Format

Send to:

Choose Destination
See comment in PubMed Commons below
Purinergic Signal. 2013 Dec;9(4):687-93. doi: 10.1007/s11302-013-9365-4. Epub 2013 May 22.

Receptor-independent effects of 2'(3')-O-(4-benzoylbenzoyl)ATP triethylammonium salt on cytosolic pH.

Author information

  • 1Department of Physiology and Pharmacology, Schulich School of Medicine and Dentistry, The University of Western Ontario, London, Ontario, N6A 5C1, Canada.

Abstract

The effect of the relatively potent P2X7 receptor agonist 2'(3')-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate triethylammonium salt (BzATP-TEA) on cytosolic pH (pHi) was studied using MC3T3-E1 osteoblast-like cells, which endogenously express P2X7 receptors. pHi was measured fluorimetrically using the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. BzATP-TEA (0.3-1.5 mM) elicited fast-onset alkalinization responses. In contrast, adenosine 5'-triphosphate disodium salt (5 mM) failed to reproduce the BzATP-TEA-induced responses, indicating a P2 receptor-independent mechanism. We speculated that triethylamine, which is present in solutions of BzATP-TEA, permeates the plasma membrane, and is protonated intracellularly, leading to an increase in pHi. Consistent with this hypothesis, triethylammonium (TEA) chloride mimicked the effects of BzATP-TEA on pHi. Moreover, measurements using a Cytosensor microphysiometer revealed that TEA chloride transiently suppressed proton efflux from cells, whereas washout of TEA transiently enhanced proton efflux. BzATP-TEA also elicited a sustained increase in proton efflux that was blocked specifically by the P2X7 antagonist A-438079. Taken together, we conclude that BzATP-TEA-induced alkalinization is unrelated to P2X7 activation, but is due to the presence of TEA. This effect may confound assessment of the outcomes of P2X7 activation by BzATP-TEA in other systems. Thus, control experiments using TEA chloride are recommended to distinguish between receptor-mediated and nonspecific effects of this widely used agonist. We performed such a control and confirmed that BzATP-TEA, but not TEA chloride, caused the elevation of cytosolic free Ca(2+) in MC3T3-E1 cells, ruling out the possibility that receptor-independent effects on pHi underlie BzATP-TEA-induced Ca(2+) signaling.

PMID:
23689980
[PubMed - indexed for MEDLINE]
PMCID:
PMC3889390
Free PMC Article
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Icon for Springer Icon for PubMed Central
    Loading ...
    Write to the Help Desk