Alain Dubois

Autoradiographic localization of D1 dopamine receptors in the rat brain with [3H]SCH 23390

The regional distribution of D1 dopamine (DA) receptors in the rat brain has been studied by quantitative autoradiography using the specific D1 antagonist [3H]SCH 23390 as a ligand. The binding of [3H]SCH 23390 to striatal sections was saturable, stereospecific, reversible and of high affinity (Kd = 2.05 nM); it occurred at a single population of sites and possessed the pharmacological features of the D1 DA receptor. The highest densities of [3H]SCH 23390 binding sites were found in the caudate-putamen, olfactory tubercle, nucleus accumbens and substantia nigra (especially in the pars compacta). High densities were also observed in the nucleus interstitialis striae terminalis, the anterior olfactory nucleus, the entopeduncular nucleus, the subthalamic nucleus, the claustrum and the amygdalohippocampal area. An intermediate labelling was found in the anteromedial and suprarhinal DA terminal fields of the cerebral cortex, the basolateral, medial and lateral amygdaloid nuclei, the endopiriform nucleus, the primary olfactory cortex, the globus pallidus, the superior colliculus (especially the superficial layer), the nucleus amygdaloideus corticalis and the dorsal hippocampus (molecular layer of the CA1 and dentate gyrus). In the anteromedial and suprarhinal cortices, [3H]SCH 23390 binding was more concentrated in layers V and VI. Moderate levels of [3H]SCH 23390 were found in the thalamus, hypothalamus, the habenula, the ventral tegmental area, the posterior cingulate and entorhinal cortices, the supragenual dopamine terminal system and the cerebellum (molecular layer). This regional distribution of [3H]SCH 23390 closely correlated (except for the cerebellum) with the reported distribution of dopaminergic terminals. The topographical distribution of [3H]SCH 23390 has also been studied in detail in striatal subregions. The density of D1 receptors was much greater in the ventrolateral sector and medial margin of the striatum than in the ventromedial and dorsolateral sectors. A rostrocaudal decrease in the densities of D1 sites was also found along the rostrocaudal axis of the caudate-putamen. These lateral to medial and anteroposterior gradients overlapped with the density of the dopaminergic afferents.

Imaging of primary and remote ischaemic and excitotoxic brain lesions. An autoradiographic study of peripheral type benzodiazepine binding sites in the rat and cat.

Seven days after unilateral middle cerebral artery occlusion in rats, peripheral type benzodiazepine binding sites (PTBBS), using [3H]PK 11195 as a specific radioligand, were greatly increased in the cortical and striatal regions surrounding the focus of infarction with smaller increases in the ventrolateral and posterior thalamic complexes and in the substantia nigra, all ipsilateral to the occlusion. Similarly, PTBBS increases were observed in the caudate nucleus and entorhinal cortex of cats likewise subjected to prior unilateral occlusion of the middle cerebral artery. Intrastriatal administration of N-methyl-D-aspartate (250 nmol) in the rat resulted in a dramatic ipsilateral increase in PTBBS levels in the striatum and in the deeper laminae of the ipsilateral frontoparietal cortex. Intrastriatal kainic acid administration (12 nmol) also elicited PTBBS increases ipsilaterally in rat striatum and cortex; a bilateral elevation of PTBBS levels was observed in the hippocampus. With all these interventions there existed a good spatial correlation between the PTBBS increase and neuronal loss as assessed either histologically or by the autoradiographic detection of the putative neuronal marker [3H]SCH 23390 (a D1 dopamine receptor ligand). Moreover, a glial proliferation of non-neuronal cells (macrophage and glial cells) was observed in brain regions noted to have increased PTBBS levels. PTBBS autoradiography thus constitutes a suitable technique for the localization of damaged areas in several experimental models of brain injury. PTBBS label not only the primary lesions but also functionally related areas and could further our understanding of phenomena such as partial neuronal loss and diaschisis. The study of PTBBS could be envisaged for the detection, localization and quantification of all neuropathological situations which engender a glial reaction or macrophage invasion and is potentially applicable to both experimental and human subjects, in which both autoradiographic and tomographic approaches could be undertaken.

Different plasticity changes in D1 and D2 receptors in rat striatal subregions following impairment of dopaminergic transmission

The precise topographical changes in striatal D1 and D2 dopamine receptor density that occurred after chronic treatment with haloperidol or SCH 23390 or after 6-hydroxydopamine-induced lesion of the mesostriatal dopaminergic pathway have been studied autoradiographically in the rat. Repeated treatment with SCH 23390 (0.5 mg/kg i.p., 21 days) caused an almost similar increase in [3H]SCH 23390 binding sites in the different striatal subregions whereas lesion of the dopaminergic pathway was ineffective. Subacute administration of haloperidol (2 mg/kg i.p., 18 days) or lesion of dopaminergic afferents provoked an increase in [3H]spiperone binding which was restricted to the ventro- and dorsolateral striatal sectors.

Denervation supersensitivity of striatal D2 dopamine receptors is restricted to the ventro- and dorsolateral regions of the striatum

The precise topographical changes in striatal D2 dopamine receptors that occur after neurotoxic lesion of the mesostriatal dopaminergic pathway have been studied autoradiographically in the rat through the use of [3H]spiperone as a ligand. 6-Hydroxydopamine-induced lesion of the dopaminergic afferents to the striatum caused an increase in [3H]spiperone binding in the ventro- and dorsolateral but not in the ventro- and dorsomedian aspects of the striatum. This lesion caused a loss of tyrosine hydroxylase-like immunoreactivity in all striatal subregions. These results demonstrate that not all striatal D2 dopamine receptors are able to proliferate after dopaminergic denervation.

The quantification of brain lesions with anω3 site ligand: a critical analysis of animal models of cerebral ischaemia and neurodegeneration

Previous investigations have indicated that the detection and quantification of omega 3 (peripheral type benzodiazepine) binding site densities that are associated with reactive astroglia and macrophages could be of widespread applicability in the localization and indirect assessment of neural tissue damage in the central nervous system. In the present study, we analyze the usefulness of this approach in a number of experimental models that are characterized by (or putatively involve) neuronal degeneration. One week after the systemic administration of the excitotoxin, kainate, a marked increase in omega 3 site densities (as assessed by [3H]PK 11195 binding) was noted, an increase that was most prominent in known regions of selective vulnerability (hippocampus and septum). However, the kainate-induced omega 3 site proliferation was not a function of the dose administered, a marked interstudy variation was observed, and the binding increase was prevented by the administration of the anticonvulsant, clonazepam. The densities of omega 3 sites were studied, by autoradiography (using [3H]PK 11195 or [3H]PK 14105 as ligands), in 4 groups of Fischer 344 rats aged 3, 12, 22 and 30 months. No age-related changes were noted except in the 30-month-old group in which discrete and focal increases (reflecting tumoral processes) were observed in various brain regions. In spontaneously hypertensive, stroke-prone rats, omega 3 binding increases were observed concomitant with the development of stroke-related neurological signs. With autoradiography, the omega 3 site increase was localized to focal increases in the boundary zones between major cerebral arteries (and corresponding to regions of ischaemic or haemorrhagic infarction). Focal cerebral ischaemia was studied in rats and mice. Subsequent to middle cerebral artery occlusion in normotensive (Wistar/Kyoto) and spontaneously hypertensive rats, the density of omega 3 sites in the ipsilateral hemisphere was markedly elevated, the increase being greater in the spontaneously hypertensive rats. The increases in omega 3 labelling in these two strains matched the absolute volumes of infarctions, determined previously. Middle cerebral artery occlusion in the mouse also increased hemispheric levels of omega 3 sites; the maximum values were obtained between 4 and 8 days following the induction of focal ischaemia. These results demonstrate the feasibility of using omega 3 sites as a marker of excitotoxic, ischaemic and proliferative damage in the rodent brain. Binding measurement in tissue homogenates is an economic and time-efficient approach, whereas the autoradiographic detection of omega 3 sites allows the localization of brain lesions with a macroscopic or microscopic level of anatomical resolution.(ABSTRACT TRUNCATED AT 400 WORDS)

Different neuronal location of [3H]SCH 23390 binding sites in pars reticulata and pars compacta of the substantia nigra in the rat

The precise neuronal localization of D1 receptors in the substantia nigra has been studied autoradiographically in the rat by measuring the alterations of [3H]SCH 23390 binding site densities in this brain area after 6-hydroxydopamine (6-OHDA) induced destruction of nigrostriatal dopaminergic neurons and after ibotenate-induced lesion of striatal afferents. 6-OHDA-induced nigral lesion provoked a total loss of [3H]SCH 23390 binding sites in the pars compacta and pars lateralis (but not in the pars reticulata) of the substantia nigra. In contrast, ibotenate-induced striatal lesion caused a large diminution of the [3H]ligand binding site density in the pars reticulata but not in the pars compacta and pars lateralis of the substantia nigra. These results suggest that D1 receptors in the pars compacta or pars lateralis of the substantia nigra are located on the dopaminergic perikarya whereas those D1 receptors present in the pars reticulata of the substantia nigra lie on the terminals of nigral afferents of striatal origin.

Heterogeneous distribution of dopamine D2 receptors within the rat striatum as revealed by autoradiography of [3H]N-n-propylnorapomorphine binding sites.

The precise topographical distribution of dopamine D2 receptors in the rat striatum has been studied autoradiographically through the use of [3H]N-n-propylnorapomorphine (NPA) as a ligand. The highest densities of [3H]NPA binding sites were found in the ventrolateral aspect and medial margin of the striatum, while sparse grain densities were present in the medial and latero-dorsal striatum. Moreover, a clear gradient of [3H]NPA binding was observed along the rostro-caudal axis of the striatum. These heterogeneities in the distribution of dopamine receptors provide further evidence for a functional compartmentalization of the striatum.

Autoradiographic localization and quantification of dopamine D2 receptors in normal human brain with [3H]N-n-propylnorapomorphine

The precise distribution of dopamine receptors has been studied autoradiographically in the normal human brain using [3H]N-n-propylnorapomorphine ([3H]NPA) as a ligand. Preliminary experiments aimed at optimizing the binding assay conditions revealed that preincubation washing of caudate nucleus sections was a prerequisite to obtain a good ratio of specific to non-specific binding. The binding of [3H]NPA to caudate-putamen sections was saturable, stereospecific, reversible, of high affinity (Kd = 0.27-0.35 nM) and occurred at a single population of sites. Competition experiments with various drugs indicated that in the caudate-putamen the specific [3H]NPA binding sites possess the pharmacological features of the dopamine D2 receptor. The highest levels of [3H]NPA binding sites were found in the caudate nucleus, putamen, globus pallidus and nucleus accumbens. There were also intermediate to low concentrations of the 3H-ligand in the hippocampus, the insular and cingular cortices and in the occipito-temporal gyrus, while almost undetectable levels of binding were found in the anterior frontal cortex. Thorough examination of the subregional distribution of [3H]NPA binding sites in the caudate-putamen-pallidum complex revealed heterogeneous patterns of radioactivity. In these brain regions, the distribution of autoradiographic grains was punctate and islands of high and low densities were observed. Moreover, in the caudate nucleus, there was a subtle high lateral to low medial gradient in the topography of the [3H]NPA binding sites and a more pronounced gradient along the rostrocaudal axis; the highest levels of binding being located at the midbody of the nucleus. No gradients of [3H]NPA binding were observed in the putamen. The present data indicate that [3H]NPA is a suitable ligand for accurate autoradiographic labeling of dopamine D2 receptors in human postmortem brain tissue and that dopamine receptors are heterogeneously distributed and topographically organized in patches and gradients in the basal ganglia regions.

Autoradiographic localization of muscarinic cholinergic receptors at various segmental levels of the human spinal cord.

Muscarinic cholinergic receptors were localized autoradiographically at different segmental levels of the normal human spinal cord after in vitro labelling using [3H]quinuclidinylbenzilate. The highest densities of muscarinic receptors were found in laminae II (substantia gelatinosa) and IX (motor neurons) of the gray matter. Lower densities of receptors were present in the other laminae and no binding sites were found in the white matter. This pattern of distribution of muscarinic receptors was similar at the cervical, thoracic, lumbar and sacral levels of the cord. These data suggest that muscarinic receptors may not only take part in motor function but also in the processing of sensory information.

Cellular distribution of ω3 (peripheral type benzodiazepine) binding sites in the normal and ischaemic rat brain: An autoradiographic study with the photoaffinity ligand [3H]PK 14105

The microscopic distribution of omega 3 (peripheral type benzodiazepine) binding sites in sections from normal and ischaemic rat brain has been determined by emulsion autoradiography with the photoaffinity ligand [3H]PK 14105. In the normal rat brain, high densities of omega 3 sites were present in the ependymal cells, the apical pole of the secretory cells in the choroid plexus and astrocyte-like cells in the outer cellular layer of the olfactory bulb. In ischaemic brain lesions caused by middle cerebral artery occlusion in spontaneously hypertensive rats or in spontaneous ischaemic and haemorrhagic lesions in spontaneously hypertensive stroke-prone rats, the proliferating omega 3 sites were associated with reactive glial cells and macrophages.