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Information about the family of protein arginine methyltransferases (PRMTs) has been growing rapidly over the last few years and the emerging role of arginine methylation involved in cellular processes like signaling, RNA processing, gene transcription, and cellular transport function has been investigated. To date, 11 PRMTs gene transcripts have been identified in humans. Almost all PRMTs have been shown to have enzymatic activity and to catalyze arginine methylation. This review will summarize the overall function of human PRMTs and include novel highlights on each family member.

The basal transcriptional activity of nuclear receptors (NRs) is regulated by interactions with additional comodulator proteins (coactivator/corepressor). Here, we describe a new androgen receptor (AR)-associated coactivator, PRMT2, which belongs to the arginine methyltransferase protein family. To search for AR-interacting proteins a fragment of the AR was used in a library screen exploiting the yeast two-hybrid technique and identifying the C-terminal region of PRMT2. We demonstrated that PRMT2 acts as a strong coactivator of the AR, had modest or none influence on transcriptional activation mediated by other NRs. Interestingly, PRMT2 interaction with the estrogen receptor (ER) was strongly dependent on the cellular background, thus, suggesting the involvement of additional, differentially expressed coregulators. We also demonstrated synergistic interaction of PRMT2 with other known nuclear receptor coactivators, such as GRIP1/TIF-2. Potentiation of AR-mediated transactivation by PRMT2 alone and in synergism with GRIP1 was prevented by a competitive inhibitor of methyltransferase activity. The PRMT2 expression profile overlaps with the distribution of AR, with strongest PRMT2 abundance in androgen target tissues. Immunofluorescence experiments showed that the intracellular localization of PRMT2 depends on the presence of the cognate receptor ligand. Under androgen-free conditions, both AR and PRMT2 are confined to the cytoplasm, whereas in the presence of androgens both proteins colocalize and translocate into the nucleus. Treatment with the AR antagonist hydroxyflutamide results in nuclear translocation of the AR, but not the coactivator PRMT2. Thus, it appears that the ligand-dependent AR conformation is essential for the recruitment and nuclear translocation of PMRT2 which acts as AR-coactivator, presumably by arginine methylation.

The androgen receptor (AR) is a ligand-dependent transcriptional regulator which belongs to the nuclear receptor superfamily. The basal transcriptional activity of the androgen receptor is regulated by interaction with coactivator or corepressor proteins. The exact mechanism whereby comodulators influence target gene transcription is only partially understood, especially for corepressors. Whereas several coactivators are described for the AR, only a few corepressors are known. Here, we describe the discovery of a new androgen receptor corepressor, FoxG1, which belongs to the forkhead family. By using a fragment of the AR (aa 325-919) as bait in a yeast two hybrid screen, the C-terminal region (aa 175-489) of FoxG1 (also known as BF1), was identified as AR-interacting protein. Binding of AR to the FoxG1 fragment was verified by one- and two-hybrid assays, and pull-down experiments. In addition, we show that the full-length form of FoxG1 functions as a strong corepressor in the AR-mediated transactivation. The FoxG1 expression profile in adult individuals is restricted to brain and testis in human and decreases during aging in the rodent brain. Both AR and FoxG1 expression are developmentally regulated. Besides its reported role in neurogenesis, the strong expression of FoxG1 in AR-abundant areas of the adult brain suggests possible involvement in neuroendocrine regulation. Taken together, the data presented suggest that, in addition to repression of transcription by direct binding to DNA, FoxG1 may interact with AR in vivo, thereby targeting its repressor function specifically to sex hormone signaling.

Evidence is accumulating in support of the view that tissue-specific effects of steroid hormones depend on the recruitment of nuclear receptor comodulator proteins. The latter interact directly with the hormone receptors and modify their transcriptional effects on specific target genes. The mechanisms of comodulator influence on nuclear receptor-controlled gene transcription is only partially understood. Here, we describe the discovery of a new AR coactivator which belongs to the JmjC containing enzyme family as a novel variant of JMJD1C (jumonji domain-containing 1C). By using a fragment of the human AR (aa 325-919) as bait in a yeast two-hybrid screen, a region of the human JMJD1C gene was identified as interacting with AR. A novel splice variant s-JMJD1C was amplified by RACE, and the binding to AR was analysed by GST-pull-down and mammalian one-hybrid experiments. As a nuclear-localized protein, the s-JMJD1C gene is expressed in a variety of human tissues. In the brain, this protein is present in several, but not confined to, AR-expressing neuronal populations and its abundance varies with the hormonal status in a region-specific fashion. Interestingly, the expression of s-JMJD1C is reduced in breast cancer tumors and significantly higher in normal breast tissues indicating a putative role in tumor suppression. As s-JMJD1C has putative demethylase activity, removal of methylation seems to be important for nuclear receptor-based gene regulation.

The transcription-intermediary-factor-2 (TIF-2) is a coactivator of the glucocorticoid receptor (GR), and its disruption would be expected to influence glucocorticoid-mediated control of the hypothalamo-pituitary-adrenal (HPA) axis. Here, we show that its targeted deletion in mice is associated with altered expression of several glucocorticoid-dependent components of HPA regulation (e.g., corticotropin-releasing hormone, vasopressin, ACTH, glucocorticoid receptors), suggestive of hyperactivity under basal conditions. At the same time, TIF-2(-/-) mice display significantly lower basal corticosterone levels and a sluggish and blunted initial secretory response to brief emotional and prolonged physical stress. Subsequent analysis revealed this discrepancy to result from pronounced aberrations in the structure and function of the adrenal gland, including the cytoarchitectural organization of the zona fasciculata and basal and stress-induced expression of key elements of steroid hormone synthesis, such as the steroidogenic acute regulatory (StAR) protein and 3beta-hydroxysteroid dehydrogenase (3beta-HSD). In addition, altered expression levels of two nuclear receptors, DAX-1 and steroidogenic factor 1 (SF-1), in the adrenal cortex strengthen the view that TIF-2 deletion disrupts adrenocortical development and steroid biosynthesis. Thus, hyperactivity of the hypothalamo-pituitary unit is ascribed to insidious adrenal insufficiency and impaired glucocorticoid feedback.

The regional distribution, developmental profiles, and gonadectomy- and estrogen-induced changes in the density of transcripts encoding the steroid receptor coactivator-1 (SRC-1) were examined in the female rat brain by semiquantitative in situ hybridization. The results demonstrate striking differences in the abundance of SRC-1 mRNA in discrete brain regions throughout ontogeny. Whereas transcript densities gradually decreased with age in the cerebral cortex, they peaked prominently during the peripubertal period in the hypothalamic medial preoptic area (MPOA) and ventromedial nucleus (VMN). Gonadectomy and estrogen substitution influenced SRC-1 mRNA levels in sexually mature animals in a region-specific fashion. Ovariectomy resulted in a down-regulation of SRC-1 mRNA levels in the VMN, a brain region richly endowed with estrogen receptors and playing a major role in neuroendocrine control of reproductive functions. In contrast, SRC-1 transcript levels were significantly up-regulated after estradiol treatment. Interestingly, SRC-1 expression in the cortex was refractory to alterations of the estrogen milieu. The obtained SRC-1 mRNA expression profiles during development clearly demonstrate brain region specificity and regulation by estrogen, thus it is proposed that SRC-1 amplifies estrogen receptor-dependent transcription in a temporally and spatially coordinated manner and therefore contributes to the functional specialization of brain areas involved in the regulation of reproduction.

The enzyme 3 alpha-hydroxysteroid dehydrogenase (3 alpha-HSD) is involved in the generation of neuroactive steroids through ring-A-reduction of hormonal precursors. We examined the developmental regulation of, gender differences in, and effects of hormonal manipulations on the expression of 3 alpha-HSD in the rat hippocampus. High levels of 3 alpha-HSD mRNA were found on postnatal day 7, coinciding with the stress hyporesponsive period in the rat. Gender differences in 3 alpha-HSD expression were documented during puberty, but not in adulthood. Adrenalectomy and gonadectomy, and supplementation with individual steroid hormones influenced 3 alpha-HSD expression in a gender-specific mode. We also demonstrate that the manifestation of behavioral and endocrine consequences of early life stress depends on the individual's gender and gonadal status. Males are liable to aftereffects of neonatal maternal deprivation, regardless of their adult gonadal status. In females, however, anxiogenic aftereffects of neonatal stress become apparent only after gonadectomy. These data suggest that (i) transient increase of neurosteroid biosynthesis may contribute to stress hyporesponsiveness during early infancy; (ii) gonadal steroids regulate 3 alpha-HSD expression in the hippocampus in a sex-specific mode; (iii) physiological sex steroid secretions in females may mask behavioral consequences of adverse early life events, and (iv) concomitant treatment with the neurosteroid THP counteracts behavioral and endocrine dysregulation induced by neonatal stress in both genders.

Fatty acid synthase (FAS), one of the main lipogenic enzymes, converts dietary calories into a storage form of energy. The transcription factors, stimulatory proteins 1 and 3 (Sp1 and Sp3), nuclear factor Y (NF-Y), upstream stimulatory factor (USF) and sterol regulatory element binding protein-1 (SREBP-1) have cognate binding sites on the promoter of the FAS gene. It was shown that Sp1 and NF-Y interact co-operatively at the diet-induced DNase I-hypersensitive site at position -500. Adjacent binding sites for NF-Y and Sp1 have also been found between -71 and -52, and -91 and -83. cAMP regulation is mediated via the inverted CAAT element (ICE) at -99 to -92, which binds NF-Y. The FAS insulin-responsive element 3 (FIRE3)-binding site at -71 to -52 is capable of binding NF-Y, USF and SREBP-1, and is required for the sterol response in conjunction with the co-activator NF-Y around -100. Surprisingly, both FIRE3 and ICE are also necessary for the response to retinoic acid that plays a role in development and is an essential component of the diet.

Background: Convergent evidence from in vivo and in vitro studies of schizophrenia have implicated the mesial temporal lobe as a primary site of pathological change in this disorder. We have previously reported decreased neurotensin receptor density in layer II of the intermediate entorhinal cortex (ERC) in schizophrenia, a finding seen elsewhere but not seen in more caudal ERC., Methods: To study neuroanatomic and diagnostic specificity, we measured the density of neurotensin receptors in the intermediate and caudal ERC and hippocampal formation of schizophrenic, affective disorder control subjects, and normal control subjects. Slide-based radioligand binding was used to perform these studies., Results: Not only schizophrenic but also affective disorder subjects had decreased neurotensin receptor density in layer II of the intermediate ERC. Affective disorder subjects had significantly decreased neurotensin receptor density in layers V/VI of the intermediate ERC, and schizophrenic subjects trended in the same direction., Conclusions: These findings demonstrate region-specific changes in neurotensin receptor binding levels in the mesial temporal lobe; however, there is no clear diagnostic specificity for these changes, because they were seen to varying degrees in both schizophrenia and affective disorders.

The transcription of the fatty acid synthase (FAS) gene is regulated by the sterol status of the cell via cleavage of the sterol regulatory element-binding protein (SREBP). When human HepG2 hepatoma cells were cotransfected with an expression plasmid for mature SREBP-1a together with FAS promoter/reporter constructs significant increases in reporter activity were observed. Deletion analysis of the FAS promoter between -151 and -52 relative to the transcription start site pinpoint two cis-elements important in sterol regulation of the FAS gene. One element, FIRE3, between -71 and -52 can bind in vitro translated and transcribed SREBP-1a whereas the other element, the inverted CCAAT element ICE(-97/-92), binds the trimeric transcription factor NF-Y/CBF as shown with rat liver extract and reconstituted, recombinant NF-Y. The results clearly show that the coactivator for SREBP-1a in this cell line is NF-Y. This finding was confirmed by using a dominant negative form of NF-YA, NF-YAm29, which interferes with the effect of ectopically expressed SREBP-1a on FAS reporter activity.

Inspection of the 5' region of the sequence of the rat fatty acid synthase (FAS) gene revealed a high GC content between -900 and +500, implying several binding sites for members of the Sp1 family of transcription factors. Using SL2 and H4IIE cells in conjunction with FAS promoter/luciferase constructs either successively deleted or containing defined deletions we characterized six GC boxes--GC-I to GC-VI--located between -557 and -83 and discovered a seventh, GC-VII, in the first intron. In vitro DNAse I-footprinting, electrophoretic mobility shift assays, and the yeast one-hybrid system indicated that Sp1 as well as Sp3 interacts with GC-I to GC-VII. Each of the GC boxes conferred Sp1-dependent transcription on the FAS-Mini promoter and in the case of GC-I, Sp1, and Sp3 exert an additive effect on FAS promoter activity.

Electrophoretic mobility shift assay (EMSA) and in vitro transcription/translation show that NF-Y binds to the inverted CCAAT-element in the promoter of the rodent spot14 gene. The NF-Y-binding sequence has been shown to be responsible for basal activity in H4IIE. Given the similar role found for the inverted CCAAT-element in the promoter of the FAS gene, NF-Y may have an important function in the control of lipogenesis.

The regulation of the rat fatty acid synthase gene by mediators such as diet, hormones, cAMP, sterols or retinoic acid is controlled by three NF-Y binding sites. All three sites have a neighbouring Sp1-binding GC-box. This NF-Y/Sp1 motif is conserved in the FAS promoters of rat, human, goose and chicken. We have previously shown cooperative binding of NF-Y and Sp1 to the promoter region at -500 coincident with a diet-induced DNAse I-hypersensitive site. Here, we show an in-vivo interaction of NF-YA with Sp1 using the yeast two-hybrid system. The interacting domains are located between amino acids 55 and 139 of the NF-Y subunit NF-YA and between amino acids 139 and 344 of Sp1. In addition, we show by co-immunoprecipitation direct interaction of NF-Y subunit NF-YA with Sp1 in extracts of rat hepatoma cells H4IIE. Furthermore, we demonstrate by the GST pull-down assay that NF-YA interacts physically with Sp1 in-vitro in the absence of DNA. Therefore, NF-Y can be added to the list of transcription factors interacting with Sp1.

Several putative insulin-responsive elements (IRE) in the fatty acid synthase (FAS) promoter have been identified and shown to be functional in adipocytes and hepatocytes. Here we report on the insulin-responsiveness in the rat hepatoma cell line H4IIE of four cis-elements in the FAS promoter: the FAS insulin-responsive elements, FIRE2 and FIRE3; the inverted CCAAT element, ICE; and the insulin/glucose-binding element, designated hepatic FIRE element, hFIRE, originally identified in rat hepatocytes. Using electrophoretic mobility shift assay (EMSA) competition experiments together with supershifts and in vitro transcription/translation we show that FIRE3 (-68/-58) binds not only the upstream stimulatory factors USF-1/USF-2 but also the CCAAT-binding factor CBF, also known as the nuclear factor Y, NF-Y. The putative IRE FIRE2, which shows sequence similarity to FIRE3, is located between -267 and -249. Gel retardation experiments indicate that USF-1 and USF-2 also bind to this element, which contains an imperfect E-box motif. Using the same approach we have shown that hFIRE binds the stimulatory proteins Sp1 and Sp3 in addition to CBF. Transient transfection experiments using FAS promoter constructs deleted for FIRE2 and FIRE3 demonstrate that neither of these elements mediates the insulin response of the FAS promoter in the rat hepatoma cell line H4IIE, however, ICE at -103/-87 is responsible for mediating the effect of the insulin antagonist cAMP. The hFIRE element located at -57/-34, in spite of its role in the glucose/insulin response in primary rat hepatocytes, is apparently not involved in the insulin regulation of the rat FAS promoter in H4IIE cells. The fact that the topology of the promoters of the FAS genes in rat, human, goose and chicken is conserved regarding CBF-binding sites and USF-binding sites implies an important role for these ubiquitously expressed transcription factors in the regulation of the FAS promoter.

Objective: To examine in vivo the density of brain monoaminergic transporters in Tourette's syndrome (TS)., Background: TS is a heritable neuropsychiatric disorder characterized by chronic vocal and motor tics and is often associated with obsessive-compulsive symptoms. Hyperstimulation of dopamine receptors and dysfunction of serotonergic transmission have been implicated in its pathogenesis, but direct evidence of involvement of these neurochemical systems has been limited., Methods: Symptom severity and the availability of presynaptic monoaminergic transporters in the basal ganglia, midbrain, and thalamus were measured using SPECT and the radioligand [I-123]2beta-carbomethoxy-3beta-(4-iodophenyl)tropane ([I-123]beta-CIT) in 10 patients with TS and in 10 age- and sex-matched normal volunteers., Results: A significant negative correlation was found between a measure of overall tic severity and beta-CIT binding in the midbrain (r = -0.73, p = 0.02) and the thalamus (r = -0.82, p < 0.01). When examined post hoc, these correlations were determined largely by vocal tic severity. No other significant correlations were found between symptom severity and beta-CIT binding in the striatum or cortex., Conclusions: These findings indicate that serotonergic neurotransmission in the midbrain and serotonergic or noradrenergic neurotransmission in the thalamus may be important factors in the expression of TS and may suggest novel targets for treatment.

In vitro DNase I footprint analysis of the rat fatty acid synthase (FAS) promoter from -568 to -468 revealed four protein binding sites: A, B, and C boxes and the FAS insulin-responsive element 1 (FIRE1). As demonstrated by gel mobility shift analysis and supershift experiments, FIRE1, located between -516 and -498, is responsible for binding NF-Y. The C box located downstream of FIRE1 was shown by in vitro footprinting to be a Sp1 binding site, and furthermore, competition with Sp1 also abolished FIRE1 binding. Since the half-life of the Sp1.NF-Y.DNA complex is significantly longer than the half-lives of the Sp1.DNA or NF-Y.DNA complexes, the two transcription factors are deemed to bind cooperatively in the FAS promoter at -500. It is unusual that NF-Y binds at this distance from the start site of transcription. NF-Y binding sites are found in the promoters of at least three other FAS genes, viz. goose, chicken, and man. A second NF-Y binding site is located in the FAS promoter at the more usual position of -103 to -87, and it too has a neighboring Sp1 site. CTF/NF-1 competes for proteins binding to the B box. The A box binds Sp1 and contains a 12/13 match of the inverted repeat sequence responsible for binding the nuclear factor EF-C/RFX-1 in the enhancer regions of hepatitis B virus and the major histocompatibility complex class II antigen promoter. The same relative positions of NF-Y and Sp1 binding sites in the promoters of FAS genes of goose, rat, chicken, and man emphasize the involvement of these transcription factors in the diet and hormonal regulation of FAS.

Using EMSA competition experiments together with supershifts and in vitro transcription/translation we show that the basal transcription factor NF-Y or a related factor binds to the cAMP-responsive inverted CCAAT box recently identified in the rat fatty acid synthase (FAS) gene from nucleotide -99 to -92 relative to the transcription start site of the FAS mRNA. This result indicates a putative novel role for NF-Y in the cAMP-dependent gene regulation in a small class of genes such as FAS and tryptophan hydroxylase. Since NF-Y is a constitutively produced factor, not surprisingly, no differences in the specific DNA/protein complex with the CCAAT(FAS) box and nuclear proteins from H4IIE cells treated with cAMP and/or insulin or not could be observed. This implies that NF-Y might be modified in response to cAMP or might interact with another factor whose properties are altered by cAMP.

In a shotgun approach we sequenced the cosmid pEOA284 containing a fragment derived from the right arm of chromosome XV of Saccharomyces cerevisiae. An analysis of the sequence revealed that it contained open reading frames (ORFs) corresponding to the known genes SLY41, SPS4, COT1, FAA1, PMT3, PRO2 and MYO2. Of the 18 unknown ORFs, five are contained totally within, and two, O6105 and O6163, partially overlap other ORFs. ORF O6116 and O6139 have putative introns. Regions of similarity with chromosomes I and XIII have been uncovered. Interestingly, most of the paired ORFs encode proteins of the same gene family. The relatedness of these ORFs suggests gene duplication.

Tourette syndrome, a chronic tic disorder with autosomal dominant inheritance, exhibits considerable phenotypic variability even within monozygotic twin pairs. The origins of this variability remain unclear. Recent findings have implicated the caudate nucleus as a locus of pathology, and pharmacological evidence supports dopaminergic involvement. Within monozygotic twins discordant for Tourette syndrome severity, differences in D2 dopamine receptor binding in the head of the caudate nucleus predicted differences in phenotypic severity (r = 0.99); this relation was not observed in putamen. These data may link Tourette syndrome with a spectrum of neuropsychiatric disorders that involve associative striatal circuitry.

Evidence from diverse sources, including postmortem investigations, in vivo imaging studies and animal models, suggests that schizophrenia has its origins in early cortical maldevelopment, which in turn may lead to dysfunctional connectivity during brain maturation and clinical symptomatology in early adulthood. Antipsychotic drugs, including the atypical agent clozapine, appear to act at key sites involved in higher cortical-limbic connectivity, possibly mediated by a variety of neurotransmitters. Studies of gene expression may provide a better understanding of how antipsychotic drug effects are integrated at the postsynaptic level. The data from schizophrenia research are discussed within a neurodevelopmental framework.

We describe the use of UV-crosslinking in combination with SDS-PAGE to determine the approximate molecular weight of DNA-binding proteins. A 5-bromo-2'-deoxyuridine (5-BrdU)-substituted, radioactively labeled double-stranded oligonucleotide representing the protein binding site is incubated with a crude nuclear extract containing the protein of interest. Following irradiation with a UV light source, the DNA/protein complex is subjected to SDS-PAGE and its molecular weight determined by comparison with appropriate protein standards.

Neurotensin, an endogenous peptide and putative neurotransmitter, exhibits a wide range of interactions with dopaminergic neurons and displays some actions akin to neuroleptics. Moreover, neurotensin receptors are abundant in specific layers of the entorhinal cortex where cytoarchitectural abnormalities have been reported in schizophrenia. We therefore examined the entorhinal cortex from postmortem specimens of five control patients and six schizophrenic patients for alterations in neurotensin receptor quantitation and distribution using receptor autoradiography. Specific 125I- neurotensin binding was concentrated in layer II cell clusters, with a 40% reduction in binding in the schizophrenic group (p < 0.05). Moderate binding was observed in both cohorts in deep layers V/VI, with negligible binding in the hippocampus. There was no statistical difference in quantitative neurotensin binding in other lamina of the entorhinal cortex of schizophrenics compared with controls. The characteristic laminar pattern of binding did not differ between cohorts. The reduction in neurotensin binding in schizophrenics is consistent with an increasing number of reports of structural abnormalities in the medial temporal lobe of schizophrenics in general and the entorhinal cortex in particular. Further studies are required to examine the evidence for neuroanatomic and neurochemical pathology in the entorhinal cortex.

Here we show that insulin may play a role in the diet-induced regulation of the rat fatty acid synthase (FAS; EC 2.3.1.85). Transient transfection of human and rat hepatoma cell lines with successively deleted FAS/CAT promoter fusion plasmids was used to determine the effect of insulin on FAS promoter activity. Our results indicate the existence of cis-acting insulin-responsive elements in the FAS promoter; the position of one of these is coincident with the position of a previously determined diet-induced DNAse I hypersensitive site (HSi-1) at approximately -500 bp relative to the transcription start site of FAS mRNA.

Two patients with schizophrenia had markedly abnormal neuroimaging studies; one was found to have a large, cystic cavum vergae, while the other demonstrated complete agenesis of the septum pellucidum with a monoventricular system. The increased prevalence of developmental abnormalities of midline forebrain structure in patients with schizophrenia suggests that dysgenesis of these structures may be contributory to the neurobiology of schizophrenia and other psychotic disorders.

The distribution of 5-HT3 receptors was examined in the human medulla using [3H]LY278584, a highly selective 5-HT3 receptor antagonist. The highest density of 5-HT3 receptors was found in the substantia gelatinosus subnucleus of nucleus of the solitary tract (NTS) throughout its rostrocaudal extent, followed by the dorsal subnucleus, the area postrema (AP), the commissural subnucleus, the medial subnucleus, and in an arc corresponding to the pars gelatinosus of the spinal trigeminal nucleus (nSp5). The distribution of 5-HT3 receptors in the brain may help explain some of the reported CNS activities of 5-HT3-selective drugs. The anti-emetic and antinociceptive activities of 5-HT3 antagonists may be mediated by receptors in sensory areas of the brainstem.

The laminar and rostro-caudal distribution of 125I-neurotensin binding sites is described in human entorhinal cortex using quantitative autoradiography. Specific binding was most prominent over the cell clusters of layer II of the entorhinal cortex throughout its rostro-caudal extent. Dense binding was also observed in the adjacent presubiculum and cortical amygdaloid transition area, whereas minimal binding was detected in the hippocampus and dentate gyrus. 125I-Neurotensin may serve as a selective probe for neurotensin receptor alterations and layer II-specific cytoarchitectural disturbances in the entorhinal cortex in neuropsychiatric diseases associated with abnormalities of the mesial temporal lobe.

Schizophrenia is the most prevalent of the major psychoses, but the underlying neurobiology of this debilitating disorder remains mysterious. Recent developments in molecular biology, neuroanatomic pathology, neurochemistry, and functional imaging suggest that a number of factors converge to produce schizophrenia. Specifically, an early neurodevelopmental "lesion," possibly within the mesial temporal lobe, may contribute to later temporolimbic-prefrontal dysfunction as the nervous system matures. Genetic factors appear to facilitate liability to schizophrenia, and dopaminergic and possibly other neurotransmitter systems may mediate clinical expression of the illness through newly recognized receptor subtypes.

The repertoire of cytokine and cytokine receptor mRNA expressed by unstimulated human thymocytes and thymic stromal cells was explored by a quantitative polymerase chain reaction (PCR) using sequence specific internal standards. Of the 18 cytokines tested we found a considerable overlap in the expression of cytokines by human thymocytes and by thymic stromal cells; both cell types express the mRNA for interleukin-1 beta(IL-1, IL-6, IL-7 and tumour necrosis factor-alpha (TNF-alpha). However, there are substantial differences in the levels of cytokine mRNA expressed in these two types of cells as revealed by the quantitative PCR assay. Stromal cells express considerably higher levels of IL-1 beta and IL-6 than thymocytes (14- and 27-fold respectively). In addition, a number of cytokines such as lymphotoxin and interferon-gamma (IFN-gamma), are expressed exclusively in thymocytes whereas others such as stem cell factor (SCF), IL-1 receptor antagonist-2 (IRAP-2) and granulocyte-macrophage colony-stimulating factor (GM-CSF) are produced only in stromal cells. There is a complete overlap in the expression of a group of cytokine receptors tested in thymocytes and thymic stromal cells; these include IL-1R, IL-2R, IL-6R, IL-7R, TNFR and stem cell growth factor receptor (c-KIT). The expression of specific cytokines by thymic stromal cells and the parallel expression of their receptors on thymocytes under physiological conditions, support the hypothesis that these cytokines participate in paracrine interactions between these two cell populations during thymocyte differentiation.

We report the binding properties and subnuclear localization of [3H]hemicholinium-3 binding sites in human amygdala using quantitative autoradiography. Specific binding was saturable and high affinity (apparent Kd 2-11 nM). Binding was highest in the basolateral nucleus which receives dense cholinergic innervation from the basal forebrain. Binding closely approximated acetylcholinesterase reactivity. These data support [3H]hemicholinium-3 as a quantitative marker for cholinergic terminals in human brain.

The binding of [125I]physalaemin to rat brain slices was investigated. Radiolabeled physalaemin bound with high affinity (Kd = 0.3 nM) to a single class of sites (Bmax = 22 fmol/mg protein). Kinetic studies indicated that binding was time-dependent and all specific binding was reversible. Pharmacology studies indicated that specific [125I]physalaemin binding was inhibited by structurally related peptides such as substance P and eledoisin. Biochemical studies indicated that specific binding of radiolabeled physalaemin was greatly reduced if the brain slices were pretreated with heat, trypsin or N-ethyl maleimide. Autoradiographic studies indicated that the [125I]physalaemin binding sites were discretely distributed throughout the brain. Highest grain densities were present in the olfactory bulb, dentate gyrus, amygdala, superficial layers of the superior colliculus, subiculum, dorsal parabrachial nucleus, locus coeruleus, nucleus tractus solitarii and dorsal horn of the spinal cord. Moderate grain densities were present in the nucleus accumbens, olfactory tubercle, pyriform cortex, striatum, hippocampus, inferior colliculus and central gray of the midbrain. Low grain densities were present in most thalamic nuclei, the substantia nigra and cerebellum. The corpus callosum and controls treated with 1 microM unlabeled physalaemin had negligible levels of binding. The unique pharmacological and regional distribution data obtained suggest that [125I]physalaemin may serve as a valuable probe to study central substance P receptors.

Binding sites in the rat forebrain were characterized using (125I-Tyr4)bombesin as a receptor probe. Pharmacology experiments indicate that gastrin releasing peptide (GRP) and the GRP fragments GRP as well as Ac-GRP inhibited radiolabeled (Tyr4)bombesin binding with high affinity. Biochemistry experiments indicated that heat, N-ethyl maleimide or trypsin greatly reduced radiolabeled (Tyr4)bombesin binding. Also, autoradiographic studies indicated that highest grain densities were present in the stria terminalis, periventricular and suprachiasmatic nucleus of the hypothalamus, dorsomedial and rhomboid thalamus, dentate gyrus, hippocampus and medial amygdaloid nucleus. The data suggest that CNS protein receptors, which are discretely distributed in the rat forebrain, may mediate the action of endogenous GRP/bombesin-like peptides.