Types and density of calbindin D28k-immunoreactive ganglion cells in mouse retina

Abstract

Single-cell injection after immunocytochemistry is a reliable technique for classifying neurons by their morphological structure and their expression of a particular protein. The aim of the present study was to classify the morphological types of calbindin D28k-immunoreactive retinal ganglion cells in the mouse using single-cell injection after immunocytochemistry, to estimate the density of calbindin D28k-immunoreactive retinal ganglion cells in the mouse retina. Calbindin D28k is an important calcium-binding protein that is widely expressed in the central nervous system. Calbindin D28k-immunoreactive retinal ganglion cells were identified by immunocytochemistry and then iontophoretically injected with the lipophilic dye, DiI. Subsequently, the injected cells were imaged by confocal microscopy to classify calbindin D28k-immunoreactive retinal ganglion cells based on their dendritic ramification depth within the inner plexiform layer, field size, and morphology. The cells were heterogeneous in morphology: monostratified or bistratified, with small to large dendritic field size and sparse to dense dendritic arbors. At least 10 different morphological types (CB1–CB10) of calbindin D28k-immunoreactive retinal ganglion cells were found in the mouse retina. The density of each cell type was quite variable (1.98–23.76%). The density of calbindin D28k-immunoreactive cells in the ganglion cell layer of the mouse retina was 562 cells/mm2, 8.18% of calbindin D28k-immunoreactive cells were axon-less displaced amacrine cells, 91.82% were retinal ganglion cells, and approximately 18.17% of mouse retinal ganglion cells expressed calbindin D28k. The selective expression of calbindin D28k in cells with different morphologies may provide important data for further physiological studies of the mouse retina.