Alpha retinal ganglion cells in pigmented mice retina: number and distribution

Abstract

Purpose: To identify and characterize numerically and topographically the population of alpha retinal ganglion cells (αRGCs) and their subtypes, the sustained-response ON-center αRGCs (ONs-αRGCs), which correspond to the type 4 intrinsically photosensitive RGCs (M4-ipRGCs), the transient-response ON-center αRGCs (ONt-αRGCs), the sustained-response OFF-center αRGCs (OFFs-αRGCs), and the transient-response OFF-center αRGCs (OFFt-αRGCs) in the adult pigmented mouse retina.Methods: The αRGC population and its subtypes were studied in flat-mounted retinas and radial sections immunodetected against non-phosphorylated high molecular weight neurofilament subunit (SMI-32) or osteopontin (OPN), two αRGCs pan-markers; Calbindin, expressed in ONs-αRGCs, and amacrines; T-box transcription factor T-brain 2 (Tbr2), a key transcriptional regulator for ipRGC development and maintenance, expressed in ipRGCs and GABA-displaced amacrine cells; OPN4, an anti-melanopsin antibody; or Brn3a and Brn3c, markers of RGCs. The total population of RGCs was counted automatically and αRGCs and its subtypes were counted manually, and color-coded neighborhood maps were used for their topographical representation.Results: The total mean number of αRGCs per retina is 2,252 ± 306 SMI32+αRGCs and 2,315 ± 175 OPN+αRGCs (n = 10), representing 5.08% and 5.22% of the total number of RGCs traced from the optic nerve, respectively. αRGCs are distributed throughout the retina, showing a higher density in the temporal hemiretina. ONs-αRGCs represent ≈36% [841 ± 110 cells (n = 10)] of all αRGCs and are located throughout the retina, with the highest density in the temporal region. ONt-αRGCs represent ≈34% [797 ± 146 cells (n = 10)] of all αRGCs and are mainly located in the central retinal region. OFF-αRGCs represent the remaining 32% of total αRGCs and are divided equally between OFFs-αRGCs and OFFt-αRGCs [363 ± 50 cells (n = 10) and 376 ± 36 cells (n = 10), respectively]. OFFs-αRGCs are mainly located in the supero-temporal peripheral region of the retina and OFFt-αRGCs in the mid-peripheral region of the retina, especially in the infero-temporal region.Conclusions: The combination of specific antibodies is a useful tool to identify and study αRGCs and their subtypes. αRGCs are distributed throughout the retina presenting higher density in the temporal area. The sustained ON and OFF response subtypes are mainly located in the periphery while the transient ON and OFF response subtypes are found in the central regions of the retina.

A newOpn4crerecombinase mouse line to target intrinsically photosensitive retinal ganglion cells (ipRGCs)

“…However, it is unclear how sensitive they are to label or recombine loxP -flanked cassettes in M4-M6s. Surveying the entire retina for M4s using the Opn4 cre(DSO ) ; Ai14 (tdTomato+ SMI32+, Fig 5a-c), we found 722 ± 58 cells, representing ∼1.44% of all RGCs similar to the reported ∼840 Alpha ON-sustained cells per retina 55 . However, with no alternative means to label the M5 and M6 population comprehensively and independently of Opn4 cre(DSO ) , it remains unclear if this, or any line captures the majority of these low melanopsin-expressing populations.…”

Role of PKN1 in Retinal Cell Type Formation

“…Analysis of adult retinal flat mounts stained with SMI-32 further showed that due to a significant reduction in SMI-32 + α-RGCs upon Pkn1 knockout ( Figure 4 b–d), fewer RGC axons left the retina at the optic nerve head. The fact that the pan-RGC marker RBPMS did not reveal differences in RGC numbers is likely explained by the much lower abundance of α-RGCs (~5% of all RGCs) compared to the total amount of RGCs [ 18 ]. SMI-32 + α-RGC axons of Pkn1 −/− animals did not show pathfinding defects to the optic nerve ( Figure 4 e) or optic tract ( Figure 4 f).…”

Vision rescue via chemically engineered endogenous retinal ganglion cells

“…Interestingly, Md -CiRGCs exhibited axon growth-enhancing genes ( Extended data Fig. 15b ) and 90% of BRN3A + Md- CiRGCs express the αRGC marker SPP1, a subtype, possess a greater propensity to regenerate their axons after axotomy 51, 52 . These results suggest a pro-regenerative state in CiRGCs, that support its regenerative ability in mice in vivo studies.…”