Abstract
Purpose :
Retinal neurons assemble into circuits to process specific types of visual information. The proper development of each cell type within the circuit is essential for its function. Recent development of new genetic tools has allowed us to label and manipulate specific neuronal subtypes to understand how they wire into functional circuits and contribute to vision. Here, we use a genetic approach to investigate a distinct and novel population of non-GABAergic non-Glycinergic (nGnG) amacrine cells to define their function in the retina and the genetic factors that drive their unique identity during development.
Methods :
We used a Gbx2creER; R26LSL-TdTom mouse line to identify a novel amacrine cell (AC) subtype. For co-localization and morphological analysis, Gbx2+ ACs were analyzed by immunochemistry in adult retina cryosections and flat-mounts. To assess gap junction coupling, single Gbx2+ ACs were filled by electroporation of 2% Neurobiotin in a cell-attached configuration. For phenotypical analysis of the Gbx2 knockout and Robo1; Robo2 double knockout retinas, we compared mutant mice, Gbx2creER/flox or Gbx2creER; Robo1-/-; Robo2F/F, with heterozygous control littermates, Gbx2creER/+ or Gbx2creER; Robo1+/-; Robo2F/+.
Results :
We identified a population of ACs that selectively express the transcription factor Gbx2. Gbx2+ ACs consist of two similar, but distinct, subtypes with dendritic fields that stratify in sublamina 3 or 5 within the inner plexiform layer. Gbx2+ ACs do not express the inhibitory neurotransmitters GABA or Glycine, or other canonical neurotransmitters, suggesting they are a novel nGnG AC subtype. At least one subtype of Gbx2+ ACs are coupled to bipolar cells (BCs) through gap junction channels, suggesting that their primary synaptic output may be through electrical synapses. Conditional deletion of Gbx2 results in the mis-stratification of Gbx2+ AC dendritic arbors. (n=4 mice/genotype, p<0.05; t-test). Likewise, deletion of Robo receptors in Gbx2+ ACs result in a similar dendritic mis-stratification, suggesting they function as critical effectors downstream of Gbx2 during development. (n=3 mice/genotype, p<0.05; t-test).
Conclusions :
These results genetically identify a novel nGnG AC subtype with an unusual connectivity through gap junctions, and provide a model for studying transcriptional control of retinal circuit development and visual system function.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.