Coexpression of Rat P2X₂and P2X₆Subunits inXenopusOocytes

Abstract

Transcripts for P2X(2) and P2X(6) subunits are present in rat CNS and frequently colocalize in the same brainstem nuclei. When rat P2X(2) (rP2X(2)) and rat P2X(6) (rP2X(6)) receptors were expressed individually in Xenopus oocytes and studied under voltage-clamp conditions, only homomeric rP2X(2) receptors were fully functional and gave rise to large inward currents (2-3 microA) to extracellular ATP. Coexpression of rP2X(2) and rP2X(6) subunits in Xenopus oocytes resulted in a heteromeric rP2X(2/6) receptor, which showed a significantly different phenotype from the wild-type rP2X(2) receptor. Differences included reduction in agonist potencies and, in some cases (e.g., Ap(4)A), significant loss of agonist activity. ATP-evoked inward currents were biphasic at the heteromeric rP2X(2/6) receptor, particularly when Zn(2+) ions were present or extracellular pH was lowered. The pH range was narrower for H(+) enhancement of ATP responses at the heteromeric rP2X(2/6) receptor. Also, H(+) ions inhibited ATP responses at low pH levels (<pH 6.3). The pH-dependent blocking activity of suramin was changed at this heteromeric receptor, although the potentiating effect of Zn(2+) on ATP responses was unchanged. Thus, the rP2X(2/6) receptor is a functionally modified P2X(2)-like receptor with a distinct pattern of pH modulation of ATP activation and suramin blockade. Although homomeric P2X(6) receptors function poorly, the P2X(6) subunit can contribute to functional heteromeric P2X channels and may influence the phenotype of native P2X receptors in those cells in which it is expressed.

Mouse Leydig cells express multiple P2X receptor subunits

“…P2X 2 homomeric receptors show only an increase in peak currents when pH is changed from control (pH 7.4) to a pH of 6.5 [26,45,51]. In addition, pH variations to 5.5 caused an increase in P2X 2 peak currents but a decrease on P2X 2/6 peak currents [27]. In mouse Leydig cells, bringing the pH to 5.5 induced a decrease on peak currents [41], corroborating the hypothesis that functional purinergic receptors are formed by P2X 2 and P2X 6 subunits.…”

“…Seven P2X receptor subunits (P2X 1-7 ) were cloned and characterized in vertebrates, and each of them can form two type of assemblies, homomeric, P2X 1 [50], P2X 2 [3], P2X 3 [8], P2X 4 [2], P2X 5 [18], P2X 7 [46], and P2X 6 [23], or heterotrimeric P2X 2/3 [31], P2X 1/5 [48], P2X 4/6 [30], P2X 2/6 [27], P2X 1/2 [5], P2X 1/4 [38], and P2X 4/7 [19]. Each subunit is composed by two transmem-brane regions (TM1 and TM2) forming the channel pore, a large extracellular domain, which contains the ATP binding site and both N and C termini located intracellularly [14,48,22].…”

“…Each purinergic receptor assembly presents a particular functional profile [31,27]. The pharmacological and biophysical properties of homomeric P2X receptors, characterized in heterologous expression systems, allowed their classification into three distinct groups: (1) α,β-methylene-ATP (αβmeATP) sensitive and rapidly desensitizing (P2X 1 and P2X 3 ); (2) αβmeATP insensitive and slowly desensitizing (P2X 2 , P2X 4 , P2X 5 ); and (3) αβmeATP insensitive, slowly desensitizing and pore formation after long-time exposure to the agonist (P2X 7 ) [15,33].…”

Determination of the architecture of ionotropic receptors using AFM imaging

“…It is known that the P2X 6 subunit can associate with both P2X 2 and P2X 4 to form heteromeric receptors, which have properties distinct from the corresponding homomers [27,41]. Furthermore, P2X 6 is co-localized with both P2X 2 and P2X 4 in many parts of the central and peripheral nervous systems [22,42,43,61,64,66,69,72].…”

“…The glutamate receptors are often heteromeric, although the subunit arrangements are again unknown [30,47]. P2X receptors are known to exist as both homomers and heteromers [2,27,34,41,54,55], and the heteromers have different characteristics from the parent homomers [27,34,41], indicating that the properties of endogenous receptors may vary in subtle ways depending on the subunit composition.…”