Sodium channel expression detected with antibody 7493 in A2B5+ and A2B5- astrocytes from rat optic nerve in vitro.
Astrocytes cultured from neonatal rat optic nerve can be classified into two subtypes, distinguished by their morphology (stellate or fibroblast-like) and their ability to bind monoclonal antibody A2B5. The presence of sodium channels in astrocytes cultured from rat optic nerve was demonstrated by indirect immunofluorescence with polyclonal antibody 7493, which is directed against purified rat brain sodium channel protein. Astrocytes cultured from postnatal day 7 (P7) rat optic nerves exhibited sodium channel immunostaining on both A2B5+ and A2B5- astrocytes up to 6 days in vitro (DIV). Staining was distributed throughout the cytoplasm and cell processes, with areas of greater intensity in the perinuclear region. At 6 DIV, the A2B5-/GFAP+ cells exhibited a loss of sodium channel immunostaining, while the A2B5+/GFAP+ cells continued to display 7493 immunoreactivity. This sodium channel staining pattern persisted for up to 28 DIV (the longest time point examined). Astrocyte cultures derived from PO rat optic nerves exhibited sodium channel immunoreactivity during the first 6 DIV. The P0 astrocyte cultures, in which, the vast majority of cells are A2B5-/GFAP+, displayed a similar staining pattern to those astrocytes with corresponding phenotype derived from P7 optic nerves. P0-derived A2B5- astrocytes showed loss of 7493 immunostaining at 6 DIV, while the rare (less than 1% of cells) A2B5+/GFAP+ cells continued to express sodium channels reactive to 7493. The reduction of sodium channel immunoreactivity in A2B5- but not A2B5+, astrocytes from both P0 and P7 optic nerves after a similar latency (approximately 6 DIV) suggests that the loss of immunostaining may result from the absence of neuronal associations in the culture environment, rather than an intrinsic biologically timed change in astrocytic expression of sodium channels.
Minturn
J E
JE
Department of Neurology, Yale University School of Medicine, New Haven, Connecticut 06510.
Black
J A
JA
Angelides
K J
KJ
Waxman
S G
SG
D000818:Animals; D000911:Antibodies, Monoclonal; D001253:Astrocytes / Q000378:metabolism*; D002470:Cell Survival; D002478:Cells, Cultured; D007150:Immunohistochemistry; D009900:Optic Nerve / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D015222:Sodium Channels / Q000378:metabolism*; D013997:Time Factors
D000911:Antibodies, Monoclonal; D015222:Sodium Channels