Formation of carnosine‐Cu(II) complexes prevents and reverts the inhibitory action of copper in P2X₄ and P2X₇ receptors

Abstract

To further analyze the action of copper on brain synaptic mechanisms, the brain dipeptide carnosine (b-alanyl-L-histidine) was tested in Xenopus laevis oocytes expressing the rat P2X 4 or P2X 7 receptors. Ten micromolar copper halved the currents evoked by ATP in both receptors; co-application of carnosine plus copper prevented the metal induced-inhibition with a median effective concentration of 12.1 3.9 and 12.0 5.5 lM for P2X 4 and P2X 7 , respectively. Zinc potentiated only the P2X 4 ATP-evoked currents; carnosine had no effect over this metal. The relative potency and selectivity of classical metal chelators to prevent the copper inhibition was compared between carnosine and penicillamine (PA), bathophenanthroline (BPh) or L-histidine (His). Their rank order of potency in P2X 4 and P2X 7 receptors was carnosine PA His > BPh > Glycine (Gly) and carnosine BPh His > PA > Gly, respectively. The potency to prevent the zinc-induced potentiation in the P2X 4 receptor was BPh > PA > His; carnosine, Gly and b-alanine were inactive. Whereas 1±100 lM carnosine or His alone did not modify the ATP-evoked currents, 10±100 lM PA augmented and 100 lM BPh decreased the ATP-evoked currents. Carnosine was able to revert the copper-induced inhibition restoring the maximal ATP gated current in a concentration-dependent manner. Electronic spectroscopy con®rm the formation of carnosine-Cu(II) complexes, mechanism that can account for the prevention and reversal of the copper inhibition, revealing its potential in copper intoxication treatment.

Histidine 140 Plays a Key Role in the Inhibitory Modulation of the P2X4 Nucleotide Receptor by Copper but Not Zinc

“…Copper and zinc also modulate other members of the P2X family of receptors, but their effects depend on the receptor subtype. For example, both zinc and copper facilitate the ATP-evoked responses on the P2X 2 receptor (14,15) whereas in the P2X 7 receptor, copper inhibits the ATP-gated currents, whereas zinc is virtually without effect (13). These findings further support the notion that the P2X purinoceptors may have separate metal-binding sites to modulate the channel activity.…”

“…Acuñ a-Castillo et al (12) and Coddou et al (13) reported that zinc potentiates the ATP-evoked currents whereas copper has an inhibitory effect on the activity of this receptor. Based on these findings, Acuñ aCastillo et al (12) proposed that trace metals modulate the activity of the P2X 4 receptor via two separate metal-binding sites.…”

Microglia Proliferation Is Controlled by P2X7 Receptors in a Pannexin-1-Independent Manner during Early Embryonic Spinal Cord Invasion

“…These functional characteristics of ATP-evoked currents are compatible with the activation of P2X4R and/or of P2X7R, but not of P2X1R (Coddou et al, 2002). To have an estimation of the pharmacological profile of these microglia P2XRs, we then analyzed the effect of two agonists and eight antagonists on the response evoked by repetitive applications of 500 M ATP in free [Ca 2ϩ , Mg 2ϩ ] out (see Materials and Methods).…”

“…We then tested the effect of Cu 2ϩ and Zn 2ϩ on ATP-evoked currents. Cu 2ϩ inhibits both P2X7Rs and P2X4Rs whereas Zn 2ϩ increases the activity of P2X4R and is virtually without effect on rat P2X7R expressed in oocytes (Coddou et al, 2002) or fully blocks the activity of these two receptors expressed in transfected human embryonic kidney (HEK) cells (Liu et al, 2008). Then 50 M Cu 2ϩ evoked a significant ( p Ͻ 0.01) Ϸ77% inhibition (Fig.…”

“…5G), but the Zn 2ϩ effect on P2X7R remains controversial and depends on the preparation used. Indeed, Zn 2ϩ has a low potency on rat P2X7R expressed in oocytes (Coddou et al, 2002;Acuña-Castillo et al, 2007), when compared with rat P2X7R expressed in transfected HEK cells (Liu et al, 2008).…”