Acetylcholine and tachykinins involvement in the caffeine‐induced biphasic change in intracellular Ca²⁺ in bovine airway smooth muscle

Abstract

1 Caffeine has been widely used as a pharmacological tool to evaluate Ca 2 þ release from the sarcoplasmic reticulum in isolated smooth muscle cells. However, in nervous tissue this drug also causes neurotransmitters release, which might cause additional effects when smooth muscle strips are evaluated. To assess this last possibility, simultaneous measurements of contraction and cytosolic Ca 2 þ concentration (using Fura -2/AM) were carried out in bovine airway smooth muscle strips during caffeine stimulation. 2 A first stimulation (S1, n ¼ 11) with caffeine (10 mm) induced a biphasic change in cytosolic Ca 2 þ , which consisted of a transient Ca 2 þ peak (254740 nm, X X7SEM) followed by a plateau (92713 nm), and a transient contraction (204.72731.56 mg tension mg tissue À1 ). A second caffeine stimulation (S2) produced a similar response but these parameters had a different magnitude. The S2/S1 ratios for these parameters were 0.6970.02, 0.8370.06 and 1.0170.03, respectively. Addition of o-conotoxin GVIA (1 mm) and tetrodotoxin (3.1 mm) before S2 significantly diminished these S2/S1 ratios (0.2670.05, 0.2670.09 and 0.6470.11, respectively, n ¼ 5, Po0.05), implicating the neurotransmitters release involvement in the response to caffeine. A similar effect (Po0.01) was observed with atropine (1 mm, n ¼ 4), the fragment 4 -11 of substance P (SP) (an SP receptor antagonist, 10 mm, n ¼ 5), and with both substances (n ¼ 4). 3 We discarded a direct effect of o-conotoxin GVIA (1 mm) plus tetrodotoxin (3.1 mm) or of atropine (1 mm) plus SP fragment 4 -11 on smooth muscle cells because they did not modify caffeine responses in isolated tracheal myocytes. 4 We confirmed by HPLC that caffeine increased the release of acetylcholine (from 0.4370.19 to 2.0770.56 nm mg tissue À1 , Po0.02) in bovine airway smooth muscle strips. Detection of substance P by ELISA was not statistically different after caffeine stimulation (geometric means before and after caffeine, 0.69 vs. 1.97 pg ml À1 mg tissue À1 , respectively, P ¼ 0.053). 5 We concluded that acetylcholine and tachykinins release are involved in the caffeine-induced biphasic changes in cytosolic Ca 2 þ concentration.

Role of 5‐HT2A, 5‐HT4 and 5‐HT7 receptors in the antigen‐induced airway hyperresponsiveness in guinea‐pigs

“…In the present study, we found that activation of either 5-HT 4 or 5-HT 7 receptors increased the contractile cholinergic responses to electrical field stimulation in guinea-pig tracheas, thus suggesting a possible pre-junctional positioning of these receptors. This is in line with the fact that both 5-HT 4 and 5-HT 7 receptors are coupled to Gs protein, and thus their activation induces an increment of cAMP, as it is well known that a cAMP increment induces neurotransmitters release in the brain [47] and in airways neural network [48]. Furthermore, Weigand et al [10] recently reported that activation of pre-junctional 5-HT 2 receptors on parasympathetic cholinergic neurons, leading to enhanced ACh release, is required in order to fully develop the antigen-induced contraction of mouse trachea.…”

LTD4induces hyperresponsiveness to histamine in bovine airway smooth muscle: role of SR-ATPase Ca2+pump and tyrosine kinase

“…Airway smooth muscle cells were obtained from bovine tracheas, as described elsewhere (26). Smooth muscle (ϳ200 mg) was minced, placed in 5 ml Hanks' solution (GIBCO-BRL, Rockville, MD) containing 2 mg cysteine and 0.05 U/ml papaine, and incubated for 10 min at 37°C.…”

Possible Beneficial Actions of Caffeine in SARS-CoV-2

“…We and other research groups have studied the pharmacological activities of caffeine in the smooth muscle of the respiratory tract, including mainly adenosine receptor antagonism, phosphodiesterase inhibition, and intracellular calcium release from the sarcoplasmic reticulum (SR), and as a taste receptor type 2 ligand (TAS2R) [ 33 , 34 , 35 , 36 ]. It is important to emphasize that these mechanisms are found in almost all tissues and cells of the body.…”