Differential expression of Ca²⁺ permeable AMPA receptors among RGC subtypes

Abstract

Purpose : Evidence suggest that specific populations of RGCs show selective vulnerability in animal models of glaucoma. This study was undertaken to determine if differences in expression of AMPA receptors between RGC subtypes might contribute to vulnerability to risk factors of glaucoma. While most neurons have AMPARs that are impermeable to Ca²⁺ (CI-AMPARs), some express significant numbers of Ca²⁺ permeable (CP) AMPARs, even into adulthood. Another goal was to determine if elevated pressure, a common feature of glaucoma models, further altered CP-AMPAR expression in RGC subtypes.

Methods : Retinas from 3 mouse lines that label ipRGCs, DS- and α RGCs were cultured for at least 28 days. We assessed the contribution of CP-AMPARs to the overall population of AMPARs using two approaches. One was to measure AMPA-evoked currents in the presence and absence of the CP-AMPAR antagonist PhTX-74. Measurements of block by PhTX were also carried out in parallel on labeled RGCs in the flatmount retina. The second was to image Ca²⁺ transients evoked by AMPA while blocking Ca²⁺ channels. Pressure was increased using a custom designed chamber.

Results : At 4-9 DIV, approximately 50% of the AMPA current was carried by CP-AMPARs in ipRGCs, and this fraction dropped to 30% at 21 DIV. Conversely, at 6-8 DIV, CP-AMPAR expression in DS-RGCs and α RGCs was about 85% of total AMPA current. Ca²⁺ imaging experiments confirmed these findings. DS-RGCs were completely lost from the culture by 10 DIV. Partial rescue of DS-RGCs was accomplished by adding PhTX to the culture media, indicating that CP-AMPARs contributed to cell loss. Pressure treatment of ipRGCs decreased CP-AMPARs, but increased CP-AMPAR in presumed OFF α RGCs and other unidentified RGC types, as determined by both Ca²⁺ imaging and PhTX application. The effect of pressure was blocked by overexpression of ADAR2, which controls Ca²⁺ permeability of the GluA2 subunit, and in a transgenic line in which GluA2 is impermeable to Ca²⁺.

Conclusions : Expression of CP-AMPARs can vary widely between RGC subtypes, and thus could be a contributing factor to susceptibility to diseases such as glaucoma. Elevated pressure may further exacerbate this problem, and the response to pressure also varies by RGC subtype. The pressure mediated increase in CP-AMPAR in DS- and α RGCs may be due to a fall in ADAR2. Conversely, ipRGCs appears to express alternate signaling pathways downstream from pressure transduction.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.