Competition of NO synthases and arginase in the airways hyperreactivity

The competition between arginases and NO synthases (NOS) for their common substrate L-arginine can be important in the airways hyperreactivity. We investigated the effect of the simultaneous modulation of arginase and NOS activities in allergen-induced airways hyperreactivity. We analysed the response of tracheal and lung tissue smooth muscle to histamine or acetylcholine after administration N(ω)-nitro-L-arginine methyl ester (L-NAME), aminoguanidine (AG) and N(ω)-hydroxy-L-arginine (NOHA) in the combinations in in vitro conditions. The results show the decrease of ovalbumin-induced hyperreactivity after inhibition of arginase activity with NOHA. A supplementation of L-arginine caused favourable effect on the airway smooth muscle response. We found the airway reactivity decrease on the whole if we used the combination of NOS and arginase inhibitors. The inhibition of both types of enzymes caused more expressive effect in tracheal smooth muscles. We recorded the difference in the response to histamine or acetylcholine. The simultaneous inhibition of iNOS (with AG) and arginase (with NOHA) evoked the most expressive effect. Results show the importance of competition of both types enzymes - NOS and arginase for the balance of theirs activities in the control of airways bronchomotoric tone in the conditions of the airways hyperreactivity.     

N-methyl-D-aspartate receptor antagonists enhance histamine neuron activity in rodent brain

The modulation of histamine neuron activity by various non-competitive NMDA-receptor antagonists was evaluated by changes in tele-methylhistamine (t-MeHA) levels and histidine decarboxylase (hdc) mRNA expression induced in rodent brain. The NMDA open-channel blockers phencyclidine (PCP) and MK-801 enhanced t-MeHA levels in mouse brain by 50-60%. Ifenprodil, which interacts with polyamine sites of NR2B-containing NMDA receptors, had no effect. PCP also increased hdc mRNA expression in the rat tuberomammillary nucleus. The enhancement of t-MeHA levels elicited by MK-801 (ED50 of approximately 0.1 mg/kg) was observed in the hypothalamus, cerebral cortex, striatum and hippocampus. Control t-MeHA levels and the t-MeHA response to MK-801 were not different in male and female mice. Double immunostaining for HDC and NMDA receptor subunits showed that histamine neurons of the rat tuberomammillary nucleus express NMDA receptor subunit 1 (NR1) with NMDA receptor subunit 2A (NR2A) and NMDA receptor 2B subunit (NR2B). In addition, immunoreactivity for the neuronal glutamate transporter EAAC1 was observed near most histaminergic perikarya. Hence, these findings support the existence of histamine/glutamate functional interactions in the brain. The increase in histamine neuron activity induced by NMDA receptor antagonists further suggests a role of histamine neurons in psychotic disorders. In addition, the decrease in MK-801-induced hyperlocomotion observed in mice after administration of ciproxifan further strengthens the potential interest of H3-receptor antagonist/inverse agonists for the symptomatic treatment of schizophrenia.     

Interleukin-1beta-induced airway hyperresponsiveness enhances substance P in intrinsic neurons of ferret airway

Interleukin (IL)-1beta causes airway inflammation, enhances airway smooth muscle responsiveness, and alters neurotransmitter expression in sensory, sympathetic, and myenteric neurons. This study examines the role of intrinsic airway neurons in airway hyperresponsiveness (AHR) induced by IL-1beta. Ferrets were instilled intratracheally with IL-1beta (0.3 microg/0.3 ml) or saline (0.3 ml) once daily for 5 days. Tracheal smooth muscle contractility in vitro and substance P (SP) expression in tracheal neurons were assessed. Tracheal smooth muscle reactivity to acetylcholine (ACh) and methacholine (MCh) and smooth muscle contractions to electric field stimulation (EFS) both increased after IL-1beta. The IL-1beta-induced AHR was maintained in tracheal segments cultured for 24 h, a procedure that depletes SP from sensory nerves while maintaining viability of intrinsic airway neurons. Pretreatment with CP-99994, an antagonist of neurokinin 1 receptor, attenuated the IL-1beta-induced hyperreactivity to ACh and MCh and to EFS in cultured tracheal segments. SP-containing neurons in longitudinal trunk, SP innervation of superficial muscular plexus neurons, and SP nerve fiber density in tracheal smooth muscle all increased after treatment with IL-1beta. These results show that IL-1beta-enhanced cholinergic airway smooth muscle contractile responses are mediated by the actions of SP released from intrinsic airway neurons.     

Protease-activated receptor 2 in regulation of bronchomotor tone: effect of tobacco smoking

Protease-activated receptors are G protein-coupled receptors activated by serine-proteases. Protease-activated receptor 2 is involved in the regulation of airway smooth muscle tone but its effects vary according to species and experimental conditions. We determined the effects of protease-activated receptor 2 activation on smooth muscle tone and airway reactivity to histamine in guinea pigs and smoking or non-smoking humans. The effects of trypsin and protease-activated receptor activating peptide on the isometric tension and response to histamine of guinea pig tracheal and human bronchial rings were studied. Human tissues were obtained from 6 smokers and 4 non-smokers. We assessed the effects of epithelial removal, inhibitors of cyclooxygenases, nitric oxide synthases, neutral endopeptidase and antagonists of acetylcholine, histamine, bradykinin and tachykinin receptors. Bronchomotor responses to protease-activated receptor 2 activation were variable in guinea pig, in half of animals PAR2 activation induced smooth muscle relaxation through the epithelial release of prostanoids but not of nitric oxide. In human airways, protease-activated receptor 2 activation reduced responsiveness to histamine in bronchial rings from smokers but increased responsiveness in bronchi from non-smokers. This study demonstrates an influence of tobacco smoking on the effect of protease-activated receptor 2 activation on airway responsiveness in humans, with an increased protection against histamine-induced contractions, probably through an increased epithelial release of prostanoids. The role of airway protease-activated receptor 2 may be to maintain smooth muscle tone homeostasis.     

Epithelium modulates the reactivity of ovalbumin-sensitized guinea-pig airway smooth muscle

Mechanical removal of the airway epithelium alters the in vitro reactivity of airway smooth muscle. The modulation of reactivity may involve the release of inhibitory and excitatory factors from epithelial cells. Guinea pigs sensitized with ovalbumin have been used as an animal model of airway hyperreactivity. We evaluated the influence of the epithelium on the reactivity of in vitro tracheal smooth muscle from control and ovalbumin-sensitized guinea pigs, and the extent to which the presence of the epithelium affects the contractile response to in vitro challenge with ovalbumin. In both control and ovalbumin-sensitized tissues, epithelium removal increased the sensitivity of the preparations to histamine, methacholine and isoproterenol to a similar extent, i.e., 2- to 2.5-fold. Epithelium removal resulted in an 8.1-fold increase in sensitivity to ovalbumin in sensitized tissues. The epithelium appears not only to modulate the reactivity of the tissues to bronchoactive agents, but it also influences the magnitude of the contractile response following antigen challenge.     

Influence of ventrolateral surface of medulla on reflex tracheal constriction

To assess the role of structures located superficially near the ventrolateral surface of the medulla on the reflex constriction of tracheal smooth muscle that occurs when airway and pulmonary receptors are stimulated mechanically or chemically, experiments were conducted in alpha-chloralose-anesthetized, paralyzed, and artificially ventilated cats. Pressure changes within a bypassed segment of the trachea were used as an index of alterations smooth muscle tone. The effects of focal cooling of the intermediate areas or topically applied lidocaine on the ventral surface of the medulla on the response of the trachea to mechanical and chemical stimulation of airway receptors were examined. Atropine abolished tracheal constriction induced by mechanical stimulation of the carina or aerosolized histamine, showing that the responses were mediated over vagal pathways. Moderate cooling of the intermediate area (20 degrees C) or local application of lidocaine significantly decreased the tracheal constrictive response to mechanical activation of airway receptors. Furthermore, when the trachea was constricted by histamine, cooling of the intermediate area significantly diminished the increased tracheal tone, whereas rewarming restored tracheal tone to the previous level. These findings suggest that under the conditions of the experiments the ventral surface of the medulla plays an important role in constriction of the trachea by inputs from intrapulmonary receptors and in the modulation of parasympathetic outflow to airway smooth muscle.     

Activation of endogenous GABAA channels on airway smooth muscle potentiates isoproterenol-mediated relaxation

Reactive airway disease predisposes patients to episodes of acute smooth muscle mediated bronchoconstriction. We have for the first time recently demonstrated the expression and function of endogenous ionotropic GABA(A) channels on airway smooth muscle cells. We questioned whether endogenous GABA(A) channels on airway smooth muscle could augment beta-agonist-mediated relaxation. Guinea pig tracheal rings or human bronchial airway smooth muscles were equilibrated in organ baths with continuous digital tension recordings. After pretreatment with or without the selective GABA(A) antagonist gabazine (100 muM), airway muscle was contracted with acetylcholine or beta-ala neurokinin A, followed by relaxation induced by cumulatively increasing concentrations of isoproterenol (1 nM to 1 muM) in the absence or presence of the selective GABA(A) agonist muscimol (10-100 muM). In separate experiments, guinea pig tracheal rings were pretreated with the large conductance K(Ca) channel blocker iberiotoxin (100 nM) after an EC(50) contraction with acetylcholine but before cumulatively increasing concentrations of isoproterenol (1 nM to 1 uM) in the absence or presence of muscimol (100 uM). GABA(A) activation potentiated the relaxant effects of isoproterenol after an acetylcholine or tachykinin-induced contraction in guinea pig tracheal rings or an acetylcholine-induced contraction in human endobronchial smooth muscle. This muscimol-induced potentiation of relaxation was abolished by gabazine pretreatment but persisted after blockade of the maxi K(Ca) channel. Selective activation of endogenous GABA(A) receptors significantly augments beta-agonist-mediated relaxation of guinea pig and human airway smooth muscle, which may have important therapeutic implications for patients in severe bronchospasm.     

Induction of rhythmic contraction in canine tracheal smooth muscle

Na(+)-K+ ATPase activity of the canine tracheal smooth muscle membrane is responsible for the electrogenic pumping of Na+ and K+ ions. It has been shown that this activity results in muscle relaxation. Based on the results of the current study, we suggest that prolonged electrical stimulation induces increased Na(+)-K+ ATPase activity in isolated tracheal smooth muscle. Tracheal smooth muscle pretreated with prolonged electrical stimulation developed graded mechanical activity when subsequently treated with histamine, serotonin, acetylcholine, or 80 mM K+. This increased isometric tension was interrupted by rhythmic activity, which was elicited by histamine or serotonin but not by acetylcholine or 80 mM K+ stimulation. The spontaneous phasic activity was not inhibited by atropine or propranolol but was totally inhibited by 10(-6) M ouabain. These results suggested that the relaxation phase of rhythmic contraction in response to histamine and serotonin stimulation could be the result of stimulated Na(+)-K+ ATPase activity.     

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.     

NMDA receptor antagonism produces antinociception which is partially mediated by brain opioids and dopamine.

Inhibition of nitric oxide synthase (NOS) activity results in opioid-mediated supraspinal analgesia in the rat, as indicated by increased reaction time in the hot plate test. It is documented that a relationship exists between NMDA receptor activation and the activity of NOS. The present investigation sought to determine if inactivation of the NMDA receptor produced antinociception of supraspinal origin, as was observed in response to inhibition of NOS, and if this response was mediated by brain opioids, by activation of receptors for the neurotransmitter, dopamine, or both. Administration of MK-801, a non-competitive antagonist of the NMDA receptor, produced significant antinociception as measured by reaction time in the hot plate test of analgesia. Antinociception resulting from treatment with MK-801 appeared to be mediated by brain opioids, as indicated by the ability of the opioid antagonist, naloxone, to partially reverse the effect of MK-801 administration. This analgesic response was also partially diminished by administration of the dopamine D1 receptor antagonist, SCH 23390 and the dopamine D2 receptor antagonist, sulpiride. The analgesia resulting from NMDA receptor antagonism was found to be only partially attributable to dopamine and brain opioids, since co-administration of naloxone and SCH 23390 or naloxone and sulpiride, were unable to completely reverse the antinociceptive response to MK-801. The present findings suggest that inhibition of NMDA receptor activity produces supraspinal analgesia. Furthermore, it appears that antinociception induced by blockade of the NMDA receptor results, at least in part, from activation of endogenous brain opioids and stimulation of D1 and D2 subtypes of the dopamine receptor.     

Interaction between histamine and vagal stimulation on tracheal smooth muscle in dogs

We examined the interaction between histamine and vagal efferent activity on airway smooth muscle reactivity in 11 anesthetized vagotomized dogs using an isolated closed segment of the intrathoracic trachea filled with Tyrode solution under an isovolumetric condition. Intratracheal pressure change was measured as an index of tracheal smooth muscle tone. The administration into the tracheal segment of histamine (0.1 or 1.0 mg/ml) in six dogs and methacholine chloride (0.001 or 0.01 mg/ml) in the other five dogs elevated intratracheal pressure by about 5 cmH2O. The electrical stimulation of the peripheral ends of both of the cut cervical vagus nerves in the presence of histamine produced significantly greater responses than the additive responses of these two stimuli applied individually (two-way analysis of variance, P less than 0.025). However, the combined effects of vagal stimulation and methacholine were not significantly different from the additive responses of these two stimuli applied individually. The average values of intratracheal pressure elevated by the combined effects of vagal stimulation and histamine were significantly higher than those obtained by the combination of vagal stimulation and methacholine (two-way analysis of variance, P less than 0.01). This suggests that histamine potentiates tracheal smooth muscle reactivity to electrical vagal stimulation, which may contribute to the hyperreactivity observed in patients with asthma.     

ACE-inhibitors and defence reflexes of the airways.

The group of angiotensin-converting enzyme (ACE) inhibitors is one of the drugs of choice for the treatment of hypertension and congestive heart disease. However, it has been reported that in some of patients ACE-inhibitors induce hyperreactivity of the airways with occurrence of a persistent dry cough, dyspnoe and wheezing. We supposed that the mechanism of these hyperreactivity is connected to accumulation of bradykinin, tachykinins and other inflammatory mediators in the airways. Increased local concentration of inflammatory neuropeptides stimulates bronchial C fibres and rapidly adapting receptors and provoke the cough reflex. Inflammatory processes in the airways could be followed by contraction of airway smooth muscle. In this study, our aim was to measure the changes of the number and intensity of mechanical induced cough in cats, which were treated for days with enalapril (5 mg/kg b.w.). After 15 days of treatment the reactivity of the lung and tracheal smooth muscles to the bronchoconstrictor mediator histamine was estimated. As to our finding 15 days of administration of enalapril results in significant increase of cough parameters measured with a more significant sensitivity of the laryngopharyngeal part. In the experimental animals we observed increased reactivity of bronchial smooth muscle to histamine after 15 days of enalapril treatment. The reactivity of the lung smooth muscle to the histamine was not significantly changed. These results confirmed the increased cough sensitivity and increased bronchial reactivity after enalapril treatment. These experimental animal model may be useful for the investigation of the pharmacological minimization of respiratory adverse effect of ACE-inhibitors.     

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.     

Differential effects of prostaglandin E2 on contractions of airway smooth muscle

In an in vitro muscle bath, the active tension generated by strips of canine tracheal smooth muscle responding to cumulative additions of either histamine (10(-8) to 10(-3) M) or acetylcholine (10(-9) to 10(-3) M) was measured in the absence and presence of prostaglandin E2 (PGE2) (10(-6) to 10(-5) M). When contractile responses of equal magnitude were compared, the contractions elicited by acetylcholine were resistant to the inhibitory effects of PGE2, relative to comparable contractions elicited by histamine. To assess the role of adenylate cyclase in determining the different responses to histamine and acetylcholine in the presence of PGE2, we assayed adenylate cyclase activity in membranes prepared from canine tracheal smooth muscle and found that acetylcholine, but not histamine, decreased PGE2-stimulated adenylate cyclase activity by 48 +/- 2% (mean +/- SE; n = 5). However, in other experiments, we found that even large pharmacological increases in tissue adenosine 3',5'-cyclic monophosphate (cAMP) content only partially inhibited muscarinic tone. Also, exogenously applied analogues of cyclic AMP inhibited contractions induced by histamine more effectively than comparable contractions induced by acetylcholine. We concluded that acetylcholine decreased adenylate cyclase activity in membranes prepared from canine tracheal smooth muscle and that this effect may have contributed to, but did not completely account for, the relative resistance of muscarinic contractions to the inhibitory effects of PGE2.     

Influenza infection causes airway hyperresponsiveness by decreasing enkephalinase

Ferret tracheal segments were infected with human influenza virus A/Taiwan/86 (H1N1) in vitro. After 4 days, the smooth muscle contractile responses to acetylcholine and to substance P were measured. The response to substance P was markedly accentuated, with a threefold increase in force of contraction at a substance P concentration of 10(-5) M, the highest concentration tested. In contrast, the response to acetylcholine was not affected by viral infection. Histological examination of tissues revealed extensive epithelial desquamation. Activity of enkephalinase (neutral metallo-endopeptidase, EC.3.4.24.11), an enzyme that degrades substance P, was decreased by 50% in infected tissues. Inhibiting enkephalinase activity by pretreating with thiorphan (10(-5) M) increased the response to substance P to the same final level in both infected and control tissues. Inhibiting other substance P-degrading enzymes including kininase II (angiotensin-converting enzyme), serine proteases, and aminopeptidases did not affect the response to substance P. Inhibiting cyclooxygenase and lipoxygenase activity using indomethacin and BW 755c did not affect hyperresponsiveness to substance P. Pretreating tissues with antagonists of alpha-adrenoceptors, beta-adrenoceptors, and H1 histamine receptors (phentolamine 10(-5) M, propranolol 5 X 10(-6) M, and pyrilamine 10(-5) M, respectively) had no effect on substance P-induced contraction. These results demonstrate that infection of ferret airway tissues with influenza virus increases the contractile response of airway smooth muscle to substance P. This effect is caused by decreased enkephalinase activity in infected tissues.     

Substance P released from intrinsic airway neurons contributes to ozone-enhanced airway hyperresponsiveness in ferret trachea.

Exposure to ozone (O3) induces airway hyperresponsiveness mediated partly through the release of substance P (SP) from nerve terminals in the airway wall. Although substantial evidence suggests that SP is released by sensory nerves, SP is also present in neurons of airway ganglia. The purpose of this study was to investigate the role of intrinsic airway neurons in O3-enhanced airway responsiveness in ferret trachea. To remove the effects of sensory innervation, segments of ferret trachea were maintained in culture conditions for 24 h before in vitro exposure to 2 parts/million of O3 or air for 1 h. Sensory nerve depletion was confirmed by showing that capsaicin did not affect tracheal smooth muscle responsiveness to cholinergic agonist or contractility responses to electrical field stimulation (EFS). Contractions of isolated tracheal smooth muscle to EFS were significantly increased after in vitro O3 exposure, but the constrictor response to cholinergic agonist was not altered. Pretreatment with CP-99994, an antagonist of the neurokinin 1 receptor, attenuated the increased contraction to EFS after O3 exposure but had no effect in the air exposure group. The number of SP-positive neurons in longitudinal trunk ganglia, the extent of SP innervation to superficial muscular plexus nerve cell bodies, and SP nerve fiber density in tracheal smooth muscle all increased significantly after O3 exposure. The results show that release of SP from intrinsic airway neurons contributes to O3-enhanced tracheal smooth muscle responsiveness by facilitating acetylcholine release from cholinergic nerve terminals.     

The role of striatal glutamate receptors in models of Parkinson's disease

Summary The aim of the study was to examine the effect of antagonists of the NMDA receptor on the parkinsonian-like muscle rigidity in rats. Reserpine and haloperidol increased the muscle resistance of the hind foot to passive movements, as well as the reflex electromyographic (EMG) activity in the gastrocnemius and tibialis anterior muscles. MK-801 (0.32-1.28 mg/kg sc), an uncompetitive antagonist of the NMDA receptor, and L-701,324 (5-40 mg/ kg ip), an antagonist of the glycine site, reduced the muscle tone and the reflex EMG activity enhanced by reserpine or haloperidol. AP-5 (2 and 5 ,g/ 0.5 pl), a competitive antagonist of the NMDA receptor, and 5,7-dichlorokynurenic acid (1.0-4.5g/0.5 pl), the glycine site antagonist injected bilaterally into the rostral striatum, inhibited the muscle rigidity induced by haloperidol. In contrast, AP-5, injected alone bilaterally into the intermediate-caudal striatum induced muscle rigidity. The present results suggest that: (1) the inhibitory effect of the NMDA receptor antagonists on the parkinsonian-like muscle rigidity depends, at least partly, on their action on the rostral striatum; (2) the blockade of NMDA receptors in the intermediate-caudal striatum may reduce the beneficial impact of these compounds.     

The Association of Cortactin with Profilin-1 Is Critical for Smooth Muscle Contraction

Profilin-1 (Pfn-1) is an actin-regulatory protein that has a role in modulating smooth muscle contraction. However, the mechanisms that regulate Pfn-1 in smooth muscle are not fully understood. Here, stimulation with acetylcholine induced an increase in the association of the adapter protein cortactin with Pfn-1 in smooth muscle cells/tissues. Furthermore, disruption of the protein/protein interaction by a cell-permeable peptide (CTTN-I peptide) attenuated actin polymerization and smooth muscle contraction without affecting myosin light chain phosphorylation at Ser-19. Knockdown of cortactin by lentivirus-mediated RNAi also diminished actin polymerization and smooth muscle force development. However, cortactin knockdown did not affect myosin activation. In addition, cortactin phosphorylation has been implicated in nonmuscle cell migration. In this study, acetylcholine stimulation induced cortactin phosphorylation at Tyr-421 in smooth muscle cells. Phenylalanine substitution at this position impaired cortactin/Pfn-1 interaction in response to contractile activation. c-Abl is a tyrosine kinase that is necessary for actin dynamics and contraction in smooth muscle. Here, c-Abl silencing inhibited the agonist-induced cortactin phosphorylation and the association of cortactin with Pfn-1. Finally, treatment with CTTN-I peptide reduced airway resistance and smooth muscle hyperreactivity in a murine model of asthma. These results suggest that the interaction of cortactin with Pfn-1 plays a pivotal role in regulating actin dynamics, smooth muscle contraction, and airway hyperresponsiveness in asthma. The association of cortactin with Pfn-1 is regulated by c-Abl-mediated cortactin phosphorylation.     

Age differences in cholinergic airway responsiveness in relation with muscarinic receptor subtypes

Children seem more susceptible to increased airway reactivity than adults. Such an age-dependent discrepancy in airway reactivity may involve different airway smooth muscle functions. Therefore, we compared the in vivo and in vitro responsiveness of airway smooth muscles between two age groups of animals. Rats of 6 and 21 weeks old were challenged in vivo with acetylcholine (ACh) infused intravenously and airway resistance (R(aw)) was measured. Tracheal muscle was also isolated and the isometric force developed to ACh or KCl was measured. Furthermore, the level of genes encoding muscarinic receptor subtypes (M(1-3)) and acetylcholinesterase (AChE) expressed in the tracheal muscle was determined by RT-PCR. In results, the basal R(aw) was similar in the two age groups. The R(aw) at each ACh dose was significantly greater in young rats than older rats (p<0.05, n=22-27). Tracheal muscles from young rats were more sensitive to ACh than older rats (p<0.05, n=20-21), while receptor-independent muscle contraction to KCl was greater in older rats (p<0.05, n=10-19). Genes encoding AChE, M(2) and M(3) muscarinic receptors were more highly expressed in the tracheal muscles from young than older rats (p<0.05, n=4-6). In conclusion, airway smooth muscle in young rat is more sensitive to cholinergic stimulation in vivo and in vitro compared to older rats, which may be due to a higher expression of M(2) and M(3) muscarinic receptors in airway smooth muscle.     

Maturation of respiratory reflex responses in the piglet

Stimulation of chemo-, irritant, and pulmonary C-fiber receptors reflexly constricts airway smooth muscle and alters ventilation in mature animals. These reflex responses of airway smooth muscle have, however, not been clearly characterized during early development. In this study we compared the maturation of reflex pathways regulating airway smooth muscle tone and ventilation in anesthetized, paralyzed, and artificially ventilated 2- to 3- and 10-wk-old piglets. Tracheal smooth muscle tension was measured from an open tracheal segment by use of a force transducer, and phrenic nerve activity was measured from a proximal cut end of the phrenic nerve. Inhalation of 7% CO2 caused a transient increase in tracheal tension in both age groups, whereas hypoxia caused no airway smooth muscle response in either group. The phrenic responses to 7% CO2 and 12% O2 were comparable in both age groups. Lung deflation and capsaicin (20 micrograms/kg iv) administration did not alter tracheal tension in the younger piglets but caused tracheal tension to increase by 87 +/- 28 and 31 +/- 10%, respectively, in the older animals (both P less than 0.05). In contrast, phrenic response to both stimuli was comparable between ages: deflation increased phrenic activity while capsaicin induced neural apnea. Laryngeal stimulation did not increase tracheal tension but induced neural apnea in both age groups. These data demonstrate that between 2 and 10 wk of life, piglets exhibit developmental changes in the reflex responses of airway smooth muscle situated in the larger airways in response to irritant and C-fiber but not chemoreceptor stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)