerves. Hyperreactivity in antigen-challenged guinea pigs is due to blockade of these M(2) autoreceptors by eosinophil major basic protein (MBP) increasing the release of acetylcholine. In vivo, substance P-induced hyperactivity is vagally mediated. Because substance P induces eosinophil degranulation, we tested whether substance P-induced hyperreactivity is mediated by release of MBP and neuronal M(2) receptor dysfunction. Pathogen-free guinea pigs were anesthetized and ventilated. Thirty minutes after intravenous administration of [Sar(9),Met(O(2))(11)]- substance P, guinea pigs were hyperreactive to vagal stimulation and M(2) receptors were dysfunctional. The depletion of inflammatory cells with cyclophosphamide or the administration of an MBP antibody or a neurokinin-1 (NK(1)) receptor antagonist (SR-140333) all prevented substance P-induced M(2) dysfunction and hyperreactivity. Intravenous heparin acutely reversed M(2) receptor dysfunction and hyperreactivity. Thus substance P releases MBP from eosinophils resident in the lungs by stimulating NK(1) receptors. Substance P-induced hyperreactivity is mediated by blockade of inhibitory neuronal M(2) receptors by MBP, resulting in increased release of acetylcholine.     

Role of macrophages in virus-induced airway hyperresponsiveness and neuronal M2 muscarinic receptor dysfunction

Viral infections exacerbate asthma. One of the pathways by which viruses trigger bronchoconstriction and hyperresponsiveness is by causing dysfunction of inhibitory M(2) muscarinic receptors on the airway parasympathetic nerves. These receptors normally limit acetylcholine (ACh) release from the parasympathetic nerves. Loss of M(2) receptor function increases ACh release, thereby increasing vagally mediated bronchoconstriction. Because viral infection causes an influx of macrophages into the lungs, we tested the role of macrophages in virus-induced airway hyperresponsiveness and M(2) receptor dysfunction. Guinea pigs infected with parainfluenza virus were hyperresponsive to electrical stimulation of the vagus nerves but not to intravenous ACh, indicating that hyperresponsiveness was due to increased release of ACh from the nerves. In addition, the muscarinic agonist pilocarpine no longer inhibited vagally induced bronchoconstriction, indicating M(2) receptor dysfunction. Treating animals with liposome-encapsulated dichloromethylene-diphosphonate depleted macrophages as assessed histologically. In these animals, viral infection did not cause airway hyperresponsiveness or M(2) receptor dysfunction. These data suggest that macrophages mediate virus-induced M(2) receptor dysfunction and airway hyperresponsiveness.     

Ozone-induced hyperresponsiveness and blockade of M2 muscarinic receptors by eosinophil major basic protein.

Control of airway smooth muscle is provided by parasympathetic nerves that release acetylcholine onto M(3) muscarinic receptors. Acetylcholine release is limited by inhibitory M(2) muscarinic receptors. In antigen-challenged guinea pigs, hyperresponsiveness is due to blockade of neuronal M(2) receptors by eosinophil major basic protein (MBP). Because exposure of guinea pigs to ozone also causes M(2) dysfunction and airway hyperresponsiveness, the role of eosinophils in ozone-induced hyperresponsiveness was tested. Animals were exposed to filtered air or to 2 parts/million ozone for 4 h. Twenty-four hours later, the muscarinic agonist pilocarpine no longer inhibited vagally induced bronchoconstriction in ozone-exposed animals, indicating M(2) dysfunction. M(2) receptor function in ozone-exposed animals was protected by depletion of eosinophils with antibody to interleukin-5 and by pretreatment with antibody to guinea pig MBP. M(2) function was acutely restored by removal of MBP with heparin. Ozone-induced hyperreactivity was also prevented by antibody to MBP and was reversed by heparin. These data show that loss of neuronal M(2) receptor function after ozone is due to release of eosinophil MBP.     

The changing role of eosinophils in long-term hyperreactivity following a single ozone exposure

Ozone hyperreactivity over 24 h is mediated by blockade of inhibitory M(2) muscarinic autoreceptors by eosinophil major basic protein. Because eosinophil populations in the lungs fluctuate following ozone, the contribution of eosinophils to M(2) dysfunction and airway hyperreactivity was measured over several days. After one exposure to ozone, M(2) function, vagal reactivity, smooth muscle responsiveness, and inflammation were measured in anesthetized guinea pigs. Ozone-induced hyperreactivity to vagal stimulation persisted over 3 days. Although hyperreactivity one day after ozone is mediated by eosinophils, AbVLA-4 did not inhibit either eosinophil accumulation in the lungs or around the nerves or prevent hyperreactivity at this time point. Two days after ozone, eosinophils in BAL, around airway nerves and in lungs, were decreased, and neuronal M(2) receptor function was normal, although animals were still hyperreactive to vagal stimulation. Depleting eosinophils with AbIL-5 prevented hyperreactivity, thus eosinophils contribute to vagal hyperreactivity by mechanisms separate from M(2) receptor blockade. Three days after ozone, vagal hyperreactivity persisted, eosinophils were again elevated in BAL in lungs and around nerves, and M(2) receptors were again dysfunctional. At this point, airway smooth muscle was also hyperresponsive to methacholine. Eosinophil depletion with AbIL-5, AbVLA-4, or cyclophosphamide protected M(2) function 3 days after ozone and prevented smooth muscle hyperreactivity. However, vagal hyperreactivity was significantly potentiated by eosinophil depletion. The site of hyperreactivity, muscle or nerve, changes over 3 days after a single exposure to ozone. Additionally, the role of eosinophils is complex; they mediate hyperreactivity acutely while chronically may be involved in repair.     

Atropine pretreatment enhances airway hyperreactivity in antigen-challenged guinea pigs through an eosinophil-dependent mechanism

Airway hyperreactivity in antigen-challenged animals is mediated by eosinophil major basic protein (MBP) that blocks inhibitory M(2) muscarinic receptors on parasympathetic nerves, increasing acetylcholine release onto M(3) muscarinic receptors on airway smooth muscle. Acutely, anticholinergics block hyperreactivity in antigen-challenged animals and reverse asthma exacerbations in the human, but are less effective in chronic asthma. We tested whether atropine, given before antigen challenge, affected hyperreactivity, M(2) receptor function, eosinophil accumulation, and activation. Sensitized guinea pigs received atropine (1 mg/kg ip) 1 h before challenge and 6 h later. Twenty-four hours after challenge, animals were anesthetized, vagotomized, paralyzed, and ventilated. Airway reactivity to electrical stimulation of the vagi and to intravenous acetylcholine was not altered by atropine pretreatment in nonsensitized animals, indicating that atropine was no longer blocking postjunctional muscarinic receptors. Antigen challenge induced airway hyperreactivity to vagal stimulation that was significantly potentiated by atropine pretreatment. Bronchoconstriction induced by acetylcholine was not changed by antigen challenge or by atropine pretreatment. M(2) receptor function was lost in challenged animals but protected by atropine pretreatment. Eosinophils in bronchoalveolar lavage and within airway tissues were significantly increased by challenge but significantly reduced by atropine pretreatment. However, extracellular MBP in challenged airways was significantly increased by atropine pretreatment, which may account for reduced eosinophils. Depleting eosinophils with antibody to IL-5 before challenge prevented hyperreactivity and significantly reduced MBP in airways of atropine-pretreated animals. Thus atropine pretreatment potentiated airway hyperreactivity by increasing eosinophil activation and degranulation. These data suggest that anticholinergics enhance eosinophil interactions with airway nerves.     

Dexamethasone prevents virus-induced hyperresponsiveness via multiple mechanisms

In the lungs, neuronal M2 muscarinic receptors inhibit acetylcholine release from the parasympathetic nerves. Parainfluenza virus infection causes loss of M2 receptor function, which increases acetylcholine release and vagally mediated bronchoconstriction. Because glucocorticoids are known to inhibit airway hyperresponsiveness, we tested whether dexamethasone (6.5 or 65 microg/kg i.p.) prevents virus-induced hyperresponsiveness and M2 receptor dysfunction in guinea pigs. In controls, pilocarpine, a muscarinic agonist, inhibited vagally induced bronchoconstriction, demonstrating functional M2 receptors. However, in virus-infected animals, pilocarpine failed to inhibit vagally induced bronchoconstriction, demonstrating M2 receptor dysfunction. Frequency-dependent bronchoconstriction was greater in virus-infected animals than in controls, indicating airway hyperresponsiveness. Low-dose dexamethasone (6.5 microg/kg i.p.) treatment prevented virus-induced airway hyperresponsiveness, ameliorated M2 receptor dysfunction, and decreased viral content in the lungs without inhibiting virus induced inflammation. High-dose dexamethasone (65 microg/kg i.p.) prevented virus-induced hyperresponsiveness, completely reversed M2 receptor dysfunction, decreased viral titers, and decreased virus-induced inflammation. This high-dose dexamethasone also increased M2 receptor function in uninfected animals. In conclusion, dexamethasone prevented virus-induced hyperresponsiveness and M2 receptor dysfunction via multiple mechanisms.     

Effect of M2 and M3 muscarinic receptors on airway responsiveness to carbachol in bronchial-hypersensitive (BHS) and bronchial-hyposensitive (BHR) guinea pigs.

The expression balance of M2 and M3 muscarinic receptor subtypes on the pathogenesis of airway hyperresponsiveness was investigated by using two congenitally related strains of guinea pigs, bronchial-hypersensitive (BHS) and bronchial-hyposensitive (BHR). CCh-induced airway responses in vivo and in vitro were investigated by comparing the effects of muscarinic receptor subtype antagonists, and the relative amounts of M2 and M3 muscarinic receptor mRNA in tracheal smooth muscle and lung tissue were investigated. After treatment with muscarinic receptor subtype antagonists, the ventilatory mechanics (VT, Raw, and Cdyn) of response to CCh aerosol inhalation were measured by the bodyplethysmograph method. The effects of these antagonists on CCh-induced tracheal smooth muscle contraction were also investigated. The effects of M2 muscarinic receptor blockade were less but the effects of M3 muscarinic receptors blockade on the airway contractile responses were greater in BHS than in BHR. In M3 muscarinic receptor blockades, CCh-induced tracheal contractions in BHS were significantly greater than those in BHR. In tracheal smooth muscle from BHS, the relative amount of M2 muscarinic receptors mRNA was less but that of M3 muscarinic receptor mRNA was more than those in BHR. These results suggest that the high ACh level as a consequence of dysfunction of M2 muscarinic autoreceptors and the excessive effect of M3 muscarinic receptors on the airway smooth muscle may play an important role in the pathogenesis of airway hyperresponsiveness.     

Three days after a single exposure to ozone, the mechanism of airway hyperreactivity is dependent on substance P and nerve growth factor

Ozone causes persistent airway hyperreactivity in humans and animals. One day after ozone exposure, airway hyperreactivity is mediated by release of eosinophil major basic protein that inhibits neuronal M(2) muscarinic receptors, resulting in increased acetylcholine release and increased smooth muscle contraction in guinea pigs. Three days after ozone, IL-1β, not eosinophils, mediates ozone-induced airway hyperreactivity, but the mechanism at this time point is largely unknown. IL-1β increases NGF and the tachykinin substance P, both of which are involved in neural plasticity. These experiments were designed to test whether there is a role for NGF and tachykinins in sustained airway hyperreactivity following a single ozone exposure. Guinea pigs were exposed to filtered air or ozone (2 parts per million, 4 h). In anesthetized and vagotomized animals, ozone potentiated vagally mediated airway hyperreactivity 24 h later, an effect that was sustained over 3 days. Pretreatment with antibody to NGF completely prevented ozone-induced airway hyperreactivity 3 days, but not 1 day, after ozone and significantly reduced the number of substance P-positive airway nerve bundles. Three days after ozone, NK(1) and NK(2) receptor antagonists also blocked this sustained hyperreactivity. Although the effect of inhibiting NK(2) receptors was independent of ozone, the NK(1) receptor antagonist selectively blocked vagal hyperreactivity 3 days after ozone. These data confirm mechanisms of ozone-induced airway hyperreactivity change over time and demonstrate 3 days after ozone that there is an NGF-mediated role for substance P, or another NK(1) receptor agonist, that enhances acetylcholine release and was not present 1 day after ozone.     

Effects of neurokinin receptor antagonists in virus-infected airways

We investigated the effects of a neurokinin-1 (NK(1)) receptor antagonist (SR-140333) and a NK(2) receptor antagonist (SR-48968) on airway responsiveness and on the function of neuronal M(2) muscarinic receptors, which normally inhibit vagal acetylcholine release, in guinea pigs infected with parainfluenza virus. Antagonists were given 1 h before infection and daily thereafter. Four days later, bronchoconstriction induced by either intravenous histamine (which is partly vagally mediated) or electrical stimulation of the vagus nerves was increased by viral infection compared with control. In addition, the ability of the muscarinic agonist pilocarpine to inhibit vagally induced bronchoconstriction was lost in virus-infected animals, demonstrating loss of neuronal M(2) receptor function. Macrophage influx into the lungs was inhibited by pretreatment with both antagonists. However, only the NK(1) receptor antagonist prevented M(2) receptor dysfunction and inhibited hyperresponsiveness (measured as an increase in either vagally induced or histamine-induced bronchoconstriction). Thus virus-induced M(2) receptor dysfunction and hyperresponsiveness are prevented by a NK(1) receptor antagonist, but not by a NK(2) receptor antagonist, whereas both antagonists had similar anti-inflammatory effects.     

Eosinophils and airway nerves in asthma

In the lungs, neuronal M2 muscarinic receptors limit the release of acetylcholine from postganglionic cholinergic nerves. However, these receptors are not functional under certain circumstances in animal models of hyperreactivity such as occurs after exposure of sensitised animals to an allergen or during a respiratory tract virus infection. This loss of M2 receptor function leads to an increase in acetylcholine release from cholinergic nerves and thus is a mechanism for the vagally mediated hyperreactivity seen in these animals. Studies in animal models of hyperreactivity have shown that eosinophils localise to the airway nerves of sensitised animals after antigen challenge. Inhibiting this localisation of eosinophils either with an antibody to the eosinophil survival cytokine IL-5 or the eosinophil adhesion molecule VLA-4 prevents loss of M2 muscarinic receptor function. It is likely that eosinophil MBP is responsible for the loss of M2 receptor function, since inhibiting eosinophil MBP with an antibody or neutralising MBP with heparin prevents this loss of function. These data are also supported by ligand binding studies where it has been shown that eosinophil MBP is an allosteric antagonist at neuronal M2 muscarinic receptors. Loss of function of lung neuronal M2 muscarinic receptors may also occur under certain circumstances in patients with asthma, although the mechanisms are not yet established.     

Functional expression of the GABAB receptor in human airway smooth muscle

gamma-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian central nervous system and exerts its actions via both ionotropic (GABA(A)/GABA(C)) and metabotropic (GABA(B)) receptors (R). In addition to their location on neurons, GABA and functional GABA(B) receptors have been detected in nonneuronal cells in peripheral tissue. Although the GABA(B)R has been shown to function as a prejunctional inhibitory receptor on parasympathetic nerves in the lung, the expression and functional coupling of GABA(B) receptors to G(i) in airway smooth muscle itself have never been described. We detected the mRNA encoding multiple-splice variants of the GABA(B)R1 and GABA(B)R2 in total RNA isolated from native human and guinea pig airway smooth muscle and from RNA isolated from cultured human airway smooth muscle (HASM) cells. Immunoblots identified the GABA(B)R1 and GABA(B)R2 proteins in human native and cultured airway smooth muscle. The GABA(B)R1 protein was immunohistochemically localized to airway smooth muscle in guinea pig tracheal rings. Baclofen, a GABA(B)R agonist, elicited a concentration-dependent stimulation of [(35)S]GTPgammaS binding in HASM homogenates that was abrogated by the GABA(B)R antagonist CGP-35348. Baclofen also inhibited adenylyl cyclase activity and induced ERK phosphorylation in HASM. Another GABA(B)R agonist, SKF-97541, mimicked while pertussis toxin blocked baclofen's effect on ERK phosphorylation, implicating G(i) protein coupling. Functional GABA(B) receptors are expressed in HASM. GABA may modulate an uncharacterized signaling cascade via GABA(B) receptors coupled to the G(i) protein in airway smooth muscle.     

O3-induced mucosa-linked airway muscle hyperresponsiveness in the guinea pig.

We investigated the effects of ozone exposure (3.0 ppm, 2 h) on the responsiveness of guinea pig airway muscle in vitro from animals developing bronchial hyperreactivity. Muscarinic reactivity in vivo was determined by measuring specific airway resistance (sRaw) in response to increasing concentrations of aerosolized acetylcholine (ACh) administered before and 30 min after exposure. Immediately after reactivity testing, multiple tracheal rings from ozone- and air-exposed animals were prepared and the contractile responses to increasing concentrations of substance P, ACh, or KCl were assessed in the presence of 10 microM indomethacin with or without 1 microM phosphoramidon, an inhibitor of neutral endopeptidase. Isometric force generation in vitro was measured on stimulation by cumulative concentrations of the agonists, and force generation (in g/cm2) was calculated after determination of muscle cross-sectional area. The smooth muscle of mucosa-intact airways from guinea pigs with ozone-induced bronchial hyper-reactivity proved to be hyperresponsive in vitro to substance P and ACh but not to KCl. Pretreatment with phosphoramidon abolished the increase in substance P responsiveness but had no effect on muscarinic hyperresponsiveness after ozone exposure. Furthermore, substance P responsiveness was not augmented in ozone-exposed airways in which the mucosa had been removed before testing in vitro. Likewise, muscarinic hyperresponsiveness was not present in ozone-exposed airways without mucosa. Our data indicate that airway smooth muscle responsiveness is increased in guinea pigs with ozone-induced bronchial hyperreactivity and suggest that this hyperresponsiveness may be linked to non-cyclooxygenase mucosa-derived factors.     

Nerve growth factor acutely potentiates synaptic transmission in vitro and induces dendritic growth in vivo on adult neurons in airway parasympathetic ganglia

Elevated levels of nerve growth factor (NGF) and NGF-mediated neural plasticity may have a role in airway diseases such as asthma and chronic obstructive pulmonary disease (COPD). Although NGF is known to affect sensory and sympathetic nerves, especially during development, little is known regarding its effect on parasympathetic nerves, especially on adult neurons. The purpose of this study was to analyze the acute and chronic effects of NGF on the electrophysiological and anatomical properties of neurons in airway parasympathetic ganglia from adult guinea pigs. Using single cell recording, direct application of NGF caused a lasting decrease in the cumulative action potential afterhyperpolarization (AHP) and increased the amplitude of vagus nerve-stimulated nicotinic fast excitatory postsynaptic potentials. Neuronal responsiveness to nicotinic receptor stimulation was increased by NGF, which was blocked by the tyrosine kinase inhibitor, K-252a, implicating neurotrophin-specific (Trk) receptors. Neurotrophin-3 and brain-derived neurotrophic factor had no effect on the synaptic potentials, AHP, or nicotinic response; inhibition of cyclooxygenase with indomethacin inhibited the effect of NGF on the cumulative AHP. Forty-eight hours after in vivo application of NGF to the trachealis muscle caused an increase in dendritic length on innervating neurons. These results are the first to demonstrate that NGF increases the excitability of lower airway parasympathetic neurons, primarily through enhanced synaptic efficacy and changes to intrinsic neuron properties. NGF also had dramatic effects on the growth of dendrites in vivo. Such effects may indicate a new role for NGF in the regulation of parasympathetic tone in the diseased or inflamed lower airways.     

Dysfunction of M2-muscarinic receptors in pulmonary parasympathetic nerves after antigen challenge.

The effect of antigen challenge on the function of neuronal M2-muscarinic autoreceptors in the lungs was studied in anesthetized guinea pigs. Guinea pigs were injected intraperitoneally with saline (control group) or ovalbumin (10 mg/kg) on days 1, 3, and 5. One group of sensitized animals was challenged on days 20-25 with aerosolized ovalbumin for 5 min/day (challenged group), while another group of the sensitized animals was not challenged (sensitized group). On day 26 the animals were anesthetized, paralyzed, tracheostomized, and artificially ventilated. Pulmonary inflation pressure (Ppi), tidal volume, blood pressure, and heart rate were recorded. Both vagus nerves were cut, and electrical stimulation of the distal portions caused bronchoconstriction (measured as an increase in Ppi) and bradycardia. In the control group, pilocarpine (1-100 micrograms/kg iv) attenuated vagally induced bronchoconstriction by stimulating inhibitory M2-muscarinic receptors on parasympathetic nerves in the lungs. Conversely, blockade of these receptors with the antagonist gallamine (0.1-10 mg/kg iv) produced a marked potentiation of vagally induced bronchoconstriction. These results confirm previous findings. In the challenged guinea pigs, pilocarpine did not inhibit vagally induced bronchoconstriction. Furthermore, gallamine did not potentiate vagally induced bronchoconstriction to the same degree as in the controls. In the group of animals that was sensitized but not challenged, the potentiation of vagally induced bronchoconstriction by gallamine was identical to the controls. There was no increase in baseline Ppi in the sensitized or challenged animals compared with the controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

U-60,257 inhibits O3-induced bronchial hyperreactivity in the guinea pig

We studied the effects on ozone-induced airway hyperreactivity of U-60,257, a pyrroloprostacyclin shown to inhibit leukotriene C/D biosynthesis in vitro. A group of 5 guinea pigs were pretreated with U-60,257 (5 mg/kg IV), and studied before and 30 min after a 15 min exposure to 3.0 ppm ozone. These animals were compared to a similarly exposed group that was untreated (n = 10). Reactivity was determined by measuring specific airway resistance (SRaw) upon intravenous acetylcholine infusion in unanesthetized, spontaneously breathing animals. Prior to ozone exposure, we found that U-60,257 treatment did not affect either SRaw or muscarinic reactivity. After exposure to 3.0 ppm, all untreated guinea pigs showed substantial muscarinic hyperreactivity. In contrast, no significant change in SRaw or muscarinic reactivity occurred after ozone in any animal pretreated with U-60,257. We conclude that ozone-induced bronchial hyperreactivity in the guinea pig rapidly develops after a brief, high level exposure. This effect may be mediated, in part, by leukotrienes generated upon ozone exposure.     

Role of insulin in antigen-induced airway eosinophilia and neuronal M2 muscarinic receptor dysfunction

In the lungs, neuronalM₂ muscarinic receptors limit AChrelease from parasympathetic nerves. In antigen-challenged animals, eosinophil proteins block these receptors, resulting in increased AChrelease and vagally mediated hyperresponsiveness. In contrast, diabeticrats are hyporesponsive and have increasedM₂ receptor function. Becausethere is a low incidence of asthma among diabetic patients, weinvestigated whether diabetes protects neuronalM₂ receptor function inantigen-challenged rats. Antigen challenge of sensitized rats decreasedM₂ receptor function, increasedvagally mediated hyperreactivity by 75%, and caused a 10-fold increase in eosinophil accumulation around airway nerves. In antigen-challenged diabetic rats, neuronal M₂receptor function was preserved and there was no eosinophilaccumulation around airway nerves. Insulin treatment of diabetic ratscompletely restored loss of M₂receptor function, vagally mediated hyperresponsiveness, andeosinophilia after antigen challenge. These data demonstrate thatinsulin is required for development of airway inflammation, loss ofneuronal M₂ muscarinic receptorfunction, and subsequent hyperresponsiveness in antigen-challenged ratsand may explain decreased incidence of asthma among diabetic humans.

     

Glutamate receptors and the airways hyperreactivity

It is proposed the link between the hyperactivity of NMDA receptors and airway hyperresponsiveness. We investigated the effect of agents modulating the activity of NMDA receptors in the ovalbumin-induced airway hyperreactivity in guinea pigs. The airways hyperreactivity was influenced by the agonist (NMDA) and selective antagonist - competitive (AP-5) and non-competitive (MK-801) of NMDA receptors. Airway responsiveness to histamine or acetylcholine was evaluated in in vitro conditions. NMDA administration caused the increase of tracheal smooth muscle response in ovalbumin-induced hyperreactivity to acetylcholine. MK 801 as well as AP-5 provoked the decrease of reactivity mainly to acetylcholine in tracheal smooth muscle, while the former, non-competitive antagonist was more effective. We recorded more pronounced response in tracheal than in lung tissue smooth muscle with more considerable response to acetylcholine than to histamine. The results of experiments show the modification of airway smooth muscles responses by agents modulating the activity of NMDA receptors. They confirm the possibility of NMDA receptors participation in experimental airway hyperreactivity. The results enlarge information regarding the link of the inflammatory diseases and glutamatergic system.