A serotonergic system in the brain of the Antarctic fish, Trematomus bernacchii

The immunohistochemical distribution of serotonin-containing nerve fibres and cells has been described in the brain of the Antarctic fish, Trematomus bernacchii. The largest serotonergic system was associated with the diencephalic and rhombencephalic ventricles. In particular, serotonin-positive cells have been found in the lateral recess and neuropile zone of the diencephalic ventricle, where we have identified the serotonergic portion of the paraventricular organ. Numerous serotonin cells were localized in the dorsal nucleus of the raphe, the dorsal tegmental nucleus and the central gray. Two large cell groups, arranged in a pair of well-defined columns and connecting the central gray with the dorsal reticular formation, were immunostained in the region of the trigeminal nuclei. In addition, few positive cells have been found in the preoptic area and the cerebellar valvula, and few serotonergic nerve fibres, probably belonging to the lateral lemniscus, have been identified. The distribution of serotonin elements in the brain of T. bernacchii has been compared with that described in other fish, where it showed some modifications in the immunoreactive pattern. Finally, the lack of a serotonergic system at the level of the reticular superior formation has been reported; however, it was not possible to rule out a phylogenetic or environmental explanation.     

Human papillomavirus type 48.

The cloning and partial characterization of the genome of human papillomavirus (HPV) type 48 is presented. Hybridization and short DNA sequence analyses permitted the alignment of the genome to the HPV genetic map.     

Studies on the (pro) insulin biosynthesis of islets of Langerhans of sand rats (Psammomys obesus).

By feeding a regular laboratory chow, sand rats (Psammomys obesus) from our breeding colony gained different body weights, though they received approximately the same quantity of calories. Sand rats, reaching a body weight above 160 g (group B) showed significantly increased blood glucose values in contrast to the animals with a body weight under 160 g (group A). Isolated pancreatic islets of these two groups of sand rats were incubated with [3H]-leucine to study the incorporation of this amino acid into proinsulin and insulin. The incorporation into proteins of pancreatic islets of sand rats of group B was stimulated by 0.45 mg and 3.0 mg/ml glucose. In group A there was no further stimulation from 0.45 mg to 3.0 mg/ml glucose. Insulin secretion could be stimulated by glucose in both groups, but the stimulation was stronger in group B than in group A.     

Charged residues are major determinants of the transmembrane orientation of a signal-anchor sequence

Uncleaved signal-anchor sequences of membrane proteins inserted into the endoplasmic reticulum initiate the translocation of either the amino-terminal or the carboxyl-terminal polypeptide segment across the bilayer. Which topology is acquired is not determined by the apolar segment of the signal but rather by the hydrophilic sequences flanking it. To study the role of charged residues in determining the membrane topology, the insertion of mutants of the asialoglycoprotein receptor H1, a single-spanning protein with a cytoplasmic amino terminus, was analyzed in transfected COS-7 cells. When the charged amino acids flanking the hydrophobic signal were mutated to residues of opposite charge, half the polypeptides inserted with the inverted orientation. When, in addition, the amino-terminal domain of the mutant protein was truncated, approximately 90% of the polypeptides acquired the inverted topology. The transmembrane orientation appears to be primarily determined by the charges flanking the signal sequence but is modulated by the domains to be translocated.     

New markers for the neurofibromatosis-2 region generated by microdissection of chromosome 22

To identify new DNA markers around the neurofibromatosis-2 gene on human chromosome 22, the critical region (22q12-q13.1) was microdissected and microcloned from GTG-banded metaphase chromosomes. Eighteen thousand recombinant clones were obtained. Twenty-seven of 55 clones tested (50%) detected single-copy DNA sequences. Nine of nine clones analyzed in detail were found to map to chromosome 22. Interestingly one clone (EAN04) is part of the leukemia inhibitory factor gene which has previously been mapped to 22q11.2-q13.1. Four clones (EAN01, EAN47, EAN57, and EAN68) detect DNA polymorphisms. These probes were used to compare constitutional and tumor genotypes of 41 patients with acoustic neurinoma. Loss of constitutional heterozygosity was identified in 17 of 31 informative cases (55%). From our data we conclude that the microdissection library is a valuable resource for physical and genetic mapping studies in neurofibromatosis-2.     

Electronmicroscopy studies on the exocrine pancreas of Wistar-Rats following treatment with Streptozotocin.

In Wistar rats the intraveneous injection of streptozotocin (65 mg/kg body weight) caused a permanent hyperglycemia. After 5 days there were lesions in the exocrine parenchyme of the pancreas and its nerve fibers. Pathological changes were found in cytoplasm, cell membrane, nucleus and all other cell organelles, too. The zymogen granules remaining after extensive degranulation may disintegrate in two different ways: 1. Shrinking of the granules and formation of a hale between granule membrane and core, the electronic density of which is decreased; indistinct demonstrability of the granule membrane and finally its decomposition. 2. Shrinking of the granules, decrease of the electron density and either homogeneous or mainly peripheral arrangement of the disintegrated material of the granules; irregular shape of the granules and splitting of their membranes.     

Tetanus toxin selectively impairs anti-tumoral but not anti-microbial macrophage-mediated effector functions

Abstract The present study was designed to establish the susceptibility of macrophage-mediated effector functions to tetanus toxin (TT). Using the murine macrophage cell line, GG2EE, generated in vitro by v- raf /v- myc oncogenes, we have previously provided evidence that TT selectively inhibits interferon gamma (IFN-γ), but not basal, lysozyme activity. Here we show that while neither phagocytic nor candidacidal activities are affected by TT treatment, antitumoral activity is significantly impaired after exposure to TT. This phenomenon, which is dose-dependent, is fully ascribed to the holotoxin, as heat inactivated TT, C or A-B fragments result ineffective. Furthermore, C but not A-B fragment competes with TT in abrogating its inhibitory effects. Overall, these data indicate that TT is not a broad-spectrum, down-regulating signal on macrophage-mediated functions, thus implying that its toxic action is exerted on specific molecular targets.     

Heterologous production and characterisation of two distinct dihaem-containing membrane integral cytochrome b561 enzymes from Arabidopsis thaliana in Pichia pastoris and Escherichia coli cells

Cytochrome (cyt) b₅₆₁ proteins are dihaem-containing membrane proteins, belonging to the CYBASC (cytochrome-b₅₆₁-ascorbate-reducible) family, and are proposed to be involved in ascorbate recycling and/or the facilitation of iron absorption. Here, we present the heterologous production of two cyt b₅₆₁ paralogs from Arabidopsis thaliana (Acytb₅₆₁-A, Acytb₅₆₁-B) in Escherichia coli and Pichia pastoris, their purification, and initial characterisation. Spectra indicated that Acytb₅₆₁-A resembles the best characterised member of the CYBASC family, the cytochrome b₅₆₁ from adrenomedullary chromaffin vesicles, and that Acytb₅₆₁-B is atypical compared to other CYBASC proteins. Haem oxidation–reduction midpoint potential (E_M) values were found to be fully consistent with ascorbate oxidation activities and Fe^3 +-chelates reductase activities. The ascorbate dependent reduction and protein stability of both paralogs were found to be sensitive to alkaline pH values as reported for the cytochrome b₅₆₁ from chromaffin vesicles. For both paralogs, ascorbate-dependent reduction was inhibited and the low-potential haem E_M values were affected significantly by incubation with diethyl pyrocarbonate (DEPC) in the absence of ascorbate. Modification with DEPC in the presence of ascorbate left the haem E_M values unaltered compared to the unmodified proteins. However, ascorbate reduction was inhibited. We concluded that the ascorbate-binding site is located near the low-potential haem with the Fe^3 +-chelates reduction-site close to the high-potential haem. Furthermore, inhibition of ascorbate oxidation by DEPC treatment occurs not only by lowering the haem E_M values but also by an additional modification affecting ascorbate binding and/or electron transfer. Analytical gel filtration experiments suggest that both cyt b₅₆₁ paralogs exist as homodimers.