Acetylcholine in isolated airways of rat, guinea pig, and human: species differences in role of airway mucosa

Abstract

Stored endogenous acetylcholine (ACh) and in vitro synthesis of [3H]ACh were measured in isolated, mucosa-intact and mucosa-denuded airways of rat, guinea pig, and humans. In addition, choline acetyltransferase (ChAT) activity and ACh content were measured in freshly isolated airway mucosa as well as in cultured epithelial cells of rat tracheas. Rat tracheas stored 25 nmol/g ACh, whereas guinea pig tracheas and human bronchi contained only 2-3 nmol/g ACh. When incubated with [3H]choline, the isolated airways of rat, guinea pig, and human synthesized significant amounts of [3H]ACh. In guinea pig and human airways, removal of the mucosa affected neither stored ACh nor in vitro synthesis of [3H]ACh. In rat tracheas, however, removal of the mucosa resulted in a 50% reduction of stored ACh. Freshly isolated mucosa wiped off from the luminal surface of rat tracheas contained large amounts of ACh (6.5 nmol/g airway), whereas in human mucosa (central bronchi) only small amounts of ACh were found. In enzymatically isolated mucosal cells of rat tracheas, a considerable ChAT activity (21 nmol.mg protein 1.h-1) was detected, blockable by bromoacetylcholine. Enzymatically isolated human mucosa contained a rather low ChAT-like activity (0.5 nmol.mg protein 1.h-1), not sensitive to bromoacetylcholine. In cultured epithelial cells of rat tracheas (4th-6th passage), neither ChAT activity nor ACh was detected. The large airways of rat, guinea pig, and humans contain considerable amounts of ACh, supporting histological evidence of a dense cholinergic innervation, particularly of rat tracheas. The mucosa of rat tracheas synthesizes and stores large amounts of ACh, whereas the low ChAT activity in human mucosa argues against the presence of cholinergic neurons able to synthesize and store ACh.

M3 Muscarinic Receptor Antagonists Inhibit Small Cell Lung Carcinoma Growth and Mitogen-Activated Protein Kinase Phosphorylation Induced by Acetylcholine Secretion

“…Previous studies from our laboratory and other laboratories have shown that acetylcholine and M3 receptors are expressed in a wide variety of normal endothelial and epithelial cell types (5)(6)(7)(8)(9)(10)(11)(12)(13). This suggests that cancers derived from these diverse cell types will similarly coexpress acetylcholine and M3 receptors.…”

“…Recently, it has been established that acetylcholine is also widely synthesized by a variety of nonneuronal cell types, including keratinocytes (5), airway epithelial cells (6,7), glia (8), vascular endothelium (9, 10), placental trophoblasts (11), and ovarian follicular cells (12) among others (13). Expression of acetylcholine has been unambiguously shown in these cells and tissues as shown by the presence of choline acetyltransferase, the enzyme that synthesizes acetylcholine, and by demonstration of acetylcholine synthesis in these cells as measured by high-performance liquid chromatography (HPLC).…”

Blocking M2 muscarinic receptor signaling inhibits tumor growth and reverses epithelial-mesenchymal transition (EMT) in non-small cell lung cancer (NSCLC)

“…The recent discovery that lung cancers synthesize and secrete acetylcholine (ACh) that acts as an autoparacrine growth factor provides potential new targets to inhibit lung cancer growth. 3,4 It is well established that ACh is an important neurotransmitter in the central and peripheral nervous systems and acts via activation of ACh receptors (AChRs).Recently, it has been found that ACh is also widely synthesized by a variety of non-neuronal cell types including airway epithelial cells, 5,6 keratinocytes, 7 small and large intestine, gall bladder, 8 glial, 9 vascular endothelial, 10 immune cells and most common cancer cells such as NSCLC, SCLC, colon, glial and ovarian carcinomas. [11][12][13][14][15][16] Although the primary function has not yet been completely elucidated, the expression of ACh in many different cell types suggests that its roles will be fundamental and will present novel therapeutic targets.…”

Activated Cholinergic Signaling Provides a Target in Squamous Cell Lung Carcinoma

“…Expression of nAChR in lung cancers derives from expression of nAChR in normal lung cells. Normal bronchial epithelial cells express nAChR as part of a cholinergic autocrine loop (13)(14)(15)(16) in which all proteins needed for cholinergic signaling are present, including the ACh-synthesizing enzyme choline acetyltransferase (ChAT), the vesicular ACh transporter (VAChT), the high-affinity choline transporter CHT1, the ACh hydrolyzing enzymes acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), nAChRs, and muscarinic ACh receptors (mAChR). We and others have also shown that the interaction of ACh with both nAChR and mAChR stimulates cell proliferation (11,(17)(18)(19)(20), and we have previously reported that SCLC similarly express this cholinergic autocrine loop and that muscarinic antagonists can inhibit SCLC cell proliferation.…”