Opioid binding sites in the fish brain: an autoradiographic study

Cryostat sections of trout brains were incubated with tritiated opioid ligands (5 nM [3H]etorphine or 4 nM D-[3H]Ala2, Met5 enkephalinamide) with the initial aim of locating opioid binding sites associated with the hypothalamo-pituitary axis. Naloxone-displaceable binding was observed in all regions of the brain, with a density ranking order of cerebellum greater than telencephalon greater than optic tectum greater than hypothalamus greater than brain stem greater than pituitary gland. Within the hypothalamus and pituitary gland binding was unexpectedly low, apart from a slightly enhanced binding anteriorly in the preoptic region and posteriorly associated with presumptive sensory fibres. A similar distribution of opioid binding sites was seen in the eel and lamprey brain. The high level of opioid binding in the cerebellum permitted a tentative identification of opioid subtypes using [3H]etorphine binding to membrane preparations from trout cerebellum. Scatchard analysis indicated the presence of a single class of high-affinity binding sites with a density of 0.38 pmol/mg protein and affinity constant (KD) of 2.6 nM. Displacement by unlabelled ligands suggested the existence of mu and/or kappa receptors.