Functional implications of the localization of arrestin-1 in rod synaptic terminals

Abstract

 

Purpose

 

Arrestin-1 is abundant in synaptic terminals in dark-adapted rods and persists there even after light adaptation. However, its function in the synapse remains poorly understood. The exact localization of arrestin-1 in this compartment can suggest its possible specific functions in the synapse.

 

Methods

 

Arrestin-1 localization in rod synaptic terminals of fixed eyes from light- or dark-adapted mice and its co-localization with synaptic proteins was determined by immunohistochemistry. Animals expressing WT arrestin-1, its monomeric form, and arrestin-1-3A mutant that binds unphosphorylated rhodopsin were compared, using both fluorescent and electron microscopy.

 

Results

 

WT arrestin-1 did not co-localize with SNAP-25 and partially co-localized with NSF, but showed complete overlap with VGLUT1 at light and electron microscopic levels. Arrestin-1 was not associated with synaptic ribbons or vesicles along the ribbon as evidenced by co-staining with Ribeye and electron microscopy. The localization of constitutive monomer resembled that of WT arrestin-1 regardless of the expression level. In contrast, 3A mutant that causes rod death was abundant in cell bodies but was not found in the synaptic terminals labeled with VGLUT1 even in the dark-adapted retina. The expression of synaptic proteins in the retina of mice expressing 3A mutant was similar to that in WT, but light adaptation did not reduce their levels, as it did in WT.

 

Conclusions

 

In healthy rod synapses, arrestin-1 is not associated with the synaptic membrane or ribbons, but localizes to the synaptic cytosol. The toxic arrestin-1-3A mutant is effectively absent from the synaptic terminals but overabundant in photoreceptor bodies, which might interfere with adaptation of synaptic protein levels to the light-dark cycle resulting in photoreceptor death. The data suggest that arrestin-1 localized to photoreceptor synaptic terminals has specific binding partners and fulfills functions that are important for rod health.

 

 

Images of the retina from light and dark-adapted wild type mice (WT) and transgenic mice expressing arrestin-1-3A mutant (1-3A) co-stained with anti-arrestin-1 (green) and anti-VGLUT1 (red) antibodies. Note co-localization of arrestin-1 and VGLUT1 in OPL of WT mice in both light and dark-adapted conditions (arrows). Also note lack of arrestin-1 in OPL in 1-3A mice and high level of VGLUT1 in these mice in the light as compared to WT (asterisks).

 

Keywords: 648 photoreceptors • 714 signal transduction • 728 synapse