Localization of substance P and leucine enkephalin in the nerve terminals of the guinea pig paracervical ganglion

Summary Immunoreactivities (IR) of substance P and leucine enkephalin have been demonstrated in the guinea-pig paracervical ganglion by an immunogold electron microscope method. Both substance P-IR and leucine enkephalin-IR were detected in large synaptic vesicles with electron-dense cores. The former neuropeptide was detected in nerve terminals and varicosities comprised mainly of large vesicles with electron-dense cores; the latter was detected in nerve terminals and varicosities that also included small, clear synaptic vesicles. In a minority of nerve terminals and varicosities coexistence of both immunoreactivities could be demonstrated within vesicles with an electron-dense core. Also present in these nerve terminals and varicosities were small, clear synaptic vesicles, though these were unreactive.     

Protease inhibitor localization in control and streptozotocin-diabetic skeletal muscles

Summary ₁-Antitrypsin and ₁-inhibitor-3 were localized for the first time inside skeletal muscle cells. Their content, especially that of ₁-inhibitor-3, was greatly reduced following streptozotocin-induced diabetes. ₁-Antitrypsin and ₁-inhibitor-3 were also observed in the vascular components and interstitial space surrounding both control and diabetic soleus muscles as revealed by immunofluorescence. In diabetic muscles, the non-myofibre locale of ₁-inhibitor-3 was reduced, and to a lesser extent, ₁-antitrypsin. Both myofibre and extracellular patterns were reversed to control levels by insulin replacement.     

Analysis of murine coronavirus surface glycoprotein functions by using monoclonal antibodies.

The murine coronavirus surface glycoprotein gene was expressed as a fusion protein in bacteria, and the expressed protein was used to generate S protein-specific monoclonal antibodies (MAbs). Three of the MAbs, 11F, 30B, and 10G, were able to neutralize virus infectivity, and two of them, 11F and 10G, were able to block virus-induced, cell-to-cell fusion. The binding sites of the 11F, 30B, and 10G MAbs were determined by Western immunoblotting and epitope mapping. The 11F and 30B MAbs bound to sites located, respectively, between amino acids 33 to 40 and 395 to 406 in the amino-terminal (S1) subunit of the S protein, and the 10G MAb bound to a site located between amino acids 1123 and 1137 in the carboxy-terminal (S2) subunit. These data define more precisely the interactions between the S1 and S2 subunits of the murine coronavirus S protein and provide further insights into its structure and function.     

Increased autophagy in chloroquine-treated tonic and phasic muscles: An alternative view

Ultrastructural and cytochemical techniques were used to investigate autophagy in the tonic anterior (ALD) and phasic posterior (PLD) latissimus dorsi muscles of the chicken following chloroquine administration. Autophagic vacuoles were seen in the ALD after 1 day of chloroquine administration while no change was seen in the PLD until 3 days. In both muscles, autophagic vacuoles and myeloid bodies were found at the level of the I band. Myeloid bodies usually were found in the longitudinal rows of mitochondria in the ALD muscle. Some, but not all, of the autophagic vacuoles and myeloid bodies were cytochemically acid phosphatase positive, while the portion of the sarcoplasmic reticulum of both muscles which is normally acid phosphatase positive was devoid of activity following chloroquine administration. These observations are discussed in regard to accepted mechanisms of autophagy and the possible inhibition of autophagy in skeletal muscle tissue by chloroquine.     

Regulation of intestinal phosphate transport. II. Metabolic acidosis stimulates Na(+)-dependent phosphate absorption and expression of the Na(+)-P(i) cotransporter NaPi-IIb in small intestine

During metabolic acidosis, P(i) serves as an important buffer to remove protons from the body. P(i) is released from bone together with carbonate buffering protons in blood. In addition, in the kidney, the fractional excretion of phosphate is increased allowing for the excretion of more acid equivalents in urine. The role of intestinal P(i) absorption in providing P(i) to buffer protons and compensating for loss from bone during metabolic acidosis has not been clarified yet. Inducing metabolic acidosis (NH(4)Cl in drinking water) for 2 or 7 days in mice increased urinary fractional P(i) excretion twofold, whereas serum P(i) levels were not altered. Na(+)-dependent P(i) transport in the small intestine, however, was stimulated from 1.89 +/- 3.22 to 40.72 +/- 11.98 pmol/mg protein (2 days of NH(4)Cl) in brush-border membrane vesicles prepared from total small intestine. Similarly, the protein abundance of the Na(+)-dependent phosphate cotransporter NaPi-IIb in the brush-border membrane was increased 5.3-fold, whereas mRNA levels remained stable. According to immunohistochemistry and real-time PCR NaPi-IIb expression was found to be mainly confined to the ileum in the small intestine, and this distribution was not altered during metabolic acidosis. These results suggest that the stimulation of intestinal P(i) absorption during metabolic acidosis may contribute to the buffering of acid equivalents by providing phosphate and may also help to prevent excessive liberation of phosphate from bone.     

Development of Panel of Monoclonal Antibodies Specific to Urochordate Cell Surface Antigens

Monoclonal antibodies are an important tool in the study of botryllid ascidians’ immunology and developmental biology. Here we describe the development of a panel of 38 monoclonal antibodies that are specific to Botryllus schlosseri (Ascidiacea; subfamily Botryllinae) cell surface antigens. Many of these hybridomas recognize (by enzyme-linked immunosorbent assay and immunohistochemistry) epitopes of Botrylloides subpopulations (SP) II and III from the Mediterranean coast of Israel and show, on blood cell smear assays, reactions with subsets of Botryllus circulating blood cells. Fluorescence-activated cell sorting analyses using antibodies positive for botryllid tissues revealed up to 3.6% positive cells. ELISA screenings were performed with 64 new monoclonal antibodies on 5 different individual botryllid ascidian colonies (B. schlosseri, Botrylloides). The positive antibodies in this panel identified a large number of different antigenic determinants, some of which distinguish Botryllus versus Botrylloides colonies, and other, different colonies within these two species, or different cell types within tissues, embryos, and buds of individual colonies. Only 21 monoclonal antibodies tested positive with all colonies. Cross-reactivity with at least one Botrylloides colony was recorded in 49 hybridomas that identified Botryllus cells. This wide panel of monoclonal antibodies is the first such detailed set of monoclonals available for studies on botryllid ascidians.     

The reaction of copper-2,9-dimethyl-1,10-phenanthroline with hydrogen peroxide

The copper complex of 2,9-dimethyl-1,10-phenanthroline(2,9-dmp) is accumulated by a variety of organisms and is highly toxic. Bioaccumulation depends on reduction of copper(II) to (I), since only the copper(I)-2,9-dmp complex is lipophilic. In the case of the marine diatom, Nitzschia closterium, it is proposed that hydrogen peroxide, produced by the algae during photosynthesis, is the in vivo reductant. Hydrogen peroxide rapidly reduces copper(II)-2,9-dmp, but an excess of H₂O₂ leads to destruction of the yellow copper(I) complex. Rate constants for the formation and degradation of the yellow complex are reported. Oxygen, light, and a hydroxylating agent are released during the degradation reaction. A reaction mechanism is proposed for the destruction of copper-2,9-dmp by excess H₂O₂, involving attack on the 5, 6 positions of the phenanthroline ring by hydroxyl radical, then further oxidation by singlet oxygen and H₂O₂. These in vivo degradation reactions are believed to be the cause of the extreme toxicity of the complex.     

PHOTOSYNTHETIC PIGMENTS IN FIFTY-ONE SPECIES OF MARINE DIATOMS1

The photosynthetic pigments of 51 species (71 isolates) of tropical and sub-tropical diatoms from 13 out of 22 families were examined. These were the Thalassiosiraceae, Melosiraceae, Coscinodiscaceae, Rhizosoleniaceae, Biddulphiaceae, Chaetoceraceae, Lithodesmiaceae, Eupodiscaceae, Cymatosiraceae, Diatomaceae, Naviculaceae, Nitzschiaceae and Phaeodactylinaceae. Pigments were analyzed by cellulose and polyethylene thin-layer chromatography (TLC) and reverse-phase high-performance thin-layer chromatography (HPTLC). All species contained chlorophylls a and c₂ and the carotenoids carotene, fucoxanthin, diatoxanthin and diadinoxanthin. In addition, 14 species (20 isolates) contained one or more of four minor carotenoids, which were not identified further. One species, Thalassiothrix heteromorpha, contained small amounts of a 19′-butanoyloxyfucoxanthin-like pigment, in addition to fucoxanthin. Chlorophyll c₂ was present in all the diatoms tested and occurred together with chlorophyll c₁ in 88% of them. The presence of both chlorophylls c₁ and c₂ therefore can no longer be considered a universal characteristic of the diatom class. Where chlorophyll c₁ was absent or occurred in trace amounts only (8 species, 11 isolates), it was usually replaced by a new chlorophyll c pigment designated chlorophyll c₃, recently characterized from several prymnesiophytes and one chrysophyte. Exceptions were Nitzschia closterium (CS-114), which contained only chlorophyll c₂, and Nitzschia bilobata (CS-47), which contained all three chlorophylls (c₁, c₂ and c₃) in approximately equal amounts. Five species that contained chlorophylls c₁ and c₂ also contained chlorophyll c₃ in trace quantities Quantitative pigment analyses of the 71 isolates showed that chlorophyll concentrations ranged from 0.02 μg. 10⁶ cells^?1 in the smallest diatom, Extubocellulus spinifer, to 174.4 μg. 10⁶ cells^?1 in one of the largest diatoms, Coscinodiscus sp. under the standard growth conditions used. The mean molar ratio of chlorophyll a:c in the 72 isolates was 3.33, with a range of 1.65–7.25. The close similarity between diatom and prymnesiophyte pigmentation was confirmed. Each class has three patterns of pigmentation: viz species with chlorophylls c₁ and c₂ and‘true’fucoxanthin, species with chlorophylls c₃ and c₂ and‘true’fucoxanthin, and species with chlorophylls c₃ and c₂ and fucoxanthin derivatives.     

The interpyramidal muscle of Aristotle's lantern: its myoepithelial structure and its growth (Echinodermata,Echinoida)

Summary The interpyramidal muscles of the lantern of Diadema setosum have been studied as an example of such muscles in regular echinoids. The light- and electron microscopic study proves that the interpyramidal muscle is nothing but a continuous, highly folded myoepithelium. Although it is a powerful and specialized comminator muscle its histological organization (a pseudostratified myoepithelium) is rather simple when compared with other echinoderm myoepithelia. It consists of only two cell types: 1) a single layer of well-developed myocytes and 2) monociliated adluminal cells that totally cover the myocytes and touch the basal lamina by thin basal processes. The interpyramidal muscle grows by addition of new folds to its upper region. Consecutive stages of the myoepithelial differentiation are found in each of the young folds. The origin of the cells which are necessarily added to the growing epithelium is unknown. The growth rate of the muscle is in accordance with the enlargement of the lantern ossicles. The respective data are discussed in detail.