Transferrin and Iron in Normal, Alzheimer's Disease, and Parkinson's Disease Brain Regions

Abstract: Oxidant-mediated damage is suspected to be involved in the pathogenesis of several neurodegenerative disorders. Iron promotes conversion of hydrogen peroxide to hydroxyl radical and, thus, may contribute to oxidant stress. We measured iron and its transport protein transferrin in caudate, putamen, globus pallidus, substantia nigra, and frontal cortex of subjects with Alzheimer's disease (n = 14) and Parkinson's disease (n = 14), and in younger adult (n = 8) and elderly (n = 8) normal controls. Although there were no differences between control groups with regard to concentrations of iron and transferrin, iron was significantly increased ( p < 0.05) in Alzheimer's disease globus pallidus and frontal cortex and Parkinson's disease globus pallidus, and transferrin was significantly increased in Alzheimer's disease frontal cortex, compared with elderly controls. The transferrin/iron ratio, a measure of iron mobilization capacity, was decreased in globus pallidus and caudate in both disorders. Regional transferrin and iron concentrations were generally more highly correlated (Pearson's correlation coefficient) in elderly controls than in Alzheimer's and Parkinson's disease. The altered relationship between iron and transferrin provides further evidence that a disturbance in iron metabolism may be involved in both disorders.     

Effect of dialysis on oxidative stress in uraemia

It has been postulated that dialysis of patients with chronic renal failure (CRF) is associated with increased lipid peroxidation which may contribute to vascular and other complications of the syndrome. In the present study, a specific and precise technique [ferrous oxidation in xylenol orange (FOX) assay] was used to measure plasma lipid hydroperoxides (ROOHs) in three groups of uraemic patients. Patients were either studied before starting dialysis (n = 12) or on continuous ambulatory peritoneal dialysis (CAPD, n = 12) or haemodialysis (HD, n = 36) and compared to healthy controls (n = 20). Plasma ROOHs were markedly elevated in HD patients compared with the controls (7.01 +/- 2.9 microM versus 4.25 +/- 2.05 microM; P < 0.005, Mann-Whitney test). Plasma ROOH concentrations in the CAPD patients were increased but not significantly higher than controls (5.36 +/- 3.56 microM versus 4.25 +/- 2.05 microM). By contrast, no differences in ROOH levels were found between controls and predialysis patients. There was no difference in plasma thiobarbituric acid reactive substances (TBARS) between control and the three CRF groups. Absolute and cholesterol standardised plasma alpha-tocopherol levels were lower in the patients (whether they were on dialysis or not) than in the controls (18.62 +/- 6.88 microM versus 22.73 +/- 5.33 microM; P < 0.01 and 1.99 +/- 1.88 microM/mM versus 5.25 +/- 1.0 microM/mM; P < 0.0005, respectively). This study provides direct evidence that enhanced oxidative stress in CRF patients is related to the dialysis treatment rather than the disease itself. Further studies will be necessary to establish the relationships between plasma measures of oxidative stress and cardiovascular complications in CRF patients under dialysis and whether treatment with antioxidants may reduce oxidative stress or reverse adverse effects associated with dialysis.     

Amyloid beta-peptide1-40 increases neuronal membrane fluidity: role of cholesterol and brain region

There is increasing evidence of an interaction between cholesterol dynamics and Alzheimer's disease (AD), and amyloid beta-peptide may play an important role in this interaction. Abeta destabilizes brain membranes and this action of Abeta may be dependent on the amount of membrane cholesterol. We tested this hypothesis by examining effects of Abeta1-40 on the annular fluidity (i.e., lipid environment adjacent to proteins) and bulk fluidity of rat synaptic plasma membranes (SPM) of the cerebral cortex, cerebellum, and hippocampus using the fluorescent probe pyrene and energy transfer. Amounts of cholesterol and phospholipid of SPM from each brain region were determined. SPM of the cerebellum were significantly more fluid as compared with SPM of the cerebral cortex and hippocampus. Abeta significantly increased (P < or = 0.01) annular and bulk fluidity in SPM of cerebral cortex and hippocampus. In contrast, Abeta had no effect on annular fluidity and bulk fluidity of SPM of cerebellum. The amounts of cholesterol in SPM of cerebral cortex and hippocampus were significantly higher (P < or = 0.05) than amount of cholesterol in SPM of cerebellum. There was significantly less (P < or = 0.05) total phospholipid in cerebellar SPM as compared with SPM of cerebral cortex. Neuronal membranes enriched in cholesterol may promote accumulation of Abeta by hydrophobic interaction, and such an interpretation is consistent with recent studies showing that soluble Abeta can act as a seed for fibrillogenesis in the presence of cholesterol.     

Oxidative Parameters in the Rat Brain of Chronic Mild Stress Model for Depression: Relation to Anhedonia-Like Responses

The chronic mild stress (CMS) protocol is widely used to evoke depression-like behaviors in the laboratory. Some animals exposed to CMS are resistant to the development of anhedonia, whereas the remaining are responsive, CMS-resilient and CMS-sensitive, respectively. The aim of this study was to examine the effects of chronic stress on oxidative parameters in the rat brain. The consumption of sweet food, protein and lipid oxidation levels and superoxide dismutase and catalase activities in the rat hippocampus, cortex and cerebellum were assessed. We found a significant increase in protein peroxidation (hippocampus and cortex), a significant increase in catalase activity (cortex, hippocampus and cerebellum) and a decrease in superoxide dismutase activity (cortex, hippocampus and cerebellum) in the CMS-sensitive group compared to the CMS-resilient group and normal controls as well as an increase in lipid peroxidation (cerebellum) in the CMS-sensitive and CMS-resilient groups compared to normal controls. However, there was no significant difference in protein peroxidation (cerebellum) and lipid peroxidation (cortex and hippocampus) among the three groups. In conclusion, our results indicate that the segregation into CMS-sensitive and -resilient groups based on sucrose intake is paralleled by significant differences in oxidative parameters. CMS induces oxidative damage and alterations in the activity of antioxidants which may lead to increased oxidative damage, irrespective of the anhedonia-like status of the stressed animals.     

Direct and continuous measurement of hydroperoxide-induced oxidative stress on the membrane of intact erythrocytes

Having minimized spectroscopic interference by hemoglobin (Hb), peroxidation processes in intact erythrocytes could be monitored in a continuous assay using the fluorescent polyunsaturated fatty acid, parinaric acid (PnA), as a peroxidation probe. Control experiments to establish the character of the method are described in detail. As a practical application, comparative studies were performed to monitor the response of normal and sickle Hb-containing human erythrocytes to oxidative stress in the PnA assay. After 10 min of incubation with 200 microM cumene hydroperoxide (cumOOH), peroxidation of PnA was found to be enhanced in erythrocytes from sickle cell disease patients (SS: 48 +/- 9% (n = 6) of initial amount had been peroxidized) compared to healthy controls (AA: 30 +/- 4% (n = 9)). PnA peroxidation in erythrocytes from sickle cell trait individuals (AS: 30 +/- 3% (n = 4)) was equal to that in control cells. The increased oxidation of PnA in sickle erythrocytes was accompanied by enhanced oxidation of Hb (metHb and hemichrome formation), indicating that sickle Hb mediates enhanced cumOOH-derived radical generation. It is concluded that PnA can be a useful tool in studying membrane peroxidation processes in intact normal and pathological erythrocytes.     

Association between an alpha(2) macroglobulin DNA polymorphism and late-onset Alzheimer's disease.

An association between a five-base-pair deletion/insertion DNA polymorphism at the alpha(2) macroglobulin gene (A2M) and late-onset Alzheimer's disease (LOAD) has been recently described. We developed a PCR assay to analyze this polymorphism in 190 LOAD patients (older than 65 years) and 400 controls from Spain. Controls were stratified into three groups: <65 years (n = 200), 65 to 80 years (n = 100), and 81 years or older (n = 100). We found a significantly higher frequency of carriers of the D allele in patients older than 81 years compared to controls older than 81 years (p = 0.0012). In addition, the frequency of the D allele was significantly lower in controls older than 81 years compared to controls younger than 65 (p = 0.048). Our work suggests that the D allele confers an age-dependent increased risk to develop late-onset Alzheimer's disease.     

Increased brain levels of 4-hydroxy-2-nonenal glutathione conjugates in severe Alzheimer's disease

In the last decade an important role for the progression of neuronal cell death in Alzheimer's disease (AD) has been ascribed to oxidative stress. trans-4-Hydroxy-2-nonenal, a product of lipid peroxidation, forms conjugates with a variety of nucleophilic groups such as thiols or amino moieties. Here we report for the first time the quantitation of glutathione conjugates of trans-4-hydroxy-2-nonenal (HNEGSH) in the human postmortem brain using the specific and very sensitive method of electrospray ionization triple quadrupole mass spectrometry (ESI-MS-MS). Levels of HNEGSH conjugates calculated as the sum of three chromatographically separated diastereomers were determined in hippocampus, entorhinal cortex, substantia innominata, frontal and temporal cortex, as well as cerebellum from patients with AD and controls matched for age, gender, postmortem delay and storage time. Neither age, nor postmortem delay, nor storage time did correlate with levels of HNEGSH conjugates which ranged between 1 and 500 pmol/g fresh weight in the brain areas examined. The brain specimen from patients with clinically and neuropathologically probable AD diagnosed according to criteria of the consortium to establish a registry for AD (CERAD) show increased levels of HNEGSH in the temporal and frontal cortex, as well as in the substantia innominata. Classification of disease severity according to Braak and Braak, which takes into consideration the amount of neurofibrillary tangles and neuritic plaques, revealed highest levels of HNEGSH in the substantia innominata and the hippocampus, two brain regions known to be preferentially affected in AD. These results substantiate the link between conjugates of glutathione with a product of lipid peroxidation and Alzheimer's disease and justify further studies to evaluate the role of HNE metabolites as potential biomarkers for disease progression in AD.     

Age-dependent variations in mitochondrial and cytosolic antioxidant enzymes and lipid peroxidation in different regions of central nervous system of guinea pigs

The age-related changes in the activities of antioxidant enzymes of mitochondrial and cytosolic fractions were measured in different regions of the central nervous system (CNS) in 10 and 32 months old guinea pigs. In old animals, the activities of superoxide dismutase (SOD) and glutathione peroxidase (GPx) were reduced (p < 0.05) in all the regions of CNS studied but catalase (CAT) declined significantly only in the cerebral cortex, hypothalamus and cerebellum. Glutathione reductase (GRd) activity declined in cerebral cortex and hypothalamus in the cytosolic fractions and only in cerebellum in the mitochondrial fraction. It is concluded that age-related decline in the activities of antioxidant enzymes is both region and enzyme specific. The endogenous lipid peroxide was found to be significantly higher (p < 0.05) in the 32 month old animals whereas, lipid peroxidation after incubating the tissue homogenate in air was found to be lower (p < 0.05). The in vitro mitochondrial lipid peroxidation decreased with age. The results indicate that accumulation of lipid peroxides takes place with ageing but the susceptibility of lipid peroxidation decreases in the older animals.     

Histological changes during development of the cerebellum in the chick embryo exposed to a static magnetic field

Few studies have been performed to evaluate the ultrastructural changes that exposure to static magnetic fields (SMF) can cause to the processes of cell migration and differentiation in the cerebellum during development. Thus, we have studied the development of the cerebellum in the chick embryo (n = 144) under a uniform SMF (20 mT). All of our observations were done on folium VIc of Larsell's classification. The cerebella of chick embryos, which were exposed solely on day 6 of incubation and sacrificed at day 13 of incubation [short exposure (S)1; n = 24], showed an external granular layer (EGL) that was less dense than the EGL in the control group (n = 24). The molecular layer (ML) exhibited a low number of migratory neuroblastic elements. Moreover, the internal granular layer (IGL) was immature, with the cellular elements less abundant and more dispersed than in controls. In chick embryos exposed on day 6 of incubation and sacrificed at day 17 (S2; n = 24), the outstanding feature was the regeneration of the different layers of the cerebellar cortex. The cerebellar cortex of chick embryos exposed continuously to an identical field from the beginning of the incubation up to day 13 [long exposure (L)1; n = 24] or day 17 (L2; n = 24) of incubation showed a higher number of alterations than that of group S1. Electron microscopy confirmed the findings from light microscopy and, at the same time, showed clear signs of cell degeneration and delay in the process of neuronal differentiation. This was more apparent in groups L1 (100%) and L2 (100%) than in groups S1 (95.4%) and S2 (65.2%). In conclusion, the present study showed that SMF can induce irreversible developmental effects on the processes of cell migration and differentiation of the chick cerebellar cortex. Bioelectromagnetics 18:36–46, 1997. © 1997 Wiley-Liss, Inc.     

Vitamin E concentrations in human brain of patients with Alzheimer's disease,fetuses with Down's syndrome,centenarians, and controls

The concentration of vitamin E (alpha-tocopherol) was measured in samples of cortex from patients with Alzheimer's disease (AD), fetuses with Down's syndrome (DS), and also in a group of centenarians. The mean tocopherol concentrations in the two patient groups did not differ significantly from appropriate controls. When expressed per lipid the mean tocopherol concentration of the centenarians was greater than that of the controls but this reflected a significant decrease in the lipid concentration of the former group. These results indicate that neither the normal aging processes, Alzheimer's disease, nor the increased in vitro lipid peroxidation reported in fetuses with Down's syndrome result from a gross lack of alpha-tocopherol, or cause a significant depletion of the vitamin.     

Plasma protein glycation in Alzheimer's disease

Recent studies have suggested that formation of advanced glycation end-products (AGEs) in some brain proteins could be associated with Alzheimer's disease.These AGEs can be produced by various sugars (hexose, pentose, glyceraldehyde and oxidative products of vitamin C). In this study, we quantified plasma protein glycation specifically derived from glucose in patients with Alzheimer's disease with different grades of cognitive disorders.Two groups of Alzheimer patients were studied: a group with moderate Alzheimer's disease (n=6, 9相似文献   

Serum levels of inflammation factors and cognitive performance in amnestic mild cognitive impairment: a Chinese clinical study

Early diagnosis of Alzheimer's disease (AD) is important for initiating timely therapy to block or slow the rate of disease progression. This study was designed to investigate the potential of inflammation-related biomarkers in peripheral blood to accurately reflect AD onset and progression. Individuals (n=150) with amnestic mild cognitive impairment (aMCI) were divided into two subgroups (low- and high-risk) based on APOEε4 allele carrier status, and administered a battery of neuropsychological tests and tested for serum levels of IL-6, IL-10, TNF-α, and IFN-γ by using specific enzyme-linked immunosorbent assays. Results were compared with those from age-matched healthy controls (n=150). The levels of IL-6 were significantly higher in the aMCI group than in controls (P<0.01). When the aMCI group was stratified by APOEε4 status, significant differences were found between the low- and high-risk groups and controls in the levels of IL-6 and IFN-γ (P<0.01 and P=0.041, respectively). Moreover, the IL-6 level in the low-risk aMCI group was higher than that in the high-risk aMCI group (P=0.028). A weak but significant negative correlation was found between IL-6 and cognitive performance. Taken together, these findings indicate that IL-6, while not useful alone, has potential in combination with other biomarkers to support early diagnosis of aMCI due to its association with the progression of cognitive impairment.     

Reduced Neuropeptide Y Concentrations in Suicide Brain

Neuropeptide Y (NPY) was measured in postmortem brain tissue from victims of suicide and from individuals dying a sudden natural or accidental death (controls). Concentrations of NPY-immunoreactivity were measured by radioimmunoassay in frontal cortex (BA 10), temporal cortex (BA 22), caudate nucleus, and cerebellum. Concentrations of NPY-immunoreactivity were significantly lower in postmortem frontal cortex (-14%) and caudate nucleus (-27%) from suicide victims compared with age-matched controls. A subgroup of suicides with evidence of a history of depression revealed more robust reductions in concentrations of NPY-immunoreactivity in frontal cortex and caudate nucleus, as did four individuals who died from natural causes and also were described as having a possible history of depression. Concentrations of NPY-immunoreactivity in temporal cortex and cerebellum from victims of suicide or from the subgroup of subjects with a possible history of depression were not significantly different from those of age-matched controls. We suggest there is a deficit in the brain NPY system leading to region-specific reductions in peptide concentrations in subjects who have a history of depression.     

Amyloid-beta deposition in the cerebral cortex in Dementia with Lewy bodies is accompanied by a relative increase in AbetaPP mRNA isoforms containing the Kunitz protease inhibitor

Deposition of amyloid-beta, the fibrillogenic product of the cell surface protein AbetaPP (amyloid-beta protein precursor), occurs in the cerebral cortex of patients with Dementia with Lewy bodies (DLB). Amyloid deposition, basically in the form of senile plaques, occurs not only in the common form (DLBc), which is defined by changes consistent with diffuse Lewy body disease accompanied by Alzheimer's disease (AD), but also in the pure form (DLBp), in which neurofibrillary tangles are absent. The present study analyses the expression of AbetaPP mRNA isoforms with (AbetaPP751 and AbetaPP770) and without (AbetaPP695) the Kunitz-type serine protease inhibitor (KPI) domain, in the cerebral cortex in DLBc (n=4), DLBp (n=4), Parkinson's disease (PD, n=5), AD (n=3 stages I-IIA, and n=4 stage VC of Braak and Braak), amyloid angiopathy (AA, n=2) and progressive supranuclear palsy (PSP, n=4) compared with age-matched controls (n=6). For this purpose, TaqMan RT-PCR assay was used on frozen post-mortem samples of the frontal cortex (area 8) obtained with short post-mortem delays (8.29+/-4.57 h) and strict RNA preservation (A260/280 of 1.78+/-0.15). A 3.66-fold, 6.67-fold, 4.28-fold and 5.24-fold increases, in the (AbetaPP751+AbetaPP770)/AbetaPP695 mRNA ratio were found in DLBc, DLBp, AD stage VC and AA, respectively, when compared with controls. No modifications in the ratio were found in PD, AD stage I-IIA and PSP. These findings suggest that alternative splicing of the AbetaPP mRNA may play a role in betaA4 amyloidogenesis in DLBp, DLBc, AD stage VC and Amyloid angiopathy.     

A Method for the In Vivo Investigation of the Serotonergic 5-HT2 Receptors in the Human Cerebral Cortex Using Positron Emission Tomography and 18F-Labeled Setoperone

Following previous validation in baboons, we have studied the characteristics of [18F]setoperone as a radioligand for investigating serotonergic 5-hydroxytryptamine2 (5-HT2) receptors in the normal, unmedicated human brain with positron emission tomography (PET); subjects orally pretreated with therapeutic amounts of ketanserin, sulpiride, or prazosin were also studied to evaluate the specificity and sensitivity of [18F]setoperone brain specific binding. In controls (n = 10), the tracer showed a clear-cut retention in both frontal cortex and striatum (known to contain a high density of 5-HT2 receptors) relative to cerebellum (known to be devoid of 5-HT2 receptors). In the seven young controls (20-39 years old), the frontal cortex/cerebellum and striatum/cerebellum ratios increased during the first hour to reach similar values of 2.53 +/- 0.12 and 2.38 +/- 0.11 (mean +/- SEM), respectively, and were essentially stable during the second hour. Pretreatment with ketanserin (a 5-HT2 blocker) significantly reduced the frontal cortex/cerebellum ratio to 0.7-1.0 at 65 min, whereas the striatum/cerebellum ratio was significantly, but only partially, reduced. During sulpiride treatment (a D2 blocker), the frontal cortex/cerebellum ratio was not altered, whereas the striatum/cerebellum ratio was significantly, but only partially, reduced. With prazosin pretreatment (an alpha 1-adrenergic blocker), neither the frontal cortex/cerebellum nor the striatum/cerebellum ratio was modified. These data in humans with PET demonstrate that [18F]setoperone labels with high sensitivity and selectivity 5-HT2 receptors in the frontal cortex; in striata, however, binding is to both 5-HT2 and D2 receptors. The deproteinated-to-whole plasma radio-activity concentration ratio increased with time following injection. The mean percentage of intact [18F]setoperone, in deproteinated plasma, was 82, 74, 53, 45, 30, and 22% at 5, 10, 20, 30, 60, and 110 min following injection, respectively. These data indicate that [18F]setoperone (a) is significantly bound to plasma proteins and (b) is significantly metabolized into several labeled metabolites that are much more hydrophilic than setoperone and, hence, presumably do not cross the blood-brain barrier. These results suggest the suitability of [18F]setoperone data for modeling of 5-HT2 receptor binding in brain.     

Forskolin Stimulation of Cyclic AMP Accumulation in Rat Brain Cortex Slices Is Markedly Enhanced by Endogenous Adenosine

Stimulation of cyclic AMP (cAMP) accumulation in rat cortex slices by 1 microM forskolin (F) was markedly reduced (96%) by treatment with adenosine deaminase (ADA). The effect of ADA was progressively less at higher concentrations of F, but still inhibited the response by 50% at 100 microM F. ADA-mediated inhibition of the cAMP response to 1 microM F was completely reversed by 5 microM 2-chloroadenosine (CA), an ADA-resistant analogue. Stimulation by F (controls) and F plus CA (ADA treated) in cortex slices was significantly inhibited by 200 microM caffeine (CAF) and by 10 microM 8-phenyltheophylline. cAMP accumulation in ADA-treated cortex slices stimulated with CA at concentrations from 5 to 100 microM was markedly enhanced by 1 microM F. Neither ADA treatment nor 200 microM CAF significantly affected cAMP accumulation in slices stimulated by 1 microM vasoactive intestinal polypeptide or adenylate cyclase in membranes stimulated by 1 microM F. CAF (1 mM) did not significantly increase basal cAMP levels in cortex slices, whereas 1 mM 3-isobutyl-1-methylxanthine caused a significant 80% increase and 100 microM rolipram enhanced cAMP levels by 4.5-fold. F-stimulated cAMP accumulation (1 microM) in cortex slices was inhibited 98% by 1 mM CAF and 49% by 1 mM 3-isobutyl-1-methylxanthine, and was enhanced 2.5-fold by 100 microM rolipram. These data have been interpreted to indicate that the stimulation of cAMP accumulation in rat cortex slices by 1 microM F is predominantly due to synergistic interaction with endogenous adenosine and that the inhibition of this response by CAF is largely due to blockade of adenosine receptors.     

Acetylcholinesterase activation and enhanced lipid peroxidation after long-term exposure to low levels of aluminum on different mouse brain regions

Aluminum (Al), oxidative stress and impaired cholinergic functions have all been related to Alzheimer's disease (AD). The present study evaluates the effect of aluminum on acetylcholinesterase (AChE) and lipid peroxidation in the mouse brain. Mice were loaded by gavage with Al 0.1 mmol/kg/day 5 days per week during 12 weeks. The mice were divided into four groups: (1) control; (2) 10 mg/mL of citrate solution; (3) 0.1 mmol/kg of Al solution; (4) 0.1 mmol/kg of Al plus 10 mg/mL of citrate solution. AChE activity was determined in the hippocampus, striatum, cortex, hypothalamus and cerebellum and lipid peroxidation was determined in the hippocampus, striatum and cortex. An increase of AChE activity was observed in the fourth group (Al + Ci) in the hippocampus (36%), striatum (54%), cortex (44%) and hypothalamus (22%) (p<0.01). The third group (Al) presented a decrease of AChE activity in the hypothalamus (20%) and an enhancement in the striatum (27%). Lipid peroxidation, measured by TBARS (thiobarbituric acid reactive substances), was elevated in the hippocampus and cerebral cortex when compared with the control (p < 0.01). The effect of aluminum on AChE activity may be due to a direct neurotoxic effect of the metal or perhaps a disarrangement of the plasmatic membrane caused by increased lipid peroxidation.     

Proteomic analysis identifies dysfunction in cellular transport, energy, and protein metabolism in different brain regions of atypical frontotemporal lobar degeneration

Frontotemporal lobar degeneration (FTLD) is an umbrella term for a heterogeneous group of young-onset dementias of uncertain prevalence and incidence worldwide. Atypical cases of FTLD with fused in sarcoma inclusions (aFTLD-U) have been described recently, but their molecular characterization is still due. Using shotgun mass spectrometry, we identified a total of 107 differentially expressed proteins in the prefrontal cortex, cerebellum and occipital lobe from aFTLD-U patients compared to controls. These proteins are involved in a range of biological pathways such as cellular transport in the prefrontal cortex, energy metabolism in the cerebellum, and protein metabolism in the occipital lobe. In addition, they were validated by selective reaction monitoring (SRM). Comparison of the aFTLD-U proteomic findings with similar studies of Alzheimer's disease and schizophrenia led to identification of proteins that may be related to dementias and psychoses, respectively. Further studies of aFTLD-U and other FTLD subtypes are warranted, although this will require intensive biobanking efforts.     

Binding sites for melanin-concentrating hormone in the human brain

Binding sites for melanin-concentrating hormone (MCH) in human brain were investigated and characterized by radioligand binding. Specific binding sites for MCH were present in every region of human brain (cerebral cortex, cerebellum, thalamus, hypothalamus, pons, and medulla oblongata) obtained at autopsy. alpha-Melanocyte stimulating hormone or ACTH was a poor inhibitor of (125)I-MCH binding (IC(50) 1 microM) compared with MCH (IC(50) = 0.3 +/- 0.07 nM, mean +/- SEM, n = 3). Scatchard plots of (125)I-MCH binding in human brain (thalamus) gave a dissociation constant of 0.2 +/- 0.06 nM and maximal binding of 5.8 +/- 0.3 fmol/mg protein (n = 3). These findings suggest that specific MCH binding sites that differ from the melanocortin receptors exist in human brain.