Daily Modifications of 3h-Naloxone Binding Sites in the Rat Brain: A Quantitative Autoradiographic Study

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Abstract
The endogenous opioid peptides, the opiate receptors and several related behaviours, like opioid-mediated analgesia, show daily variations in different animal species including rats. The attempt to correlate the daily rhythm of opiate receptors in the central nervous system (CNS) to opiate related rhythmic phenomena requires an experimental approach with a high anatomical resolution, as the opioid distribution is very heterogeneous. In this paper we present the study of daily variations of 3H-naloxone binding sites in the different regions of the adult male rat brain, performed by means of quantitative autoradiography. Five rats are sacrificed at each investigated time of the day (0200, 0600,1000,1400,1800 and 2200). The ligand is 3H-naloxone(4nM), the quantification is performed by means of densitometric procedures (image analyzer Tesak VDC 501, computer Digital PDP11,3H-microscale). The statistical analysis is performed according to the single Cosinor method and the one-way analysis of variance followed by the multiple range test of Duncan. We analysed 33 different regions of the rat CNS, and the daily variations of opiate receptors are regionally selective. A circadian rhythm is found in the anterior cingulate cortex, hippocampal cortex, periventricular, medial, ventral, reticular and posterior nuclei of the thalamus, rhomboid, gelatinosus and rheuniens nuclei, lateral hypothalamus, locus coeruleus, grey substance of the pons, reticular formation of medulla oblongata, inferior olivary complex, medial part of the nucleus of the solitary tract and nucleus of the spinal tract of the trigeminal nerve. An ultradian rhythm is found in the medial and lateral preoptic areas, in the medial hypothalamus, in the medial and in the lateral nuclei of habenula. No significant variations during 24 hr according to the Cosinor analysis are found in the dorsal and lateral cerebral cortex, striatum, globus pallidus, bed nucleus of the stria terminalis, septal nuclei, lateral nucleus of the thalamus, cochlear nuclei, nucleus of the solitary tract, lateral and caudal parts, dorsal motor nucleus of the vagal nerve, XII and IX nerve nuclei. The amplitude of the daily variations observed ranges from 10 to 40%. Our results demonstrate the high anatomical selectivity of the daily modifications of 3H-naloxone binding sites in the rat CNS. They also indicate that quantitative autoradiography is a suitable and sensitive technique for these studies.