Quantitative autoradiographic localization in rat brain of α2-adrenergic and non-adrenergic I-receptor binding sites labelled by [ 3H]rilmenidine

Abstract

α 2A-Adrenergic receptor (AR) and non-adrenergic imidazoline receptor (I-R) binding sites have been previously characterized in rat cerebral cortex membranes using the N-substituted oxazoline, [ 3H]rilmenidine ([ 3H]Ril) [King, P.R. et al, Eur. J. Pharmacol., 218 (1992) 101–108]. In the present study, in vitro autoradiography was used to quantify the regional distribution of these receptors throughout the rat neuroaxis. The distribution and relative density (fmol/mg tissue) of I-Rs was examined in the presence of 1 μM adrenaline to block the adrenergic component of 40 nM [ 3H]Ril binding and non-specific binding was measured in the presence of another oxazoline, Bay a6781 (10 μM). Both α 2A-ARs and I-Rs were broadly, but heterogeneously, distributed. In forebrain, high levels of [ 3H]Ril-labelled α 2A-AR sites were observed in the anterior olfactory nucleus, the piriform, entorhinal and perirhinal cortices, lateral septum, bed nucleus of the stria terminalis, several thalamic nuclei, the amygdala and the arcuate, dorsomedial and posterior hypothalamic nuclei. In hindbrain, α 2A-AR sites were concentrated in locus coeruleus, lateral parabrachial nucleus, nucleus of the solitary tract and area postrema. I-R sites accounted for 50% or more of specific [ 3H]Ril binding (40 nM) in most cortical and hypothalamic nuclei, nucleus of the solitary tract, cranial motor nuclei and most spinal cord layers. The highest densities of I-Rs were found in the arcuate, dorsomedial and posterior hypothalamic nuclei, the locus coeruleus, the area postrema, the cranial motor nuclei and associated with spinal motor neurones. A very high concentration of I-Rs was also detected in the pineal gland. The distribution of α 2-AR sites determined resembled that reported with [ 3H] p-aminoclonidine which appears to specifically label α 2-ARs and not I 1-R sites in rat brain sections, and [ 3H]methoxyidazoxan which is a selective α 2-AR antagonist. The regional and cellular distribution of I-R binding sites was unlike the distribution of putative I 1-R sites labelled by [ 3H]clonidine in human brain, although comparable autoradiographic mapping studies in rat brain have not been done using this ligand. The regional and cellular distribution of [ 3H]Ril-labelled I-R binding sites had both similarities and differences to that reported using the imidazoline ligand, [ 3H]idazoxan, with common labelling of areas such as area postrema, arcuate and interpeduncular nuclei and pineal gland with the two ligands, and differential relative binding levels ([ 3H]Ril > [ 3H]idazoxan) associated with hippocampal pyramidal cells and brainstem and spinal motor neurones. Thus, according to the current classification of I-Rs (I 1-, I 2A- or I 2B-subtypes) based on various pharmacological, biochemical, anatomical and functional characteristics of the sites, our results suggest that in rat brain [ 3H]Ril labels a population of binding sites most similar to the described I 2B-subtype of I-R (i.e. a site-labelled by imidazolines and derivatives, oxazolines and guanidinium compounds; with low affinity for clonidine and imidazoles; insensitive to amiloride) but may also label additional oxazoline-preferring binding sites in some brain regions.