Effects of Hypoxia on the Activity of the Dopaminergic Neuron System in the Rat Striatum as Studied by In Vivo Brain Microdialysis

The purpose of the present study is to clarify the effects of hypoxia on the activity of the dopaminergic neurons in the brain and its mechanism of action. For this purpose, the effects of hypoxia on the extracellular levels of 3,4-dihy-droxyphenylethylamine (dopamine) were examined in the rat Striatum using in vivo brain microdialysis in the presence or absence of pretreatment with either tetrodotoxin (a blocker of voltage-dependent sodium channels) or nomifensine (a blocker of dopamine reuptake). Exposure to various degrees of hypoxia (15, 10, and 8% O₂ in N₂) increased dopamine levels in striatal dialysates to 200, 400, and 1,100%, respectively, of the control value. On reoxygenation, dopamine levels in the dialysates rapidly returned to the control level. Reexposure to hypoxia increased the dopamine levels to the same extent as during the first exposure. After addition of tetrodotoxin (40 mUM) to the perfusion fluid or pretreatment with nomifensine (100 mg/kg, i.p.), exposure to hypoxia no longer increased the dopamine levels. These results suggest that although hypoxia induces an increase in the extracellular dopamine levels (hence, an apparent increase in the activity of the dopaminergic neurons), this increase is not the result of an increase in dopamine release itself, but rather the result of inhibition of the dopamine reuptake mechanism.     

Identification of a novel S-phase-specific gene during the cell cycle in synchronous cultures of Catharanthus roseus cells.

The cell-cycle specific cDNAs were isolated from a cDNA library prepared from cells in the S phase in the synchronous cultures of Catharanthus roseus. One of the isolated genes, which we refer to as cyc07, was analyzed in detail. The full-length cDNA of cyc07 contains an open reading frame of 735 nucleotides, encoding a protein of 245 amino acids with a molecular weight of 28,356 Da. The protein predicted from the nucleotide sequence is highly basic, as are mammalian histones. cyc07 mRNA was detected specifically in cells at the S phase in synchronous cultures. The induction and accumulation of mRNA in the S phase were suppressed when DNA synthesis was inhibited by aphidicolin. In the intact plant, cyc07 mRNA was found preferentially in root tips that contained meristematic tissue. A databank search revealed that a sequence homologous to the nucleotide sequence of cyc07 cDNA is present in the downstream region of the SIR3 gene in the yeast genome. The amino acid sequence predicted from the corresponding region of the yeast genome exhibited significant homology with that of cyc07 protein. These similarities between cyc07 and the corresponding region in yeast suggest that the homologous sequence in yeast is a novel gene that is functionally homologous to cyc07. Our results presented here suggest the possibility that cyc07 may play a role in the proliferation of higher plant cells, in particular in the entry into or progression of the S phase of the cell cycle.     

A child with pituitary gigantism and precocious adrenarche: does GH and/or PRL advance the onset of adrenarche?

We describe a female child with pituitary gigantism and precocious adrenarche. From two years of age she showed unusual overgrowth, and at 5 years old she was 133.5 cm (+ 5.5 SD) tall and weighed 40.5 kg. Her precocious manifestations were public hair, acne vulgaris, hirsutism, and advanced bone age. Endocrinological examination revealed markedly increased serum growth hormone (GH) and prolactin (PRL), which responded paradoxically to a TRH test. In addition, the concentrations of serum dehydroepiandrosterone (DHA) and its sulfate (DHAS) were increased to adult levels, moving in accordance with changes in ACTH, which suggested that these androgens were secreted from the adrenal glands functionally. These androgens seemed to be responsible for her partial precocity. Prior reports have suggested that GH and/or PRL overproduction might have played a role in the induction of adrenarche. Also, in previous reports of 9 gigantism patients under 10 years old, the manifestation of precocious adrenarche was suggested in 8. Further investigation of the influence of GH and PRL on adrenal androgen production in children with pituitary gigantism is required. On the other hand, in short children with normal GH secretion, attention should be paid to whether or not the GH therapy in early childhood induces precocious adrenarche.     

Micronucleated reticulocyte induction by ethylating agents in mice.

Six model ethylating agents were tested for clastogenic potency by means of a new technique of the micronucleus assay with mouse peripheral blood cells using acridine orange (AO)-coated slides, to evaluate the test. The alkylating agents were: N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG), N-ethyl-N-nitrosourea (ENU), diethylsulfate (DES), ethyl methanesulfonate (EMS), epichlorohydrin (ECH) and ethylene dibromide (EDB). The animals were given a single intraperitoneal injection of the following doses of the chemicals: ENNG and ENU, 25, 50 and 100 mg/kg; EMS and DES, 100, 200 and 400 mg/kg body weight. For EDB and ECH, the doses were 50, 100 and 200 mg/kg, given twice, 24 h apart. Before and after the injection, blood samples were taken from the tails at 24-h intervals up to 72 h and preparations were made on AO-coated slides. For each dose group, 4 animals were used and 1000 reticulocytes were examined per slide for the presence of micronuclei. At the optimum induction time of 48 h, ENU induced micronucleated reticulocytes (MNRETs) at all 3 doses. ENNG and EMS induced MNRETs significantly at 2 dose levels each and DES only at the highest dose. ECH and EDB failed to induce MNRETs. On the basis of the dose of chemical needed to double the spontaneous frequency, the order of clastogenic potency was ENU greater than ENNG greater than EMS greater than DES. The results obtained compared favorably with those from other in vivo methods. The present technique proves to be simple, flexible and relatively sensitive. Shifts in the optimum induction peak in individual animals and by some chemicals can be picked up easily which is important when testing weak mutagens and chemicals with an unknown mechanism of action.     

Micronucleus induction in mouse peripheral reticulocytes by 7,12-dimethylbenz[a]anthracene.

Micronucleus assays using mouse peripheral blood stained vitally on acridine orange (AO)-coated slides were evaluated at two laboratories with 7,12-dimethylbenz[a]anthracene (DMBA) and compared with the standard bone marrow assay. DMBA was administered by single intraperitoneal injection to CD-1 mice at doses ranging from 5 to 80 mg/kg, then 5 microliters of peripheral blood was sampled from a tail vein at 24, 48, 72, 96, and 120 h after treatment. Similar incidences of micronucleated young erythrocytes were observed in peripheral blood reticulocytes and bone marrow polychromatic erythrocytes. The dose response of micronucleated reticulocytes was delayed compared to that of micronucleated polychromatic erythrocytes. The dose-response curves after treatment with DMBA differed depending on the sampling times, which revealed the difficulty of obtaining accurate dose-response relations in the micronucleus assay. The present result demonstrated that the simple and rapid AO supravital staining method is a valuable and easier method for obtaining dose- and time-response data for quantification of micronucleus induction by chemicals.     

Scyliorhinus tokubee sp. nov. from Izu Peninsula,southern Japan (Scyliorhinidae,Elasmobranchii)

A new catshark,Scyliorhinus tokubee sp. nov., is described based on specimens from the coast of Shirahama, eastern Izu Peninsula, southern Japan. The present species is distinguished from other congeners in having a particular coloration with dark saddles, blotches and numerous small light spots, a wide oral cleft, the anterior nasal flap not reaching the oral cleft, a short interspace between the dorsal fins, developed clasper hooks, and some meristic characters (number of vertebrae, jaw teeth, and spiral valve turns). This species has been bred under captivity for several years in Shimoda Floating Aquarium.     

Experimental stomach-ulcer on pika

Experimental gastric ulcer formation was performed in the pika and compared with that in the rat. Gastric ulcers were formed in pika that were subjected to water restraint for 4-5 days for 2 hours each day. Gastric ulcers were also formed under conditions of 1-4 days for 3 hours each day and 1-2 days for 5 hours each day. The severest (widest) ulcers were obtained under the condition of 5 hours' water restraint. Histopathologically, the ulcers were mostly erosions, but those formed under 5 hours' restraint reached the tunica muscularis mucosae. In addition, inflammatory changes were recognized. In contrast, while gastric ulcers in the rat formed within a short time, they were histopathologically less severe than those in the pika. Therefore, water restraint for 4 hours performed 4-5 times is suitable to obtain gastric ulcer formation in the pika and may result in more severe gastric ulcers than in the rat. Compared with the rat, the pika showed differences in the appearance and degree of gastric ulcers formed by the injection of serotonin and reserpine.     

Comparison of renal ornithine transcarbamylase activities within different chicken breeds

Comparisons were made of the renal ornithine transcarbamylase (OTC) activities within different groups of chickens including Japanese native breeds. OTC activities varied markedly within these groups. The Cochin Bantam breed and White Leghorn B line had an especially high activity, about 400 units/g of kidney, in contrast to two Japanese native breeds, Japanese Game (white variety) and Banshuu Gashiwa, and the California Gray breed, which showed a very low activity, the values being almost undetectable. In crossing experiments using the California Gray breed as a tester strain, Cochin Bantam OTC represents a simple autosomal incompletely dominant trait similar to the White Leghorn B line OTC. Kinetic studies using partially purified OTC preparations from the White Leghorn B line and Cochin Bantam breed revealed that both enzymes were identical for a variety of enzymic characteristics. In light of these results, the physiological significance of chick kidney OTC is discussed.     

Arylsulfatases of human-lung tumors transplanted into athymic mice. Cancer-associated modification of arylsulfatase B variant

The activities and properties of arylsulfatase A and B from human lung carcinoma transplanted into athymic mice were demonstrated. The activities of arylsulfatase A and B from transplanted carcinomas with four histological types were more than twofold higher as compared to those from surgical tumors, except for arylsulfatase A activity in blastoma. Arylsulfatase B in transplanted tumors was almost completely replaced, except for blastoma, by an anionic B variant (B1) which was a minor component of arylsulfatase B in surgical lung tumor and absent in normal human lung. The properties of arylsulfatases A and B from transplanted tumors were essentially identical, respectively, with those from normal lung or surgical tumors in respect of molecular weight, heat stability, pH optimum, isoelectric point (pI), Km, time course profile and substrate specificity. Arylsulfatase B1 showed the properties similar to B enzyme except for net charge. The cause of the negative charge of tumor B1 enzyme was investigated. By the action of phosphatase, which was added exogenously or had been persistently included in the partially purified enzyme preparation, B1 enzyme (pI 7.5) shifted to about pI 8.2. Treatment of B1 enzyme with neuraminidase, concomitant with the endogenous phosphatase, resulted in marked increase (pI 9.5) of the isoelectric point, identical to that of arylsulfatase B. Thus, it is most probable that tumor B1 enzyme is modified by additional sialic acid and phosphate bound to arylsulfate B.     

13C Nuclear magnetic resonance study of salt induced conformational change of basic polypeptides: Poly (L-lysine), poly (L-arginine), and poly (L-ornithine)

A ¹³C-nmr study of the salt-induced helix–coil transition of the basic polypeptides poly(L -lysine) [(Lys)_n], poly(L -arginine) [(Arg)_n], and poly (L -ornithine) [(Orn)_n] was performed to serve as a reference of the helical portion of histones and other proteins. As is the case with pH-induced helix–coil transition, the downfield displacement of the C^α and carbonyl carbon signals are observed in the helical state. The upfield shift of the C^β signals, on the other hand, is noted in the salt-induced transition. Regardless of the differences in the side chains and also the salts used, very similar helix-induced chemical shifts are obtained for (Lys)_n and (Arg)_n. However, the displacement of the C^α, C^β, and carbonyl carbons of (Orn)n in the presence of 4M NaClO₄ is found to be almost 50% of that of (Lys)_n and (Arg)_n. This is explained by the fact that the maximum helical content is about 50%, consistent with the ORD result. Further, the motion of the backbone and side chains of the helical from was estimated by measuring the spin-lattice relaxation time (T₁), nuclear Overhauser enhancement (NOE), and line width. In the case of (Lys)_n, the motion of the side chains is charged very little in comparison with that of the random coil. Indicating that the aggregation of the salt-induced helix is small in contrast to that of the pH-induced helix. For (Arg)_n, however, the precipitate of the helical polymers is mainly due to aggregation.