Neurochem. Int. Vol. 9, No. 1, pp. 35-42, 1986 0197-0186/86 $3.00+ 0.00 Printed in Great Britain PergamonJournals Ltd NEUROTENSIN IN THE RETINA: IDENTIFICATION, RELEASE AND RECEPTOR ANALYSIS NEVILLE N. OSBORNE Nuffield Laboratory of Ophthalmology, University of Oxford, Walton Street, Oxford OX2 6AW, U.K. (Received 4 September 1985; accepted 23 November 1985) Abstract--1. Neurotensin-like immunoreactivity is associated with a subpopulation of multistratified amacrine cells situated in the inner nuclear and plexiform layers of the chick retina. 2. HPLC and radioimmunoassay have shown that the neurotensin immunoreactive material in the chick retina is similar to neurotensin ~ ~3, previously identified in bovine and human tissues. 3. Isolated chick retina does not take up 3H or ~2~I-labelled neurotensin, as revealed by autoradiography. 4. By increasing the external K +-concentration, it was shown that the release of endogenous neurotensin from the retina was stimulated. This release is Ca2+-dependent. 5. The binding of [3H]neurotensinto crude membrane preparations of chick retina was saturable and revealed a single population of binding sites with a KD-valueof 3.4 nM and a Bma x of 115 pmol/g protein. The characteristics of these binding sites suggest the presence of neurotensin receptors. 6. It is proposed that neurotensin is a likely transmitter in the avian retina. Neurotensin is a tridecapeptide first isolated and characterised in the central nervous system by Car- raway and Leeman (1973, 1975a, b). It has been reported as having a wide variety of physiological actions, among which are effects on glucoregulation, thermoregulation and neuroendocrinregulation (Carraway and Leeman, 1976; Carraway et al., 1976; Nagyi and Frohman, 1976; Andersson et al., 1976; Brown and Miller, 1982; Nemeroffet al., 1977; Rivier et al., 1977; Maeda and Frohman, 1978). Several lines of evidence also suggest that neurotensin subserves a neurotransmitter role in the CNS. It is preferentially localised in synaptosomal fractions of brain hom- ogenates (Uhl and Snyder, 1977) and it is released from hypothalamic fragments after depolarisation by potassium; this release is calcium-dependent (Iversen et al., 1978; Maeda and Frohman, 1981) and neurotensin appears to interact with specific high affinity binding sites on neuronal elements. These binding sites exhibit a pattern of distribution in the CNS similar to that of neurotensin-like immuno- reactivity (Uhl and Snyder, 1977; Young and Kunhar, 1981). In the retina, neurotensin-like activity has been clearly observed in bird (Brecha et al., 1980; Torn- quist et al., 1981; Fakuda et al., 1981), turtle (Eldred and Karten, 1983; Weiler and Ball, 1984) and goldfish (Li et al., 1985). In the bird retina immunoreactivity is associated with a subpopulation of multistratified amacrine cells. These neurones are strikingly similar in appearance to enkephalin and somatostatin immu- noreactive amacrine cells. Double label immuno- histochemical experiments have demonstrated that there is no overlap of staining of each cell-type in the same section, thus showing that neurotensin- immunoreactive cells in bird retinas represent a dis- tinct population of neurones (see Brecha et al., 1983). In the turtle and goldfish retinas, some neurotensin amacrine cells display properties which suggest that these neurones may utilize more than one putative transmitter. These cells in the turtle retina can take up exogenous glycine (Weiler and Ball, 1984), while in the goldfish retina certain neurotensin positive cells also contain substance P-like immunoreactivity (Li et al., 1985). The restriction of neurotensin immunoreactivity to selective retinal cell populations in itself suggests that the peptide plays a specific role in the retina, perhaps that of a neurotransmitter, or neuromodulator, or both. The only other data to support such a role for neurotensin in the retina come from the electro- physiological studies by Dick and Miller (1981) who showed that neurotensin excites retinal ganglion cells. In the present study some further data are provided to lend credence to the idea that neurotensin may have a transmitter function in certain (bird)retinas. There are findings on the binding of [3H]neurotensin to retinal membranes and on the release, localization and characterization of neurotensin immuno- reactivity in the chick retina. 35