The first synaptic relay in the primary visual pathway occurs between terminals of photoreceptors and second-order neurons within the outer plexiform layer of the retina. During development, one of these types of second-order neurons, the type A horizontal cell, differentiates and assumes mature characteristics several days before any other cells ramifying in that synaptic layer. In neonates, horizontal cells appear to be GABAergic during the first 5 days of postnatal life and in addition they also are responsive to kainic acid. We have previously suggested that they may play a pioneering role in the postnatal development of the outer plexiform layer, perhaps providing structural guidance or trophic substances such as GABA, for synaptic development. To test this hypothesis, we first demonstrated that a single intraocular injection of kainic acid within 24 hr of birth results in a permanent and selective loss of type A horizontal cells in the outer retina. Retinas from animals maintained for 5 days postinjection were harvested for analysis of postnatal development of the outer plexiform layer in the absence of horizontal cells. One of the major findings was that kainic acid treatment caused a reversal of the normal complement of photoreceptor cell types, resulting in an abnormally high rod/cone ratio. The distribution of cell processes within the outer plexiform layer was also altered and normal synaptic connections were not made. In spite of these changes in the constituents of the outer plexiform layer, the normal position of the synaptic layer was not affected by the loss of horizontal cells. These results rule out the possibility that horizontal cells provide a structural barrier which is an absolute requirement for establishing the location of the outer plexiform layer. Rather, these cells may be more involved in cell differentiation and synaptogenesis.