Cross talk between P2 purinergic receptors modulates extracellular ATP-mediated interleukin-10 production in rat microglial cells
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Updated: Feb 1, 2026
Magnetic Isolation of Microglial Cells from Neonate Mouse for Primary Cell Cultures
Published on: July 25, 2022
Dong Reoyl Seo1, Soo Yoon Kim, Kyung You Kim
1Neuroscience Graduate Program, Ajou University School of Medicine, Suwon 443-721, Korea.
Abstract:
Previously we demonstrated that ATP released from LPS-activated microglia induced IL-10 expression in a process involving P2 receptors, in an autocrine fashion. Therefore, in the present study we sought to determine which subtype of P2 receptor was responsible for the modulation of IL-10 expression in ATP-stimulated microglia. We found that the patterns of IL-10 production were dose-dependent (1, 10, 100, 1,000 microM) and bell-shaped. The concentrations of ATP, ATP-gammaS, ADP, and ADP-betaS that showed maximal IL-10 release were 100, 10, 100, and 100 microM respectively. The rank order of agonist potency for IL-10 production was 2'-3'-O-(4-benzoyl)-benzoyl ATP (BzATP)=dATP>2-methylthio-ADP (2-meSADP). On the other hand, 2-methylthio-ATP (2-meSATP), alpha,beta-methylene ATP (alpha,beta-meATP), UTP, and UDP did not induce the release of IL-10 from microglia. Further, we obtained evidence of crosstalk between P2 receptors, in a situation where intracellular Ca(2+) release and/or cAMP-activated PKA were the main contributors to extracellular ATP-(or ADP)-mediated IL-10 expression, and IL-10 production was down-regulated by either MRS2179 (a P2Y(1) antagonist) or 5'-AMPS (a P2Y(11) antagonist), indicating that both the P2Y(1) and P2Y(11) receptors are major receptors involved in IL-10 expression. In addition, we found that inhibition of IL-10 production by high concentrations of ATP-gammaS (100 microM) was restored by TNP-ATP (an antagonist of the P2X(1), P2X(3), and P2X(4) receptors), and that IL-10 production by 2-meSADP was restored by 2meSAMP (a P2Y(12) receptor antagonist) or pertussis toxin (PTX; a Gi protein inhibitor), indicating that the P2X(1), P2X(3), P2X(4)receptor group, or the P2Y(12) receptor, negatively modulate the P2Y(11) receptor or the P2Y(1) receptor, respectively.
Insights
This study identifies specific P2 receptor subtypes, P2Y(1) and P2Y(11), as key mediators of Interleukin-10 (IL-10) release from microglia stimulated by extracellular ATP. Findings reveal complex P2 receptor crosstalk influencing IL-10 production.
Area of Science:
- Neuroimmunology
- Cellular Signaling
- Purinergic Signaling
Background:
- Extracellular ATP released from activated microglia stimulates Interleukin-10 (IL-10) expression via P2 receptors in an autocrine manner.
- Understanding the specific P2 receptor subtypes involved is crucial for elucidating the mechanisms of microglial immune modulation.
Purpose of the Study:
- To identify the specific P2 receptor subtypes responsible for modulating IL-10 expression in ATP-stimulated microglia.
- To investigate the complex interactions and crosstalk between different P2 receptor subtypes in regulating IL-10 production.
Main Methods:
- Microglia were stimulated with various extracellular nucleotides (ATP, ADP, and their analogs) at different concentrations.
- IL-10 production was measured, and specific P2 receptor antagonists (MRS2179, 5'-AMPS, TNP-ATP, 2meSAMP) and pertussis toxin were used to probe receptor function.
- Intracellular calcium (Ca2+) release and cAMP-activated protein kinase A (PKA) pathways were assessed as contributors to IL-10 expression.
Main Results:
- IL-10 production exhibited a dose-dependent, bell-shaped response to extracellular nucleotides.
- P2Y(1) and P2Y(11) receptors were identified as major contributors to IL-10 expression, indicated by down-regulation with specific antagonists.
- Evidence of negative modulation was observed, with P2X(1), P2X(3), P2X(4) receptor group, or P2Y(12) receptor, negatively impacting P2Y(11) or P2Y(1) receptor function, respectively.
Conclusions:
- Both P2Y(1) and P2Y(11) receptors play significant roles in mediating IL-10 production in response to extracellular ATP and ADP.
- Complex crosstalk exists between P2 receptor subtypes, including inhibitory interactions, which collectively regulate microglial IL-10 expression.
- These findings provide critical insights into the purinergic signaling pathways governing microglial inflammatory responses and IL-10 regulation.
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