TRPM8 Channel Activation Reduces the Spontaneous Contractions in Human Distal Colon.

Antonella Amato,Pierenrico Marchesa,Laura Lentini,Flavia Mulè,Simona Terzo

doi:10.3390/ijms21155403

Abstract

The transient receptor potential-melastatin 8 (TRPM8) is a non-selective Ca2+-permeable channel, activated by cold, membrane depolarization, and different cooling compounds. TRPM8 expression has been found in gut mucosal, submucosal, and muscular nerve endings. Although TRPM8 plays a role in pathological conditions, being involved in visceral pain and inflammation, the physiological functions in the digestive system remain unclear as yet. The aims of the present study were: (i) to verify the TRPM8 expression in human distal colon; (ii) to examine the effects of TRPM8 activation on colonic contractility; (iii) to characterize the mechanism of action. Reverse transcriptase-polymerase chain reaction (RT-PCR) and western blotting were used to analyze TRPM8 expression. The responses of human colon circular strips to different TRPM8 agonists [1-[Dialkyl-phosphinoyl]-alkane (DAPA) 2–5, 1-[Diisopropyl-phosphinoyl]-alkane (DIPA) 1–7, DIPA 1–8, DIPA 1–9, DIPA 1–10, and DIPA 1–12) were recorded using a vertical organ bath. The biomolecular analysis revealed gene and protein expression of TRPM8 in both mucosal and smooth muscle layers. All the agonists tested, except-DIPA 1–12, produced a concentration-dependent decrease in spontaneous contraction amplitude. The effect was significantly antagonized by 5-benzyloxytryptamine, a TRPM8 antagonist. The DIPA 1–8 agonist resulted in the most efficacious and potent activation among the tested molecules. The DIPA 1–8 effects were not affected by tetrodotoxin, a neural blocker, but they were significantly reduced by tetraethylammonium chloride, a non-selective blocker of K+ channels. Moreover, iberiotoxin, a blocker of the large-conductance Ca2+-dependent K+-channels, but not apamin, a blocker of small-conductance Ca2+-dependent K+ channels, significantly reduced the inhibitory DIPA 1–8 actions. The results of the present study demonstrated that TRPM8 receptors are also expressed in human distal colon in healthy conditions and that ligand-dependent TRPM8 activation is able to reduce the colonic spontaneous motility, probably by the opening of the large-conductance Ca2+-dependent K+-channels.

Highlights

Transient receptor potential (TRP) channels constitute a large family of non-selective cation channels involved in diverse cellular functions [1,2]
The results of the present study demonstrated that TRPM8 receptors are expressed in human distal colon in healthy conditions and that ligand-dependent TRPM8 activation is able to reduce the colonic spontaneous motility, probably by the opening of the large-conductance Ca2+-dependent K+-channels
Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed the presence of a 621 bp product relative to TRPM8 mRNA expression, in both mucosa and smooth muscle layer (Figure 1A)

Summary

Introduction

Transient receptor potential (TRP) channels constitute a large family of non-selective cation channels involved in diverse cellular functions [1,2]. They display large diversity in activation mode, ion selectivity and, physiological functions. The TRPM (transient receptor potential melastatin) represents the largest and most variable subfamily of the TRP channels, being constituted by eight members, from TRPM1 to TRPM8, and responding to various stimuli such as changes in the concentration of ions, small molecules, and lipids [2]. TRPM8 is a member of the temperature-sensitive TRP channels It is activated by mild cold temperatures, membrane depolarization, changes in extracellular osmolarity, cooling compounds such as menthol and icilin, and it shows a modest permeability to calcium [5,6]. TRPM8 channels have been implicated in oropharyngeal dysphagia [13] and chronic cough [14], while the downregulation of TRPM8 by angiotensin II may be involved in hypertension [15]

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