Although there are many glutamate receptors in the retina, 2-amino-4-phosphonobutyrate (L-AP4) is an agonist that acts selectively at metabotropic glutamate receptors (mGluR6) of ON bipolar cells. We explored the properties of agonists that activate this receptor. The effects of various glutamate analogs on the b-wave of the electroretinogram (ERG) were used as a measure of their activity. Conformational comparisons among agonists suggest that ligands in an extended conformation preferentially bind to the ON bipolar synaptic receptor. But this property is insufficient to explain the selectivity of mGluR6 because some inactive glutamate analogs could also match this extended conformation. Comparative molecular field analysis (CoMFA) was used to compare the electrostatic and steric potentials of agonists with their action at the ON bipolar synapse. Steric potentials beneath a plane defined by the three putative binding sites plays a key role in determining agonist activity. The CoMFA model was used to predict the activity of glutamate analogs and correlations between predicted and measured activity support the model.