The contributions of synaptic ribbons to retinal circuit function and assembly

PROJECT SUMMARY The retina faces the considerable challenge of continuously and reliably encoding a wide dynamic range of light intensities. At the first two synapses of the retina are ribbons, large planar structures often described as ‘conveyer belts’ that continuously supply active zones of release with glutamate vesicles. As such, ribbons are thought to mediate high rates of glutamate release that allow for dynamic encoding of visual stimuli. In the retina, visual information flows from photoreceptors to bipolar cells to ganglion cell (GC) output neurons that send information to the brain in the form of spikes. In mice, there are at least 40 functionally distinct retinal GCs, each tuned to different features of the light environment and exhibiting unique circuit organizations. Therefore, our understanding of how the retina informs the brain is based on understanding how ribbons mediate the transfer of visual information through retinal circuits to shape GC output. The goal of this project is to determine how the function of two different GCs—ON-sustained and OFF-sustained alpha—are shaped by ribbons. As lighting conditions change, glutamate release probability at ribbon synapses is dynamically adjusted to match sensitivity needs—termed light adaptation. However, whether ribbons are directly responsible for regulating glutamate release during light adaptation is unclear. To answer this question, by measuring synaptic inputs to ON-sustained and OFF-sustained alpha GCs in a ribbon loss-of-function (Ribeye-ko) mouse model, I will first determine if ribbons regulate two forms of glutamate release that are thought to be important for adaptation: multivesicular release (MVR), the simultaneous release of multiple vesicles, and synaptic depression, a transient reduction in release probability following a stimulus. I will then use more dynamical stimulation methods to test whether ribbons set the time-course of adaptation to changes in luminance (mean light intensity) and contrast (variance about the mean) in GCs. Finally, I will identify how ribbons contribute to the formation of GC synaptic inputs using near-infrared branding and electron microscopy, thus correlating structure to function. GCs possess receptive fields that integrate information across space and enable the detection of specific features of the visual scene, such as edges, texture, or motion. Such spatial integration is contingent on tunable excitatory inputs to GCs, raising the possibility that ribbons are key determinants of the emergence of certain receptive field properties. To test this, using the Ribeye-ko model, I will measure excitatory synaptic inputs and spike outputs of ON-sustained and OFF-sustained alpha GCs in response to spatial stimulation paradigms that probe basic receptive field properties, including center-surround organization, nonlinear spatial integration, and feature sensitivity (i.e., to stimuli that exhibit texture or motion). I will complement functional studies with measurements of circuit anatomy, including presynaptic bipolar cell morphology and the organization of synaptic inputs, to both GC types via two-photon and confocal microscopy. This combinatorial structure-function approach will allow me to disentangle how ribbons shape overall circuit organizations.