Novel aspects of cholinergic regulation of colonic ion transport

Abstract

Nicotinic receptors are not only expressed by excitable tissues, but have been identified in various epithelia. One aim of this study was to investigate the expression of nicotinic receptors and their involvement in the regulation of ion transport across colonic epithelium. Ussing chamber experiments with putative nicotinic agonists and antagonists were performed at rat colon combined with reverse transcription polymerase chain reaction (RT-PCR) detection of nicotinic receptor subunits within the epithelium. Dimethylphenylpiperazinium (DMPP) and nicotine induced a tetrodotoxin-resistant anion secretion leading to an increase in short-circuit current (Isc) across colonic mucosa. The response was suppressed by the nicotinic receptor antagonist hexamethonium. RT-PCR experiments revealed the expression of α2, α4, α5, α6, α7, α10, and β4 nicotinic receptor subunits in colonic epithelium. Choline, the product of acetylcholine hydrolysis, is known for its affinity to several nicotinic receptor subtypes. As a strong acetylcholinesterase activity was found in colonic epithelium, the effect of choline on Isc was examined. Choline induced a concentration-dependent, tetrodotoxin-resistant chloride secretion which was, however, resistant against hexamethonium, but was inhibited by atropine. Experiments with inhibitors of muscarinic M1 and M3 receptors revealed that choline-evoked secretion was mainly due to a stimulation of epithelial M3 receptors. Although choline proved to be only a partial agonist, it concentration-dependently desensitized the response to acetylcholine, suggesting that it might act as a modulator of cholinergically induced anion secretion. Thus the cholinergic regulation of colonic ion transport – up to now solely explained by cholinergic submucosal neurons stimulating epithelial muscarinic receptors – is more complex than previously assumed.

Stimulation of Na+‐K+‐pump currents by epithelial nicotinic receptors in rat colon

“…This series of experiments and all subsequently described experiments were performed in the continuous presence of the neurotoxin TTX (10 −6 mol·L −1 at the serosal side) in order to prevent effects of nicotine mediated by cholinergic receptors on secretomotor neurons of the enteric nervous system. However, under these conditions, nicotine (10 −4 mol·L −1 at the serosal side; for a complete concentration–response curve describing the effect of nicotine on Cl − secretion across rat colonic epithelium, see Bader and Diener, ) was completely ineffective (Figure ). As positive control, at the end of the experiment, the activator of adenylate cyclase(s) forskolin (5·10 −6 mol·L −1 at the mucosal and the serosal side) was administered, which induced a prompt positive I sc , as expected, when the cystic fibrosis transmemembrane regulator, that is, the dominant anion channel in the apical membrane of colonic epithelium (Verkman and Galietta, ), is activated by cAMP‐dependent phosphorylation.…”

“…For example, in murine tracheal epithelium, nicotinic receptors are involved in the regulation of the production of airway lining fluid (Hollenhorst et al, ). Also, colonic epithelium expresses – besides muscarinic receptors of the M 1 and the M 3 subtype (Haberberger et al, , Bader and Diener, ) – different subunits of nicotinic receptors (α2, α4, α5, α6, α7, α10 and β4), and stimulation of these receptors with nicotinic agonists induces a Cl − secretion (Bader and Diener, ). In contrast to stimulation of muscarinic receptors, which activates anion secretion via hyperpolarization of the membrane due to opening of basolateral (and apical) Ca 2+ ‐dependent K + channels (Strabel and Diener, ) and transient stimulation of apical Ca 2+ ‐dependent Cl − channels (Hennig et al, ), nothing is known about the mechanism of action of nicotinic receptors in intestinal epithelium.…”

Evidence for metabotropic function of epithelial nicotinic cholinergic receptors in rat colon

“…This textbook view was challenged by the discovery of nicotinic receptor subunits in different epithelial cells (see Wessler & Kirkpatrick, 2008) including the colonic epithelium (Bader & Diener, 2015). While muscarinic receptors are known to transmit cholinergic signals in tissues innervated by the autonomic nervous system, nicotinic receptors have been thought to be expressed only in electrically excitable tissues.…”

“…Abbreviations: Isc, short-circuit current; nAChR, nicotinic ACh receptor; TTX, tetrodotoxin In rat colonic epithelium, the different subunits (α 2 , α 4 , α 5 , α 6 , α 7 , α 10 , and β 4 ) of nicotinic receptors are expressed (Bader & Diener, 2015).…”

Acetylcholine‐related proteins in non‐neoplastic appearing colonic mucosa from patients with colorectal neoplasia

“…Following synthesis, non‐neuronal ACh is exported by three different transport mechanisms, each dependent on different transporter proteins; vesicular ACh transporter (VAChT), OCT1‐2 and CTL4 . Outside the cell, ACh binds to muscarinic (M) and nicotinic (N) receptors on epithelial cells or is quickly degraded by acetylcholinesterase (AChE) and butyryl‐cholinesterase (BChE) and re‐cycled …”