Our lab is interested in neuronal circuit assembly in development, circuit disassembly in degeneration and circuit reassembly upon cellular regeneration. Our studies are based on the vertebrate retina of zebrafish and mice. We apply a diversity of approaches including in vivo and in vitro confocal and multiphoton imaging, electron microscopy, transgenic methods and electrophysiology to investigate neuronal structure, function and connectivity in normal and perturbed retinas.
Circuits of the vertebrate retina (click images for full description)
The vertebrate retina, the light-sensitive part of the eye, comprises a complex network of synaptic connections involving five major classes of neurons. Rod and cone photoreceptors form an outer nuclear layer that contacts bipolar cells and horizontal cells. Bipolar cells relay information from the outer retina to the inner retina where they contact retinal ganglion cells and amacrine cells. The cell bodies and synaptic connections between the various retinal cell types are organized into distinct layers. This laminar arrangement, together with the rich diversity of morphological and functional subtypes of cells within each major cell class, make the retina an excellent model for investigating how pre- and postsynaptic cells acquire their appropriate morphology and synaptic connectivity.
Zebrafish (Danio rerio) develop rapidly and are visually responsive by about 4-5 days postfertilization. Because the embryo and young larvae can be maintained relatively transparent, we are able to track structural changes in zebrafish retinal neurons from the time of their genesis until they form circuits in the living animal. Retinal neurons are visualized by expression of fluorescent protein driven by cell-specific promoters. Labeling is achieved either by injection of DNA at the one-cell stage, or in stable transgenic lines.
Currently, we are investigating the cellular mechanisms that regulate the generation of cone photoreceptor subtypes and how the various cone types form specific patterns of converging input onto a common postsynaptic cell.
Taking advantage of the ability of the zebrafish retina to regenerate after damage and of molecular tools to target specific cell populations for in vivo ablation, we are also exploring how retinal neurons rewire upon cellular regeneration.
Circuit development in mice
By combining biolistic methods, transgenic animals with labeled retinal cell types, and immunohistochemistry, we are reconstructing detailed connectivity maps of neurons in the inner and outer retina. We are interested in the mechanisms that regulate the development of excitatory connections between photoreceptors and bipolar cells and between bipolar cells and retinal ganglion cells.
We are also investigating the mechanisms that shape the development of inhibitory connections onto the dendrites
and axons of retinal neurons. We are currently focusing on the role of neurotransmission in establishing and maintaining synaptic connectivity between retinal neurons.
Together with Phil Horner’s lab, we are exploring the steps of neuronal circuit disassembly in a mouse model of glaucoma using multielectrode array recording together with anatomical techniques. The image below shows a piece of retina placed on the 60 electrode array. Using white-noise stimuli, one can generate a movie of the stimulus over time that evoked spikes in a recorded cell (more information on this technique). The displayed movie shows the response of a ganglion cell that prefers dark stimuli within its receptive field.