Date on Master's Thesis/Doctoral Dissertation

5-2018

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Biochemistry and Molecular Biology

Degree Program

Biochemistry and Molecular Biology, PhD

Committee Chair

Gregg, Ronald

Committee Co-Chair (if applicable)

Ellis, Steven

Committee Member

Clark, Barbara

Committee Member

Klinge, Carolyn

Committee Member

Metz, Cynthia

Committee Member

Hasan, Nazarul

Author's Keywords

mouse retina; leucine-rich repeat protein; LRIT1; night blindness; electroretinogram; retinal ganglion cell

Abstract

Mutations in genes encoding the leucine-rich repeat (LRR) proteins nyctalopin and LRIT3 lead to complete congenital stationary night blindness because they are critical to depolarizing bipolar cell function in the retina. LRIT3 has two closely related family members, LRIT1 and LRIT2. In silico analyses of publicly available RNA-Seq data showed that Lrit1 was highly expressed in the retina. Here I describe the expression pattern and impact of loss of LRIT1 on retinal function. To enable these studies, we used CRISPR/Cas9 technology to create an Lrit1-/- mouse line. Retinal morphology and morphometry analyses showed no gross changes in retinal structure or retinal layer thickness. Immunohistochemistry (IHC) shows photoreceptor, ON bipolar cell, and horizontal cell proteins localize normally in the absence of LRIT1. These data suggest LRIT1 is unnecessary for normal retinal and synaptic development. IHC also reveals LRIT1 localizes to the OPL with punctate staining similar to proteins expressed in invaginating horizontal cells at rod spherules but does not co-localize with ON or OFF bipolar cell proteins. RNA in situ hybridization shows Lrit1 expression in the ONL and INL, suggesting LRIT1 may also be expressed in photoreceptors. We used electroretinogram analyses to assess retinal function in Lrit1-/- mice and demonstrate that the a- and b-waves are decreased in amplitude under both scotopic and photopic conditions. Multi-electrode array recordings of Lrit1-/- retinal ganglion cells demonstrated abnormal ganglion cell responses and rhythmic oscillations. Taken together, our data localize LRIT1 to the OPL of the mouse retina where the loss of LRIT1 results in abnormal outer and inner retinal function without altering retinal structure. Thus, LRIT1 is critical for normal retinal signaling.

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