Autoradiographic localization of cholecystokinin receptors in rodent brain

Cholecystokinin (CCK) receptor binding sites have been localized by autoradiography in the guinea pig and rat central nervous system. [125I]CCK-triacontatriapeptide labeled the sites in brain slices with an observed association constant equal to 0.041 min-1 and a dissociation constant equal to 0.008 min-1. CCK-triacontatriapeptide (CCK-33) and the C-terminal octapeptide of CCK-33 (CCK-8) potently inhibited [125I]CCK-33 binding with Ki's of 2 nM, whereas desulfated CCK-8 (CCK8-ds) and the C-terminal tetrapeptide of CCK-33 (CCK-4) were much weaker. Receptors were concentrated in the olfactory bulb, in the superficial laminae of the primary olfactory cortex, in the deep laminae of the cerebral cortex, and in the pretectal area. Substantial numbers of sites were also found in the basal ganglia, in the amygdala, and in the hippocampal formation. [125I]CCK-33 binding sites appear to be located on fibers of the optic tract and probably on olfactory tract fibers as well. These results are discussed in terms of physiological functions associated with CCK, presynaptic receptors, and axonal flow of CCK receptors.     

Strain differences in central nervous system concentrations of cholecystokinin between normotensive Wistar Kyoto (WKY) and spontaneously hypertensive (SH) rats

The regional brain and spinal cord concentrations of cholecystokinin-octapeptide (CCK-8) were measured in age-matched normotensive Wistar-Kyoto (WKY) and spontaneously hypertensive (SH) rats. The relative order of distribution of CCK-8 in the WKY strain was hippocampus (20.5 +/- 1.3 pmol/g) greater than cortex greater than striatum = hypothalamus greater than midbrain = thalamus greater than spinal cord greater than medulla oblongata/pons (MO/P, 1.6 +/- 0.2 pmol/g) whereas in the SH strain this order was hippocampus (12.9 +/- 0.8 pmol/g) greater than cortex = striatum greater than hypothalamus greater than midbrain greater than thalamus = spinal cord greater than MO/P (1.4 +/- 0.2 pmol/g). The concentrations of CCK-8 in the cerebellum were at the level of assay sensitivity (0.5 pmol/g in both strains). In comparison to the WKY rats, the SH strain had significantly lower levels of CCK-8 in the hippocampus (-37%), cortex (-28%), spinal cord (-23%) and pituitary (-57%). The lowered levels of CCK-8 in the brain of the SH rat may be causally related to, or result from, the cardiovascular, behavioural or morphological abnormalities of this strain.     

Localization of cholecystokinin binding sites in canine brain using quantitative autoradiography

1. CCK receptors have been characterized and localized in various mammalian species and significant species-specific differences in their distribution have been identified. In the present study, we report the first autoradiographic localization of CCK binding sites in the canine brain. 2. High densities of [125I]BH-CCK-8 binding sites were found in the cortex, cerebellum, hippocampus, caudate nucleus, olfactory bulb and nucleus accumbens. Moderate densities were present in the putamen, amygdala, and substantia gelatinosa. Low binding densities were observed in the globus pallidus, inferior colliculus, hypothalamus and thalamus. 3. Although the distribution profile of CCK binding sites in canine brain is similar to those previously reported in the rodent, primate and human brain, notable differences were observed in the hippocampus, cortex and cerebellum.     

Identification of extrastriatal dopamine D2 receptors in post mortem human brain with [I]epidepride

The regional distribution of striatal and extrastriatal dopamine D₂ receptors in human brain was studied in vitro with(S)-N-[(1-ethyl-2-pyrrolidinyl)methyl]-5-[¹²⁵I]iodo-2,3-dimethoxybenzamide, [¹²⁵I]epidepride, using post mortem brain specimens from six subjects. Scatchard analysis of the saturation equilibrium binding in twenty-three regions of post mortem brain revealed highest levels of binding in the caudate (16.5 pmol/g tissue) and putamen (16.6 pmol/g tissue) with lower levels seen in the globus pallidus (7.0 pmol/g tissue), nucleus accumbens (7.2 pmol/g tissue), hypothalamus (1.8 pmol/g tissue), pituitary (1.3 pmol/g tissue), substantia innominata (1.0 pmol/g tissue), and amygdala (0.87 pmol/g tissue). Of note was the presence of dopamine D₂ receptors in the four thalamic nuclei studied, i.e. anterior nucleus (1.0 pmol/g tissue), dorsomedial nucleus (0.96 pmol/g tissue), ventral nuclei (0.72 pmol/g tissue), and pulvinar (0.86 pmol/g tissue), at levels comparable to the amygdala (0.87 pmol/g tissue) and considerably higher than levels seen in anterior cingulate (0.26 pmol/g tissue) or anterior hippocampus (0.36 pmol/g tissue). The frontal cortex had very low levels of dopamine D₂ receptors (0.17–0.20 pmol/g tissue) while the inferior and medial temporal cortex had relatively higher levels (0.31–0.46 pmol/g tissue). Inhibition of [¹²⁵I]epidepride binding by a variety of neurotransmitter ligands to striatal, ventral thalamic and inferior temporal cortical homogenates demonstrated that [¹²⁵I]epidepride binding was potently inhibited only by dopamine D₂ ligands. The present study demonstrates that dopamine D₂ receptors are present in basal ganglia, many limbic regions, cortex and in the thalamus. The density of thalamic D₂ receptors is comparable to many limbic regions and is considerably higher than in cortex. Very few frontal lobe D₂ receptors are present in man.     

Role of the amygdaloid cholecystokinin (CCK)/gastrin-2 receptors and terminal networks in the modulation of anxiety in the rat. Effects of CCK-4 and CCK-8S on anxiety-like behaviour and [3H]GABA release

The amygdala plays a key role in fear and anxiety. The intercalated islands are clusters of glutamate-responsive GABAergic neurons rich in cholecystokinin (CCK)-2 receptors which control the trafficking of nerve impulses from the cerebral cortex to the central nucleus of amygdala. In this study, the nature of the CCK–glutamate–GABA interactions within the rat rostral amygdala, and their relevance for anxiety, were studied. CCK/gastrin-like immunoreactive nerve terminals were found to be mainly restricted to the paracapsular intercalated islands and the rostrolateral part of the main intercalated island. Behaviourally, the bilateral microinjection of CCK-4 (0.043–4.3 pmol/side) or CCK-8S (4.3 pmol/side) into the rostrolateral amygdala reduced the open-arm exploration in the elevated plus-maze without affecting locomotion. In contrast, neither CCK-4 nor CCK-8S (0.043–4.3 pmol/side) had any effects in the shock-probe burying test as compared with their saline-treated controls. Biochemically, CCK-4 (0.3 and 1.5 µ m ), unlike CCK-8S, enhanced significantly the K⁺-stimulated release of [³H]GABA from amygdala slices. These effects were fully prevented by prior superfusion of the slices with either the selective CCK-2 receptor antagonist CR2945 (3 µ m ), or 6,7-dinitroquinoxaline-2,3(1H,4H)-dione (DNQX), 10 µ m , a glutamatergic (+/–)-α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)/kainate receptor antagonist. It is suggested that CCK modulates glutamate-GABA mechanisms by acting on CCK-2 receptors via volume transmission occurring at the level of the basolateral amygdaloid nucleus and/or by synaptic or perisynaptic volume transmission in the region of the rostrolateral main and paracapsular intercalated islands, resulting in subsequent disinhibition of the central amygdaloid nucleus and anxiety or panic-like behaviour.     

Characterization of [3H] CCK4 binding sites in mouse and rat brain

We have investigated the possible occurrence of distinct CCK8 and CCK4 binding sites in the brain by comparing the binding characteristics of [3H] CCK4 to those of the CCK8 analogue, [3H] Boc (Nle28,31]CCK27-33 (BDNL-CCK7). [3H] CCK4 and [3H] BNDL-CCK7 were shown to interact with mouse brain membranes with very similar maximal binding capacities 31.7 +/- 2.1 fmol/mg prot (KD = 3.78 +/- 0.47 nM) and 38.9 +/- 2.2 fmol/mg prot (KD = 0.26 +/- 0.02 nM) respectively. The apparent affinities of five CCK analogues for the sites labelled by both probes were almost identical. Autoradiographic studies revealed that the distribution of [3H] CCK4 binding sites in rat forebrain was the same as that of [3H] BDNL-CCK7, with high densities of receptors in the cortex, nucleus accumbens, olfactory bulb and the medial striatum, moderate densities in the amygdala, the hippocampus, several nuclei of the thalamus and hypothalamus. However in the interpenduncular nucleus where there was moderate binding of [3H]BDNL-CCK7, no [3H]CCK4 labelling was observed. These studies demonstrated the occurrence of one class of high affinity binding sites for [3H] CCK4 in mouse and rat brain, with characteristics similar to those already reported with CCK33, CCK8 and pentagastrin probes. Nevertheless the presence of a small amount of very high affinity binding sites for [3H]CCK4 cannot be excluded.     

Expression, but failing maturation of procholecystokinin in cerebellum.

The cerebellum is the only region of the central nervous system which has been found to be devoid of cholecystokinin (CCK). The assays used, however, have been directed against the alpha-amidated C-terminus of fully processed CCK peptides. Using Northern blot analysis and a library of radioimmunoassays specific for different sequences of proCCK in combination with chromatography and enzyme cleavage, we have now examined the expression and processing of proCCK in fetal, neonatal and adult cerebellar tissue from man, pig and rat. In rat cerebellum CCK mRNA was present already in the fetal state. Two weeks after birth the concentrations declined. Also proCCK was found in significant concentrations in the fetal human and rat cerebellum (approximately 20 pmol/g); but already before birth the expression began to decrease towards low concentrations in adults. The adult porcine cerebellum contained 3.2 pmol proCCK and glycine-extended processing intermediates per gram (range less than 0.1-10.4 pmol/g), and 0.8 pmol carboxyamidated CCK per gram (range 0.1-4.1 pmol/g) varying in size from CCK-58 to CCK-5. For comparison, the adult porcine cerebral cortex contained 757 pmol carboxyamidated CCK/g, 20 pmol glycine-extended CCK/g and no proCCK. We conclude that cerebellum expresses proCCK with the highest level of expression in fetal life. In comparison with other regions of the brain, the maturation to transmitter-active, carboxyamidated CCK peptides is, however, attenuated in both fetal and adult cerebellar tissue.     

Autoradiographic Localization of Cholecystokinin A and B Receptors in Rat Brain Using [125I]d-Tyr25 (Nle28,31)-CCK 25 - 33S

The distribution of receptors for the sulphated octapeptide cholecystokinin 26 - 33 (CCK - 8S) in rat brain was investigated by radioligand binding in conjunction with autoradiography using the novel iodinable, non-oxidizable, amino- and thiolendopeptidase-resistant CCK analogue, d-Tyr25(Nle28,31)-CCK 25 - 33S. Labelling of the peptide was achieved by synthesis utilizing Na125I and Chloramine-T. [125I]d-Tyr25(Nle28,31)-CCK 25 - 33S (100 pM) bound rapidly and reversibly to a single population of sites on slide-mounted coronal sections of rat forebrain with a dissociation constant of 34 pM. Specific binding was fully inhibited by CCK-8S, CCK-8, CCK-4, L-365,260 and L-364,718, with inhibition constants 2.7, 9.8, 35, 7.0 and 130 nM, respectively. These inhibition data may indicate that the [125I] ligand binds preferentially to a CCKB subtype of receptor, but may also reflect the relative paucity of CCKA receptors in the rat forebrain. Optimum conditions for autoradiography combined the preincubation of brain sections in unlabelled 10 pM d-Tyr25(Nle28,31)-CCK 25 - 33S with a 60-min wash after incubation with the [125I] ligand. Analyses of the autoradiograms obtained from the use of coronal and horizontal brain sections were aided by the high levels of specific binding (80 - 90%), and revealed that CCK receptors were topographically distributed through the neuroaxis. High densities of receptor-associated silver grains were found in the olfactory bulb (internal plexiform layer), neocortex (layer III), nucleus accumbens, parasubiculum, subbrachial nucleus, parabigeminal nucleus, dorsal vagal complex, area postrema and the A2 region. Moderate labelling was observed in many telencephalic and diencephalic nuclei. The majority of these receptors were of the CCKB subtype, as shown by the use of subtype-selective antagonists, although CCKA receptors were present in moderate to high densities in the A2 area, area postrema and nucleus tractus solitarii, and at low density in the interpeduncular nucleus and central amygdala. These findings provide further evidence for the widespread, topographic distribution of CCK receptors and indicate that [125I]d-Tyr25(Nle28,31)-CCK 25 - 33S is very suitable for autoradiographic investigations because of its low non-specific binding.     

Ontogeny of procholecystokinin processing in rat hypothalamus

The concentration of procholecystokinin (pro-CCK) in the fetal hypothalamus was 126 +/- 41 pmol/g (mean +/- SEM; n = 20), 22 +/- 9 pmol/g at day 7 postpartum and 3 +/- 2 pmol/g in the adult. In contrast, the concentration of bioactive carboxyamidated CCK rose from 6 +/- 2 pmol/g in the fetal hypothalamus to 52 +/- 10 pmol/g in the adult. The concentration of glycine-extended processing intermediates first decreased from 21 +/- 5 pmol/g in the fetus to 5 +/- 1 pmol/g at day 21 postpartum. Subsequently, the concentration rose to 21 +/- 4 pmol/g in the adult. The results show that the CCK gene is well expressed in the fetal hypothalamus. However, only a small fraction of pro-CCK reaches maturation before weaning. We conclude that expression of the CCK gene in the hypothalamus as bioactive peptide to a large degree is regulated at the posttranslational level.     

High-affinity binding of [125I]RTI-55 to dopamine and serotonin transporters in rat brain.

RTI-55 (3 beta-(4-iodophenyl)tropan-2 beta-carboxylic acid methyl ester), one of the most potent inhibitors of dopamine uptake reported to date, was radioiodinated and tested as a probe for the cocaine receptor in Sprague-Dawley rat brain. Saturation and kinetic studies in the striatum revealed that [125I]RTI-55 bound to both a high- and low-affinity site. The Kd for the high-affinity site was 0.2 nM, while the Kd for the low-affinity site was 5.8 nM. The corresponding number of binding sites in the striatum was 37 and 415 pmol/g protein. The pharmacological profile of specific [125I]RTI-55 binding in the striatum was consistent with that of the dopamine transporter. Additionally, [125I]RTI-55 was found to bind with high affinity to the cerebral cortex. Scatchard analysis revealed a single high-affinity component of 0.2 nM with a density of 2.5 pmol/g protein. The pharmacological profile demonstrated by [125I]RTI-55 in the cerebral cortex matched that of the serotonin transporter. Autoradiographic analysis of sagittal brain sections with [125I]RTI-55 binding was consistent with these findings. Specific binding of [125I]RTI-55 was blocked by dopamine uptake inhibitors in areas rich in dopaminergic nerve terminals. Conversely, serotonin uptake inhibitors blocked the binding of [125I]RTI-55 in brain areas rich in serotonergic neurons. These results demonstrate that [125I]RTI-55 may be a very useful ligand for the dopamine and serotonin transporters.     

Regional distribution of somatostatin-like immunoreactivity in the human brain

The regional distribution and chromatographic characteristics of somatostatin-like immunoreactivity (SLI) were studied in autopsy specimens of 8 human brains. SLI concentrations in 8 different regions of these brains ranged from (mean +/- S.E.) 756.3 +/- 363.4 pmol/g tissue in anterior hypothalamus to 1.6 +/- pmol/g in cerebellum. Chromatography of the extracts of human brain cortex, anterior hypothalamus, thalamus and amygdala on Sephadex G-50 disclosed one major peak which corresponded to the elution peak of synthetic somatostatin. Since the human brains were obtained at autopsy 11-36 h after death, the effects of temperature and time lapse between death and tissue extraction on SLI concentration and chromatographic pattern in rat brain were examined. After storage at 4 degrees C or 23 degrees C for 2 h and 8 h, a significant increase in SLI concentration was noted, although by 24 h in increase was no longer observed. A gradual loss in the eluting forms of SLI on gel chromatography was observed with storage at 4 degrees C or 23 degrees C.     

Differentiation of 5-hydroxytryptamine2 receptor subtypes using 125I-R-(-)2,5-dimethoxy-4-iodo-phenylisopropylamine and 3H-ketanserin

The radioligand binding characteristics of 125I-R-(-)4-iodo-2,5-dimethoxyphenylisopropylamine [125I-R-(-)DOI] and 3H-ketanserin were compared in rat and bovine cortical membranes. In rat cortex, 125I-R-(-)DOI labels a relatively low density of binding sites (Bmax = 2.5 +/- 0.2 pmol/gm tissue) with high affinity (KD = 0.63 +/- 0.09 nM). In bovine cortex, specific binding of 125I-R-(-)DOI represents less than 20% of total binding at radioligand concentrations above 0.6 nM, and, therefore, the data cannot be analyzed adequately by Scatchard transformation. By contrast, 3H-ketanserin displays saturable, specific high-affinity binding in both rat cortex (KD = 1.0 +/- 0.1 nM; Bmax = 11 +/- 0.4 pmol/gm tissue) and bovine cortex (KD = 1.2 +/- 0.2 nM; Bmax = 5.3 +/- 0.4 pmol/gm tissue). Ki values for 30 drugs were determined for 125I-R-(-)DOI-labeled sites in rat cortex and 3H-ketanserin-labeled sites in bovine cortex. 5-Hydroxytryptamine (5-HT) displays 250-fold higher selectivity for the 125I-R-(-)DOI-labeled sites (Ki = 3.0 +/- 0.7 nM) than for the 3H-ketanserin-labeled sites (Ki = 750 +/- 50 nM). Structural congeners of R-(-)DOI display 80- to 160-fold higher affinity for the 125I-R-(-)DOI binding site than for the 3H-ketanserin-labeled binding site. d-LSD and putative 5-HT2 antagonists are approximately equipotent at both sites. Significant correlations were found between drug affinities for 125I-R-(-)DOI-labeled sites in rat cortex and putative 5-HT2A sites labeled previously by 77Br-R-(-)DOB (r = 0.93, p less than 0.01), putative 5-HT2B sites labeled by 3H-ketanserin in bovine cortex (r = 0.63, p less than 0.01), and 5-HT1C binding sites that have been characterized by other investigators (r = 0.78, p less than 0.01). No significant correlations were found between drug affinities for 125I-R-(-)DOI-labeled sites in rat cortex and 5-HT1A, 5-HT1B, 5-HT1D, or 5-HT3 sites, as determined by previous investigators. We conclude that 125I-R-(-)DOI labels a novel 5-HT binding site subtype (tentatively designated the 5-HT2A binding site) that is present in rat cortex but is either absent or minimally present in bovine cortex. By contrast, 3H-ketanserin labels both the putative 5-HT2A site in rat cortex as well as a separate, distinct recognition site that is present in both rat and bovine cortex, tentatively designated the 5-HT2B site.     

Autoradiographic localization and biochemical characterization of peripheral type CCK receptors in rat CNS using highly selective nonpeptide CCK antagonists

Two potent and highly selective nonpeptide antagonists, L-365,031 [1-methyl-3-(4-bromobenzoyl)amino-5-phenyl-3H-1,4 benzodiazepin-2-one] and 3H-L-364,718 [1-methyl-3-(2-indoloyl)amino-5-phenyl-3H-1,4 benzodiazepin-2-one] were used to localize "peripheral" CCK receptors in rat brain. In autoradiographic experiments, L-365,031 displaced 125I-Bolton Hunter CCK-8 binding from the interpeduncular nucleus (IPN) (IC50 = 7 X 10(-8) M), the area postrema (AP), and the nucleus tractus solitarius (NTS) without influencing specific binding to other areas, such as the cerebral cortex or the spinal tract of the trigeminal nerve. Desulfated CCK preferentially inhibited 125I-CCK binding to cerebral cortex (IC50 = 7 X 10(-8) M) rather than IPN (IC50 greater than 1 X 10(-6) M) or AP-NTS. In the medulla the localization of 3H-L-364,718 binding was similar to L-365,031-sensitive 125I-CCK-8 binding and was found in the AP and medial, but not lateral, aspects of the NTS. In membranes prepared from IPN, NTS, and AP, 3H-364,718 binding was of high affinity (Kd = 0.14 nM), saturable (Bmax = 20 fmol/mg protein), and inhibited by compounds previously shown to act at pancreatic CCK receptors. The receptors labeled by 3H-364,718 were modulated by guanyl nucleotide, which reduced agonist affinity 10-fold without affecting antagonist binding. The localization and high density of CCK receptors in AP and NTS suggest that these receptors may play an important role in processing sensory afferent information.     

Brain regions where cholecystokinin suppresses feeding in rats

The gut-brain peptide, cholecystokinin (CCK), inhibits food intake when injected either systemically or within the brain. To determine whether CCK's effect in the brain is anatomically specific, CCK-8 (0.8, 4, 20, 100, 500 pmol) was microinjected into one of 14 different brain sites of rats, and its impact on subsequent food intake was measured. CCK-8 at 500 pmol significantly suppressed intake during the first hour post-injection following administration into six hypothalamic sites (anterior hypothalamus, dorsomedial hypothalamus, lateral hypothalamus, paraventricular nucleus, supraoptic nucleus, ventromedial hypothalamus) and two hindbrain sites (nucleus tractus solitarius, fourth ventricle). Although lower doses were sometimes effective (anterior hypothalamus, dorsomedial hypothalamus, nucleus tractus solitarius), there appeared to be no significant difference in potency among sites. Injections into the medial amygdala, nucleus accumbens, posterior hypothalamus, dorsal raphe, and ventral tegmental area were either ineffective or produced a delayed response. The higher doses required for most sites, as well as the widespread effectiveness of CCK-8 within the hypothalamus, suggest that spread of CCK-8 to adjacent brain sites, and (or) to the periphery, may have been required for anorexia to occur. Findings reported in an accompanying paper provide strong evidence that paraventricular nucleus injection of CCK-8 (500 pmol) did not increase plasma CCK-levels sufficiently to suppress feeding by a peripheral mechanism. Together, these results suggest that CCK may be acting as a neurotransmitter or neuromodulator within two different brain regions to produce satiety — one region which includes the nucleus tractus solitarius in the hindbrain, and another more distributed region within the medial–basal hypothalamus.     

Selective reduction of [125I]apamin binding sites in Alzheimer hippocampus: a quantitative autoradiographic study.

[125I]Apamin binding sites were examined using quantitative autoradiography in the hippocampus of 9 patients with Alzheimer's disease and 8 age-matched controls. Within the hippocampal formation from control subjects, [125I]apamin binding sites were highly concentrated in the subiculum and CA1. In Alzheimer's disease there was a marked and discrete loss of [125I]apamin binding sites in the subiculum (control = 1.10 +/- 0.10 pmol/g; Alzheimer = 0.71 +/- 0.09 pmol/g) and CA1 (control = 1.41 +/- 0.09 pmol/g; Alzheimer = 0.85 +/- 0.11 pmol/g; values are mean +/- S.E.M.). This reduction of [125I]apamin binding sites in the subiculum correlated with cell density but not neuritic plaque density. These results indicate that an anatomically discrete loss of Ca(2+)-dependent K+ channels within the hippocampal formation occurs in Alzheimer's disease.     

125I]Iodomethyllycaconitine binds to alpha7 nicotinic acetylcholine receptors in monkey brain

We examined the binding of the novel nicotinic acetylcholine receptor (nAChR) ligand [125I]iodomethyllycaconitine (iodoMLA) in the brains of M. cynomologous (macaque) monkeys. [125I]iodoMLA bound throughout the brain with the greatest density in the thalamus and moderate intensity in the basal ganglia and cortical regions. The Kd and Bmax in whole brain tissue were similar whether 1 mM nicotine (Kd 33.25 +/- 15.17 nM, Bmax 5.80 +/- 1.06 fmol/mg) or 2 microM of the alpha7-selective antagonist alpha-bungarotoxin (Kd 46.12 +/- 18.45 nM, Bmax 6.30 +/- 1.06 fmol/mg) was used for nonspecific binding. The subtype-selectivity of this ligand was further studied with competition binding studies using nicotine, alpha-bungarotoxin and noniodinated MLA. Each ligand completely inhibited [125I]iodoMLA binding throughout the monkey brain, with Ki values of 2.23 +/- 0.85 microM for nicotine, 2.72 +/- 1.71 nM for alpha-bungarotoxin and 1.83 +/- 0.35 nM MLA in the caudate and 2.03 +/- 1.14 microM, 2.65 +/- 0.86 nM and 3.32 +/- 0.71 nM, respectively, in the putamen. The alpha3beta2/alpha6*-selective antagonist alpha-conotoxin MII failed to inhibit [125I]iodoMLA binding in any brain region. In monkeys with cognitive deficits resulting from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine administration, [125I]iodoMLA binding was significantly increased in the striatum, similar to results previously observed for [125I]alpha-bungarotoxin. These results suggest that, under the present experimental conditions, [125I]iodoMLA was selective for alpha7-containing nAChRs and did not bind to alpha6-containing nAChRs. This radioligand may be a useful tool for selectively imaging alpha7-containing nAChRs in vivo.     

In vivo quantification of blood-brain transfer and binding of [125I]HEAT, an alpha 1-adrenoceptor antagonist

The uptake and binding constants of [125I]iodo-2-[beta-(4-hydroxyphenyl)-ethyl-amino-methyl]tetralone ( [125I]HEAT) in rat brain were determined in vivo. The initial clearance of the radioligand from blood to brain, K1, was calculated from the initial uptake of the radioligand; it averaged 0.21 +/- 0.01 (SD) ml g-1 min -1, consistent with an initial extraction of 25% (i.e., one-quarter of the blood flow). The most strongly binding regions included the olfactory bulb, thalamic nuclei, medial geniculate body, and cerebral cortical layers. We identified saturable, specific binding in frontal cortex layers 1, 5a, and 5c (motor region), frontal cortex layers 3+4, ventral thalamic nuclei, medial geniculate body, striatum, cerebellum, and olfactory bulb. Addition of unlabeled ligand depressed binding in all regions to the same low level (partition coefficient) of 0.8 ml g-1. Displacement of [125 I]HEAT binding by unlabeled HEAT yielded a global affinity constant (KDVd) of 34 +/- 8 pmol g-1 and receptor densities (Bmax) that varied from 50 pmol g-1 in cerebellar cortex and caudate nucleus to 200 pmol g-1 in the region of highest specific binding, the medial geniculate body.     

Central 5-HT(1) and 5-HT(2) binding sites in transgenic mice with reduced glucocorticoid receptor number

Transgenic mice bearing a transgene coding for a glucocorticoid receptor antisense mRNA, which partially blocks glucocorticoid receptor expression, were used in order to clarify the role of glucocorticoid receptors in the regulation of 5-HT(1A), 5-HT(1nonA) and 5-HT(2) binding sites labelled by quantitative autoradiography in the frontal and prefrontal cortex, striatum, hypothalamus, amygdala and raphe nuclei. We found that 1 nM [3H]8-hydroxy-2-[di-N-propylamino]tetralin ([3H]8-OH-DPAT) binding to 5-HT(1A) sites was decreased in strata oriens (-15.1+/-3.5%) and radiatum-lacunosum-moleculare (-13.3+/-4.3%) of the hippocampal CA(3) area, and 2 nM [3H]5-hydroxytryptamine binding to 5-HT(1nonA) sites in the presence of 100 nM 8-OH-DPAT and mesulergine was decreased in the dorsal subiculum (-17.8+/-6.9%). By contrast, 5-HT(2) sites labelled by 0.5 nM of (+/-)-1-(2, 5-dimethoxy-4-[125I]iodophenyl)-2-aminopropane was increased in the dorsal subiculum (+35.2+/-11.5%) and CA(2) area (+29.2+/-11.3%). The observed differences in binding to 5-HT(1) and 5-HT(2) sites were all located in areas of the hippocampus that contain both gluco- and mineralo-corticoid receptors, and no difference was observed in anatomical structures which contain only glucocorticoid receptors. Therefore, it seems that the important factor for the regulation of these 5-HT receptors is the interaction between gluco- and mineralo-corticoid receptors rather than the absolute density of glucocorticoid receptors. These results suggest that some of the alterations of the serotonergic neurotransmission observed in depressed patients might be secondary to an altered glucocorticoid receptor function.     

Influence of cholecystokinin octapeptide sulfate ester on brain monoamine metabolism in rats

Summary The effects of intracerebroventricular administration of an 80 pmole dose of cholecystokinin octapeptide sulfate ester (CCK-8-SE) were tested on the dopamine (DA), norepinephrine (NE) and serotonin (5-HT) turnovers of the hypothalamus, mesencephalon, amygdala, septum, striatum and cerebral cortex in rats.CCK-8-SE in an 80 pmole dose decreased the DA turnovers of the hypothalamus, mesencephalon, amygdala and septum, while it increased that of the striatum. The NE turnovers were increased in the hypothalamus and amygdala, but decreased in the striatum. The 5-HT turnover decreased only in the hypothalamus.