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Fig. 1. Expression of P2X₇ receptors in macrophages, ATP-sensitive (ATP-s), and ATP-insensitive (ATP-i) J774 cells. (A) ATP-s (second lane) but not ATP-i (fourth lane) cells express P2X₇ receptor mRNA. cDNAs obtained from both cells were amplified by PCR using a set of primers specific for either P2X₇ (top) or glyceraldehyde-3-phosphate dehydrogenase (GAPDH), used as control of cDNA loading (bottom). RT-PCR product was separated on a 1% agarose gel and stained with ethidium bromide. In mock reactions (first and third lanes), RNA samples obtained from the same preparations were substituted for cDNA as a control for genomic DNA contamination. (B) Western blots showing that peritoneal macrophages (lane 1) and ATP-s cells (lane 2), but not ATP-i cells (lane 3) express P2X₇ protein. Each lane was loaded with 100 µg of protein and separated by 10% SDS-PAGE before transfer to nitrocellulose membrane. The bar histograms represent the mean±s.d. (n=5) densitometric values relative to those obtained for macrophages. The asterisk indicates that the intensities of the bands for ATP-s cells are significantly different from those of ATP-i cells (P<0.05). Whole-cell patch-clamp recordings show that ATP-s (C) but not ATP-i (D) cells display a biphasic current after iontophoretic application of ATP (arrows). The spikes in (D) are passive currents sensed by the recording electrode during each iontophoretic application of ATP. Holding potential was maintained at -40 mV. Data are representative of at least ten recordings for each cell type.

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