The photoreceptor synapse triad is a synaptic complex of great importance;physiologically, this is the site of the initial transfer of visual information, and the fidelity of information transfer is critically important for visual processing. The synaptic triad is characterized by the photoreceptor terminal, ON-bipolar cell dendrite and laterally distributed horizontal cell processes. Horizontal cells play a central role in generating the inhibitory receptive-field surrounds of photoreceptors and bipolar cells, as well as other, downstream retinal neurons. Horizontal cells participate in light adaptation, formation of center-surround receptive fields and generation of color opponency in retinal neurons. Vesicular-mediated transmitter release and proton secretion are leading candidates underlying horizontal cell output in the outer retina, although the cellular mechanisms and their sites of action are not yet fully understood. The long-term objective of this program is to understand the role of mammalian horizontal cells in visual information processing. This objective will be addressed by testing the hypotheses that 1) a regulated vesicular mechanism underlies GABA release and proton secretion from horizontal cell processes and their endings, and 2) horizontal cells participate in a pH-sensitive action on photoreceptors via V- ATPase, and an inhibitory GABA action on horizontal and bipolar cells.
Specific aim 1 will test if a) vesicle fusion with the plasma membrane is Ca2+-dependent, and determine the b) cellular localization and functional properties of the Ca2+ channel subtypes mediating Ca2+ signaling in horizontal cells.
Specific aim 2 will test if a) vesicle fusion with the plasma membrane is SNARE-protein dependent, and determine the b) cellular localization and binding partners of key vesicular and SNARE proteins mediating vesicle exocytosis, including V-ATPase, in horizontal cells.
Specific aim 3 will test if horizontal cell vesicular exocytosis mediates a) a pH-sensitive action on photoreceptors via V-ATPase insertion into the synaptic vesicle and plasma membrane and b) GABA action at horizontal and bipolar cells. Experiments will a) evaluate V-ATPase activity in horizontal cell endings, b) determine if pH-sensitive horizontal cell action is mediated by V-ATPase dependent protons and/or other pH-regulating membrane proteins and c) determine if GABA mediates horizontal cell autocrine and bipolar cell feed forward action. Studies using morphological, electrophysiological and cellular imaging approaches will initially elucidate the molecular and cellular basis of regulated vesicular release and proton secretion from horizontal cells, and will determine horizontal cell action on the response properties of outer retinal neurons. Relevance these studies will further the understanding of the cellular mechanisms underlying visual processing in the outer retina, which will aid in the development of therapeutic strategies for the treatment of eye disease and injury. These objectives are consistent with the health-related goals of the National Eye Institute for the development of therapeutic approaches for the treatment and prevention of retinal disease.

Public Health Relevance

Horizontal cells modulate the transfer of visual information in the outer retina from photoreceptors to second order retinal neurons in a process that is critically important for optimizing visual image formation under a broad range of ambient conditions, but our understanding of this process is incomplete and it appears to combine conventional and novel cellular mechanisms. Findings from the proposed studies will provide new and fundamental information concerning the regulation of horizontal cell synaptic signaling via transmitter exocytosis and proton secretion, and will increase our understanding of the molecular and cellular mechanisms underlying light adaptive processes and receptive field formation in the retina. This information is prerequisite for the understanding of normal retinal function and for developing retinal therapies to save or restore vision loss due to outer retina disease.

National Institute of Health (NIH)
National Eye Institute (NEI)
Research Project (R01)
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Special Emphasis Panel (ZRG1-BDPE-N (09))
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Greenwell, Thomas
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University of California Los Angeles
Schools of Medicine
Los Angeles
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Travis, Amanda M; Heflin, Stephanie J; Hirano, Arlene A et al. (2018) Dopamine-Dependent Sensitization of Rod Bipolar Cells by GABA Is Conveyed through Wide-Field Amacrine Cells. J Neurosci 38:723-732
Hirano, Arlene A; Liu, Xue; Brecha, Nicholas C et al. (2018) Analysis of Feedback Signaling from Horizontal Cells to Photoreceptors in Mice. Methods Mol Biol 1753:179-189
Sun, Xiaoping; Hirano, Arlene A; Brecha, Nicholas C et al. (2017) Calcium-activated BKCa channels govern dynamic membrane depolarizations of horizontal cells in rodent retina. J Physiol 595:4449-4465
Hirano, Arlene A; Liu, Xue; Boulter, Jim et al. (2016) Targeted Deletion of Vesicular GABA Transporter from Retinal Horizontal Cells Eliminates Feedback Modulation of Photoreceptor Calcium Channels. eNeuro 3:
Liu, Xue; Grove, James C R; Hirano, Arlene A et al. (2016) Dopamine D1 receptor modulation of calcium channel currents in horizontal cells of mouse retina. J Neurophysiol 116:686-97
Hoon, Mrinalini; Sinha, Raunak; Okawa, Haruhisa et al. (2015) Neurotransmission plays contrasting roles in the maturation of inhibitory synapses on axons and dendrites of retinal bipolar cells. Proc Natl Acad Sci U S A 112:12840-5
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Liu, Xue; Hirano, Arlene A; Sun, Xiaoping et al. (2013) Calcium channels in rat horizontal cells regulate feedback inhibition of photoreceptors through an unconventional GABA- and pH-sensitive mechanism. J Physiol 591:3309-24
Stella Jr, Salvatore L; Vila, Alejandro; Hung, Albert Y et al. (2012) Association of shank 1A scaffolding protein with cone photoreceptor terminals in the mammalian retina. PLoS One 7:e43463
Zampighi, Guido A; Schietroma, Cataldo; Zampighi, Lorenzo M et al. (2011) Conical tomography of a ribbon synapse: structural evidence for vesicle fusion. PLoS One 6:e16944

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