Circuit mechanisms underlying embryonic retinal waves

Spontaneous activity is a hallmark of developing neural systems. In the retina, spontaneous activity comes in the form of retinal waves, comprised of three stages persisting from embryonic day 16 (E16) to eye opening at postnatal day 14 (P14). Though postnatal retinal waves have been well characterized, little is known about the spatiotemporal properties or the mechanisms mediating embryonic retinal waves, designated Stage 1 waves. Using a custom-built macroscope to record spontaneous calcium transients from whole embryonic retinas, we show that Stage 1 waves are initiated at several locations across the retina and propagate across a broad range of areas. A gap junction antagonist, meclofenamic acid, reduced the frequency and size of Stage 1 waves, nearly abolishing them. The general nAChR antagonist, hexamethonium similarly nearly abolished Stage 1 waves. Application of the α4β2 nAChR antagonist dihydro-ß-erythroidine only slightly reduced the frequency of waves but significantly reduced the number of cells that participated in waves. Thus, Stage 1 waves are mediated by a complex circuitry involving subtypes of nAChRs and gap junctions. Stage 1 waves in mice lacking the β2 subunit of the nAChRs (β2-nAChR-KO) were reduced, but in contrast to WT mice, they persisted in the hexamethonium and were completely blocked by meclofenamic acid. To assay the impact of Stage 1 waves on retinal development, we compared the spatial distribution of a subtype of retinal ganglion cells, intrinsically photosensitive retinal ganglion cells (ipRGCs), which undergo a significant amount of cell death, in WT and β2-nAChR-KO mice. We found that the developmental decrease of ipRGC density is preserved between WT and β2-nAChR-KO mice, indicating that processes regulating ipRGC distribution are not influenced by spontaneous activity.