High temperature ferromagnetism in GaAs-based heterostructures with Mn delta doping.

We show that suitably designed magnetic semiconductor heterostructures consisting of Mn delta (delta)-doped GaAs and p-type AlGaAs layers, in which the locally high concentration of magnetic moments of Mn atoms are controllably overlapped with the two-dimensional hole gas wave function, realized remarkably high ferromagnetic transition temperatures (T(C)). A significant reduction of compensative Mn interstitials by varying the growth sequence of the structures followed by low-temperature annealing led to high T(C) up to 250 K. The heterostructure with high T(C) exhibited peculiar anomalous Hall effect behavior, whose sign depends on temperature.

Stereoselective synthesis of the neolignan, (+ )-dehydrodiconiferyl alcohol.

A stereo controlled synthesis of the biologically active neolignan, (+)-dehydrodiconiferyl alcohol (1) was achieved. This synthetic method was also efficient for preparing its enantiomer and other derivatives with biological activity.

Improved procedure for the enantiometric synthesis of 1-hydroxy/acetoxy-2,6-diaryl-3,7-dioxabicycl.

Short enantiomeric syntheses of the 1-hydroxy/acetoxy-3,7-dioxabicyclo[3.3.0]octane lignans, paulownin, and (+)-phrymarin I and II, were accomplished by starting from the chiral synthon, (R)-(+)-3-hydroxybutanolide, and employing photocyclization as the key step.

Generation of polyclonal antiserum against the growth hormone secretagogue receptor (GHS-R): evidence that the GHS-R exists in the hypothalamus, pituitary and stomach of rats.

Growth hormone (GH) secretagogues (GHSs), which stimulate GH secretion, are synthetic compounds that act through the GHS receptor (GHS-R) which has been recently cloned. We raised an antiserum in a rabbit against a synthetic peptide corresponding to amino acid residues 248-260 of the third intracellular loop of the rat GHS-R. A competitive immunoassay showed that the antiserum had a specific affinity for the target peptide. To confirm the specificity of the antiserum, the GHS-R cDNA was stably expressed in COS-7 cells. In Western blot analysis, the band was detected at 44 kDa in the extracts from COS-7 cells expressing GHS-R (COS-7/tf3-2) but not in those from wild-type COS-7 cells. Furthermore, while COS-7/tf3-2 cells were strongly immunostained for GHS-R, no GHS-R-like immunoreactivity was observed in wild-type COS-7 cells. Immunoreactive bands were also observed at approximately 46 kDa in the extracts from rat hypothalamus, pituitary and stomach by Western blot analysis. These studies are the first to show the existence of GHS-R protein in the stomach. The antiserum for the GHS-R is sensitive and specific, and it would be useful for clarifying the roles of GHS/ghrelin.

Reduced growth hormone receptor messenger ribonucleic acid in an aged man with chronic malnutrition and growth hormone resistance.

A severely malnourished 87-yr-old man presented with hypoglycemia. Serum GH levels were elevated, and serum levels of insulin-like growth factor I (IGF-I), IGF-binding protein-3, and GH-binding protein were extremely reduced. The patient's GH was biologically active. Administration of GH for 4 consecutive days resulted in a slight increment in serum IGF-I levels, but no elevation of serum IGF-binding protein-3. The expression of GH receptor messenger ribonucleic acid in the liver was greatly reduced. An autopsy revealed a Rathke's cleft cyst confined to the sella turcica. Immunohistochemical studies for GH showed that there was nothing to suggest a tumor overproducing GH. In addition, TSH levels were elevated in the presence of normal thyroid hormone levels, and there was a cluster of cells showing strong immunohistochemical staining for the TSH beta-subunit in the pituitary. In this patient, the decreased expression of GH receptor messenger ribonucleic acid in the liver may have been responsible for the GH resistance, which was probably caused by malnutrition.

Prohormone convertases 1 and 2 process ProPACAP and generate matured, bioactive PACAP38 and PACAP27 in transfected rat pituitary GH4C1 cells.

Pituitary adenylate cyclase-activating polypetide (PACAP) exists in two amidated forms, PACAP38 and PACAP27, which are expressed in the magnocellular and parvocellular neurons of the paraventricular nucleus (PVN) and the magnocellular neurons of the supraoptic nucleus (SON) of the hypothalamus. The prohormone convertases PC1 and PC2, subtilisin-like PCs of the Kex2 family, are expressed in neuroendocrine cells. Immunocytochemistry and in situ hybridization of PC1 and PC2 in the hypothalamus have shown that PC1 and PC2 are also present in the PVN and SON. Therefore, it is possible that the precursor of PACAP is processed by PC1 and/or PC2 in the hypothalamic nuclei and then converted to its mature forms. To test this hypothesis, rat pituitary GH4C1 cells were supertransfected with human PACAP cDNA and either rat PC1 or PC2 cDNA. The acid extracts of these cells were analyzed by reversed-phase HPLC for proPACAP, PACAP38 and/or PACAP27 radioimmunoassays using three antibodies with different recognition sites, and then bioassayed for the ability to stimulate adenylate cyclase. The cells transfected with PACAP cDNA alone yielded PACAP-like immunoreactivity (PACAP-li) corresponding to molecular weights between 15 and 20 kDa without PACAP bioactivity. Cotransfection of these cells with PC1 or PC2 generated PACAP-li, which coeluted with synthetic PACAP38 and PACAP27, respectively. Western blot also revealed 4.5- and 3.0-kDa PACAP-li bands, which correspond to the molecular weights of PACAP38 and PACAP27, respectively. The HPLC fractions containing PACAP-li, which were coeluted with synthetic PACAP38 and PACAP27, showed marked bioactivities. These findings suggest that the precursor of PACAP expressed in the PVN and SON of the hypothalamus could be efficiently processed by PC1 and PC2, and then converted to mature, bioactive PACAP38 and PACAP27.

Effect of Optically Active Ethyl 2-Phthalimidooxypropionate on the Growth of Cress, Lepidium sativum.

The effects of 2-phthalimidooxyalkanoic acid derivatives on the germination and root-growth of cress were examined. Since 2-phthalimidooxypropionates were most effective, the optically active ethyl esters were prepared. As the result of biological testing, the (S)-(-)-isomer exhibited stronger activity than the (R)-(+)-isomer. This result is contrary to those from commercial herbicides with similar structures, phenoxy- and oxyphenoxy-propionate-type compounds, where the (R)-isomers are generally known to be the active principles.

Testis-specific prohormone convertase PC4 processes the precursor of pituitary adenylate cyclase-activating polypeptide (PACAP).

The physiological substrate for proprotein convertase (PC) 4, which is expressed only in the testis, has remained unknown. Pituitary adenylate cyclase activating polypeptide (PACAP), originally isolated from the hypothalamus, exists as two amidated forms with 38 (PACAP38) and 27 (PACAP27) residues. PACAP-like immunoreactivity (PACAP-li) is found not only in the brain, but also in the peripheral tissues, and is especially abundant in the testis. Immunohistochemistry of the rat testis demonstrated strong PACAP-li in spermatids in the cap and acrosome phases. The nearly simultaneous expression of PC4 transcripts and PACAP-li in spermatids during spermatogenesis led to the hypothesis that PACAP precursor is processed by PC4. To investigate this possibility, rat pituitary GH4C1 cells were stably transfected with human PACAP cDNA, and some of these cells were co-transfected with mouse PC4 cDNA. The acid extracts of the cells were fractionated by reversed-phase HPLC. Each fraction was examined for PACAP-li using three antisera which recognize PACAP precursor, PACAP38 and/or PACAP27. Negligible PACAP-li that eluted with synthetic PACAP38 or PACAP27 was detected from cells transfected with PACAP cDNA; however, PC4 co-transfected cells showed marked PACAP-li peaks with the retention times for both PACAP38 and PACAP27. Moreover, Western blot analysis revealed immunostained bands, corresponding to the Mr for PACAP38 and PACAP27, in the PC4 co-transfected cells. Bioactivity, as indicated by stimulation of cAMP production in pituitary cell cultures, was found only in the extracts of PC4 co-transfected cells. These results provide evidence that PACAP precursor in the testis is a substrate for PC4. The processing of PACAP precursor by PC4 at a critical time in spermatogenesis suggests an important regulatory role of PC4 and PACAP in the maturation of germ cells in the testis.

Effects of Batatasin III and Its Analogs on Gibberellic Acid-Dependent α-Amylase Induction in Embryoless Barley Seeds and on Cress Growth.

The effects of batatasin III and its analogs on gibberellic acid (GA3)-dependent α-amylase induction in embryoless barley seeds and on cress root-growth were examined. Batatasin III was most effective and caused 68% inhibition of α-amylase induction at 4×10(-4) M, but its potency was low compared with that of abscisic acid. In the cress test, p-hydroxybibenzyl had high activity.

Cushing's syndrome due to bilateral adrenocortical adenomas with different pathological features.

A 48-year-old woman with Cushing's syndrome due to bilateral adrenocortical adenomas is reported. The patient presented with a typical Cushingoid appearance. The serum cortisol level was elevated with loss of the diurnal rhythm and the plasma adrenocorticotropic hormone (ACTH) level was undetectable. Dynamic testing showed no suppression of urinary 17-OHCS by high-dose dexamethasone and no stimulation by metyrapone. An abdominal computed tomography (CT) scan showed bilateral adrenal tumors. Bilateral adrenalectomy was performed. The right adrenal gland contained a tumor that was encapsulated and consisted mainly of compact cells. The surrounding cortex was atrophic. The left adrenal gland contained an encapsulated tumor composed predominantly of clear cells. There were numerous small adrenocortical nodules in the surrounding cortex. Immunohistochemical analysis of steroidogenic enzymes (P450scc, 3beta-HSD, P450c21, P450c17 and P450c11) was performed. Immunoreactivity of all the enzymes was intense in the compact cells of the right adrenocortical adenoma, while the adjacent non-neoplastic cortex was negative for the enzymes. In the left adrenal tumor, the immunoreactivity of 3beta-HSD was intense, while that of P450c17 was weak. In the adrenocortical nodules, 3beta-HSD activity was sporadically observed. G protein genes encoding Gs alpha and Gi2 were examined for activating mutations at codons 201 and 227 (Gs alpha) and codons 179 and 205 (Gi2 alpha) in the bilateral adrenal tumors, but no mutations were found. The bilateral adenomas of this patient showed marked differences in microscopic and immunohistochemical studies, suggesting that the capacity of steroidogenesis differs between the right and left tumors.

Response of respiratory epithelium of BN and F344 rats to formaldehyde inhalation.

BN rats are well-known for their high capacity for IgE production and hyperresponsiveness to exposure to allergens or other chemicals. We examined the histological changes in the nasal cavity, trachea and lungs of BN and F344 rats after the inhalation of aerosol formaldehyde (HCHO), which exerts direct toxic effects on the respiratory system. The incidence of clinical signs such as sneezing and abnormal respiration in HCHO-treated F344 rats was higher than that in HCHO-treated BN rats. The mean body weight of HCHO-treated F344 rats apparently decreased in comparison with control F344 rats, but that of HCHO-treated BN rats was not significantly different from that of control BN rats. Changes such as squamous metaplasia, stratification, degeneration and desquamation were observed by light microscopy in nasal, tracheal and bronchial mucosa in the lungs of the HCHO-treated F344 rats. In the HCHO-treated BN rats, similar but milder lesions were restricted to the nasal mucosa. Scanning electron microscopic observation supported these light microscopic observations. These results suggest that BN rats have lower sensitivity to HCHO inhalation than F344 rats.

Localization and gene expression of the receptor for pituitary adenylate cyclase-activating polypeptide in the rat brain.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a recently identified member of the secretin/vasoactive intestinal polypeptide (VIP) family. There are at least two types of receptor for PACAP: type I (PACAPR), which specifically binds PACAP; and type II (VIP/PACAPR), which binds both PACAP and VIP. The localization of PACAPR in the rat brain was determined by in situ hybridization and immunocytochemistry. We raised antisera against a synthetic peptide that corresponds to the carboxy-terminal cytoplasmic domain which is found in all subtypes of PACAPR in order to localize PACAPR-like immunoreactivity (PACAPR-LI) in the rat brain. In general, the distribution of PACAPR-LI correlated well with the distribution of PACAPR transcripts. Particularly strong PACAPR mRNA expression was detected in the olfactory bulb, hippocampus, cerebellum and hypothalamus and moderate labeling was detected in other scattered regions. At the cellular level, PACAPR-LI appeared to be concentrated predominantly in neuronal perikarya and dendrites. At the ultrastructural level, strong immunostaining for the PACAPR was found in plasma membranes, rough endoplasmic reticulum, cytoplasmic matrix, and at synapses. This study provides the basis for a better understanding of the functions of PACAP in the rat brain.

Ontogeny of pituitary adenylate cyclase activating polypeptide and its receptor mRNA in the mouse brain.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide which was first isolated from ovine hypothalamic tissue by screening for pituitary adenylate cyclase stimulating activity. Our previous data showed that radioimmunoassayable PACAP and PACAP-binding sites were detected in the whole rat brain as early as embryonic day 14(E14). In order to understand more precisely the developmental pattern of the synthesis of PACAP and its receptors in the brain, we studied the expression of PACAP and its receptor genes in the prenatal and postnatal mouse brain using RNase protection assay. The mRNAs for both PACAP and its receptor were detected as early as 9.5 days of gestation (E9.5) in the whole head of mouse embryos. The levels of PACAP mRNA in the brain increased during the prenatal period peaking at postnatal day 0 (P0). On the other hand, the levels of PACAP receptor mRNA gradually increased after E9.5. The levels sharply increased at P6 (479.0 +/- 82.5% of P0 levels), and then fell to the P3 levels at P10. These data together with our previous study on the ontogeny of PACAP immunoreactivity and its binding sites in the rat brain support the view that PACAP plays an important regulatory role in the development of brain.

Optical Analysis of Reduction Products of 2-Methylcyclohexanone by Aspergillus repens MA0197.

The products of reduction of 2-methylcyclohexanone by Aspergillus repens MA0197 were analyzed quantitatively by GC after conversion to corresponding diastereoisomeric ( -)-menthyl carbonate derivatives. Although the contents varied considerably with time, the predominant production of S-alcohol was observed, that is, Prelog's rule was obeyed.

Oxidation of acetophenone by Aspergillus species and their possible contribution to Katsuobushi flavor.

Acetophenone was converted to phenol by A. glaucus MA0200. Production of phenol, which has a pungent flavor, seemed to give a contrary effect on the creation of flavor of molded Katsuobushi. Production of phenol is the process of degradation of acetophenone, also with a pungent flavor. It would play a role in the decreasing of the pungent flavor of Katsuobushi.

Effect of hypophysectomy on pituitary adenylate cyclase-activating polypeptide gene expression in the rat testis.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a member of the secretin/glucagon/vasoactive intestinal peptide (VIP) family. Our immunohistochemical and in situ hybridization histochemical studies indicated that PACAP-like immunoreactivity (PACAP-LI) and its mRNA were present in the germ cells in the rat testis. Because the testicular function is regulated by the pituitary gonadotropins, effect of hypophysectomy on the PACAP gene expression was investigated in the rat testis as an attempt to reveal the regulation of the testicular PACAP by the pituitary. The levels of testicular PACAP mRNA, which were determined by RNase protection assay, increased 2 weeks after hypophysectomy. In contrast, the levels of radioimmunoassayable PACAP decreased 2 weeks after the surgery. Immunohistochemistry showed that hypophysectomy did not change the distribution of PACAP-LI, although the number of immunopositive cells was markedly reduced after hypophysectomy. The replacement treatments of hypophysectomized animals with FSH or LH+FSH restored testicular PACAP mRNA to the levels in the control animals. On the other hand, all of these treatments (testosterone, LH, FSH, or LH+FSH) significantly increased radioimmunoassayable PACAP in the hypophysectomized rat testis. The results suggest that both testicular PACAP and its mRNA expression are regulated by the hypothalamic-pituitary-gonadal activity, and that FSH may play a major role in this regulation.

Effect of hypophysectomy on pituitary adenylate cyclase activating polypeptide gene expression in the rat hypothalamus.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a new member of the secretin/glucagon/vasoactive intestinal peptide (VIP) family. It stimulates adenylate cyclase in cultured rat pituitary cells, which have PACAP-specific receptors and expression of pituitary hormones. Therefore, PACAP is considered as a hypophysiotropic hormone. If so, there might be a feedback regulatory mechanism between pituitary hormones and hypothalamic PACAP. In the present study, we used nuclear run-on and RNase protection assays to examine whether transcription of the PACAP gene in the rat hypothalamus would change after hypophysectomy. PACAP levels in the hypothalamus were also determined by radioimmunoassay. The transcriptional rate of the PACAP gene and PACAP mRNA content decreased 1 and 2 weeks after hypophysectomy. Radioimmunoassayable PACAP levels in the hypothalamus also decreased after hypophysectomy. These findings suggest that the reduced rate of PACAP gene transcription after hypophysectomy causes the decreased mRNA and peptide levels in the hypothalamus. Replacement with GH, PRL, T4, corticosterone, and testosterone significantly restored PACAP mRNA levels in hypophysectomized rats to those in control animals. The results suggest that feedback regulation takes place between pituitary hormones or pituitary-dependent factors and hypothalamic PACAP.