Retinas of macaque monkeys were immunostained for glycogen phosphorylase (glypho). Glypho was localized to regular and displaced amacrine cells. Their processes occupied two narrow strata within the inner plexiform layer (IPL). The labeling pattern is reminiscent of cholinergic amacrine cells; however, double immunostaining of the retinas for choline acetyltransferase and glypho revealed two different cell populations. Intracellular injection of DiI showed that glypho-immunoreactive amacrine cells are wide-field amacrine cells with straight, radially oriented, and sparsely branched dendrites. The density of the cells increased from approximately 70/mm(2) in the peripheral retina to approximately 700/mm(2) in the central retina. The regular glypho-immunoreactive amacrine cells branch in sublamina 2 of the IPL, where they receive input from OFF-cone bipolar cells. The displaced cells branch in sublamina 3/4 and receive input from ON-cone bipolar cells. This suggests that the regular cells are OFF-cells and the displaced cells are ON-cells. The cells express gamma-aminobutyric acid (GABA)-like immunoreactivity and receive glycinergic input through synapses expressing preferentially the glycine receptor alpha2 subunit. The close proximity of the dendritic strata of glypho-immunoreactive amacrine cells, cholinergic amacrine cells, and direction-selective ganglion cells suggests a possible role of the cells in the generation of direction-selective light responses of the monkey retina.