A dual function for LRIT3 – control of retinal photoreceptor synaptic morphology and depolarizing bipolar cell signaling

Abstract

Purpose : The human disease, complete congenital stationary night blindness (cCSNB) is a heterogeneous disorder of the retina characterized by impairment of low light vision and loss of the b-wave of the electroretinogram (ERG). Mutations in Grm6, Trpm1, Nyx and Gpr179, which encode proteins critical to signal transmission in one class of retinal interneurons called ON bipolar cells (BCs) cause cCSNB. LRIT3, a leucine rich repeat protein, expressed at the dendrites of ON BCs, was recently found mutated in cCSNB patients (Zeitz et al.). The function of LRIT3 in the mGluR6 signaling cascade remains unknown. Here we have created a Lrit3^-/- mouse line and investigated the function of LRIT3 in the mGluR6 cascade.

Methods : Using a zinc-finger nuclease strategy, we constructed a mouse line with a 40 base pair deletion within a highly conserved region in exon 2 of Lrit3. We characterized retinal function of Lrit3^-/- using the ERG. Localization and protein expression levels of LRIT3, nyctalopin and TRPM1, were examined by immunohistochemistry and immunoblotting, respectively. Glutamate release in the IPL was measured by infecting retinas with AAV2/1.hSynapsin.iGluSnFR (Borghuis et al.) and retinal function assessed by electrophysiological recordings from type 4 OFF BCs and using a multi-electrode array to record from ganglion cells.

Results : The dark- and light-adapted ERGs from Lrit3^-/- mouse have a normal a-wave, while their b-wave is absent. Nyctalopin and TRPM1 expression was absent from the dendritic terminals of ON BCs of Lrit3^-/- retina and PNA staining was reduced in the cone synaptic pedicles. LRIT3 expression was not effected in retinas lacking GRM6, TRPM1, GPR179 or Nyctalopin, although TRPM1 total protein expression was significantly reduced in Lrit3^-/- retinas. Glutamate release in the IPL and the amplitude of the light responses was decreased in Lrit3^-/- retina, as measured by OFF BCs and OFF ganglion cell responses.

Conclusions : Lrit3^-/- mice manifest a common phenotype of cCSNB, including the absence of the b-wave. They lack Nyctalopin and TRPM1 expression at the dendritic terminals of all ON BCs. LRIT3 expression and localization is independent of GRM6, TRPM1, GPR179 or Nyctalopin expression. LRIT3 is required for light responses in OFF BCs and ganglion cells.

References:
Zeitz C, et al. Am J Hum Genet. 2013; 92(1): 67
Borghuis BG, et al. J Neurosci. 2013; 33(27): 10972

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