Subchronic haloperidol increases CB(1) receptor binding and G protein coupling in discrete regions of the basal ganglia

The present study was designed to test whether chronic neuroleptic treatment, which is known to alter both expression and density of dopamine D(2) receptors in striatal regions, has effects upon function and binding level of the cannabinoid CB(1) receptor in the basal ganglia by using receptor autoradiography. As predicted, subchronic haloperidol treatment resulted in increased binding of (3)H-raclopride and quinpirole-induced guanosine 5'-O-(gamma-[(35)S]thio)triphosphate ([(35)S]GTPgammaS) in the striatum when compared to that measured in control animals. This increased D(2) receptor binding and function after 3 days washout was normalized after a 2-week washout period. Effect of haloperidol treatment was studied for CB(1) receptor binding and CP55,940-stimulated [(35)S]GTPgammaS in the striatum, globus pallidus, and substantia nigra. (3)[H]CP55,940 binding levels were found in rank order from highest to lowest in substantia nigra > globus pallidus > striatum. Furthermore, subchronic haloperidol treatment resulted in elevated binding levels of (3)[H]CP55,940 in the striatum and the substantia nigra and CB(1) receptor-stimulated [(35)S]GTPgammaS bindings in the substantia nigra after 3 days washout. These increased binding levels were normalized at 1-4 weeks after termination of haloperidol treatment. Haloperidol treatment had no significant effect on CB(1) receptor or [(35)S]GTPgammaS binding levels in globus pallidus. The results help to elucidate the underlying biochemical mechanism of CB(1) receptor supersensitivity after haloperidol treatment.     

Autoradiographic mapping of 5-HT(1B) and 5-HT(1D) receptors in the post mortem human brain using [(3)H]GR 125743

The distribution of 5-HT(1B) and 5-HT(1D) receptors in the human post mortem brain was examined using whole hemisphere autoradiography and the radioligand [(3)H]GR 125743. [(3)H]GR 125743 binding was highest in the substantia nigra and the globus pallidus. Lower levels were detected in the striatum, with the highest densities in the ventromedial parts. In the amygdala, the hippocampus, the septal region and the hypothalamus, lower [(3)H]GR 125743 binding was observed, reflecting low densities of 5-HT(1B/1D) receptors. In the cerebral cortex, binding was similar in most regions, although restricted parts of the medial occipital cortex were markedly more densely labeled. Binding densities were very low in the cerebellar cortex and in the thalamus. Two methods were used to distinguish between the two receptor subtypes, the first using ketanserin to block 5-HT(1D) receptors and the second using SB 224289 to inhibit 5-HT(1B) receptor binding. The autoradiograms indicated that in the human brain, the 5-HT(1B) receptor is much more abundant than the 5-HT(1D) receptor, which seemed to occur only in low amounts mainly in the ventral pallidum. Although [(3)H]GR 125743 is a suitable radioligand to examine the distribution of 5-HT(1B) receptors in the human brain in vitro, the selectivities of ketanserin and SB 224289 are not sufficiently high to give definite evidence for the occurrence of the 5-HT(1D) receptor in the human brain.     

Lesion Study of the Distribution of Serotonin 5-HT4 Receptors in Rat Basal Ganglia and Hippocampus

The regional distribution of 5-hydroxytryptamine (5-HT₄) receptors labelled with [³H]GR113808 was examined in rat basal ganglia and hippocampus after specific lesions. Lesion of serotonin neurons induced by injections of 5,7-dihydroxytryptamine into the dorsal and medial raphe nuclei resulted in increased 5-HT₄ receptor binding in most regions examined, compared with controls. More precisely, there was a 78% increase in the rostral but no change in the caudal part of caudate-putamen, and 83% and 54% increases in the shell and core of the nucleus accumbens respectively. In the substantia nigra, the increase in 5-HT₄ binding was larger (72%) than that in the globus pallidus (32%). In the hippocampus, 63%, 30% and 28% increases were measured in CA2, CA1 and CA3 respectively. Following lesion of dopamine neurons by intranigral injection of 6-hydroxydopamine, increased 5-HT₄ receptor binding was observed in the caudal (59%), but not the rostral part of caudate-putamen, as well as in the globus pallidus (93%). Since no decreases in 5-HT₄ receptor density were detected after the dopamine lesion, it was concluded that these receptors are not expressed in dopamine neurons. Kainic acid lesions of the caudate-putamen were associated with dramatic local decreases in 5-HT₄ receptor binding on the injected side (-89%), which suggested that striatal neurons express 5-HT₄ receptors. Corresponding decreases of 72 and 20% in receptor density were detected in globus pallidus and substantia nigra, consistent with a presumed localization of 5-HT₄ receptors on striatal GABA neurons projecting to these regions. In the substantia nigra, the decrease in [³H]GR113808 binding was localized to the pars lateralis, indicating that striatal neurons belonging to the cortico-striato-nigrotectal pathway, and containing GABA and dynorphin, express 5-HT₄ receptors.     

5-HT1D receptors in the guinea pig brain: pre- and postsynaptic localizations in the striatonigral pathway

The caudate-putamen, globus pallidus and substantia nigra pars reticulata of the guinea pig contain high densities of the 5-HT1D receptor subtype. The cellular localization of these sites in the striatonigral pathway was investigated using receptor autoradiography and selective neurotoxin lesions. In guinea pigs with unilateral 6-hydroxydopamine lesions of the nigral dopaminergic cells, no significant decrease was observed in any of the components of the striatonigral pathway. In contrast, when quinolinic acid was injected in the caudate-putamen, marked reductions in [3H]5-HT binding were seen in the caudate-putamen, the globus pallidus and the substantia nigra pars reticulata, on the side ipsilateral to the lesion. These data, which are comparable to previous results in human pathologies where similar cell populations are known to degenerate (Parkinson disease and Huntington's chorea), indicate a presynaptic localization of 5-HT1D receptors on the terminals of the striatal neurons projecting to the pars reticulata of the substantia nigra. In addition, these receptors could be located on the cell bodies or dendrites of these neurons in the striatum, postsynaptically to serotoninergic fibers.     

Autoradiographic identification and topographical analyses of high affinity serotonin receptor subtypes as a target for the novel putative anxiolytic TVX Q 7821

TVX Q 7821 is a putative non-benzodiazepine anxiolytic which has a high affinity to 5-HT1 receptors. In this study some of the binding characteristics of the radiolabeled drug using rat brain cryostat sections and the autoradiographic localization of binding sites using the LKB-Ultrofilm technique have been investigated. Parallel experiments have been performed using [3H]serotonin ( [3H]5-HT). Both [3H]TVX Q 7821 and [3H]5-HT bound specifically and in a saturable manner to tissue sections, the Kd values being 6.8 and 3.7 nmol/l, respectively. Quantitative autoradiography using computer-assisted image analysis revealed a mean inhibition by TVX Q 7821 of [3H]5-HT binding of 56% in many brain areas. The inhibition ranged from 80% in the hippocampus and entorhinal area to practically none in the substantia nigra and the dorsal subiculum. Color coded autoradiograms obtained either with [3H]5-HT or [3H]TVX Q 7821 showed a nearly identical pattern of labeling with high receptor densities in the hippocampus, the entorhinal area, the septum, the interpeduncular nucleus and the dorsal raphe. However, in some brain areas striking differences in the intensity of labeling were found. [3H]5-HT but not [3H]TVX Q 7821 bound strongly in the substantia nigra, the dorsal subiculum and the globus pallidus. It is proposed that TVX Q 7821 binds to a subtype of 5-HT1 receptor (the so-called 5-HT1A sites as recently proposed). Thus, the putative anxiolytic TVX Q 7821 may provide a means for the study of the functional role of 5-HT1 receptors.     

Quantitative autoradiographic mapping of serotonin receptors in the rat brain. I. Serotonin-1 receptors

The distribution of serotonin-1 (5-HT1) receptors in the rat brain was studied by light microscopic quantitative autoradiography. Receptors were labeled with [3H]serotonin (5-[3H]HT), 8-hydroxy-2-[N-dipropylamino-3H]tetralin (8-OH- [3H]DPAT), [3H]LSD and [3H]mesulergine, and the densities quantified by microdensitometry with the aid of a computer-assisted image-analysis system. Competition experiments for 5-[3H]HT binding by several serotonin-1 agonists led to the identification of brain areas enriched in each one of the three subtypes of 5-HT1 recognition sites already described (5-HT1A, 5-HT1B, 5-HT1C). The existence of these 'selective' areas allowed a detailed pharmacological characterization of these sites to be made in a more precise manner than has been attained in membrane-binding studies. While 5-[3H]HT labeled with nanomolar affinity all the 5-HT1 subtypes, the other 3H-labeled ligands labeled selectively 5-HT1A (8-OH-[3H]DPAT), 5-HT1C ([3H]mesulergine) and both of them ([3H]LSD). Very high concentrations of 5-HT1 receptors were localized in the choroid plexus, lateroseptal nucleus, globus pallidus and ventral pallidum, dentate gyrus, dorsal subiculum, olivary pretectal nucleus, substantia nigra, reticular and external layer of the entorhinal cortex. The different fields of the hippocampus (CA1-CA4), some nuclei of the amygdaloid complex, the hypothalamic nuclei and the dorsal raphé, among others, also presented high concentrations of sites. Areas containing intermediate densities of 5-HT1 receptors included the claustrum, olfactory tubercle, accumbens, central grey and lateral cerebellar nucleus. The nucleus caudate-putamen and the cortex, at the different levels studied, presented receptor densities ranging from intermediate to low. Finally, in other brain areas--pons, medulla, spinal cord--only low or very low concentrations of 5-HT1 receptors were found. From the areas strongly enriched in 5-HT1 sites, dentate gyrus and septal nucleus contained 5-HT1A sites, while globus pallidus, dorsal subiculum, substantia nigra and olivary pretectal nucleus were enriched in 5-HT1B. The sites in the choroid plexus, which presented the highest density of receptors in the rat brain, were of the 5-HT1C subtype. The distribution of 5-HT1 receptors reported here is discussed in correlation with the distribution of serotoninergic neurons and fibers, the related anatomical pathways and the effects which appear to be mediated by these sites.     

Excitatory amino acid receptor regulation after subthalamic nucleus lesions in the rat

The subthalamic nucleus plays a pivotal role in the regulation of basal ganglia output. Recent electrophysiologic, lesion and immunocytochemical studies suggest that the subthalamic nucleus uses an excitatory amino acid as a neurotransmitter. After complete ablation of the subthalamic nucleus, we have examined the NMDA, AMPA, kainate and metabotropic subtypes of excitatory amino acid receptors in two major subthalamic projection areas (globus pallidus and substantia nigra pars reticulata) with quantitative autoradiography. Two weeks after ablation, binding sites for [³H]AMPA and [³H]kainate increased in substantia nigra pars reticulata ipsilateral to the lesion. In globus pallidus on the lesioned side, [³H]glutamate binding to the NMDA recognition site decreased. The results suggest that glutamate receptors regulate after interruption of subthalamic nucleus output.     

Autoradiographic distribution in rat brain of a non-conventional binding site for the opiate []diprenorphine

[³H]Diprenorphine, in the presence of saturating concentrations of μ, δ and opiate receptor blocking agents, was found to label an additional binding site in rat brain, for which various opiates showed moderate affinity. Using light microscopic autoradiography, a unique, highly discrete distribution of binding sites was found, with a high density only in the globus pallidus, substantia nigra, interpeduncular nucleus, parabrachial nuclei and locus coeruleus. The identity of this binding site remains unclear.     

Distribution of the histamine H(2) receptor in monkey brain and its mRNA localization in monkey and human brain

The distribution of histamine H(2) receptor mRNA was determined by in situ hybridization histochemistry in human and monkey brain. In the case of monkey brain, we combined this technique with receptor ligand autoradiography to compare the distribution of mRNA and receptor binding sites. [(125)I]Iodoaminopotentidine ([(125)I]-APT), a reversible, high specific activity antagonist with high affinity and selectivity for the H(2) receptor, was used for receptor autoradiography. Radiolabeled oligonucleotides derived from the human mRNA sequence encoding this receptor were used as hybridization probes. The highest density of the H(2) receptor mRNA in human and monkey brain was found in caudate and putamen nuclei and external layers of cerebral cortex. Moderate levels were seen in the hippocampal formation and lower densities in the dentate nucleus of cerebellum. Areas such as globus pallidus, amygdaloid complex, cerebellar cortex, and substantia nigra were devoid of hybridization signal. The distribution of H(2) receptor mRNA in monkey brain is generally in good agreement with that of the corresponding binding sites: prominent in caudate, putamen, accumbens nuclei, and cortical areas. The hippocampus showed lower densities of receptors and low levels were detected in the globus pallidus pars lateralis. No binding sites were seen in amygdaloid complex and substantia nigra. The distribution of histaminergic innervation is in good correlation with the areas of high density for H(2) receptors: caudate, putamen, and external layers of cerebral cortex in monkey and human brain. The presence of mRNA in caudate and putamen nuclei, together with its absence from substantia nigra, suggests that the H(2) receptors found in the striatum are synthesized by intrinsic cells and not by nigral dopaminergic cells. These striatal H(2) receptors may be located on short circuit striatal interneurons or somatodendritically on striatal projection neurons which project to the globus pallidus pars lateralis. In conclusion, the present results, which constitute, to our knowledge, the first report of the regional distribution of mRNA encoding H(2) receptors detected by in situ hybridization, define the sites of synthesis of H(2) receptors and are the basis for future, more detailed studies that should result in a better understanding of H(2) receptor function.     

Distribution and modulation of histamine H(3) receptors in basal ganglia and frontal cortex of healthy controls and patients with Parkinson's disease

Parkinson's disease (PD) is a brain degenerative disorder with unknown etiology, and specific degeneration of mesencephalic dopaminergic cells is a morphological manifestation of the disease. The central histaminergic system appears to be activated in PD, since the histaminergic innervation is increased in the substantia nigra. The aim of the present study was to investigate the expression and function of histamine H(3) receptors in PD, using receptor mRNA in situ hybridization with oligonucleotide probes, receptor binding assay with a specific radioactive agonist, and GTP-gamma-[(35)S]-binding assay as a tool to study the activation of the receptor G-protein. H(3) receptor binding sites were detected using N-alpha-methylhistamine autoradiography in the basal ganglia and cortex, being most abundant in the substantia nigra and striatum. In PD substantia nigra we detected an increase of the receptor binding density. In situ hybridization study of the receptor mRNA revealed prominent sites of H(3) receptor synthesis in the putamen, cortex, and globus pallidus, whereas very low mRNA expression was seen in the substantia nigra. In the PD pallidum externum, H(3) receptor mRNA expression was elevated as compared with the normal brains. GTP-gamma-[(35)S]-binding assay did not reveal any significant difference between PD and normal brains, although the density values in PD substantia nigra tended to be lower than in the normal brain, and density values in PD striatum were higher. The dopaminergic neurons did not express significant amount of H(3) receptor mRNA, suggesting that the effects of H(3) receptor-mediated modulation of dopamine release are indirect. Our data indicates modulation of the histamine H(3) receptor in PD at the level of the mRNA expression in the striatum and receptor density in the substantia nigra. The receptor activity seems to be unchanged or decreased, as revealed by GTP-gamma-[(35)S]-binding assay. Modulation of the histamine H(3) receptor may influence the activity of other neurotransmitter systems, e.g., the GABAergic one, in the substantia nigra.     

Substance P receptors: localization by light microscopic autoradiography in rat brain using [3H]SP as the radioligand

Substance P (SP) is a putative neurotransmitter in both the peripheral and central nervous systems. In the present report we have used a modification of the Young and Kuhar technique to investigate some of the SP receptors binding properties and the distribution of SP receptors in rat brain. Tritiated SP [( 3H]SP) absorbed extensively to glass but this adsorbtion was greatly reduced by preincubating the slide-mounted tissue sections in a solution containing the cationic polymer polyethylenimine. [3H]SP was found to bind to rat tissue in a saturable fashion with a Bmax of 14.7 fmol/mg tissue wet weight and a Kd of 1.1 nM. The rank order of potencies for displacing [3H]SP binding from rat tissue sections was SP greater than SP sulphoxide greater than DiMeC7 greater than Eledoisin greater than SP(5-11) greater than SP(COOH) greater than SP(1-9) amide. Using autoradiography coupled with LKB tritium-sensitive Ultrofilm or the dry emulsion-coated coverslip technique the distribution of [3H]SP binding sites was found to be very dense within olfactory bulb, amygdalo-hippocampal area and the nucleus of the solitary tract. Heavy concentrations of receptors were observed in the septum, diagonal band of Broca, striatum subiculum, hypothalamus, locus coeruleus, parabrachial nucleus and lobule 9 and 10 of the cerebellum. Moderate to low concentrations of receptors were observed in the cerebral cortex, globus pallidus, raphe nuclei and the trigeminal nucleus. Very low densities were observed in most aspects of the dorsal thalamus, substantia nigra and cerebellum (other than lobule 9 and 10). Comparisons of the present data with SP peptide levels indicate that in some areas of the brain there is a rough correlation between peptide and receptor levels. However, in other brain areas (olfactory bulb, globus pallidus and substantia nigra) there is little obvious correlation between the two.     

5-HT2C receptor binding is increased in the substantia nigra pars reticulata in Parkinson's disease.

The involvement of abnormalities in nondopaminergic transmitter systems in Parkinson's disease is noteworthy because of the complications, such as dyskinesia, associated with long-term dopamine replacement therapy. The output regions of the basal ganglia, the substantia nigra pars reticulata, and the medial segment of the globus pallidus are overactive in Parkinson's disease but underactive in dyskinesia. 5-HT2C receptors are localized in these regions and are excitatory. A 5-HT2C receptor binding assay using [3H]-mesulergine and SB 200646A to define nonspecific binding was applied to postmortem tissue from patients with Parkinson's disease and from age-matched control patients. [3H]-mesulergine binding was increased in the substantia nigra pars reticulata by 108% in Parkinson's disease tissue as compared with control tissue. These data suggest abnormalities of 5-HT2C transmission in the basal ganglia of patients with Parkinson's disease.     

Changes in [3H]zolpidem and [3H]Ro 15-1788 binding in rat globus pallidus and substantia nigra pars reticulata following a nigrostriatal tract lesion

Changes in GABA(A) receptor alpha(1) subunit gene expression occur in the globus pallidus and substantia nigra pars reticulata following lesions of the nigrostriatal tract. To determine whether these changes are translated at the protein level, we performed quantitative autoradiography with the alpha(1) selective ligand, [3H]zolpidem, and the non-selective benzodiazepine site ligand, [3H]Ro 15-1788. Binding of both [3H]zolpidem and [3H]Ro 15-1788 was significantly increased in the substantia nigra pars reticulata (13. 5+/-4.1 and 26.3+/-2.9%, respectively) and significantly reduced in the globus pallidus (20.9+/-0.8 and 18.3+/-1.3%, respectively). These changes in alpha(1) subunit protein expression may help to compensate for the pathological changes in GABAergic activity that occur after striatal dopamine depletion.     

Non 5-HT1A/5-HT1C [3H]5-HT binding sites in the hamster, opossum, and rabbit brain show similar regional distribution but different sensitivity to beta-adrenoceptor antagonists.

We have used receptor autoradiography to investigate the distribution and pharmacological profile of non 5-HT1A/5-HT1C[3H]5-hydroxytryptamine binding sites in the brain of rabbits, hamsters and opossums. These data were compared to those found under similar conditions in the brain of rats and guinea pigs, species which are known to possess 5-HT1B and 5-HT1D receptors, respectively. In the presence of 100 nM 8-OH-DPAT and mesulergine, the regional distribution of [3H]5-hydroxytryptamine binding sites was very similar in the brain of all species investigated; densest labelling was observed in the globus pallidus, substantia nigra and superior colliculus. In all species, 5-carboxamidotryptamine competed for the labelled sites in a biphasic manner and metergoline displayed a subnanomolar affinity. In contrast, the beta-adrenoceptor blocking agents (-)propranolol, (-)pindolol, and (+/-)SDZ 21009 were potent displacers only in the rat, hamster and opossum brains. These data indicate that non 5-HT1A/5-HT1C[3H]5-HT binding sites display a high affinity for these agents in a particular rodent suborder as well as in opossum, a phylogenetically unrelated species.     

Quantitative autoradiography of [H]forskolin binding sites in the rat brain

The binding sites for a radiolabeled form of the potent activator of adenylate cyclase, forskolin, have been localized in the rat brain, pituitary and spinal cord. Using the quantitative technique of in vitro autoradiography, a high density of [3H]forskolin binding was detected in brain structures such as the caudate-putamen, nucleus accumbens, olfactory tubercle, globus pallidus, substantia nigra and the hilus of the area dentata. A comparison of the distribution of [3H] forskolin binding sites with those reported for several neurotransmitter receptor types indicated that forskolin identified adenylate cyclase was probably not linked to any single type of neurotransmitter receptor. These results also presented several new brain areas in which to investigate the neuronal role of adenylate cyclase.     

The distribution of 5-HT(6) receptors in rat brain: an autoradiographic binding study using the radiolabelled 5-HT(6) receptor antagonist [(125)I]SB-258585

We used the highly selective 5-HT(6) receptor radioligand [(125)I]SB-258585 (4-iodo-N-[4-methoxy-3-(4-methylpiperazin-1-yl)phenyl]benzene-sulfonamide) to perform autoradiographic binding studies on the rat brain. High levels of specific binding occurred in the corpus striatum, nucleus accumbens, Islands of Calleja and the olfactory tubercle. A high level of binding also appeared in the choroid plexus. Moderate levels occurred in several regions of the hippocampal formation and in certain regions of the cerebral cortex, thalamus, hypothalamus, and substantia nigra; and very low levels in the globus pallidus, cerebellum, other mesencephalic regions, and the rhombencephalon. Displacement of total binding with 10 microM unlabelled SB-214111 (4-bromo-N-[4-methoxy-3-(4-methylpiperazin-1-yl)phenyl]benzene-sulfonamide), another selective 5-HT(6) receptor antagonist, or 10 microM unlabelled methiothepin, reduced binding to barely discernible levels. Some animals received unilateral injections of 6-hydroxydopamine (6-OHDA) into the median forebrain bundle to lesion the nigro-striatal pathway before autoradiographic examination. Effectiveness of the 6-OHDA lesions in the substantia nigra and striatum was confirmed with tyrosine hydroxylase immunohistochemistry. Such lesions resulted in no significant changes in [(125)I]SB-SB258585 binding in any brain region examined, suggesting that 5-HT(6) receptors in the striatum are not located on dendritic, somatic or terminal elements of dopaminergic neurones. Thus, the striatal binding sites seen in this study may be on intrinsic GABAergic or cholinergic neurones, or on terminals of projection neurones from the thalamus or cerebral cortex. The 5-HT(6) receptor ligand binding seen here in the striatum, accumbens, olfactory tubercle, Islands of Calleja, cerebral cortex and hippocampus are in concordance with previous immunohistochemical studies, and suggest a possible involvement of 5-HT(6) receptors in locomotor control, cognition, memory, and control of affect. The high levels of binding observed in the choroid plexus in this study have not been reported before. This finding suggests that 5-HT(6) receptors could play a role in the control of cerebrospinal fluid dynamics.     

Quantitative autoradiography of multiple 5-HT1 receptor subtypes in the brain of control or 5,7-dihydroxytryptamine-treated rats

The distribution of the 2 main types (A and B) of 5-HT1 binding sites in the rat brain was studied by light-microscopic quantitative autoradiography. The 5-HT1A sites were identified using 3H-8-hydroxy-2-(N-dipropylamino)tetralin (3H-8-OH-DPAT) or 3H-5-HT as the ligand. In the latter case, it was shown that 3H-5-HT binding to 5-HT1A sites corresponded to that displaceable by 0.1 microM 8-OH-DPAT or 1 microM spiperone. The "non-5-HT1A" sites labeled by 3H-5-HT in the presence of 0.1 microM 8-OH-DPAT corresponded mainly to 5-HT1B sites. 5-HT1A binding was notably high in limbic regions (dentate gyrus, CA1 and CA3 hippocampal regions, lateral septum, frontal cortex), whereas 5-HT1B binding was particularly concentrated in extrapyramidal areas (caudate nucleus, globus pallidus, substantia nigra). Except in the latter regions, where only one class of 5-HT1 sites was found, both 5-HT1A and 5-HT1B sites existed in all areas examined. The selective degeneration of serotoninergic neurons produced by an intracerebral injection of 5,7-dihydroxytryptamine was associated only with a significant loss of 5-HT1A binding to the dorsal raphe nucleus (-60%) and of 5-HT1B binding to the substantia nigra (-37%). These results are discussed in relation to the possible identity of 5-HT1A and/or 5-HT1B sites with the presynaptic 5-HT autoreceptors controlling nerve impulse flow and neurotransmitter release in serotoninergic neurons.     

Species differences in the distribution of central 5-HT1 binding sites: a comparative autoradiographic study between rat and guinea pig.

In this study, we compared the localization of central 5-HT1 binding sites of rat and guinea pig. The 5-HT1B sites were absent in the guinea pig brain. Good correlations were found between species in the regional distribution of 5-HT1 sites labelled with [3H]5-HT (r = 0.73), 5-HT1A sites labelled with [3H]8-OH-DPAT (r = 0.87), and 5-HT1B versus 5-HT1D sites labelled with [3H]5-HT in the presence of ipsapirone and DOI (r = 0.76). Despite the overall similarities, species differences were observed in many brain regions. The CA1/CA2 fields of the hippocampus and the dorsal subiculum displayed significantly more 5-HT1A receptor binding in guinea pig than in rat. Conversely, the 5-HT1A binding in dorsolateral septum, cingulate cortex and laminae IV-V of the neocortex, was more pronounced in rat. Areas almost exclusively containing 5-HT1B or 5-HT1D sites, such as the ventral pallidum, globus pallidus and substantia nigra, expressed markedly more [3H]5-HT binding in rat as compared to guinea pig, while the opposite occurred in claustrum, dorsal endopiriform nucleus, lateral geniculate nucleus, and superficial grey layer of the superior colliculus. The implications of the species differences are illustrated by the binding of [3H]eltoprazine. The distribution of [3H]eltoprazine binding sites showed a good correlation with that of the 5-HT1B sites in rat (r = 0.89), and with that of the 5-HT1A sites in guinea pig (r = 0.97). The data give rise to the possibility that differences in the presence and distribution of 5-HT1 receptor sites are related to species differences in behavioural, neurochemical and physiological responses to drugs with 5-HT1 receptor affinity.     

Cannabinoid receptor and WIN-55,212-2-stimulated [35S]GTPγS binding and cannabinoid receptor mRNA levels in the basal ganglia and the cerebellum of adult male rats chronically exposed to Δ9-tetrahydrocannabinol

The inhibition of motor behavior in rodents caused by the exposure to plant or synthetic cannabinoids has been reported to develop tolerance after repeated exposure. This tolerance seems to have a pharmacodynamic basis, since downregulation of cannabinoid receptors in motor areas, basal ganglia and cerebellum, has been demonstrated in cannabinoid-tolerant rats. The present study was designed to further explore this previous evidence by analyzing simultaneously in several motor areas of delta 9-tetrahydrocannabinol- (delta 9-THC)-tolerant rats: 1. Cannabinoid receptor binding, by using [3H]WIN-55,212-2 autoradiography; 2. Cannabinoid receptor activation of signal transduction mechanisms, by using WIN-55,212-2-stimulated [35S]-guanylyl-5'-O-(gamma-thio)-triphosphate ([35S]-GTP gamma S) autoradiography; 3. Cannabinoid receptor mRNA expression, quantitated by in situ hybridization. Results were as follows. As expected, the exposure to delta 9-THC for 5 d resulted in a decrease of cannabinoid receptor binding in the molecular layer of the cerebellum, medial, and lateral caudate-putamen and, in particular, entopeduncular nucleus. We also found decreased cannabinoid receptor binding in the superficial and deep layers of the cerebral cortex, two regions used as a reference to test the specificity of changes observed in motor areas. There were only two brain regions, the globus pallidus and the substantia nigra, where the specific binding for cannabinoid receptors was unaltered after 5 d of a daily delta 9-THC administration. However, in the substantia nigra, the magnitude of WIN-55,212-2-stimulated [35S]-GTP gamma S binding was lesser in delta 9-THC-tolerant rats than controls, thus suggesting a possible specific change at the level of receptor coupling to GTP-binding proteins. This was not seen neither in the globus pallidus nor in the lateral caudate-putamen, where agonist stimulation produced similar [35S]-GTP gamma S binding levels in delta 9-THC-tolerant rats and controls. Finally, animals chronically exposed to delta 9-THC also exhibited a decrease in the levels of cannabinoid receptor mRNA in the medial and lateral caudate-putamen, but there were no changes in the cerebellum (granular layer) and cerebral cortex. In summary, the chronic exposure to delta 9-THC resulted in a decrease in cannabinoid receptor binding and mRNA levels in the caudate-putamen, where cell bodies of cannabinoid receptor-containing neurons in the basal ganglia are located. However, this decrease particularly affected the receptor binding levels in those neurons projecting to the entopeduncular nucleus, but not in those projecting to the globus pallidus and substantia nigra, although, in this last region, a specific decrease in the efficiency of receptor activation of signal transduction mechanisms was seen in delta 9-THC-tolerant rats. The chronic exposure to delta 9-THC also resulted in decreased cannabinoid receptor binding in the cerebellum, although without affecting mRNA expression.