Conserved extracellular cysteines differentially regulate the inhibitory effect of ethanol in rat P2X4 receptors

Yi, Chu-Li; Liu, Yuwei; Xiong, Keming; Stewart, Randall R.; Peoples, Robert W.; Xiang, Tian; Zhou, Li; Ai, Yong-Xun; Li, Zhiwang; Wang, Qin-Weng; Li, Chaoying

doi:10.1016/j.bbrc.2009.02.018

Cited by 14 publications

(24 citation statements)

References 13 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Previous investigations suggest that the interface between the ectodomain and the TM segments in P2X4Rs contains targets for ethanol and IVM action or modulation [17,23,24]. We have identified residues Asp331 and Met336 in the ectodomain-TM2 segment interface to be important for ethanol modulation, whereas residues Asn338, Ser341, Gly342, Leu346, Gly347, Ala349, and Ile356 in the TM2 segment were found to play a role in IVM modulation of the receptor [25,26].…”

Section: Introductionmentioning

confidence: 71%

Tryptophan 46 is a site for ethanol and ivermectin action in P2X4 receptors

Popova

Trudell

et al. 2013

Purinergic Signalling

View full text Add to dashboard Cite

ATP-gated purinergic P2X4 receptors (P2X4Rs) are the most alcohol-sensitive P2XR subtype. We recently reported that ivermectin (IVM), an antiparasitic used in animals and humans, antagonized ethanol inhibition of P2X4Rs. Furthermore, IVM reduced ethanol intake in mice. The first molecular model of the rat P2X4R, built onto the Xray crystal structure of zebrafish P2X4R, revealed an action pocket for both ethanol and IVM formed by Asp331, Met336 in TM2 and Trp46, and Trp50 in TM1 segments. The role of Asp331 and Met336 was experimentally confirmed. The present study tested the hypothesis that Trp46 plays a role in ethanol and IVM modulation of P2X4Rs. Trp46 was mutated to residues with different physicochemical properties and the resultant mutants tested for ethanol and IVM responses using Xenopus oocyte expression system and twoelectrode voltage clamp. Nonaromatic substitutions at position 46 reduced ethanol inhibition at higher concentrations and switched IVM potentiation to inhibition. Simultaneous substitution of alanine at positions Trp46 and Met336 also resulted in similar changes in ethanol and IVM responses. Furthermore, a new molecular model based on the open pore conformation of zebrafish P2X4R suggested a role for Tyr42 that was further supported experimentally. Our previous and current findings, combined with our preliminary evidence of increased ethanol consumption in P2X4R knockout mice, suggest that the ethanol and IVM action pocket in P2X4Rs formed by positions 42, 46, 331, and 336 presents a potential target for medication development for alcohol use disorders.

show abstract

Section: Introductionmentioning

confidence: 71%

Tryptophan 46 is a site for ethanol and ivermectin action in P2X4 receptors

Popova

Trudell

et al. 2013

Purinergic Signalling

View full text Add to dashboard Cite

show abstract

“…It was shown that conserved cysteine residues in the extracellular loop of the human P2X(1) receptor form disulfide bonds and are involved in receptor trafficking to the cell surface (Ennion and Evans, 2002). Also, a recent study showed that conserved extracellular cysteines and disulfide bonds may differentially regulate the inhibitory effect of ethanol in rat P2X4 receptors (Yi et al, 2009). In the present study, ATP, a purinergic receptor agonist, or suramin, a purinergic receptor antagonist, significantly inhibited the effect of L-cysteine in neonatal bladders.…”

Section: Discussionmentioning

confidence: 99%

Differential Effect of l-Cysteine in Isolated Whole-Bladder Preparations from Neonatal and Adult Rats

Buyuknacar

Göçmen²,

Groat³

et al. 2010

J Pharmacol Exp Ther

View full text Add to dashboard Cite

The present study was undertaken to compare the effects of the thiol reagents L-cysteine and (diazene dicarboxylic acid bis 5N,N-dimethylamide) diamide on contractile activity of neonatal and adult rat bladders. In vitro whole-bladder preparations from Wistar rats were used to study the modulation of spontaneous bladder contractions by thiol reagents. After blocking cholinergic and adrenergic transmission with atropine and guanethidine, L-cysteine facilitated spontaneous bladder contractions in neonatal rat bladders. The effect of L-cysteine was suppressed by diamide. Diamide alone did not change basal activity of the neonatal rat bladder. The facilitatory effects of L-cysteine were reduced by the L-type Ca 2ϩ channel-blocking agent nifedipine and the calciumactivated K ϩ channel opener NS1619 [1,3-dihydro-1-[2-hydroxy-5-(trifluoromethyl)phenyl]-5-(trifluoromethyl)-2H-benzimidazol-2-one]. ATP or suramin, a purinergic receptor antagonist, significantly inhibited the effect of L-cysteine in neonatal bladders, whereas the nitric-oxide synthase inhibitor N -nitro-L-arginine was ineffective. L-cysteine did not elicit any detectable effects in the adult rat bladder; whereas diamide caused a large-amplitude sustained tonic contraction. The contraction induced by diamide in adult bladder did not occur when the preparation was pretreated with L-cysteine. Also, L-Cysteine administered during the diamide-evoked contraction completely inhibited the contraction to diamide. In conclusion, our results suggest that L-cysteine has markedly different effects in isolated wholebladder preparations from neonatal and adult rats. Thus thiol-sensitive mechanisms may modulate contractility by regulation of Ca 2ϩ and K ϩ channels and/or purinergic transmission in the neonatal bladder. The effects of L-cysteine and diamide were reversed in adult bladders, indicating that the regulation of bladder contractility by thiols is markedly altered during postnatal development.

show abstract

“…Previous findings suggested that the ectodomain and the TM interfaces in P2X4Rs contained targets for ethanol action (Xiong et al, 2005;Asatryan et al, 2008;Yi et al, 2009). Additional support for this hypothesis comes from a report that mutations at positions 331 or 336 in the ectodomain-TM2 region of P2X4Rs significantly reduced or eliminated the modulatory effects of ethanol .…”

Section: Introductionmentioning

confidence: 93%

Ethanol Is a Fast Channel Inhibitor of P2X4 Receptors

Ostrovskaya

Asatryan

Wyatt

et al. 2011

J Pharmacol Exp Ther

Self Cite

View full text Add to dashboard Cite

P2X receptors (P2XRs) are ion channels gated by synaptically released ATP. The P2X4 is the most abundant P2XR subtype expressed in the central nervous system and to date is the most ethanol-sensitive. In addition, genomic findings suggest that P2X4Rs may play a role in alcohol intake/preference. However, little is known regarding how ethanol causes the inhibition of ATP-gated currents in P2X4Rs. We begin to address this issue by investigating the effects of ethanol in wild-type and mutant D331A and M336A P2X4Rs expressed in human embryonic kidney (HEK) 293 cells using whole-cell patch-clamp methods. The results suggest that residues D331 and M336 play a role in P2X4R gating and ethanol inhibits channel functioning via a mechanism different from that in other P2XRs. Key findings from the study include: 1) ethanol inhibits ATP-gated currents in a rapid manner; 2) ethanol inhibition of ATP-gated currents does not depend on voltage and ATP concentration; 3) residues 331 and 336 slow P2X4 current deactivation and regulate the inhibitory effects of ethanol; and 4) ethanol effects are similar in HEK293 cells transfected with P2X4Rs and cultured rat hippocampal neurons transduced with P2X4Rs using a recombinant lentiviral system. Overall, these findings provide key information regarding the mechanism of ethanol action on ATP-gated currents in P2X4Rs and provide new insights into the biophysical properties of P2X4Rs.

show abstract

Conserved extracellular cysteines differentially regulate the inhibitory effect of ethanol in rat P2X4 receptors

Cited by 14 publications

References 13 publications

Tryptophan 46 is a site for ethanol and ivermectin action in P2X4 receptors

Tryptophan 46 is a site for ethanol and ivermectin action in P2X4 receptors

Differential Effect of l-Cysteine in Isolated Whole-Bladder Preparations from Neonatal and Adult Rats

Ethanol Is a Fast Channel Inhibitor of P2X4 Receptors

Contact Info

Product

Resources

About