P2Y₁₃ and P2X₇ receptors modulate mechanically induced adenosine triphosphate release from mast cells

Abstract

Subcutaneous mast cells (MCs) are vulnerable to mechanical stimulation from external environment. Thus, MCs immune function could be modulated by their mechanosensitivity. This property has been identified as the trigger mechanism of needling acupuncture, a traditional oriental therapy. Previously we have demonstrated the release of adenosine triphosphate (ATP), a stress-responsive signalling molecule, from mechanical-perturbed MCs. The current work explores its underlying mechanisms. We noticed that propagation of intracellular free Ca 2+ occurred among HMC-1 cells in response to 50% hypotonic shock. Additionally, amplifying cascade of ATP-induced ATP release was observed in RBL-2H3 cells stimulated by medium displacement, which could be mimicked by exogenous ATP (exoATP). Secondary ATP liberation induced by low level (50 nmol/L) of exoATP was reduced by inhibiting ecto-ATPase-dependent ADP production with ARL67156, or blocking P2 receptors with suramin or PPADS, or with specific P2Y 13 receptor antagonist MRS2211, or siRNA. Secondary ATP release induced by higher dose (200 μmol/L) of exoATP, sufficient to stimulate P2X 7 receptor, was attenuated by suramin, PPADS or specific P2X 7 receptor antagonist BBG, or siRNA. Finally, RT-PCR confirmed mRNA expression of P2Y 13 and P2X 7 in RBL-2H3 cells. Additionally, such secondary ATP release was attenuated by DPCPX, specific antagonist of adenosine A1 receptor, but not by MRS2179, specific inhibitor of P2Y 1 receptor. In summary, mechanosensitive ATP release from MCs is facilitated by paracrine/autocrine stimulation of P2Y 13 and P2X 7 receptors. This multi-receptor combination could mediate transmission of information from a local site to distal areas, enabling communication with multiple surrounding cells to coordinate and synchronize their function. K E Y W O R D SATP secretion, mast cells, mechanical stress, purinergic receptor P2X 7 , purinergic receptor P2Y 13

Does Cholinergic Stimulation Affect the P2X7 Receptor-Mediated Dye Uptake in Mast Cells and Macrophages?

“…The activation of P2X7 receptors and associated K + efflux is a key event in the activation of inflammasome (Muñoz-Planillo et al, 2013 ). We and others have shown that mast cells express functional proinflammatory P2X7 receptors (Kurashima et al, 2012 ; Wareham and Seward, 2016 ; Nurkhametova et al, 2019 ; Shen et al, 2020 , current study). P2X7 receptor signaling was proposed to be enhanced in sensitized (primed) mast cells (Di Virgilio et al, 2018 ).…”

Purinergic signaling as a basis of acupuncture-induced analgesia

“…Due to their location, they are sensitive to mechanical stimulation from the external environment. Mast cells contain ATP which may be released as a result of AP needling [21,22]. Four pieces of evidence support this notion: Firstly, mast cell-deficient rats (with c-kit gene mutant) exhibit less mechanical analgesia than their wild-type counterparts [22].…”

“…Secondly, mechanical stimuli lead to a rise in intracellular Ca 2+ of mast cells and release ATP in a Ca 2+ -dependent manner [23]. Thirdly, non-specific P2R antagonists or specific P2X7 or P2Y13R antagonists attenuate the release of ATP from mast cells suggesting the occurrence of an ATPinduced ATP release [21]. Fourthly, the outflow of ATP and a whole range of ATP metabolites (ADP/AMP/adenosine) have been measured in the neighboring interstitium by a microdialysis probe after AP of the Zusanli point (ST36) of mice [24,25].…”

“…In this respect, it is important to refer to findings already presented previously (see Section "Local release of ATP by AP needling"). Paracrine P2Y13Rs have been shown to facilitate the mechanosensitive release of ATP from the mast cell-like rat basophilic leukemia cell line RBL-2HR, in a manner inhibited by the selective P2Y13R antagonist MRS 2211 [21].…”

Tribute to Prof. Geoffrey Burnstock: his contribution to acupuncture

“…Subsequently, AP research in the field of purinergic signalling started to flourish and a large set of data has been accumulated on changes in purine (ATP, adenosine) concentrations, both locally and in the CNS, induced by manual acupuncture, electroacupuncture, or moxibustion [12][13][14][15][16][17]. The participation of purinergic receptors (P1: A1, A2a, A2b, A3) [18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34], (P2: P2X2, P2X3, P2X7, P2Y1, P2Y13) [35][36][37][38][39][40][41][42][43][44][45][46][47][48][49][50][51][52], and neuronal pathways bearing these receptors [53,54] has been confirmed in different AP-sensitive conditions (pain, inflammation, myocardial ischemia, obesity, cerebral ischemia). The functionalized acupuncture needle as a SERS (surfaceenhanced Raman spectroscopy)-active platform was developed for rapid and sensitive determination of ATP concentration [55], and led further support to the purinergic basis of AP.…”