Identification of neuronal populations that express calcium-permeable AMPA receptors in mouse and macaque retina

Abstract

Purpose : Calcium-permeable AMPA receptors (CP-AMPARs) play important roles in retinal neurotransmission, synaptic plasticity and development, but the neuronal populations that express these receptors have not been completely elucidated. The goal of this study was to determine the localization and functional properties of CP-AMPARs in the adult mouse and macaque retina.

Methods : Neurons that express CP-AMPARs were identified by a modification of the cobalt (Co²⁺) staining method described previously (Pruss et al., 1991; Osswald et al, 2007). Light-adapted retinae were incubated in an assay buffer containing AMPA (50μM) and cyclothiazide (100μM). ACET (1 μM) and D-AP5 (50 μM) were added to block kainate and NMDA receptors, respectively. Incubations were performed in the presence or absence of the AMPA receptor antagonist, GYKI-53655 (50 μM). After incubation, retinas were sectioned, silver-intensified and then immunolabeled with cell-type specific markers. Voltage-clamp recordings were made in macaque retinal slices to study CP-AMPAR mediated currents in horizontal cells.

Results : Strong Co²⁺ uptake was observed in mouse B-type horizontal cells (calbindin+) and macaque H1 & H2 horizontal cells (parvalbumin+). A subset of OFF bipolar cells showed weak Co²⁺ uptake in mouse (secretagogin+, Islet1-), but not macaque retina. In both species, a sub-population of amacrine cells were strongly Co²⁺ loaded. Of these, ~39.4% and ~51.2% were immunoreactive for the glycine transporter, GlyT1, in the macaque and mouse retina, respectively. In the macaque retina, double labeling for GlyT1 and the AII amacrine cell (AII-AC) marker, calretinin, showed that ~56.4% of the Co²⁺ labeled/GlyT1+ cells were AII-ACs. AII-ACs constituted ~22.2% of all of the Co²⁺ loaded amacrine cells. Approximately 72.9% of RBPMS-labeled ganglion cells showed Co²⁺ uptake in macaque retina. Co²⁺ uptake was completely blocked by GYKI-53655 in all cell types. In voltage-clamp recordings of macaque horizontal cells, the CP-AMPAR antagonist IEM-1460, suppressed glutamate evoked currents by 72±9.6% (mean±s.d. n=6 cells, p< 0.0001).

Conclusions : Our results suggest that CP-AMPARs are expressed in similar neuronal populations in mouse and macaque retina. In addition to AII-ACs, CP-AMPARs are expressed in other glycinergic and GABAergic amacrine cell types. At least one type of OFF bipolar cell expresses CP-AMPARs in mouse retina.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.