Heterogeneity of intrinsically photosensitive retinal ganglion cells in the mouse revealed by molecular phenotyping

Abstract

Intrinsically photosensitive retinal ganglion cell (ipRGC) types can be distinguished by their dendritic tree stratification and intensity of melanopsin staining. We identified heavily stained melanopsin-positive M1 cells branching in the outermost part of the inner plexiform layer (IPL) and weakly melanopsin-positive M2 cells branching in the innermost layer of the IPL. A third type can be distinguished by the displacement of the soma to the inner nuclear layer and has morphological similarities with either M1 cells or M2 cells, and is termed here displaced or M-d cells. The aim of the present study was to examine the phenotypic traits of ipRGC types. Using whole retinae from adult mice, we performed immunohistochemistry using melanopsin immunostaining and a number of antibodies directed against proteins typically expressed in retinal ganglion cells. The majority of M1 and M2 ipRGCs expressed Isl-1, microtubule associated protein-2 (MAP2), γ-synuclein, and NeuN, whereas Brn3 transcription factor and the different neurofilaments (NF68, NF160, NF200) were able to discriminate between ipRGC subtypes. Brn3 was expressed preferentially in M2 cells and in a small subpopulation of weakly melanopsin-positive M-d cells with similarities to M2 cells. All three neurofilaments were primarily expressed in large M2 cells with similarities to the recently described alpha-like M4 cells, but not in M1 cells. Expression of NF68 and NF160 was also observed in a few large M-d ipRGCs. These findings show that ipRGCs are not a phenotypically homogenous population and that specific neuronal markers (Brn3 and neurofilament) can partly distinguish between different ipRGC subtypes.

Intrinsic Photosensitive Retinal Ganglion Cells in the Diurnal Rodent, Arvicanthis ansorgei

“…In general, the morphology of the different types of melanopsin-immunopositive cells in the retina of diurnal Arvicanthis closely resemble those described previously in nocturnal rodents [20,36–38]. However, the respective proportions of each subtype were significantly different, with a high percentage of M1 cells (74%) in Arvicanthis compared to only 44% in the mouse retina [20].…”

“…In the mouse retina displaced cells were shown to comprise M1-like displaced and M2-like displaced ipRGCs [20,35]. In Arvicanthis , all displaced cells (termed M-d cells) were strongly melanopsin-immunoreactive and stratified in the outer IPL, similar to M1 ipRGCs (Figure 2).…”

“…The somata of M-d cells were 9.6–14 µm in diameter (median value 11.9 µm, n = 20). M-d cells were very rare and constituted only ~1% ipRGCs compared to ~6% in the mouse retina [20]. Because of the clear stratification in the OFF-sublamina of the IPL and the strong melanopsin immunoreactivity, all displaced cells in Arvicanthis seem to be of the M1-like type (M1-d).…”

“…The characteristics of primary antibodies used in the present study were extensively described in a previous study [20]. For the detection of melanopsin we used a rabbit polyclonal antibody PA1-781 (1:1000; Affinity Bioreagents) that was shown to cross-react with rat and mouse melanopsin (manufacturer’s information [9,21]).…”

Neurochemical Markers in the Mammalian Brain: Structure, Roles in Synaptic Communication, and Pharmacological Relevance

“…The expression (or lack thereof) of molecular markers can yield information about cellular populations and shed light on both normal biological processes and pathogenic events. More specifically, seminal studies used molecular markers to (1) simultaneously classify and sub-classify assemblages of cells while providing informative signatures about their functional mechanisms [1–9]; (2) estimate direct and indirect numbers and/or densities of cellular populations [10,11]; and (3) detect and diagnose abnormal cells (e.g. cancerous tumors) or possible pathological changes in association with certain diseases [12–14].…”