Binding and uptake of the GABA analogue, 3H-muscimol, in the retinas of goldfish and chicken.

Gamma-aminobutyric acid (GABA) is a candidate as a neurotransmitter in the vertebrate retina. The GABA analogue muscimol has been used to probe the properties of GABA receptors in other parts of the vertebrate central nervous system (CNS). We thus used 3H-muscimol to investigate potential GABA receptors in the retinas of goldfish and chick by means of biochemical assay techniques and light microscopic autoradiography. In both animals 3H-muscimol shows specific and saturable binding with a dissociation constant (KD) of about 10 nM. GABA effectively inhibits 3H-muscimol binding at 50% inhibitory concentration (IC50) of 10(-6) M. The labeling pattern of 7 x 10(-7) M 3H-muscimol shows common features for both species in that amacrine cell bodies are intensely labeled, horizontal cells are much less so, and there is a laminar pattern throughout the inner plexiform layer (IPL). A 1 mM concentration of GABA abolishes 3H-muscimol labeling in the chick retina and throughout much of the goldfish retina except for some label over amacrine cells and the distal two thirds of the IPL. The intense somatic labeling suggests neuronal uptake of 3H-muscimol, and indeed, virtually all 3H-muscimol labeling is abolished with the addition of 0.4 mM ouabain. The uptake pattern of 3H-GABA differs from that of 3H-muscimol and is largely unaffected by the addition of 1 mM muscimol. We conclude that 3H-muscimol binding in retinas can be adequately demonstrated biochemically but that only 3H-muscimol uptake is observed with autoradiography from tissue conventionally processed through Epon. The fact that GABA can inhibit 3H-muscimol uptake whereas the reverse is not the case shows that the transport carriers for muscimol and GABA are different. Finally, the strong degree of 3H-muscimol uptake by retinal neurons raises serious questions about the use of 3H-muscimol as a probe for GABA synaptic receptors in the retina with autoradiography.