Neuron-specific enolase (NSE)-like and neurofilament protein (NFP)-like immunoreactivities in the rat dorsal root ganglia and sciatic nerve

The presence of neuron-specific enolase (NSF) and neurofilament proteins (NFP) immunoreactivities (IR) was investigated in dorsal root ganglia (DRG) of adult rats at cervical, thoracic, lumbar and sacral levels. All neurons display NSE-like IR with a variable intensity of immunostain which is not related to the neuronal size. Conversely, the antibody against all three proteic subunits of NFP no labelled the primary sensory neurons, whereas the intraganglionic axons and dorsal root of spinal nerves result positives. In the sciatic nerve the immunoreactivity was similar for NSE- and NFP-like IR. No regional differences were found among the different levels of DRG for NSE-like IR. The present results demonstrate heterogeneity in the neurons of the rat. DRG for NSE-like IR, and differences between sensory neurons and fibers in the distribution of NFP-like IR.     

Nucleotide regulation of vasoactive intestinal peptide binding to bovine thyroid plasma membranes

The specific binding of vasoactive intestinal peptide (VIP) to bovine thyroid plasma membranes is inhibited by guanine nucleotides. Guanosine 5-triphosphate (GTP) and the non-hydrolyzable GTP analogs guanosine 5-,-imidotriphosphate (Gpp(NH)p) and guanosine 5-O-(3-thiotriphosphate) (GTP--S) inhibited markedly the binding of VIP to its receptors. This inhibition was higher with GTP than with Gpp(NH)p and GTP--S and was due to an increase of the rate of dissociation of peptide bound to membranes. Other nucleotides did not show any effect.     

Somatostatin binding to dissociated cells from rat cerebral cortex

A method has been developed for the study of somatostatin (SS) binding to dissociated cells from rat cerebral cortex. Binding of [¹²⁵I][Tyr¹¹]SS to cells obtained by mechanical dissociation of rat cerebral cortex was dependent on time and temperature, saturable, reversible and highly specific. Under conditions of equilibrium, i.e., 60 min at 25°C, native SS inhibited tracer binding in a dose-dependent manner. The Scatchard analysis of binding data was linear and yielded a dissociation constant of 0.60±0.08 nM with a maximal binding capacity of 160±16 fmol/mg protein. The binding of [¹²⁵I][Tyr¹¹]SS was specific as shown in experiments on tracer displacement by the native peptide, SS analogues, and unrelated peptides.     

Degree of correspondence between contractile and oxidative capacities in horse muscle fibres: a histochemical study

Samples taken from the middle gluteal muscle of 95 untrained adult horses of different ages and sex were subjected to histochemical analysis using the myosin adenosine triphosphatase (m-ATPase) and nicotinamide adenine dinucleotide tetrazolium reductase (NADH-TR) staining techniques. Fibres were classified into types I, IIA and IIB according to m-ATPase activity after preincubation at pH 4.4. The percentage of FT (Fast-Twitch Glycolytic) fibres and the proportion of IIB fibres with "high" and "low" oxidative capacity were determined in serial sections stained for NADH-TR. Statistical analysis revealed a significantly higher proportion of IIB fibres than FT fibres (P less than 0.001), though both percentages were correlated. Thus, 72.2 +/- 17.6% of type IIB fibres showed low oxidative capacity, but the remaining 27.8 +/- 17.6% showed high aerobic potential, and thus did not correspond to FT fibres. These results confirm that the contractile capacity of a muscle fibre does not determine its oxidative profile. The different types of muscle fibre should thus be classified solely according to m-ATPase activity, since this characteristic is related to the molecular structure of contractile proteins. Oxidative capacity should be assessed separately, and not be used as a criterion for fibre classification in horses.     

DNA Segments Sensitive to Single-Strand-Specific Nucleases Are Present in Chromatin of Mitotic Cells

It was observed before that DNAin situin chromatin of mitotic cells is more sensitive to denaturation than DNA in chromatin of interphase cells. DNA sensitivity to denaturation, in these studies, was analyzed by exposing cells to heat or acid and using acridine orange (AO), the metachromatic fluorochrome which can differentially stain double-stranded (ds) vs single-stranded (ss) nucleic acids, as a marker of the degree of DNA denaturation. However, without prior cell treatment with heat or acid no presence of single-stranded DNA in either mitotic or interphase cells was detected by this assay. In the present experiments we demonstrate that DNAin situin mitotic cells, without any prior treatment that can induce DNA denaturation, is sensitive to ss-specific S1 and mung bean nucleases. Incubation of permeabilized human T cell leukemic MOLT-4, promyelocytic HL-60, histiomonocytic lymphoma U937 cells, or normal PHA-stimulated lymphocytes with S1 or mung bean nucleases generated extensive DNA breakage in mitotic cells. DNA strand breaks were detected using fluorochrome-labeled triphosphonucleotides in the reaction catalyzed by exogenous terminal deoxynucleotidyl transferase. Under identical conditions of the cells’ exposure to ss-specific nucleases, DNA breakage in interphase cells was of an order of magnitude less extensive compared to mitotic cells. The data indicate that segments of DNA in mitotic chromosomes, in contrast to interphase cells, may be in a conformation which is sensitive to ss nucleases. This may be a reflection of the differences in the torsional stress of DNA loops between interphase and mitotic chromatin. Namely, greater stress in mitotic loops may lead to formation of the hairpin-loop structures by inverted repeats; such structures are sensitive to ss nucleases. The present method of detection of such segments appears to be more sensitive than the use of AO. The identification of mitotic cells based on sensitivity of their DNA to ss nucleases provides an additional method for their quantification by flow cytometry.     

Heterotrophic bacteria, activity and bacterial diversity in two coastal lagoons as detected by culture and 16S rRNA genes PCR amplification and partial sequencing

Activity and numbers of heterotrophic bacteria have indicated that, as expected, Prevost Lagoon is more eutrophic than Arcachon Bay. Amplification and sequence analysis of the 16S rRNA genes from DNA samples extracted directly from the environment allow the determination of phylogenetic relationships among members of microbial communities in natural ecosystems without the need for cultivation. Analysis of partial 16S rRNA gene sequences obtained from Stations A and 11 revealed that, in both environments, a relatively large number of clones related to Cytophaga/FlexibacterBacteroides as well as to -Proteobacteria were found. One hundred percent similarity with the sequences of the data bases were not found for any of the more than a hundred clones studied. In fact for most clones maximum similarity was below 95% for the nucleotide series sequenced. Similarity was not higher with any of the sequences found for the 14 isolates (pure cultures) obtained from the same samples. Redundancy, i.e. number of identical sequences, was higher in the samples from Arcachon. In addition, sequences related to representatives of ten major phylogenetic branches of Eubacteria were obtained from Prevost Lagoon, however only five branches were represented by the data from Arcachon. These findings indicate a higher bacterial diversity in Prévost Lagoon.     

Fusion and histocompatibility in Rhodophyta

Fully developed thalli of Chondrus crispus, Gracilaria chilensis, Gymnogongrus furcellatus and Mazzaella laminarioides were used to assess tissue compatibility. The effect of thallus polarity on grafting and regeneration was also evaluated. Fusion did occur between fragments of the same life history phase in C. crispus, G. chilensis, G. furcellatus and M. laminarioides. Fusion between sporophytic and gametophytic tissue occurred in C. crispus, G. chilensis and M. laminarioides. Intergeneric fusion was observed between C. crispus and M. laminarioides, but not between G. chilensis and G. furcellatus.Outer cell wall, cortex and medulla were continuous at the contact face in compatible combinations. Medullary cells in the attached fragments were thinner and longer than normal cells, forming an interwoven scar plate. Thallus polarity did not modify fusion and regeneration.