Deciphering the regulation of P2X4 receptor channel gating by ivermectin using Markov models

Abstract

The P2X4 receptor (P2X4R) is a member of a family of purinergic channels activated by extracellular ATP through three orthosteric binding sites and allosterically regulated by ivermectin (IVM), a broad-spectrum antiparasitic agent. Treatment with IVM increases the efficacy of ATP to activate P2X4R, slows both receptor desensitization during sustained ATP application and receptor deactivation after ATP washout, and makes the receptor pore permeable to NMDG+, a large organic cation. Previously, we developed a Markov model based on the presence of one IVM binding site, which described some effects of IVM on rat P2X4R. Here we present two novel models, both with three IVM binding sites. The simpler one-layer model can reproduce many of the observed time series of evoked currents, but does not capture well the short time scales of activation, desensitization, and deactivation. A more complex two-layer model can reproduce the transient changes in desensitization observed upon IVM application, the significant increase in ATP-induced current amplitudes at low IVM concentrations, and the modest increase in the unitary conductance. In addition, the two-layer model suggests that this receptor can exist in a deeply inactivated state, not responsive to ATP, and that its desensitization rate can be altered by each of the three IVM binding sites. In summary, this study provides a detailed analysis of P2X4R kinetics and elucidates the orthosteric and allosteric mechanisms regulating its channel gating.

Synthetic testosterone derivatives modulate rat P2X2 and P2X4 receptor channel gating

“…Thus, upon binding, testosterone analogues acting as allosteric modulators may change the P2XR protein conformation and subsequently alter the binding of orthosteric ligands, enhancing receptor function. A similar mechanism has been suggested for IVM, a P2X4R-specific allosteric modulator (Khakh et al, 1999;Priel and Silberberg, 2004;Jelinkova et al, 2006;Mackay et al, 2017). Testosterone derivatives and IVM also potentiated ATP-stimulated ethidium uptake by HEK293 cells expressing the P2X4R, further showing that these compounds share common mechanisms of action.…”

“…Based on these results, testosterone derivatives reduce the rate of P2X4R desensitization and accelerate the recovery of the desensitized peak current response. A similar effect was observed for IVM, a P2X4R-specific allosteric modulator (Khakh et al, 1999;Mackay et al, 2017).…”

Simulation of P2X‐mediated calcium signalling in microglia

“…; Mackay et al . ), including 12‐ and 13‐state models from Khadra et al . and Zemkova et al ., for the sake of parsimony we merged gating states that were in rapid equilibrium relative to state transitions that follow much slower kinetics (Smith & Crampin, ).…”

“…; Mackay et al . ). These models consisted of at least eight and as many as 32 states to capture nuances of desensitization and changes in channel gating arising from pharmacological treatment.…”

P2X4R silence suppresses glioma cell growth through BDNF/TrkB/ATF4 signaling pathway

“…6 There are seven different members of P2Xs family proteins in mammals, such as P2X1, P2X2, P2X3, P2X4, P2X5, P2X6, and P2X7, and receptors were either homotrimer (constituted with one type of subunit) or heterotrimer (constituted with more than one type of subunit). 6 There are seven different members of P2Xs family proteins in mammals, such as P2X1, P2X2, P2X3, P2X4, P2X5, P2X6, and P2X7, and receptors were either homotrimer (constituted with one type of subunit) or heterotrimer (constituted with more than one type of subunit).…”