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Articles in PresS, published online ahead of print February 14, 2002
Am J Physiol Heart Circ Physiol, 10.1152/ajpheart.00892.2001
Submitted on October 12, 2001
Accepted on February 12, 2002
1 Rammelkamp Center for Education and Research, MetroHealth Systems, Cleveland, OH, USA; Neurosciences, Case Western Reserve University, Cleveland, OH, USA
* To whom correspondence should be addressed. E-mail: dkunze{at}metrohealth.org.
We examined bovine aortic endothelial cells (BAECs) for the functional expression of P2X receptors, the ATP-gated cation channels. We identified the P2X subtypes present in the BAECs using RT-PCR. mRNA was present for only three of the seven family members, P2X4, P2X5 and P2X7. We then characterized agonist-activated currents in whole cell and outside-out patch recording using 2-methyl-thio-ATP (MeSATP) as a P2X4 and P2X5 receptor agonist and 2'3'-O-(4-benzoylbenzoyl)ATP (BzATP), as a P2X7 receptor agonist. MeSATP (10-20 µM) produced current with characteristics of P2X4 receptors. The current was inwardly rectifying current, reversed near 0 mV, slowly desensitized, was not blocked by suramin (300µM) or reactive blue (60µM) and had a single channel conductance of 36 pS. BzATP (10-100 µM), on the other hand, activated a 9 pS channel with sustained activity in the continued presence of the agonist. BzATP-activated current was blocked by reactive blue (60 µM) and by suramin (~50% block at 300µM). We confirmed, by immunocytochemistry, the presence of P2X4 and P2X7 protein. The agonists failed, however, to induce significant uptake of the large molecule, YO-PRO, indicating the lack of pore development that has been demonstrated for P2X7 and P2X4 in response to agonist in some cell types.
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