Heterogeneity of GABAA receptor in goldfish retina.
The possibility of GABAA receptor heterogeneity in goldfish retina was studied with immunocytochemical and biochemical approaches: 1) immunoblotted membrane particulates of goldfish retina with mAb 62-3G1; 2) immunoprecipitation of the detergent-solubilized membrane proteins with mAb 62-3G1 followed by the receptor binding assay; 3) photoaffinity labeling of the membrane particulates with 3H-flunitrazepam (FNZ) and visualization of the labeled receptors by SDS-PAGE and fluorography; 4) dry autoradiography of 3H-muscimol and 3H-FNZ binding sites on frozen sections. Immunoblots showed that 62-3G1 reacted with 55-57.5 kDa M(r) polypeptides, similar to the muscimol-binding subunit of the receptor complex in bovine brain; while 3H-FNZ photoaffinity labeled the 52.5 kDa and 41-43 kDa M(r) polypeptides. Immunoprecipitated receptors bound only 3H-muscimol, not 3H-FNZ. An attempt to precipitate the 3H-FNZ photolabeled polypeptides failed. Dry autoradiography showed 3H-FNZ binding only in the inner plexiform layer (IPL); the binding was enhanced with gamma-aminobutyric acid (GABA) and blocked by clonazepam. In contrast, 3H-muscimol was bound in both the outer plexiform layer (OPL) and IPL, similar to that observed with 62-3G1 immunocytochemistry. We suggest that there are two subtypes of GABAA receptor in the goldfish retina: 1) GABAA receptors that are not linked to a benzodiazepine (BZD) receptor are located in the OPL and at amacrine-to-amacrine and amacrine-to-ganglion cell synapses in the IPL and are recognized by 62-3G1; 2) GABAA receptors that are linked to a BZD receptor are located only in the IPL, largely at amacrine-to-bipolar cell synapses and are not recognized by mAb 62-3G1.
Lin
Z S
ZS
Department of Neurobiology and Behavior, University at Stony Brook, New York 11794-5230.
Yazulla
S
S
D000345:Affinity Labels; D000818:Animals; D000911:Antibodies, Monoclonal; D001345:Autoradiography; D002417:Cattle; D005445:Flunitrazepam / Q000493:pharmacokinetics; D006054:Goldfish / Q000378:metabolism*; D015151:Immunoblotting; D007150:Immunohistochemistry; D007202:Indicators and Reagents; D008970:Molecular Weight; D009118:Muscimol / Q000493:pharmacokinetics; D011963:Receptors, GABA-A / Q000378:metabolism*; D012160:Retina / Q000166:cytology / Q000378:metabolism*; D013570:Synaptic Membranes / Q000378:metabolism; D009435:Synaptic Transmission / Q000502:physiology; D005680:gamma-Aminobutyric Acid / Q000502:physiology
D000345:Affinity Labels; D000911:Antibodies, Monoclonal; D007202:Indicators and Reagents; D011963:Receptors, GABA-A; D009118:Muscimol; D005680:gamma-Aminobutyric Acid; D005445:Flunitrazepam