IDENTIFICATION AND CHARACTERIZATION OF ALPHA-2D-ADRENERGIC RECEPTORS IN BOVINE PINEAL-GLAND

The vertebrate mechanisms for generating circadian rhythms, regulating the synthesis of melatonin, and modulating the activities of pineal indole-synthesizing enzymes differ significantly among species. The synthesis of melatonin in mammalian pineal glands is stimulated by beta-1-adrenergic receptor agonists and is potentiated by alpha-1-adrenergic receptor agonists, whereas in the avian pineal gland alpha-2-adrenergic receptor agonists inhibit its synthesis. By using [H-3]rauwolscine, a selective alpha-2-adrenergic receptor antagonist, we have identified for the first time alpha-2-adrenergic receptor sites in a mammalian pineal gland. [H-3] Rauwolscine bound in a saturable manner to a single class of receptors, with an equilibrium dissociation constant (K(D)) of 1.4 nM and a density (B(max)) of 71 fmol/mg of protein, in crude synaptic membrane preparations from bovine pineal gland. Competition studies carried out with various adrenergic antagonists supported the conclusion that [H-3]rauwolscine binding sites were alpha-2-adrenergic receptors. The bovine pineal alpha-2-adrenergic receptor appears to represent a pharmacological subtype distinct from the three currently proposed subtypes, i.e., alpha-2A found in a human colonic adenocarcinoma cell line (HT29 cell), alpha-2B found in rat lung, and alpha-2C found in an opossum kidney cell line (OK cell). However, the pharmacologic profile of the pineal alpha-2 receptor resembles that found in the rat submaxillary gland. We suggest that the bovine pineal receptor may represent a fourth pharmacological subtype, which would be designated as alpha-2D.