9723151 CHALUPA In the vertebrate retina the ON and OFF pathways, which signal increments and decrements of light, are anatomically segregated from each other into separate layers at maturity, but not early in development. During the previous period of NSF funding for this project, Dr. Chalupa's laboratory discovered that treating the developing retina with 2-amino-4-phosphonobutyrate (APB), the selective blocker of bipolar cell activity, prevents the normal stratification of retinal ganglion cell (RGC) dendrites into ON and OFF sublaminae in the inner plexiform layer, where synaptic contacts are made between the bipolar cells and RGCs. This finding suggested the hypothesis that activity in the bipolar cells regulates the retraction of RGC dendrites, which are initially present in both strata, or "multistratified." The current project will extend these findings by addressing three specific aims. The first aim is to establish whether the multistratified RGCs, resulting from APB treatment, respond to both the onset and offset of visual stimuli. This would provide functional evidence that blockade of the afferent activity during development results in an intermingling of ON and OFF retinal pathways. The second aim is to determine the temporal relationship between the ingrowth and segregation of bipolar cell terminals and the time course of RGC dendritic stratification. Segregation of ON and OFF bipolar cell afferents prior to the stratification of RGC dendrites would lend further support to the hypothesis that retinal afferents regulate the normal stratification of RGC dendrites. The third specific aim is to investigate synaptic connectivity between developing bipolar cells and multistratified RGCs by using the whole-cell patch-clamp technique to record synaptic activity in these neurons while stimulating their retinal afferents. These recordings will provide a direct test of the hypothesis that such retinal synapses are functional early in development. The successful completion of this research project will further our understanding of the mechanisms underlying the formation of ON and OFF retinal pathways.
