Expression, subcellular localization, and regulation of sigma receptor in retinal muller cells.
OBJECTIVE
Sigma receptors (sigmaRs) are nonopioid, nonphencyclidine binding sites with robust neuroprotective properties. Type 1 sigmaR1 (sigmaR1) is expressed in brain oligodendrocytes, but its expression and binding capacity have not been analyzed in retinal glial cells. This study examined the expression, subcellular localization, binding activity, and regulation of sigmaR1 in retinal Müller cells.


METHODS
Primary mouse Müller cells (MCs) were analyzed by RT-PCR, immunoblotting, and immunocytochemistry for the expression of sigmaR1, and data were compared with those of the rat Müller cell line (rMC-1) and the rat ganglion cell line (RGC-5). Confocal microscopy was used to determine the subcellular sigmaR1 location in primary mouse MCs. Membranes prepared from these cells were used for binding assays with [3H]-pentazocine (PTZ). The kinetics of binding, the ability of various sigmaR1 ligands to compete with sigmaR1 binding, and the effects of donated nitric oxide (NO) and reactive oxygen species (ROS) on binding were examined.


RESULTS
sigmaR1 is expressed in primary mouse MCs and is localized to the nuclear and endoplasmic reticulum membranes. Binding assays showed that in primary mouse MCs, rMC-1, and RGC-5, the binding of PTZ was saturable. [3H]-PTZ bound with high affinity in RGC-5 and rMC-1 cells, and the binding was similarly robust in primary mouse MCs. Competition studies showed marked inhibition of [3H]-PTZ binding in the presence of sigmaR1-specific ligands. Incubation of cells with NO and ROS donors markedly increased sigmaR1 binding activity.


CONCLUSIONS
MCs express sigmaR1 and demonstrate robust sigmaR1 binding activity, which is inhibited by sigmaR1 ligands and is stimulated during oxidative stress. The potential of Müller cells to bind sigmaR1 ligands may prove beneficial in retinal degenerative diseases such as diabetic retinopathy.
Jiang
Guoliang
G
Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta, GA 30912-2000, USA.
Mysona
Barbara
B
Dun
Ying
Y
Gnana-Prakasam
Jaya P
JP
Pabla
Navjotsin
N
Li
Weiguo
W
Dong
Zheng
Z
Ganapathy
Vadivel
V
Smith
Sylvia B
SB
D000818:Animals; D001667:Binding, Competitive; D015153:Blotting, Western; D018929:Cell Culture Techniques; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation / Q000502:physiology*; D008024:Ligands; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal; D009457:Neuroglia / Q000187:drug effects / Q000378:metabolism*; D018384:Oxidative Stress; D010423:Pentazocine / Q000378:metabolism / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D017480:Receptors, sigma / Q000235:genetics* / Q000378:metabolism*; D012160:Retina / Q000187:drug effects / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000097605:Sigma-1 Receptor
D008024:Ligands; D010423:Pentazocine; D012333:RNA, Messenger; D017480:Receptors, sigma; D000097605:Sigma-1 Receptor