IntroductionThere are numerous data showing the importance of mast cells (MC) in bronchial hyperreactivity [1,2]. Much evidence also supports the involvement of cholinergic mechanisms in bronchial contractility [3]. The two mechanisms could be interconnected, as there are data available showing the influence of the cholinergic system on MC secretion [4][5][6]. Therefore MC may be involved in bronchial contractions evoked by muscarinic agonists. The aim of our work was to establish whether the depletion of MC histamine by the histamine liberator compound 48/80 (C48/80) could influence contractions of guinea pig isolated trachea evoked by the muscarinic agonist carbachol.
Materials and methodsPieces of 5 mm long tubes of guinea pig isolated trachea were transversely mounted in an organ bath with Tyrode solution (37°C). After 45 min equilibration, contractions, induced by carbachol or histamine, were recorded isometrically. On each preparation, a set of cumulative concentrations of either carbachol or histamine was added twicebefore and after C48/80 (100 mg/ml), which was washed out 15 min after the addition.The parameters of concentration-response curve (ED 50 , E max and the slope, p) were calculated for each set of concentrations, [A], according to the logistic function:The parameters obtained before and after C48/80 were statistically compared by t-test for repeated observations. In 3 separate groups of experiments, pieces of tracheae were examined histologically for MC degranulation. They were first placed in Tyrode solution for 30 min and then removed for preparation of histological samples (Group 1). In groups 2 and 3, C48/80 (50 mg/ml in Group 2 and 100 mg/ml in Group 3) was added in the bath for 15 min before tracheal tubes were removed for histological examination. The samples were fixed in Carnoy¢s solution and in basic lead acetate solution (1%), sectioned and stained with toluidine blue (0.5%). MC were counted and the percentage of degranulated MC was determined in 200 cells for each preparation. MC were identified and their degranulation assessed by morphological criteria [7].
Results and discussionAt histological examination, MC were found in the submucosa and also in deeper layers of the paries membranaceus tracheae, where they lay close to the smooth muscular cells. C48/80 evoked statistically significant degranulation of MC, although the difference between the two concentrations was not statistically significant (Table 1). Manipulation and exposure to Tyrode solution probably caused the rather high percentage of degranulated MC in all preparations, including the controls. The highest concentration of C48/80 provoked nearly 80% degranulation of MC and this concentration was used in the experiments where contractions were measured.C48/80 (100 mg/ml) evoked contractions that were comparable to the maximal effects of carbachol. The effect of carbachol was significantly diminished after C48/80: The maximal force of contraction (E max ) was shifted from 2.8 to 2.2 g and ED 50 from 0.086 to 0.13 mM. Howev...