CBED contrast in the lower order Laue zone.

Contrast in a systematic arrangement of lower order Laue zone (LOLZ) beams is reported and analysed using a Bloch wave description. Observations are reported for hexagonal barium ruthenium zirconate (Ba4Ru3ZrO12) and barium ruthenium titanate (Ba3Ti2RuO9), both near the c-axis orientation. The specific scattering dynamics invoked by this diffraction geometry may have novel uses in the exploration of crystallographic parameters.

Energetics of nanocrystalline TiO2.

The energetics of the TiO(2) polymorphs (rutile, anatase, and brookite) were studied by high temperature oxide melt drop solution calorimetry. Relative to bulk rutile, bulk brookite is 0.71 +/- 0.38 kJ/mol (6) and bulk anatase is 2.61 +/- 0.41 kJ/mol higher in enthalpy. The surface enthalpies of rutile, brookite, and anatase are 2.2 +/- 0.2 J/m(2), 1.0 +/- 0.2 J/m(2), and 0.4 +/- 0.1 J/m(2), respectively. The closely balanced energetics directly confirm the crossover in stability of nanophase polymorphs inferred by Zhang and Banfield (7). An amorphous sample with surface area of 34,600 m(2)/mol is 24.25 +/- 0.88 kJ/mol higher in enthalpy than bulk rutile.

The human mineralocorticoid receptor gene promoter: its structure and expression.

The structure and expression of a clone containing the promoter region, all of exon 1, and part of the first intron of the human mineralocorticoid receptor (hMR) gene is presented. The clone has three sets of CAAT and TATA elements, one located at the very 5'-end of the clone, one located just 5'- to the start of transcription, and one set located in intron A, approximately 300 bp into the intron. The major start of transcription site by primer extension analysis and ribonuclease protection assays is located 26 bp downstream of a TATA-like box (TTTAA) and 90 and 143 bp downstream, respectively, of two CCAAT boxes. Putative cis-transcription factor binding sites are as follows: two potential AP1 sites, one potential AP2 site, two ATF/CREB sites, six potential GC boxes or SP1 sites, one potential perfect half-palindromic estrogen response element, and three potential PEA3 sites. Therefore, the hMR promoter region contains elements characteristic of both regulated genes and "housekeeping" genes. CAT assays of overlapping deletions of the promoter region demonstrated tissue-specific regulation in human neuroepithelioma (SK-N-MC-IXC) and non-neuronal, peripheral choriocarcinoma cell lines (JEG-3).

Solid state NMR characterisation of crystalline Na2O-ZrO2-SiO2 phases.

The NMR interactions of crystalline phases in the system Na2O-ZrO2-SiO2 have been studied by a combination of static and magic angle spinning NMR methods for the first time. A full multinuclear (17O, 23Na, 29Si and 91Zr) approach has been employed that allows the phases to be clearly identified. NMR interactions such as 29Si isotropic chemical shift correlate with the known structural units present. For 23Na the the different sites can often be distinguished on the basis of differing quadrupolar interactions.

17O magic-angle spinning nuclear magnetic resonance of CaCO3.

The first 17O magic-angle spinning nuclear magnetic resonance (MAS-NMR) from a carbonate ion in an inorganic compound is reported. The 17O MAS centreband of CaCO3 can be simulated with parameters CQ = 6.97 MHz, eta approximately 1 and an isotropic chemical shift of 204 ppm.

Structural characterisation of Na2ZrO3.

A combination of X-ray and electron diffraction, electron microscopy and solid-state nuclear magnetic resonance (NMR) has been used to elucidate the structure and the ordering of Na2ZrO3. The diffraction data confirm a monoclinic crystal structure. A sample prepared by a conventional solid-state reaction of the components is shown by both X-ray diffraction and electron microscope imaging to have an extremely high concentration of planar defects associated with stacking disorder of the planes along the c-axis. The incidence of these defects is significantly reduced in a sample recrystallised from a bismuth oxide flux. NMR indicates that the local coordinations are well defined in both samples but with some sharpening of the spectra from the recrystallised sample indicative of the increase of long-range order. The 23Na magic angle spinning (MAS) NMR spectra clearly show three distinct sites with widely differing quadrupolar interaction parameters that can be related to the known site symmetries. Two distinct oxygen resonances are observed in the MAS NMR spectrum from an 17O-enriched sample while the static 91Zr NMR spectrum can be simulated with one set of interaction parameters.

Schizophrenic deficits: neuroleptics and the prefrontal cortex.

Modern techniques have been applied to brain modeling, based on recent approaches in the artificial intelligence field that use brain-like "connectionistic" computational architectures. The model proposed by Cohen and Servan-Schreiber uses a gain parameter which they identify with dopamine function. They apply their model to neuroleptically treated schizophrenia patients who show improved task performance which they link to increased dopamine function and increased gain in the prefrontal cortex. However, evidence indicates that antipsychotic medications block dopamine (especially D2) receptors, decreasing mesolimbic and mesocortical dopamine function. If therapeutic dosages of neuroleptics diminish dopamine function, this would decrease gain in context modules needed for adequate task performance. Schizophrenia patients would perform more poorly by further reducing gain in their already compromised context modules. The current investigators suggest three possible ways to resolve this difficulty, to explain why normals perform more poorly when taking neuroleptics, although acute schizophrenia patients' performance may be enhanced in several areas. Evidence would suggest that multiple processes occur simultaneously in neuroleptically treated patients with some processes counterbalancing others.

Differential mineralocorticoid (type 1) and glucocorticoid (type 2) receptor expression in Lewis and Fischer rats.

Lewis (LEW/N) and Fischer (F344/N) rats represent two extremes of the spectrum of corticosterone responses to stressful stimuli, from the chronical hyporesponsiveness of LEW/N to the chronical hyperresponsiveness of F344/N. It might be expected that the amount of mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) binding, and the levels of their corresponding mRNAs in various tissues in LEW/N and F344/N rats might reflect the overall integrated levels of corticosterone to which these receptors have been exposed. We have found that while the binding affinity (Kd) of MR and GR varies between tissues, there was no strain difference in any tissue. Receptor binding number (Bmax), however, varied not only between tissues, but also between strains. MR Bmax in the hippocampus and pituitary was lower in LEW/N than in F344/N, whereas the GR Bmax in the LEW/N thymus was greater than that found in F344/N rats. The hippocampal levels of MR mRNAs in Adx LEW/N and F344/N rats were in good agreement with, and paralleled, the functional levels of these receptors as determined by binding assays. On the other hand, the number of hippocampal GR binding sites and the level of GR nRNA while similar were not identical in the two strains: the hippocampal GR Bmax did not differ between strains, while the hippocampal GR mRNA level was slightly, but significantly, lower in Adx LEW/N compared to F344/N rats.

Effects of the glucocorticoid antagonist RU 486 on pituitary-adrenal function in patients with anorexia nervosa and healthy volunteers: enhancement of plasma ACTH and cortisol secretion in underweight patients.

To further explore whether the hypercortisolism of anorexia nervosa reflects an alteration in the set point for corticotropin-releasing hormone (CRH) secretion or is a manifestation of glucocorticoid resistance, we examined plasma ACTH and cortisol responses to the competitive glucocorticoid antagonist RU 486 (10 mg/kg, p.o. at 8.00 h) versus placebo (PBO) in 7 healthy female volunteers and 8 patients with DSM-III-R anorexia nervosa, all of whom were studied while underweight [64.3 +/- 2.1% average body weight (ABW), mean +/- SE] and 5 of whom were restudied longitudinally following refeeding (> or = 85% ABW, mean 87.4 +/- 0.4% ABW). Blood samples were obtained from 16.00 to 16.30 h and from 4.00 to 8.00 h following dosing. Underweight anorexics were significantly hypercortisolemic by 24 h urinary free cortisol excretion compared with controls (239 +/- 37 vs. 119 +/- 12 nmol/day, p < 0.01). Both controls and underweight anorexics had robust early morning (4.00-8.00 h) plasma cortisol responses to RU 486 (465 +/- 61 and 719 +/- 49 nmol/l) compared with PBO (370 +/- 52 and 451 +/- 31 nmol/l; p < 0.02 and p < 0.01, respectively). The underweight anorexics showed a significant mean early morning plasma ACTH response to RU compared with placebo (3.28 +/- 0.63 vs. 2.01 +/- 0.24 pmol/l, p < 0.05), while the controls showed a trend toward an increase in mean plasma ACTH after RU (3.11 +/- 0.36 pmol/l) compared with PBO (2.31 +/- 0.41 pmol/l, p < 0.13); plasma ACTH means were greater on the RU day than the placebo day at 20 of 25 sampling points (p < 0.001). However, the increment in ACTH on the RU day compared to the placebo day was greater in the underweight anorexics at the first 20 of 25 consecutive time points of the early morning sampling period (p < 0.001). Moreover, underweight anorexics showed a significant plasma ACTH and cortisol response to RU 486 at 16.00-16.30 h (8-8.5 h following administration), while the controls showed no significant response of plasma ACTH or cortisol at this time. When restudied following weight recovery, anorexic patients showed reductions in 24-hour urinary free cortisol excretion (to 191 +/- 40 nmol/day) which were no longer significantly elevated compared with control values.(ABSTRACT TRUNCATED AT 400 WORDS)

Intrahippocampal colchicine alters hypothalamic corticotropin-releasing hormone and hippocampal steroid receptor mRNA in rat brain.

The hippocampus appears to be an important modulator of the negative feedback effects of glucocorticoids on the hypothalamic-pituitary-adrenal axis. It is not known if hippocampal subfields CA1-4 or the dentate gyrus differentially alter gene expression of corticotropin-releasing hormone (CRH) in the paraventricular nucleus (PVN) of the hypothalamus. We, therefore, examined the effects of selective destruction of dentate gyrus granule cells, which send excitatory glutaminergic inputs to subfields CA4, CA3 and CA2, on CRH expression in the PVN. To determine the possible involvement of steroid receptors in the regulation of CRH expression, we examined the effects of intrahippocampal colchicine on gene expression of the mineralocorticoid (MR; type I) and glucocorticoid (GR; type II) receptors in hippocampal CA fields and dentate gyrus. Colchicine produced a selective loss of dentate gyrus granule cells without affecting pyramidal cells in CA1-4 as early as 1 day after injection; granule cells were completely destroyed after 3 days. CRH mRNA levels were reduced by 38-48% in the PVN 2-14 days after colchicine. MR mRNA levels were decreased in dorsal and ventral CA fields 1-7 days after colchicine. GR mRNA levels were relatively unchanged, showing a slight decrease only in dorsal CA fields on days 2-7. Unexpectedly, CRH was transiently expressed in dorsal and ventral CA fields 1-3 days after colchicine. In the same time period, mRNA levels of inositol 1,4,5-trisphosphate kinase were decreased, suggesting that increases in neural metabolic activity, indicated by this marker, are not responsible for the transient CRH effect. The results suggest that the dentate gyrus is important for maintenance of steroid hormone receptor mRNA levels in the hippocampus and CRH expression in the hypothalamic PVN, and that CRH gene expression is differentially regulated in the hypothalamus and hippocampus.

The antidepressants fluoxetine, idazoxan and phenelzine alter corticotropin-releasing hormone and tyrosine hydroxylase mRNA levels in rat brain: therapeutic implications.

Various classes of antidepressant drugs with distinct pharmacologic actions are differentially effective in the treatment of classic melancholic depression--characterized by pathological hyperarousal and atypical depression--associated with lethargy, hypersomnia, and hyperphagia. All antidepressant agents exert their therapeutic efficacy only after prolonged administration. In situ hybridization histochemistry was used to examine in rats the effects of short-term (2 weeks) and long-term (8 weeks) administration of 3 different classes of activating antidepressant drugs which tend to be preferentially effective in treating atypical depressions, on the expression of central nervous system genes thought to be dysregulated in major depression. Daily administration (5 mg/kg, i.p.) of the selective 5-hydroxytryptophan (5-HT) reuptake inhibitor fluoxetine, the selective alpha 2-adrenergic receptor antagonist idazoxan, and the nonspecific monoamine oxidase A and B inhibitor phenelzine increased tyrosine hydroxylase mRNA levels by 70-150% in the locus coeruleus after 2 weeks of drug and by 71-115% after 8 weeks. The 3 drugs decreased corticotropin-releasing hormone mRNA levels by 30-48% in the paraventricular nucleus of the hypothalamus. The decreases occurred at 8 weeks but not at 2 weeks. No consistent change in steroid hormone receptor mRNA levels was seen in the hippocampus with the 3 drugs, but fluoxetine and idazoxan increased the level of mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) mRNA, respectively, after 8 weeks of drug administration. Proopiomelanocortin (POMC) mRNA levels in the anterior pituitary and plasma adrenocorticotropic-hormone (ACTH) levels were not altered after 2 or 8 weeks of drug treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Arthritis-susceptible Lewis rats fail to emerge from the stress hyporesponsive period.

Susceptibility to streptococcal cell wall (SCW)-induced arthritis in 4- to 6-week-old Lewis (LEW/N) rats is associated with blunted glucocorticoid production secondary to a profound defect in inflammatory mediator-induced hypothalamic corticotropin-releasing hormone (CRH) biosynthesis and secretion. The relative SCW arthritis resistance in histocompatible Fischer (F344/N) rats, on the other hand, is associated with robust hypothalamic-pituitary-adrenal (HPA) axis responses to inflammatory mediators. In this study, we investigated HPA axis responses to SCW during the postnatal developmental period in LEW/N and F344/N rats. We found that SCW-induced plasma corticosterone (CORT) responses do not significantly increase during development in LEW/N, while such responses clearly appear at postnatal day 14 in F344/N and outbred Harlan-Sprague-Dawley (HSD) rats. Additionally, LEW/N rats fail to exhibit the normal ontogenic increase in CRH mRNA levels in the paraventricular nucleus (PVN), whereas their SCW-induced PVN CRH mRNA responses are blunted compared to F344/N at postnatal day 14. Taken together, these results suggest that LEW/N rats fail to emerge completely from their stress hyporesponsive period. This may account for the lack of stress responsiveness in young adult LEW/N rats, and consequently, for their susceptibility to SCW-induced arthritis and other inflammatory diseases.

Long-term antidepressant administration alters corticotropin-releasing hormone, tyrosine hydroxylase, and mineralocorticoid receptor gene expression in rat brain. Therapeutic implications.

Imipramine is the prototypic tricyclic antidepressant utilized in the treatment of major depression and exerts its therapeutic efficacy only after prolonged administration. We report a study of the effects of short-term (2 wk) and long-term (8 wk) administration of imipramine on the expression of central nervous system genes among those thought to be dysregulated in imipramine-responsive major depression. As assessed by in situ hybridization, 8 wk of daily imipramine treatment (5 mg/kg, i.p.) in rats decreased corticotropin-releasing hormone (CRH) mRNA levels by 37% in the paraventricular nucleus (PVN) of the hypothalamus and decreased tyrosine hydroxylase (TH) mRNA levels by 40% in the locus coeruleus (LC). These changes were associated with a 70% increase in mRNA levels of the hippocampal mineralocorticoid receptor (MR, type I) that is thought to play an important role in mediating the negative feedback effects of low levels of steroids on the hypothalamic-pituitary-adrenal (HPA) axis. Imipramine also decreased proopiomelanocortin (POMC) mRNA levels by 38% and glucocorticoid receptor (GR, type II) mRNA levels by 51% in the anterior pituitary. With the exception of a 20% decrease in TH mRNA in the LC after 2 wk of imipramine administration, none of these changes in gene expression were evident as a consequence of short-term administration of the drug. In the light of data that major depression is associated with an activation of brain CRH and LC-NE systems, the time-dependent effect of long-term imipramine administration on decreasing the gene expression of CRH in the hypothalamus and TH in the LC may be relevant to the therapeutic efficacy of this agent in depression.

Cerebrospinal fluid immunoreactive corticotropin-releasing hormone and adrenocorticotropin secretion in Cushing's disease and major depression: potential clinical implications.

To explore whether possible differences in central nervous system neuromodulators contribute to the differential presentation of affective symptomatology in Cushing's disease and major depression, we examined the levels of immunoreactive CRH and ACTH in the cerebrospinal fluid (CSF) of 11 patients with Cushing's disease, a patient with ectopic ACTH secretion, 34 patients with major depression, and 60 healthy subjects. We elected to measure these peptides not only because both are classically involved in pituitary-adrenal regulation, but also because their primarily arousal-producing and anorexigenic behavioral effects in experimental animals suggest that they may play a role in the symptom complex of depressive syndromes. We also explored whether the CSF levels of these peptides were more helpful in determining the often difficult differential diagnosis between major depression and Cushing's disease than the plasma ACTH response to ovine CRH, a currently used but somewhat insensitive laboratory means of distinguishing these disorders. CSF levels of immunoreactive CRH and ACTH were significantly lower in Cushing's disease patients [21.9 +/- 2.7 and 15.4 +/- 1.8 pg/mL, (mean +/- SEM), respectively] compared to patients with major depression [38.4 +/- 2.3 pg/mL (P less than 0.01) and 24.5 +/- 1.6 pg/mL (P less than 0.01), respectively] and controls [38.4 +/- 1.6 pg/mL (P less than 0.001) and 26.3 +/- 1.1 pg/mL (P less than 0.001), respectively]. The coexistence of high plasma ACTH and low CSF ACTH in Cushing's disease yielded a CSF/plasma ACTH ratio consistently less than that in depressed patients, with only 2 of 31 subjects comprising both groups showing values that overlapped. In contrast, 9 of the combined patients showed ACTH responses to ovine CRH that overlapped. These data suggest that differences in centrally directed CRH secretion may account for the differential presentation of the dysphoric syndromes seen in major depression and Cushing's disease. Hence, the classic form of major depression (melancholia), is often associated with evidence of pathological hyperarousal, such as intense anxiety, sleeplessness, and anorexia, while that of Cushing's disease is associated with evidence of pathological hyperarousal, including hyperphagia, fatigue, and inertia. Moreover, measurement of the CSF/plasma ACTH ratio may serve as a clinically useful adjunct to the ovine CRH stimulation test and other laboratory measures in determining the differential diagnosis between major depression and Cushing's disease.

Optimization of cRNA probe in situ hybridization methodology for localization of glucocorticoid receptor mRNA in rat brain: a detailed protocol.

1. We have described a general ribonucleotide probe in situ hybridization methodology for localization of mRNA in frozen, unfixed tissue sections of brain. 2. The most important steps in obtaining consistent and reproducible autoradiographs with ribonucleotide probes were tissue acetylation and application of the radiolabeled probe to tissue sections under unsealed, glass coverslips. 3. Variability of the hybridization signal in tissue sections has been minimized to achieve a high degree of reproducibility within a given experiment as determined by densitometric analysis of rat glucocorticoid and mineralocorticoid receptor mRNA hybridization autoradiographs. 4. Tissue quality has been optimized for high-resolution anatomical localization of mRNA species by nuclear track emulsion. 5. The protocol is amenable to rapid, batchwise processing of tissue samples.

Effects of glucocorticoid antagonism with RU 486 on pituitary-adrenal function in patients with major depression: time-dependent enhancement of plasma ACTH secretion.

Data from our group and others suggest that pituitary-adrenal activation in major depression reflects a defect at or above the hypothalamus which results in the hypersecretion of corticotropin-releasing hormone (CRH); some have suggested, however, that elevated indices of cortisol secretion and lack of suppressibility to dexamethasone may be a manifestation of a primary defect in glucocorticoid receptor activation. We report here a study of early morning pituitary-adrenal responses to the glucocorticoid antagonist RU 486 in patients with major depression and healthy volunteers. Previous data suggested that the response to RU 486 could represent an index of endogenous CRH secretory activity. RU 486 produced a robust increase in plasma corticotropin (ACTH) and cortisol secretion in both control subjects and depressed patients. In the controls, however, the increase was confined to the last 2 hours of sampling (6 to 8 am), whereas in the depressed patients the increase occurred throughout the sampling period (3 to 8 am). The ACTH response in the depressed patients exceeded that in the controls during most of the sampling period, including a significant (p less than .005) increase between 3 and 4:30 am. These results are compatible with the idea that hypercortisolism in major depression represents an alteration in the overall set point for hypothalamic CRH secretion rather than a primary alteration at the level of the glucocorticoid receptor.

Corticotropin-releasing hormone: from endocrinology to psychobiology.

Corticotropin-releasing hormone (CRH) is a brain neuropeptide which coordinates the endocrine, autonomic and behavioral responses to stress. We review the abnormal response to exogenous CRH in various psychiatric syndromes, including major depression and anorexia nervosa. We also contrast pituitary responses to CRH in patients with depression versus Cushing's disease. We hypothesize that CRH may play a role in the pathogenesis of various psychiatric syndromes which are characterized during their course by the symptom of depression.

Coordinate developmental regulation of high and low molecular weight mRNAs for rat insulin-like growth factor II.

Insulin-like growth factor II (IGF-II) is a mitogenic polypeptide that is thought to play a role in fetal growth and development. To study the hormonal and developmental regulation of IGF-II gene expression, we have isolated a cDNA clone for rat IGF-II (rIGF-II) from a 12S [1.2-kilobase-pair (kbp)] fraction of mRNA from a rat liver cell line (BRL-3A) that directs the cell-free synthesis of pre-pro-rIGF-II. In the present study, the rIGF-II probe was used to determine the size of IGF-II RNA. Surprisingly, in BRL-3A cells and in neonatal liver, the probe hybridized under stringent conditions 10-20 times more strongly to a larger (4 kbp) RNA than to 1.2-kbp RNA. The 4-kbp RNA is almost exclusively cytoplasmic and is colinear with a 551-base fragment of the rIGF-II cDNA insert containing coding and 3' noncoding regions. The 4-kbp and 1.2-kbp RNA species are regulated coordinately with developmental age, being high in liver from neonatal rats but not detectable in liver from older animals, suggesting that both IGF-II mRNA species arise from a single primary transcript by alternative RNA processing. Although oligodeoxynucleotide hybridization and S1 nuclease protection experiments suggest that the 4-kbp RNA contains an intact protein-coding region, fractions enriched in 4-kbp RNA do not direct the translation of pre-pro-rIGF-II in vitro. This may indicate that the 4-kbp RNA specifies an altered protein product that has not yet been recognized, or alternatively that it contains a normal protein-coding region but requires further RNA processing to be activated for translation.

Isolation of a cDNA clone encoding rat insulin-like growth factor-II precursor.

Insulin-like growth factor-I (IGF-I) and IGF-II are mitogenic polypeptides of relative molecular mass (Mr) approximately 7,500 isolated from human plasma each containing four peptide domains in a single chain and identical at more than 60% of their amino acid loci. The B- and A-domains of the IGFs are approximately 40% identical to the B- and A-chains of human insulin. IGF-I and IGF-II have similar in vitro biological activities and receptor reactivity, but are immunologically distinct. IGF-I appears to mediate the effects of growth hormone on cartilage to promote skeletal growth whereas IGF-II may have a special role in fetal development and in the central nervous system. To investigate the in vivo role of IGF-II, we have studied IGF-II biosynthesis in the BRL-3A rat liver cell line. BRL-3A cells synthesize and secrete a 7,484 Mr protein 93% identical to human IGF-II and representing rat IGF-II (rIGF-II). Rat IGF-II is synthesized as a approximately 22,000 Mr prepro-rIGF-II (ref. 12) from 12 S poly(A)+mRNA. In addition, approximately 20,000 Mr pro-rIGF-II has been identified in lysates of biosynthetically labelled intact BRL-3A cells. We report here the isolation of an almost complete cDNA clone for rIGF-II. Our results indicate that pro-rIGF-II is synthesized as a 156 amino acid peptide precursor (17,619 Mr) containing mature rIGF-II 1-67 at its amino-terminus and an 89-residue carboxy-terminal peptide extension.