Whole Number, Distribution and Co-Expression of Brn3 Transcription Factors in Retinal Ganglion Cells of Adult Albino and Pigmented Rats

Abstract

The three members of the Pou4f family of transcription factors: Pou4f1, Pou4f2, Pou4f3 (Brn3a, Brn3b and Brn3c, respectively) play, during development, essential roles in the differentiation and survival of sensory neurons. The purpose of this work is to study the expression of the three Brn3 factors in the albino and pigmented adult rat. Animals were divided into these groups: i) untouched; ii) fluorogold (FG) tracing from both superior colliculli; iii) FG-tracing from one superior colliculus; iv) intraorbital optic nerve transection or crush. All retinas were dissected as flat-mounts and subjected to single, double or triple immunohistofluorescence The total number of FG-traced, Brn3a, Brn3b, Brn3c or Brn3 expressing RGCs was automatically quantified and their spatial distribution assessed using specific routines. Brn3 factors were studied in the general RGC population, and in the intrinsically photosensitive (ip-RGCs) and ipsilateral RGC sub-populations. Our results show that: i) 70% of RGCs co- express two or three Brn3s and the remaining 30% express only Brn3a (26%) or Brn3b; ii) the most abundant Brn3 member is Brn3a followed by Brn3b and finally Brn3c; iii) Brn3 a-, b- or c- expressing RGCs are similarly distributed in the retina; iv) The vast majority of ip-RGCs do not express Brn3; v) The main difference between both rat strains was found in the population of ipsilateral-RGCs, which accounts for 4.2% and 2.5% of the total RGC population in the pigmented and albino strain, respectively. However, more ipsilateral-RGCs express Brn3 factors in the albino than in the pigmented rat; vi) RGCs that express only Brn3b and RGCs that co-express the three Brn3 members have the biggest nuclei; vii) After axonal injury the level of Brn3a expression in the surviving RGCs decreases compared to control retinas. Finally, this work strengthens the validity of Brn3a as a marker to identify and quantify rat RGCs.

MicroRNA regulation in an animal model of acute ocular hypertension

“…Then, retinas were dissected as whole mounts (Nadal‐Nicolás et al. 2012; Rovere et al. 2015) and immunodetected following previously described methods (Salinas‐Navarro et al.…”

“…2013): Brn3a is expressed by the vast majority of the general population of RGCs (approximately 97%), except the melanopsin‐expressing RGCs and one half of the ipsilaterally projecting RGCs (Nadal‐Nicolás et al. 2012, 2014, 2015). The monoclonal antibody RT97 recognizes the phosphorylated heaviest subunit of the neurofilament triplet (pNFH), and its abnormal expression is an index of axonal injury (Vidal‐Sanz et al.…”

“…The total number of Brn3a + RGCs was automatically quantified using the Image‐Pro Plus (IPP 5.1, Media Cybernetics, Silver Spring, MD, USA) software, as reported (Nadal‐Nicolás et al. 2012). With these quantitative data, isodensity maps of Brn3a + RGCs were constructed using a graphing software ( sigmaplot © r ) as reported (Vidal‐Sanz et al.…”

Metabolomic Changes in the Rat Retina After Optic Nerve Crush

“…5 IONC triggers a reproducible massive RGC degeneration, and by day 14 more than 80% of their population is lost. [5][6][7] In the early phase of degeneration, there is a diffuse loss of RGCs throughout the entire retina, which is significantly detected 3 days after the lesion. 6,7 At the molecular level, as early as 12 hours after the injury there are significant changes in the regulation of prodeath genes and proteins.…”

“…[5][6][7] In the early phase of degeneration, there is a diffuse loss of RGCs throughout the entire retina, which is significantly detected 3 days after the lesion. 6,7 At the molecular level, as early as 12 hours after the injury there are significant changes in the regulation of prodeath genes and proteins. 8,9 However, at 12 hours, anatomical RGC loss is not yet detected, suggesting that the commitment to death occurs earlier than previously anticipated.…”