June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Cellular interactions during closure of the optic fissure in the embryonic mouse eye
Author Affiliations & Notes
  • Amrita Pathak
    Department of Ophthalmology and Visual Sciences, Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Katrina S Hofstetter
    Department of Ophthalmology and Visual Sciences, Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Jonathon Kuntz
    Department of Ophthalmology and Visual Sciences, Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Dylan Burnette
    Cell and Developmental Biology, Vanderbilt University, Nashville, Tennessee, United States
  • Sabine Fuhrmann
    Department of Ophthalmology and Visual Sciences, Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Footnotes
    Commercial Relationships   Amrita Pathak, None; Katrina Hofstetter, None; Jonathon Kuntz, None; Dylan Burnette, None; Sabine Fuhrmann, None
  • Footnotes
    Support  NIH Grant R01EY024373, Core Grant EY008126, Unrestricted grant from Research to Prevent Blindness to the Dept of Opthalmology and Visual Sciences
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 3005. doi:
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      Amrita Pathak, Katrina S Hofstetter, Jonathon Kuntz, Dylan Burnette, Sabine Fuhrmann; Cellular interactions during closure of the optic fissure in the embryonic mouse eye. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3005.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Optic cup morphogenesis is a critical step for proper eye development. The morphogenetic process includes invagination of the ventral optic cup and the optic stalk, which leads to the formation of the optic fissure. The fissure margins subsequently fuse together leaving only a small opening for ganglion cells axons exiting the neural retina. Defects in closure of the optic fissure result in coloboma, which accounts for up to 10% of childhood blindness. However, the cellular and molecular mechanisms underlying the closure process are still obscure. Early electron microscopic studies suggest that thin cytoplasmic extensions arise from the fissure margins across the gap during optic fissure closure. We hypothesized that these extensions could represent filopodia-like structures. As the Rho GTPase CDC42 can induce filopodia formation, we predicted that CDC42 is critical for proper contact between the fissure margins through regulation of filopodia assembly.

Methods : We used a tamoxifen-inducible mouse line, Hes1CreERT2, for temporally controlled and tissue-specific CDC42 inactivation. Normal optic fissure starts closing around embryonic day 11 (E11) and fusion is completed by E12.5. To avoid potential effects on other functions of CDC42 such as apicobasal polarity, we disrupted CDC42 as late as possible by activating Hes1CreERT2 for 18-24 hours before closure. Analysis of markers for tissue patterning and apicobasal polarity was performed using immunohistochemistry. To observe the formation of filopodia extensions and cell-cell interactions during optic fissure closure, we used high-resolution confocal microscopy with Airyscan.

Results : Our immunohistochemistry data with a variety of markers shows that tissue patterning and apicobasal polarity appears unchanged during the closure process at E11.5. However, CDC42 mutant eyes consistently exhibit coloboma, detectable at E12.5. Using Airyscan imaging, our data in control eyes so far has revealed that we can visualize cytoplasmic bridges and filopodia-like structures extending between the optic fissure margins. Currently, we are analyzing CDC42 mutant optic cups to determine whether the formation of cytoplasmic bridges and filopodia-like structures is affected.

Conclusions : Our studies will reveal how the optic fissure margins make contact and whether Cdc42 is required during closure of the optic fissure in the mammalian eye.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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