Both local cell-cell interactions and lineage bias have roles in determining the different retina cell phenotypes. In this study, subpopulations of amacrine cells that dually express GABA or serotonin (5-HT) and dopamine (DA) are identified in the early Xenopus tadpole (stages 42-48) retina. GABA is first detected by immunocytochemistry in amacrine cells at stage 35/36, 5-HT at stage 39, and DA at stage 41. As the number of these subtypes of amacrine cells increases by differentiation, a subset of them transiently express two neurotransmitters. GABA/DA double-labeled amacrine cells are detected first at stage 42, at which time they constitute 52% of the DA-containing population; this percentage decreases to only 3% by stage 48. 5-HT/DA amacrine cells are detected only at stage 44, constituting about 20% of the DA-containing cells and 4% of the small-dim 5-HT-containing cells. Regional location does not differentially affect the differentiation of these three types of amacrine cells (DA only, GABA/DA, and 5-HT/DA cells); each type is found more in the anterior and dorsal than the posterior and ventral quadrants, and their overall distribution patterns are statistically indistinguishable. However, these subtypes of amacrine cells reside in different sublamina of the inner nuclear layer. DA-only amacrine cells are located predominantly in the inner sublayer of the 2-3 cell thick amacrine cell layer, closest to the inner plexiform and the ganglion cell layers. Both types of double-labeled cells are located mostly in the outer sublayer of the amacrine cell layer, closest to other interneurons in the inner nuclear layer. This distinct sublaminar location of different neurotransmitter phenotypes suggests that local laminar cues influence the coexpression of neurotransmitters in amacrine cells.