Human neutrophils express two structurally distinct receptors for the Fc region of IgG, FcγRIIa and FcγRIIIb. Although earlier studies have suggested that the functional properties of these receptors are similar, mounting evidence suggests that these receptors are capable of inducing distinct functional responses. Accordingly, we have examined the regulation of intracellular Ca2+ transients induced by each of these receptors alone (homotypic receptor cross-linking) and together (heterotypic receptor cross- linking). The glycosylphosphatidylinositol-anchored FcγRIIIb induces a rise in [Ca2+] after homotypic cross-linking that is independent of ligand- mediated engagement of the transmembrane FcγRIIa. Both receptors were sensitive to the protein-tyrosine kinase inhibitor methyl 2,5- dihydroxycinnamate, but FcγRIIa-induced signaling was uniquely sensitive to the protein-tyrosine kinase inhibitor genistein. FcγRIIa but not FcγRIIIb engages a cAMP-sensitive and inositol 1,4,5-triphosphate-dependent pathway(s) that results in the Ca2+-transient. When these receptors are cross-linked into heterotypic clusters, a synergistic rise in [Ca2+] is observed that is accompanied by synergistic increases in the phospholipase Cγ-breakdown products inositol 1,4,5-triphosphate and diglyceride. These data provide a mechanism for the functional differences between these two receptors and suggest the possibility that they can be differentially modulated.