Influence of the ventral surface of the medulla on tracheal responses to CO2

These studies investigated the role of the intermediate area of the ventral surface of the medulla (VMS) in the tracheal constriction produced by hypercapnia. Experiments were performed in chloralose-anesthetized, paralyzed, and artificially ventilated cats. Airway responses were assessed from pressure changes in a bypassed segment of the rostral cervical trachea. Hyperoxic hypercapnia increased tracheal pressure and phrenic nerve activity. Intravenous atropine pretreatment or vagotomy abolished the changes in tracheal pressure without affecting phrenic nerve discharge. Rapid cooling of the intermediate area reversed the tracheal constriction produced by hypercapnia. Graded cooling produced a progressive reduction in the changes in maximal tracheal pressure and phrenic nerve discharge responses caused by hypercapnia. Cooling the intermediate area to 20 degrees C significantly elevated the CO2 thresholds of both responses. These findings demonstrate that structures near the intermediate area of the VMS play a role in the neural cholinergic responses of the tracheal segment to CO2. It is possible that neurons or fibers in intermediate area influence the motor nuclei innervating the trachea. Alternatively, airway tone may be linked to respiratory motor activity so that medullary interventions that influence respiratory motor activity also alter bronchomotor tone.     

Influence of ventrolateral surface of medulla on tracheal gland secretion.

Airway secretion can be modified reflexly as well as locally. Previous studies indicate that neurons in a circumscribed region near the ventral surface of the medulla (VMS) can substantially modify airway tone and reflex responses to vagal inputs. In the present studies we assessed the importance of these neurons on tracheal gland secretion. We examined the changes in the number of hillocks of secretion appearing from submucosal glands in an exposed field of tracheal epithelium (1.2 cm2) coated with tantalum dust before and after interventions on the VMS. Experiments were performed in alpha-chloralose-anesthetized dogs paralyzed and ventilated with 40% O2. Stimulation of nicotinergic receptors by application of a pledget containing nicotine in 11 dogs caused a significant elevation in tracheal gland secretion in the subsequent 60 s, compared with a control period in which buffered saline was applied. Prior application of lidocaine or hexamethonium bromide to the VMS blocked the effect of topically applied nicotine. The central effects of nicotine were diminished by atropine methylnitrate given intravenously. In addition, lidocaine application to the VMS or focal cooling of intermediate areas to between 20 and 15 degrees C significantly decreased secretion rates reflexly produced by capsaicin-induced stimulation of pulmonary C-fiber receptors and by mechanical stimulation of the carina and larynx. These findings suggest that the ventral medulla contains cells near its surface that influence tracheal fluid secretion and modulate reflex responses of airway submucosal glands, probably by altering the level of general excitation within the central respiratory integrating circuits.     

Effect of N-methyl-D-aspartate applied to the ventral surface of the medulla on the trachea

Structures located near the ventral surface of the medulla (VMS) affect both cardiovascular tone and respiratory activity. In addition cooling the intermediate area of the VMS blocks the increases in parasympathetic activity and tracheal tone resulting from ventilation with hypercapnic or hypoxic gas mixtures, or due to stimulation of mechanoreceptors within the lung. Since cooling the surface of the VMS may affect fibers of passage as well as cell bodies, we performed studies in which pledgets containing N-methyl-D-aspartic acid (NMDA), a synthetic excitatory amino acid, were applied to intermediate area of the VMS. The studies were performed in chloralose-anesthetized, artificially ventilated cats. Application of pledgets containing NMDA (10(-7) mol at 10(-3) M) caused increases in tracheal pressure and the onset of phasic phrenic activity, but application of 10(-8) mol at 10(-4) M of NMDA could produce tracheal constriction without the appearance of phasic phrenic activity. Applying to the entire VMS either 2-amino-5-phosphonovalerate (2-APV, 10(-6) M), a specific antagonist to NMDA, or lidocaine (2%), a local anesthetic, 60 s before the application of pledgets containing NMDA, prevented the increase in tracheal tone and phasic phrenic activity. Intravenous administration of atropine methyl nitrate 0.5 mg/kg, a cholinergic antagonist, blocked tracheal responses to local application of pledgets containing NMDA but did not affect the increase in phasic phrenic nerve activity. These findings suggest that when stimulated, neurons near the surface of the VMS in the vicinity of the intermediate area increase the activity of parasympathetic fibers to the airway.     

Absence of nonadrenergic noncholinergic relaxation in the cat cervical trachea

Published in vivo experiments have not supported in vitro reports of the presence of nonadrenergic noncholinergic (NANC) inhibitory pathways in the cat trachea. We therefore examined these pathways, measuring tension in an innervated tracheal segment, flow resistance in more distal airways, and dynamic compliance, in 10 anesthetized mechanically ventilated cats. Initially, cervical vagal stimulation evoked contraction followed by relaxation of smooth muscle of trachea and lower airways; sympathetic stimulation evoked relaxation only. After muscarinic blockade and restoration of smooth muscle tone with 5-hydroxytryptamine (5-HT) applied topically to the tracheal mucosa, vagal stimulation did not affect tracheal segment tension, whereas sympathetic-evoked relaxation was preserved. Similar results were found when tone was restored with intravenous 5-HT, with vagal stimulation also decreasing resistance and increasing compliance. We conclude that NANC pathways are present in lower airways but not in the cervical trachea of the cat. We hypothesize that parasympathetic constriction of cat airway smooth muscle can occur without simultaneous NANC activation, whereas NANC activity occurs only in tandem with parasympathetic stimulation.     

Influence of respiratory drive on airway responses to excitation of lung C-fibers

To determine whether the responses of tracheal smooth muscle and the nasal vasculature to stimulation of lung C-fiber receptors depend on the level of respiratory drive, the effects of right atrial injection of capsaicin and phenyldiguanide were studied in chloralose-anesthetized, paralyzed, artificially ventilated cats. Studies were performed while the animals were hyperventilated to apnea and, in addition, when breathing was stimulated by inhalation of 7% CO2 or by N-methyl-D-aspartic acid (NMDA) applied to the ventral surface of the medulla. When the cats were hyperventilated to apnea with O2, injection of capsaicin into the right atrium increased tracheal tone and slightly raised nasal resistance. However, when the animals were ventilated with 7% CO2 in O2 or respiratory activity was stimulated by the application of NMDA, administration of capsaicin eliminated spontaneous phrenic nerve activity and caused an abrupt decrease in tracheal tone but still increased nasal resistance. Similar responses were also obtained with right atrial injection of phenyldiguanide. These results showed for the first time that in the cat the direction of the reflex effects on tracheal tone but not nasal resistance depends on the preexisting level of respiratory drive and on cholinergic activity to airway smooth muscle.     

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.     

Functional anatomy of the vagal innervation of the cervical trachea of the dog.

The canine cervical trachea has been used for numerous studies regarding the neural control of tracheal smooth muscle. The purpose of the present study was to determine whether there is lateral dominance by either the left or right vagal innervation of the canine cervical trachea. In anesthetized dogs, pressure in the cuff of the endotracheal tube was used as an index of smooth muscle tone in the trachea. After establishment of tracheal tone, as indicated by increased cuff pressure, either the right or left vagus nerve was sectioned followed by section of the contralateral vagus. Sectioning the right vagus first resulted in total loss of tone in the cervical trachea, whereas sectioning the left vagus first produced either a partial or no decrease in tracheal tone. After bilateral section of the vagi, cuff pressure was recorded during electrical stimulation of the rostral end of the right or left vagus. At the maximum current strength used, stimulation of the left vagus produced tracheal constriction that averaged 28.5% of the response to stimulation of the right vagus (9.0 +/- 1.8 and 31.6 +/- 2.5 mmHg, respectively). In conclusion, the musculature of cervical trachea in the dog appears to be predominantly controlled by vagal efferents in the right vagus nerve.     

Studies of mediator involvement in cooling-induced alterations of guinea pig tracheal smooth muscle contractility

Heat loss from airway smooth muscle is a potent stimulus which causes substantial, but poorly understood, alterations in muscle tension. This study considered the involvement of endogenous mediators in cooling-induced tension changes in incubated guinea pig trachea. Smooth muscle tension was monitored in tracheal cylinders which were carefully cooled from 37 to 30 degrees C in the presence or absence of various inotropic mediators. In our study, cooling alone, at a rate of 1 degree C/min, was associated with an average loss of smooth muscle tension of 88.2 mg. Cooling tracheal tissue that had been previously exposed to 3 X 10(-6) M histamine, however, caused an additional increase in tracheal tension of 133 mg, over and above that caused by histamine alone. In the presence of 10(-5) M prostaglandin F2 alpha, or 10(-5) M thromboxane B2, cooling was associated with respective losses of smooth muscle tension of 211.4 and 211.2 mg, as compared to the tension associated with these mediators when they were used alone under control conditions. When the speed of tracheal cooling was increased to 40 degrees C/min, there was a slight increase in tension for 20 sec followed by a pronounced and sustained relaxation. The mechanisms involved in the response of airway smooth muscle to cooling are complex. The results of our study, however, suggest that mediators may play a role in the cooling-induced alterations of airway smooth muscle tension.     

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.     

Multiple mechanisms of reflex bronchospasm in guinea pigs.

The mechanisms of histamine- and bradykinin-induced reflex bronchospasm were determined in anesthetized guinea pigs. With intravenous administration, both autacoids evoked dose-dependent increases in tracheal cholinergic tone. Vagotomy or atropine prevented these tracheal reflexes. When delivered as an aerosol, bradykinin readily increased tracheal cholinergic tone, whereas histamine aerosols were much less effective at inducing tracheal reflexes. Also, unlike histamine, bradykinin could evoke profound increases in cholinergic tone without directly or indirectly (e.g., prostanoid dependent) inducing measurable airway smooth muscle contraction resulting in bronchospasm. Neither autacoid required de novo synthesis of prostanoids or nitric oxide to induce reflex tracheal contractions. Combined cyclooxygenase inhibition and tachykinin-receptor antagonism did, however, abolish all effects of bradykinin in the airways, whereas responses to histamine were unaffected by these pretreatments. The data indicate that histamine and bradykinin initiate reflex bronchospasm by differential activation of vagal afferent nerve subtypes. We speculate that selective activation of either airway C fibers or airway rapid adapting receptors can initiate reflex bronchospasm.     

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.     

Nasal and tracheal responses to chemical and somatic afferent stimulation in anesthetized cats

Respiratory chemical and reflex interventions have been shown to affect nasal resistance or tracheal tone, respectively. In the present study, nasal caliber (assessed from pressure at a constant flow) and tracheal tone (assessed from pressure in a fluid-filled balloon within an isolated tracheal segment) were monitored simultaneously in anesthetized, paralyzed, artificially ventilated (inspired O2 fraction = 100%) cats. We examined the effect of CO2 inhalation and sciatic nerve stimulation as well as the application of nicotine (6 X 10(-4) mol/l) or lidocaine (2% solution) to the intermediate area of the ventral medullary surface (VMS). CO2 and VMS nicotine resulted in a significant increase in tracheal pressure [147 +/- 73 and 91 +/- 86% (SD), respectively]; and a significant reduction in nasal pressure (-35 +/- 10 and -20 +/- 13%, respectively). In contrast, sciatic nerve stimulation resulted in a significant fall in both tracheal (-50 +/- 36%) and nasal pressure (-21 +/- 13%). Application of 2 or 4% lidocaine to the VMS reduced tracheal pressure but did not significantly affect nasal pressure. After VMS lidocaine, nasal and tracheal responses to CO2, sciatic nerve stimulation, or VMS nicotine, when present, were negligible. These results suggest a role for the VMS in the regulation and coordination of nasal and tracheal caliber responses.     

Effect of histamine on duration of phases of the respiration cycle

Histamine produced either a bronchodilation or a bronchoconstriction in rats. In a 0.01-1.0 mcg/ml concentration histamine augmented the contractions amplitude in electrical stimulation of the trachea, in a 10-100 mcg/ml concentration histamine enhances the muscle tone thus decreasing the induced contractions. The histamine effects seems to be connected with its prevailing influence on different structures of the airways depending on the concentration. Its high concentrations act directly on the smooth muscle whereas its lower concentration affects receptors signaling the functional modules of the metasympathetic nervous system within the intramural ganglia of the trachea.     

Is the epithelium-derived inhibitory factor in guinea-pig trachea a prostanoid?

To examine further the possible prostanoid involvement in the influence of the epithelium on guinea-pig tracheal smooth muscle responsiveness, we have analyzed the effects of LTD4, methacholine and histamine on the level of airway smooth muscle tone and on the amounts of PGE2, PGF2 alpha and PGI2 (determined by radioimmunoassay) in the presence and absence of the epithelium. Removal of the epithelium increased the sensitivity of guinea-pig trachea to the contractile effects of LTD4, methacholine and histamine. LTD4 (3-100 nM), methacholine (0.1-10 microM) or histamine (0.3-30 microM) did not increase prostanoid release above control values in either the presence or absence of the epithelium. The unstimulated release of PGE2 and PGF2 alpha, but not PGI2, was decreased in tissues lacking epithelium. Indomethacin (1 microM) reduced the baseline tone to a smaller extent in the absence of epithelium. In the presence but not the absence of the epithelium, indomethacin increased the sensitivity of preparations to the contractile effect of methacholine. The results support the postulate of an epithelium-derived inhibitory factor modulating guinea-pig tracheal smooth muscle responsiveness. The identity of this factor is not known but is not PGI2 and is unlikely to be PGF2 alpha or PGE2. However, the possibility remains that the basal release of PGE2 and/or PGF2 alpha derived from the epithelium may markedly affect the responsiveness of guinea-pig tracheal smooth muscle. Furthermore, the epithelium is a significant source of PGE2 and PGF2 alpha which may be involved in the maintenance of baseline tone.     

Neuronal histamine release elicited by hyperthermia mediates tracheal dilation and pressor response

Whether brain histaminergic neurons contribute to the regulation of tracheal tone and peripheral vascular tone under hyperthermia was investigated in anesthetized rabbits. Histamine release from the rostral ventrolateral medulla (RVLM), the raphe nuclei, and the solitary nucleus of the medulla oblongata was significantly increased by hyperthermia. The increased histamine was significantly suppressed by 10(-6) M tetrodotoxin microdialyzed in each area. Tracheal pressure and mean arterial pressure were significantly decreased and increased by hyperthermia, respectively. An H(1)-receptor antagonist, 5 x 10(-6) M (+)-chlorpheniramine, bilaterally microdialyzed in the RVLM significantly enhanced histamine release in the RVLM as well as significantly suppressed tracheal dilation and pressor response caused by hyperthermia. These data indicate that histamine release in the medulla oblongata is enhanced by hyperthermia. The enhanced histamine is the neuronal origin and the cause of tracheal dilation and pressor response at least via H(1) receptors in the RVLM. Brain histaminergic neurons play important roles in tracheal tone and peripheral vascular tone via H(1) receptors in the RVLM and homeostasis on body temperature.     

Receptors and pathways mediating the effects of prostaglandin E2 on airway tone

Prostaglandin E(2) (PGE(2)) has complex effects on airway tone, and the existence of four PGE(2) [E-prostanoid (EP)] receptors, each with distinct signaling characteristics, has provided a possible explanation for the seemingly contradictory actions of this lipid mediator. To identify the receptors mediating the actions of PGE(2) on bronchomotor tone, we examined its effects on the airways of wild-type and EP receptor-deficient mice. In conscious mice the administration of PGE(2) increased airway responsiveness primarily through the EP1 receptor, although on certain genetic backgrounds a contribution of the EP3 receptor was detected. These effects of PGE(2) were eliminated by pretreatment with either atropine or bupivacaine and were undetectable in anesthetized mice or in denervated tracheal rings, where only EP2-mediated relaxation of airway smooth muscle was observed. Together, our findings are consistent with a model in which PGE(2) modulates airway tone by activating multiple receptors expressed on various cell populations and in which the relative contribution of these receptors might depend on the expression of modifier alleles. PGE(2)/EP1/EP3-induced airway constriction occurs indirectly through activation of neural pathways, whereas PGE(2)-induced bronchodilation results from direct activation of EP2 receptors on airway smooth muscle. This segregation of EP receptor function within the airway suggests that PGE(2) analogs that selectively activate the EP2 receptor without activating the EP1/EP3 receptors might prove useful in the treatment of asthma.     

Tracheal hysteresis in sleep apnea

The collapsibility of pharyngeal walls, characteristic of patients with obstructive sleep apnea, likely results from reduced tone of the pharyngeal muscles. This reduction in the upper airway muscle tone may not end at the pharynx but may extend further distally, e.g., into the trachea. Because tracheal tone cannot be measured directly in conscious humans, we inferred the tone from the relative hysteresis of the tracheal area compared with the lung. Relative hysteresis was measured by plotting the cross-sectional area of a tracheal segment obtained by the acoustic reflection technique vs. lung volume. All measurements were performed during wakefulness. We found that in 42 patients with obstructive sleep apnea (apnea/hypopnea index greater than 10), relative hysteresis of the proximal trachea was predominantly clockwise, i.e., smaller than that of the lung parenchyma; in the 33 nonapneic patients (apnea/hypopnea index less than or equal to 10), it was predominantly counter-clockwise, i.e., larger than that of the lung parenchyma. For the distal trachea all patients, apneic and nonapneic, had similar, clockwise, relative hysteresis. We conclude that reduction in the upper airway muscle tone in patients with obstructive sleep apnea extends into the trachea.     

Nicotine-induced respiratory effects of cigarette smoke in dogs

We report that nicotine is responsible for both a blood-borne stimulation of the respiratory center and a direct effect on intrathoracic airway tone in dogs. We introduced cigarette smoke into the lungs of donor dogs and injected arterial blood obtained from them into the circulation of recipient dogs to show that a blood-borne material increased breathing and airway smooth muscle tone. Smoke from cigarettes containing 2.64 mg of nicotine was effective; that from cigarettes containing 0.42 mg of nicotine was not. Nicotine, in doses comparable to the amounts absorbed from smoke, also increased breathing and tracheal smooth muscle tension when injected into the vertebral circulation of recipient dogs. Finally, blockade of nicotine receptors in the central nervous system and in the airway parasympathetic ganglia inhibited the effects of inhaled cigarette smoke and intravenous nicotine on the respiratory center and on bronchomotor tone. We conclude that nicotine absorbed from cigarette smoke is the main cause of cigarette smoke-induced bronchoconstriction. It caused central respiratory stimulation, resulting in increased breathing and airway smooth muscle tension, and had a direct effect on airway parasympathetic ganglia as well.     

Developmental expression of neurokinin A and functional neurokinin-2 receptors in lung

Peribronchial smooth muscle constriction causes airway stretch, an important mechanical force in developing lung. Little is known about factors influencing these spontaneously active muscle elements. We measured contractile activity of neurokinin (NK) receptors on fetal intrapulmonary smooth muscle by tracheal perfusion assay (n = 11). Injecting either capsaicin or the NK(2) receptor agonist [NLE(10)]NKA resulted in significant (P < 0.05) bronchoconstriction. A specific NK(2) receptor antagonist inhibited constriction caused by endogenous tachykinins released by capsaicin. We then examined NK(2) receptor (n = 44) and NKA (n = 23) ontogeny in human lung. NKA immunostaining was identified in peribronchial nerves in samples with gestational age >12 wk. NK(2) receptor protein was identified in peribronchial and perivascular smooth muscle. These results indicate that endogenous tachykinins released by the developing lung act via NK(2) receptors to cause smooth muscle constriction. We speculate that tachykinins could modulate lung development.