Modulation of intercellular communication in macrophages: possible interactions between GAP junctions and P2 receptors

Abstract

Gap junctions are connexin-formed channels that play an important role in intercellular communication in most cell types. In the immune system, specifically in macrophages, the expression of connexins and the establishment of functional gap junctions are still controversial issues. Macrophages express P2X7 receptors that, once activated by the binding of extracellular ATP, lead to the opening of transmembrane pores permeable to molecules of up to 900 Da. There is evidence suggesting an interplay between gap junctions and P2 receptors in different cell systems. Thus, we used ATP-sensitive and -insensitive J774.G8 macrophage cell lines to investigate this interplay. To study junctional communication in J774-macrophage-like cells, we assessed cell-to-cell communication by microinjecting Lucifer Yellow. Confluent cultures of ATP-sensitive J774 cells (ATP-s cells) are coupled, whereas ATP-insensitive J774 cells (ATP-i cells), derived by overexposing J774 cells to extracellular ATP until they do not display the phenomenon of ATP-induced permeabilization, are essentially uncoupled. Western-blot and reverse-transcription polymerase chain reaction assays revealed that ATP-s and ATP-i cells express connexin43 (Cx43), whereas only ATP-s cells express the P2X7 receptor. Accordingly, ATP-i cells did not display any detectable ATP-induced current under whole-cell patch-clamp recordings. Using immunofluorescence microscopy, Cx43 reactivity was found at the cell surface and in regions of cell-cell contact of ATP-s cells, whereas, in ATP-i cells, Cx43 immunoreactivity was only present in cytosolic compartments. Using confocal microscopy, it is shown here that, in ATP-s cells as well as in peritoneal macrophages, Cx43 and P2X7 receptors are co-localized to the membrane of ATP-s cells and peritoneal macrophages.

The P2X7 Nucleotide Receptor Mediates Skeletal Mechanotransduction

“…While this mechanism has some experimental support (44), our data suggest that the membrane pore formation is mediated by ATP and the major mechanism is the P2X 7 R. We observed pore formation after treatment with the P2X 7 R agonist BzATP and pore formation in osteoblasts, and MLO-Y4 osteocytes was blocked with apyrase, which hydrolyzes extracellular ATP thus interrupting its signaling. Others have suggested that Cx43 and P2X 7 R might have interacting functions (45). In peritoneal macrophages, Cx43 and P2X 7 R are co-localized at the cell membrane.…”

The P2X7 Receptor and Pannexin-1 Are Both Required for the Promotion of Multinucleated Macrophages by the Inflammatory Cytokine GM-CSF

“…In addition, the P2X 7 R reportedly interacts with connexin-43 (CX-43) (31), another hemichannel protein linked to ATP release (32). It has been suggested that the presence of functional P2X 7 at the cell surface may facilitate membrane insertion of CX-43 and the formation of gapjunction channels between MA (31). Therefore, the possibility exists that P2X 7 -coupled Panx-1 and/or CX-43 may participate in MA-MA interaction and fusion.…”

Cholesterol-Binding Peptides and Phagocytosis

“…Gap-junction proteins are also known to accumulate in cholesterol-rich rafts (Schubert et al, 2002;Lin et al, 2003;Lin et al, 2004;Dunina-Barkovskaya, 2005). Connexin Cx43 is found in macrophages (Beyer & Steinberg, 1991;Beyer & Steinberg, 1993;Anand et al, 2008); colocalization of purinoreceptors P2X7 with connexin Cx43 in macrophages was reported (Beyer & Steinberg, 1991;Beyer & Steinberg, 1993;Fortes et al, 2004). Figure 1 shows the fragments of the proteins aligned versus the CRAC sequence.…”