Immunohistological Localization of Mel1a Melatonin Receptor in Pigeon Retina

Abstract

Background Melatonin (N-acetyl-5-methoxytryptamine), a significant indoleamine neuromodulator implicated in circadian rhythms and sleep patterns, regulates diverse rhythmic functions via activating its high-affinity G-protein-coupled receptors. However, the detailed cellular expression of the Mel1a receptor in the retina is still a research gap. Methods The expression of the Mel1a receptor in pigeon retina was assessed using Western blot analysis and immunofluorescent staining. The cellular localization of the Mel1a receptor was studied using double immunofluorescent staining and laser-scanning confocal microscopy. Results Our data suggested that the Mel1a receptor was extensively expressed in the outer segment of Rho4D2-labeled rod and L/M-opsin-labeled red/green cone and in the somata of the CB-labeled horizontal cell, TH-labeled dopaminergic amacrine cell, ChAT-labeled cholinergic amacrine cell, PV-labeled AII amacrine cell, Brn3a-labeled conventional ganglion cell, melanopsin-containing ganglion cell and CRALBP-labeled Müller glial cell. In addition, the Mel1a receptor was diffusely distributed throughout the full thickness of the inner plexiform layer. However, the outer segment of S-opsin-labeled blue cone, the somata of ChX-10-labeled bipolar cell and outer plexiform layer seemed to lack immunoreactivity of the Mel1a receptor. Conclusion The finding that multiple types of retinal cells express the Mel1a receptor provides a new neurobiological basis for the participation of melatonin in the regulation of retinal functions through activating the Mel1a receptor.

Expression of SH3 and Multiple Ankyrin Repeat Domains Protein 3 in Mouse Retina

“…Figures 2a-a" shows localization of the SHANK3 in red/green cones. Double labeling revealed that the SHANK3 immunoreactivity was only found on the L/M-opsin-labeled outer segment (OS) of the red/green cone (81 out of 195 positive segments from four sections, 40.9% positive rate, Sheng et al, 2021). Figures 2b-b" shows the localization of the SHANK3 in blue cones.…”

“…Figures 2b-b" shows the localization of the SHANK3 in blue cones. Double labeling shows the SHANK3 signals were weekly expressed on the inner segment (IS) and somata of S-opsin-labeled blue cones (481 somat out of 481 cells from four sections, 100% positive rate, Sheng et al, 2021). Figures 2c-c" shows the localization of the SHANK3 in rods.…”

“…The immunocytochemistry procedures were performed according to a method described previously (Sheng et al, 2021). Briefly, the retinal sections were preincubated in 0.1 M PBS containing 6% normal donkey serum, 1% bovine serum albumin, and 0.2% Triton X-100 (PBST) for 2 h at 4 • C. Rabbit polyclonal antibody against mouse SHANK3 (corresponding to amino acid residues 841-855) (1:1,000 dilution, APZ-013, Alomone Labs, Israel) was used for labeling the SHANK3.…”

“…Western blot analysis was performed as described previously in detail (Sheng et al, 2021). Mice retinal extract samples were loaded, subjected to 8% sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS/PAGE), and then transferred onto polyvinylidene fluoride (PVDF) membranes (Merck Millipore, Ireland).…”

Pan-retinal ganglion cell markers in mice, rats, and rhesus macaques

“…BRN3A expression in adult RGCs has been described in fish ( Sato et al., 2007 ), birds ( chickens and pigeons, Galindo-Romero et al., 2016 ; Sheng et al., 2021 ), and mammals ( mice, rats, cats, ground squirrels, humans, monkeys, pigs, dogs, alpacas, and blind moles, Ávila-García et al., 2012 ; Bouskila et al., 2020 ; Esquiva et al., 2016 ; Galindo-Romero et al., 2011 ; Kim et al., 2006 ; Nadal-Nicolás et al., 2009 ; Ruzafa et al., 2017 ; Wang et al., 2015 ; Xiang et al., 1995 ; Xiao et al., 2021 ; Zhang et al., 2010 ).…”