The results of our experiments should be interpreted within the context of the test materials used. Our study was simple and effective. An intact tracheal ring was an important component of our technique.9,10
Such an intact tracheal ring is much more representative of a physiological setting than smooth muscle strips. Although it was difficult to determine which tissue component of the trachea was responsible for the drug-induced contraction, the nature of specific tissues and their responses to specific drugs provided some indication. First, the tracheal strips used in our study were crude preparations containing cartilage and tracheal smooth muscle. The smooth muscle of the trachea appeared to be the main tissue component responsible for contraction, because the other components (epithelium, glands, connective tissue, nerves, and cartilage) did not contract to a significant extent. Because this method involved cross-contraction, changes in tension were caused by radial contraction of the tracheal ring. Although responses to drugs and electrical stimulation have been verified for similar preparations,9–13
the contractile response observed in this study was probably an aggregate of the responses of various types of muscle tissue. Second, the isolated tracheal preparations used in our experiments were excised from rats without damaging the endothelium or smooth muscle. Therefore, it is reasonable to assume the tracheal responses to test agents in our study are comparable with those observed after applying an inhaler to the trachea during an asthma attack.
The cholinergic contracting agent tested in this preparation is commonly used for research purposes. Note, cromolyn resulted in no relaxation of the trachea smooth muscle when introduced after applying methacholine. However, the total relaxation of the contracted tracheal strip was observed when adding 10×6
M of atropine. The previously indicated cromolyn had no cholinergic or anticholinergic effect. Thus, it should be possible to assess the effects of common drugs and potential therapeutic agents supposedly not responsible for relieving acute asthma attacks. In addition, basal tension had a minimal effect at various concentrations of cromolyn. There are reports about initial mild bronchoconstriction when using cromolyn in some patients, and it was believed to be secondary to local irritation after inhaling cromolyn.4,5
There was no such effect in this study. This result was compatible with cromolyn basic and clinical entities: it has no effect on airway smooth muscle tone and was ineffective in reversing asthmatic bronchospasm.5
EFS is a common experimental tool activating the nerve terminals within the tissue to be tested and inducing the release of endogenous neurotransmitters, thereby causing the smooth muscle to contract. EFS-induced spike contraction of canine nasal mucosa, which is believed to result from the contraction of vascular smooth muscles, disappeared after ipsilateral cervical sympathetic ganglionectomy.14
Thus, EFS-induced spike contraction of isolated canine nasal mucosa has been proven to be mediated by sympathetic innervation.14
In this study, EFS-induced spike contraction of the tracheal smooth muscle was believed to be caused by stimulation of parasympathetic innervation. Therefore, EFS-induced contraction of the trachea decreased as the cromolyn concentration was high. These findings suggested a mast cell–stabilizing drug, cromolyn, could antagonize the parasympathetic innervation responsible for trachea smooth muscle contraction. Commercial Intal Nebulizer Solution (2 mL, King Pharmaceuticals, Inc, Bristol, TN) contains 20 mg of cromolyn sodium, which is ~2 × 10−2
M cromolyn sodium. When applying an inhalation, one gets immediately a
dilution resulting in a concentration of 2 × 10−3
M of cromolyn sodium at the nasal mucosal side. It remains to be shown that a concentration of 10−4
M can be reached at the tracheal smooth muscles. Therefore, commercial Intal Nebulizer Solution could inhibit parasympathetic function of the trachea. The actual concentration of cromolyn sodium in tracheal smooth muscle when used in an inhalation requires further investigation. Note, a low dose of cromolyn resulted in mildly increased EFS-induced contraction of the trachea. Clearly, what was observed in this study is very interesting, but further study is needed to clarify these phenomena.
Another interesting finding is the experimental responses for methacholine and the EFS differed in the study administering cromolyn. Cromolyn had no cholinergic or anticholinergic effect in the methacholine experiment and caused no further contraction in the contracted tracheal smooth muscle induced by methacholine. It revealed no effect in postsynaptic events. On the other hand, a higher dose of cromolyn inhibited EFS-induced spike contraction of the trachea. It can be concluded that there is interference with transmitter release, so electrical stimulation causes decreased contraction. The presynaptic events may be responsible for the pharmacologic mechanism, so cromolyn played a role in the prophylactic treatment of asthma, stabilizing the presynaptic nerve. It was not easy to obtain human tissue for similar studies. The effect of this drug on isolated human tracheal smooth muscle still requires further investigation. Because this was an in vitro study, there are reservations as to its comparability with an in vivo situation in humans. In the in vivo situation, the response might be much more complicated than that in the in vitro situation.
A generally believed mechanism of the action of cromolyn is its ability to inhibit allergen-induced mediator release from mast cells. The presence of mast cells in rat trachea had been reported in the previous investigation. In rat trachea, two types of mast cells have been identified: connective tissue mast cells and mucosal mast cells. They have been observed in the submucosa region and epithelial layer, respectively.15
However, there are other statements for its possible association or additional mechanisms. A neural pathway mechanism for its effectiveness in asthma has been proposed where cromolyn could suppress the excitatory actions of afferent vagal C-fiber endings, suggesting part of its inhibitory capacity in vivo
may result from suppressing the vagal reflex pathways.16
Alvarez et al.
suggested the existence of a double site of pharmacologic action of cromolyn, one on the mast cell system and the second on the smooth muscle of the trachea or intestinal tract.17
Cromolyn inhibits guinea pig ileum contractions induced by electrical stimulation, which is mediated by acetylcholine release from Auerbach's plexus. They concluded cromolyn has a protective effect on smooth muscle fibers as well as on myenteric plexus.17
In their further studies, cromolyn inhibited contractions induced by electrical stimulation in both guinea pig ileum and the trachea as well as atropine does. Either this shows an inhibition of acetylcholine release from postganglionic parasympathetic fibers or an anticholinergic effect is involved in the mode of action of cromolyn.18
Our study can illustrate their statement of the inhibition of acetylcholine release from postganglionic parasympathetic fibers by cromolyn.