In this proposal the rabbit retina will be used as a model of CNS tissue to study the pharmacological characteristics of DA receptors involved in functional and biochemical responses. The release of DA elicited by electrical stimulation from pieces of rabbit retina is modulated through DA autoreceptors of the D-2 subtype. The presence of D- and D-2 receptors subtypes was been confirmed with studies of stimulated-cyclase and 3H-spiperone binding respectively. I will investigate the role of calcium and cyclic nucleotides on the release of 3H-DA evoked by several calcium-dependent and -independent stimuli from pieces of retina, as well as their participation on the mechanism by which DA agonists decrease 3H-DA release. The release of 3H-DA from the isolated intact rabbit retina maintained in vitro will be elicited with the normal physiological stimulus light and its modulation through DA autoreceptors investigated. The effect of putative neurotransmitters and neuropeptides on the depolarization evoked release of 3H-DA from retinal pieces will be also studied. The D-2 receptor subtype will be characterized using the binding of the selective D-2 antagonist, 3H-S-sulpiride, and compared with the binding of 3H-spiperone, defined with the isomers of sulpiride. In addition I will attempt to demonstrate that the D-2 receptor subtype of the retina is negatively linked to adenylate cyclase, and its pharmacological profile to DA ergic drugs determined. The pharmacological profile of various DA ergic agents on the D-2 receptor subtype as determined using biochemical methods (binding, adenylate cyclase) will be correlated with the profile of the DA autoreceptor, using functional studies (3H-DA release), in order to prove that biochemical methods could be useful to characterize DA-receptors. The localization of D-2 receptors within the layers of the retina will be assessed with studies on a) 3H-DA release from synaptosomes, b) binding of 3H-spiperone and c) autoradiography, in control retinas and after lesion with neurotoxins. The possible physiological role of DA autoreceptors in regulating the responses of the effector organ will be determined by studying the ability of S-sulpiride to potentiate the increase in cyclic AMP formation elicited by depolarization. Changes in sensitivity of retinal DA receptors linked to functional and biochemical responses will be studied after long-term neuroleptic treatment.
