Interaction between Calcium Chelators and the Activity of P2X7 Receptors in Mouse Motor Synapses

Abstract

The ability of P2X7 receptors to potentiate rhythmically evoked acetylcholine (ACh) release through Ca2+ entry via P2X7 receptors and via L-type voltage-dependent Ca2+ channels (VDCCs) was compared by loading Ca2+ chelators into motor nerve terminals. Neuromuscular preparations of the diaphragms of wild-type (WT) mice and pannexin-1 knockout (Panx1−/−) mice, in which ACh release is potentiated by the disinhibition of the L-type VDCCs upon the activation of P2X7 receptors, were used. Miniature end-plate potentials (MEPPs) and evoked end-plate potentials (EPPs) were recorded when the motor terminals were loaded with slow or fast Ca2+ chelators (EGTA-AM or BAPTA-AM, respectively, 50 μM). In WT and Panx1−/− mice, EGTA-AM did not change either spontaneous or evoked ACh release, while BAPTA-AM inhibited synaptic transmission by suppressing the quantal content of EPPs throughout the course of the short rhythmic train (50 Hz, 1 s). In the motor synapses of either WT or Panx1−/− mice in the presence of BAPTA-AM, the activation of P2X7 receptors by BzATP (30 μM) returned the EPP quantal content to the control level. In the neuromuscular junctions (NMJs) of Panx1−/− mice, EGTA-AM completely prevented the BzATP-induced increase in EPP quantal content. After Panx1−/− NMJs were treated with BAPTA-AM, BzATP lost its ability to enhance the EPP quantal content to above the control level. Nitrendipine (1 μM), an inhibitor of L-type VDCCs, was unable to prevent this BzATP-induced enhancement of EPP quantal content to the control level. We propose that the activation of P2X7 receptors may provide additional Ca2+ entry into motor nerve terminals, which, independent of the modulation of L-type VDCC activity, can partially reduce the buffering capacity of Ca2+ chelators, thereby providing sufficient Ca2+ signals for ACh secretion at the control level. However, the activity of both Ca2+ chelators was sufficient to eliminate Ca2+ entry via L-type VDCCs activated by P2X7 receptors and increase the EPP quantal content in the NMJs of Panx1−/− mice to above the control level.

Activation and expansion of presynaptic signaling foci drives presynaptic homeostatic plasticity

“…Finally, we returned to perform a similar analysis at the NMJ of wild type and ITGB1 cKO mice. As previously documented at the mouse NMJ (Miteva et al, 2020), application of EGTA-AM had no effect on wild type EJC amplitude or short-term plasticity (Figures S1E-S1G). We find that the same is true for the ITGB1 cKO (Figures S1E-S1G).…”

ProBDNF and Brain-Derived Neurotrophic Factor Prodomain Differently Modulate Acetylcholine Release in Regenerating and Mature Mouse Motor Synapses

“…Pannexin 1 hemichannels are capable of releasing intracellular ATP into the synaptic cleft. We have previously shown the obligatory role of endogenous ATP, released into the synaptic cleft via pannexins, in the activation of inhibitory presynaptic A1- and P 2 Y 13 purinoreceptors (along with vesicular ATP released with synaptic vesicles) in motor synapses ( Miteva et al, 2017 , 2018 , 2020 ). In Panx1 –/– mice the “pannexin source” of ATP is abolished.…”

Special Issue of International Journal of Molecular Sciences (IJMS) “Purinergic P2 Receptors: Structure and Function”

“…Miteva et al investigated interaction between intracellular calcium chelators and the activity of P2X7 receptors in mouse neuromuscular junction. This study shows that the activation of P2X7 receptors in motor nerve endings may provide additional Ca 2+ entry into motor nerve terminals, which, independent of the modulation of L-type voltage-dependent calcium channel activity, can provide sufficient Ca 2+ signals for ACh secretion [ 6 ]. Koldej et al found that there is an association between P2X7 polymorphisms and post-transplant outcomes in allogeneic hematopoietic stem cell transplantation, which represents a highly effective treatment method for hematologic malignancies.…”