The scaffold protein calcium/calmodulin‐dependent serine protein kinase controls ATP release in sensory ganglia upon P2X3 receptor activation and is part of an ATP keeper complex

Abstract

P2X3 receptors, gated by extracellular ATP, are expressed by sensory neurons and are involved in peripheral nociception and pain sensitization. The ability of P2X3 receptors to transduce extracellular stimuli into neuronal signals critically depends on the dynamic molecular partnership with the calcium/calmodulin-dependent serine protein kinase (CASK). The present work used trigeminal sensory neurons to study the impact that activation of P2X3 receptors (evoked by the agonist a,b-meATP) has on the release of endogenous ATP and how CASK modulates this phenomenon. P2X3 receptor function was followed by ATP efflux via Pannexin1 (Panx1) hemichannels, a mechanism that was blocked by the P2X3 receptor antagonist A-317491, and by P2X3 silencing. ATP efflux was enhanced by nerve growth factor, a treatment known to potentiate P2X3 receptor function. Basal ATP efflux was not controlled by CASK, and carbenoxolone or Pannexin silencing reduced ATP release upon P2X3 receptor function. CASK-controlled ATP efflux followed P2X3 receptor activity, but not depolarization-evoked ATP release. Molecular biology experiments showed that CASK was essential for the transactivation of Panx1 upon P2X3 receptor activation. These data suggest that P2X3 receptor function controls a new type of feed-forward purinergic signaling on surrounding cells, with consequences at peripheral and spinal cord level. Thus, P2X3 receptor-mediated ATP efflux may be considered for the future development of pharmacological strategies aimed at containing neuronal sensitization.

Glial pannexin1 contributes to tactile hypersensitivity in a mouse model of orofacial pain

“…In particular, activation of Panx1 through the P2x71230, P2x331, and P2x432 receptors have been shown to lead to maturation and release of pro-inflammatory mediators, thereby sensitizing peripheral and spinal cord neurons3132. We found that behavioral hypersensitivity in our orofacial pain model is accompanied by increased Panx1 expression and that neurons and GFAP-positive glia in the trigeminal ganglia become hyper-excitable.…”

In situ imaging reveals properties of purinergic signalling in trigeminal sensory ganglia in vitro

“…Because R192Q KI cultures at baseline express a higher number of potential pore-forming cells [18] and release cytokines, ATP and CGRP [16,18,40], the overall higher membrane permeability in R192Q KI trigeminal cultures was consistent with the expected R192Q KI phenotype. In addition, CGRP, which is a trigger for migraine pain [41] and neuronal sensitisation [26] via immunomodulatory functions [42], potentiated fluorescence responses in a wider number of cells in R192Q KI culture rather than in WT one.…”

“…Because α,β-meATP evokes the release of endogenous ATP from neurons via a Pannexin 1-mediated mechanism [40], the application of this agonist induced a fluorescence signal that we may attribute to activation of a wider P2X receptor population following by ATP priming. The reason for the smaller effect of Dashed line indicates WT control values, taken as 100%.…”

P2X receptor channels in chronic pain pathways

“…Interestingly, ATP release through pannexin was also shown to be induced downstream of P2X3 activation in trigeminal sensory neurons. This calcium/calmodulin‐dependent ATP efflux pathway potentially drives feedforward potentiation of painful signals (Bele and Fabbretti, ).…”