Catecholaminergic innervation of GRF-containing neurons in the rat hypothalamus revealed by electron-microscopic cytochemistry

Summary The Catecholaminergic innervation of neurons containing growth hormone-releasing factor (GRF) was examined by use of a method which combined either 5-hydroxydopamine (5-OHDA) uptake or autoradiography after intraventricular injection of ³H-noradrenaline with immunocytochemistry for GRF in the same tissue sections at the electron-microscopic level. In the ventrolateral part of the arcuate nucleus of the rat hypothalamus a large number of immunonegative axon terminals were found to make synaptic contact with GRF-like immunoreactive (GRF-LI) cell bodies and processes. ³H-noradrenaline autoradiography or 5-OHDA-labeling combined with GRF immunocytochemistry revealed that axon terminals labeled with ³H-noradrenaline or 5-OHDA make synaptic contact with the GRF-LI nerve cell bodies and processes. These findings indicate that catecholamine-containing neurons innervate GRF neurons to regulate GRF secretion via synapses in the rat arcuate nucleus.     

Structural characterization of lipid A component of Erwinia carotovora lipopolysaccharide

The chemical structure of the lipid A component of lipopolysaccharide excreted into the liquid medium by the plant pathogenic enterobacterium Erwinia carotovora FERM P-7576 was characterized. It consists of a -1, 6-linked glucosamine disaccharide which carries ester-and amide-bound fatty acids and phosphate similar to the lipid A from other gram-negative bacteria. The lipid A preparation was not uniform in the number and composition of the fatty acids linked to the disaccharide. Four prominent lipids A were involved, they were composed of five to seven residues of fatty acid. Among them the major component was hexa-acyl lipid A, in which the hydroxyl group at position 3 and the amino group of the non-reducing glucosamine unit carry 3-dodecanoyl-oxytetradecanoyl residues. Positions 2 and 3 of the reducing glucosamine unit were substituted by 3-hydroxytetradecanoic acid. In the hepta-acyl lipid A, an additional hexadecanoic acid was linked to the hydroxyl group of the 3-hydroxytetradecanoyl residue at position 2 of the hexa-acyl lipid A. Two penta-acyl lipids A were the homologs of the hexa-acyl lipid A with decreasing acylation. Dodecanoic acid was missing from one, and 3-hydroxytetradecanoic acid from another. 3-Dodecanoyloxytetradecanoyl residue at position 3 differentiates E. carotovora lipid A from that of other gram-negative bacteria.Abbreviations LPS lipopolysaccharide - GlcN glucosamine - KDO 3-deoxy-d-manno-octulosonic acid - FAB-MS fast atom bombardment mass spectrometry - u atomic mass unit     

Distribution of Leu 7 (HNK-1) antigen in human digestive organs: an immunohistochemical study with monoclonal antibody

Summary Leu 7 immunoreactivity was demonstrated with the indirect peroxidase-labelled antibody method on frozen and paraffin-embedded tissue sections of human digestive organs. Anti-Leu 7 monoclonal antibody, which allegedly detects mononuclear cells with natural killer or killer activity, recognized lymphoid cells among intestinal epithelial cells and in the germinal centres of solitary lymphoid follicles of small and large intestine, and a few in gallbladder, liver and the lamina propria of the intestine. In addition, peripheral nerve fibres, endocrine cells in the gut and pancreas and carcinoid and islet cell tumours were also positively stained. At the ultrastructural level, Leu 7 antigen was localized on the plasma membrane of granulated lymphoid cells in the gut mucosa and on the secretory granules of intestinal endocrine cells. In normal pancreas, Leu 7 immunoreactivity was demonstrated in most cells containing pancreatic polypeptide and in many cells containing somatostatin or glicentin. Insulin-containing cells, however, lacked Leu 7 immunoreactant. These findings were obtained in both frozen sections and paraffin-embedded sections. The possible cross-reactivities of monoclonal antibodies are discussed as they raise an important caveat in immunohistochemical studies using these antibodies.     

Involvement of colchicine-sensitive cytoplasmic element in premitotic nuclear positioning ofAdiantum protonemata

Summary When the red-light grown protonema ofAdiantum capillus-veneris was transferred to the dark, the nucleus ceased its migration ca. 5 hours before cell plate formation (Mineyuki andFuruya 1980). To see whether the nucleus was held by some cytoplasmic structure during nuclear positioning, protonemata were treated with various centrifugal forces at different stages of the cell cycle. Nuclei of G₁ phase were easily displaced by centrifugation at 360×g for 15 minutes, but those of G₂ or M phase were not displaced by it, suggesting that the nuclei were held by some cytoplasmic elements in G₂ or M phase. This nuclear anchoring was not detectable in protonemata that were treated with 5mM colchicine. With this treatment, the nucleus did not stop its migration at late G₂ and moved even in prophase. And the retardation of organelle movement which was observed in cytoplasm on the lateral side of the nucleus after the cessation of premitotic nuclear migration (Mineyuki andFuruya 1984) was not observed in the presence of colchicine. Thus the nuclei appear to be held by colchicine-sensitive structure in cytoplasm between the lateral surface of the nucleus and cell wall during the premitotic nuclear positioning. Electron micrographs showing cytoplasmic microtubules were consistent with the idea.Abbreviations PPN Premitotic positioning of the nucleus - L region Cytoplasm between the lateral surface of the nucleus and cell wall (seeMineyuki et al. 1984)     

Nucleosomelike structures associated with chromosomes of the archaebacterium Halobacterium salinarium.

Chromosomes of the halophilic archaebacterium Halobacterium salinarium were examined by electron microscopy after being spread onto water. The major part of the chromosomal DNA was associated with protein particles with diameter of 9.4 nm, arranged tandemly along the DNA fibers. Thus, the primary structure of the chromosome resembles that of eucaryote chromosomes.     

W-heterochromatin of chicken; its unusual DNA components,late replication,and chromatin structure

About 65% of DNA in the chicken W chromosome has been shown to consist ofXhoI andEcoRI family repetitive sequences. These sequences showed remarkable delay in the electrophoretic mobility at low temperature on a polyacrylamide gel. Three dimensional structures of the 0.7-kbXhoI and the 1.2-kbEcoRI family repeating units were estimated to be irregular solenoids using a computer program based on wedge angles of all the 16 dinucleotide steps. Fluorescencein situ hybridization demonstrated that these two family sequences were localized in a major heterochromatic body in an interphase nucleus. Incorporation of bromodeoxyuridine into the W chromosome in the synchronous culture of MSB-1 cells occurred about 1 h later than the peak of S phase. The chromatin structure formed alongXhoI andEcoRI family sequences was suggested to be different from the total chromatin or chromatin containing the β-actin gene sequence in that the linker DNA lengths of the former were significantly longer. Fractionation of theHaeIII-digested MSB-1 nuclei yielded a chromatin fraction in whichXhoI family sequences were partially enriched. Several DNA-binding proteins showing higher affinity for theXhoI family sequence were present in this fraction.     

Some Electrical Properties of a Nuclear Membrane Examined with a Microelectrode

Electrical potential and resistance were measured with microelectrodes in in situ and isolated nuclei of gland cells of Drosophila flavorepleta. The nucleus-cytoplasm boundary was found to be rather impermeable to ion diffusion. It presents a resistance of the order of 1 Ω cm² and sustains a "resting" potential, the nucleoplasm being about 15 mv negative with respect to the cytoplasm. Both the resistance and potential appear to be associated with the nuclear membrane: the potential declines to zero and the resistance to a fraction of its original value, when the membrane is perforated experimentally.     

In vivo sequence variability of human immunodeficiency virus type 1 envelope gp120: association of V2 extension with slow disease progression.

According to the rate of depletion of CD4 cell counts, we grouped 12 cases of human immunodeficiency virus type 1 (HIV-1) infection as 6 rapid (21.0 to 33.8 cells per microl per month) and 6 slow (0.9 to 7.9 cells per microl per month) progressors and determined the individual viral quasispecies patterns by sequencing the genome region encoding the V1, V2, and V3 loops of envelope protein. Although the quasispecies structures varied widely from one individual to another, a strong correlation was observed between a low rate of disease progression and a high degree of genetic diversity of HIV-1. Furthermore, the V2 loop extension was observed specifically in individuals with slow or no disease progression, whereas basic amino acid substitutions in V3 characteristic of a viral phenotype shift from non-syncytium inducing to syncytium inducing were observed in patients with advanced stages of disease regardless of their rate of disease progression. Studies with recombinant viruses suggested that elongation of V2 potentially restricts the capacity of HIV-1 to replicate in macrophages. Thus, our results suggest the association of distinct sequence features of both V3 and V2 with particular patterns of disease progression. Elongation of the V2 loop may be a good predictor of slow disease progression, while basic substitutions of V3 without elongation of V2 are characteristic of rapid progression.     

Changes in Organelle Movement in the Nuclear Region during the Cell Cycle of Adiantum Protonema

Movements of organelles in the nuclear region as the cell cycleprogresses in single-celled protonemata of Adiantum capillus-veneriswere examined by digital image processing techniques and microscopyof particle movement. Organelles in the nuclear region werenot very crowded and moving directionally along the longitudinalaxis of the filamentous cell in the G₁ and S phases. They beganto gather and accumulate in the nuclear region in early G₂ phase,after which directional movement changed to undirectional Brownianmotion-like movement in late G₂ phase. Movement of organelleslocated on the lateral surface of the nucleus slowed after premitoticpositioning of nucleus and lasted until the nucleolus disappeared.Movement of organelles in the cytoplasm surrounding the nucleoplasmresumed just after the nucleolus disappeared, whereas organelleslocated in the outer regions of the apical and basal surfacesof the nucleus moved rapidly during prophase but did not moveduring metaphase, movement being resumed after chromosome separation.Thus, organelle movement in the nuclear region showed temporaland spatial change during the cell cycle. (Received August 24, 1983; Accepted December 28, 1983)