[3H]Kainate and L-[3H]glutamate binding sites in a rich source of kainate binding sites, fish brain, have been thoroughly analysed here for the purpose of studying the correlation between kainate binding sites and L-glutamate receptors in vertebrate CNS. The brain of a freshwater fish, Telapilia monsanbica, was found to contain three types of kainate binding sites: Type 1 sites (Kd = 1050 +/- 380 microM, Bmax = 4 +/- 4 pmol/mg), Type 2 sites (Kd = 133 +/- 20 nM, Bmax = 190 +/- 20 pmol/mg), and Type 3 sites (Kd = 23 +/- 15 nM, Bmax = 28 +/- 19 pmol/mg). The dissociation constants of L-glutamate to Type 1, 2 and 3 sites were, respectively, 0.28 +/- 0.04, 5.5 +/- 0.2 and 137 +/- 28 microM. Pharmacological characterization of these binding sites showed that Type 1 and 2 sites, respectively, corresponded to N-methyl-D-aspartate-subtype L-glutamate receptors and non-N-methyl-D-aspartate L-glutamate receptors. Autoradiographic studies showed that Type 1 and 2 sites were distributed widely in fish brain, indicating the involvement of L-glutamate receptors in various brain functions. Type 3 sites, on the other hand, were relatively insensitive to most endogenous amino acids and were only found in the molecular layer of cerebellum and torus longitudinalis. Type 3 sites possibly representing a distinctive class of receptor has been suggested by the results.