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Updated: Jun 4, 2026
Mechanical Stimulation-induced Calcium Wave Propagation in Cell Monolayers: The Example of Bovine Corneal Endothelial Cells
Published on: July 16, 2013
ATP induces contraction mediated by the P2Y(2) receptor in rat intestinal subepithelial myofibroblasts
Tatsuro Nakamura1, Koichi Iwanaga, Takahisa Murata
1Department of Veterinary Pharmacology, Graduate School of Agriculture and Life Sciences, The University of Tokyo 113-8657, Japan.
Abstract:
Intestinal subepithelial myofibroblasts (IMFs) exist just under the epithelial membrane directly facing the mucosal microvascular capillary surface distributed in the lamina propria. In the gastrointestinal tract, ATP is released from epithelial and endothelial cells in response to mechanical stimuli. Although it has been reported that mechanical stimuli evoke synchronized Ca(2+) waves in cultured IMFs, the contractile responses by ATP stimulation have not been examined. The aim of this study was to clarify the mechanism of the contraction of IMFs in response to ATP. ATP (1-30μM) induced contraction in a concentration-dependent manner. These contractions were inhibited by LaCl(3) (100-300μM) and by Ca(2+)-free solution (0.5mM EGTA). Fura-2/Ca(2+) signals indicated that ATP (1-10μM) elicited transient increases in intracellular Ca(2+) concentration ([Ca(2+)](i)). In addition, αβ-methylene-ATP (10, 30 and 300μM), a broad spectrum P2X agonist at a concentration higher than 100μM, induced neither contraction nor [Ca(2+)](i) rise. UTP (1-30μM), a selective P2Y(2) and P2Y(4) agonist in rodent, induced concentration-dependent contractions and [Ca(2+)](i) increases, whereas ADP and UDP (10μM) did not induce contractions. Pretreatment with suramin (30-100μM), a relatively selective P2Y(2) antagonist, strongly inhibited ATP- and UTP-induced contractions and [Ca(2+)](i) increases. However, pyridoxal-phosphate-6-azophenyl-2',4'-disulfonate (PPADS: 10-30μM), a receptor antagonist for several P2X and P2Y but less effective to P2Y(2) receptor, failed to inhibit ATP- and UTP-induced contractions and [Ca(2+)](i) increases. By RT-PCR, mRNA expressions of the P2Y(1) and P2Y(2) receptors, but not P2Y(4) or P2Y(6), were detected in IMFs. These results suggest that ATP induces [Ca(2+)](i)-dependent contraction in IMFs, which is mediated through the P2Y(2) receptor.
Insights
Adenosine triphosphate (ATP) triggers intestinal myofibroblast contraction via intracellular calcium. This process is mediated by the P2Y2 receptor, highlighting a novel mechanism in gastrointestinal function.
Area of Science:
- Gastroenterology
- Cell Biology
- Pharmacology
Background:
- Intestinal subepithelial myofibroblasts (IMFs) are crucial cells in the lamina propria, interacting with the mucosal microvasculature.
- ATP, released by epithelial and endothelial cells, is known to induce calcium waves in IMFs, but its contractile effects were uncharacterized.
Purpose of the Study:
- To investigate the mechanism by which ATP induces contraction in intestinal subepithelial myofibroblasts.
- To identify the specific purinergic receptors involved in ATP-mediated IMF contraction.
Main Methods:
- ATP and various purinergic receptor agonists/antagonists (e.g., UTP, αβ-methylene-ATP, suramin, PPADS) were used to stimulate isolated IMFs.
- Contractile responses were measured, and intracellular calcium ([Ca(2+)](i)) levels were monitored using Fura-2 imaging.
- RT-PCR was employed to detect the expression of purinergic receptor mRNA in IMFs.
Main Results:
- ATP induced a concentration-dependent contraction and transient increase in intracellular calcium in IMFs.
- Contractions and calcium increases were dependent on extracellular calcium and were inhibited by LaCl(3).
- UTP, a P2Y2/P2Y4 agonist, mimicked ATP's effects, while αβ-methylene-ATP, a P2X agonist, did not. Suramin, a P2Y2 antagonist, inhibited ATP/UTP responses, whereas PPADS did not. RT-PCR confirmed P2Y2 receptor mRNA expression in IMFs.
Conclusions:
- ATP-induced contraction of IMFs is a calcium-dependent process.
- The P2Y2 purinergic receptor is the primary mediator of ATP-induced contraction in intestinal subepithelial myofibroblasts.
- These findings reveal a novel signaling pathway involving IMFs and purinergic receptors in gastrointestinal physiology.
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