Expression of AMPA receptor subunit proteins in purified retinal ganglion cells.
OBJECTIVE
To determine whether alpha-amino-3-hydroxy-5-methylisoxazole-4-propioate (AMPA) receptor (AMPAR) subunit proteins are expressed in cultured retinal ganglion cells (RGCs).


METHODS
RGCs were purified from dissociated rat retinal cells (postnatal days 6-8), using a modified two-step panning method and cultured in serum-free medium containing neurotrophic factors and forskolin. Immunohistochemistry was performed on cultured RGCs on days 1, 3, and 7 in vitro (1 DIV, 3 DIV, and 7 DIV) using specific antibodies against AMPAR subunits GluR1 to 4 and microtubule-associated protein (MAP) 2, which is a neuronal marker. Glutamate-induced Ca(2+) influx was measured with fura-2 acetoxymethyl ester fluorescence.


RESULTS
GluR1 to 4 proteins were expressed in the cell body of RGCs on 1 DIV. RGCs showed strong GluR1 to 4 immunoreactivity in both cell bodies and processes on 3 DIV and 7 DIV, with the gradual spreading of expression and the growth of processes. At all time points examined, GluR2 immunoreactivity was equal to that of the other subunits. Accumulation of intracellular Ca(2+) levels in RGCs induced by glutamate occurred equally on both 3 DIV and 7 DIV.


CONCLUSIONS
All AMPAR subunits are almost equally expressed in cultured RGCs.
Miki
Atsuya
A
Department of Ophthalmology and Visual Science, Osaka University Medical School, Suita, Osaka, Japan.
Otori
Yasumasa
Y
Okada
Masaki
M
Tano
Yasuo
Y
D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D018698:Glutamic Acid / Q000494:pharmacology; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D017136:Ion Transport / Q000187:drug effects; D051381:Rats; D018091:Receptors, AMPA / Q000096:biosynthesis*; D012165:Retinal Ganglion Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D002118:Calcium; D016895:Culture Media, Serum-Free; D018698:Glutamic Acid; D018091:Receptors, AMPA