Functional ion channel formation by mouse macrophage IgG Fc receptor triggered by specific ligands

The mouse macrophage Fc gamma 2b/gamma 1R has previously been purified with the aid of the monoclonal antibody 2.4G2. That this Fc gamma R functions as a ligand-dependent ion channel is supported by the following evidence. Employing [3H]tetraphenylphosphonium ([3H]Ph4P+) as a probe for membrane potential changes in intact cells, we found a biphasic change in membrane potential following treatment with immune complexes, monoclonal antibody 2.4G2 IgG and 2.4G2 Fab-Sephadex particles. We observed an immediate depolarization followed by prolonged hyperpolarization. [3H]Ph4P+ uptake experiments with plasma membrane vesicles prepared from J774 macrophages showed that binding of ligands to the FcR led to transmembrane monovalent cation flow. Similar [3H]Ph4+ uptake experiments were done with phospholipid vesicles containing purified and reconstituted Fc gamma 2b/gamma. Following challenge with specific ligands, transmembrane monovalent cation flow was observed. Purified FcR was reconstituted into planar lipid bilayers; exposure to ligands led to transient bilayer conductance increase. THe conductance change was resolved into single channel events. Quin-2 measurements showed an increase of free cytosolic calcium levels in macrophages following exposure of cells to different ligands of the FcR. An optimal range of calcium was found to be required for phagocytosis, below and above which inhibition of ingestion occurred.