Intermediate filaments in nervous tissues

Intermediate filaments have been isolated from rabbit intradural spinal nerve roots by the axonal flotation method. This method was modified to avoid exposure of axons to low ionic strength medium. The purified filaments are morphologically 75-80 percent pure. The gel electrophoretogram shows four major bands migrating at 200,000, 145,000, 68,000, and 60,000 daltons, respectively. A similar preparation from rabbit brain shows four major polypeptides with mol wt of 200,000 145,000, 68,000, and 51,000 daltons. These results indicate that the neurofilament is composed of a triplet of polypepetides with mol wt of 200,000, 145,000, and 68,000 daltons. The 51,000-dalton band that appears in brain filament preparations as the major polypeptide seems to be of glial origin. The significance of the 60,000- dalton band in the nerve root filament preparation is unclear at this time. Antibodies raised against two of the triplet proteins isolated from calf brain localize by immunofluorescence to neurons in central and peripheral nerve. On the other hand, an antibody to the 51,000-dalton polypeptide gives only glial staining in the brain, and very weak peripheral nerve staining. Prolonged exposure of axons to low ionic strength medium solubilizes almost all of the triplet polypeptides, leaving behind only the 51,000- dalton component. This would indicate that the neurofilament is soluble at low ionic strength, whereas the glial filament is not. These results indicate that neurofilaments and glial filaments are composed of different polypeptides and have different solubility characteristics.     

Limits of gas secretion by the salting-out effect in the fish swimbladder rete

Summary The high dissolved gas tensions required for the secretion of gases into deep-sea fish swimbladders are thought to be produced in the rete mirabile by a countercurrent multiplication mechanism, the capacity of which is theoretically limited by the physical characteristics of the rete and by the magnitudes of minute gas solubility changes in the blood plasma. These gas solubility changes are presumably induced through the salting-out effect following the addition of lactic acid to rete venous blood as it circulates through the gas gland. In order to estimate the maximum swimbladder gas pressures attainable by this mechanism, the effects of lactic acid on N₂ and Ar solubilities in water were determined at 5 and 25°C with a new volumetric method. The results show that the salting-out effect with lactic acid is much smaller than with NaCl, and that the agreement between predicted and observed swimbladder gas pressures is more critically dependent on the physical properties of the rete vasculature than indicated by previous theoretical treatments. When augmented by the release of hemoglobin-bound O₂, the salting-out effect with lactic acid appears large enough to account for the production of even the highest swimbladder O₂ pressures, provided the rete characteristics lie within certain reasonable limits. However, successful theoretical explanation of observed swimbladder N₂ pressures in some deep-sea species will require rigorous attention to such theoretically neglected factors as dissolved gas backdiffusion along the rete and the unequal size and number of the rete arterial and venous capillaries.     

Moderately thermophilic Myxobacteria: novel potential for the production of natural products isolation and characterization

Novel moderately thermophilic Myxobacteria from both known suborders (Cystobacterineae and Sorangineae) were isolated from soil samples of semiarid and warm climates. The addition of the anthelmintic and amoebizidal agent levamisole was a new and crucial improvement for the fast isolation of overall 37 strains, which grew very fast at temperatures of 42-44 degrees C. When the 16S rDNAs were compared with GenBank data of common Myxobacteria, identities were 98-99%, thus not reflecting the physiological differences. Similar to the Myxobacteria described so far, the new isolates are multiresistant against a variety of antibiotics and are producers of typical myxobacterial secondary metabolites. Analysis of our previous strain collection isolated from soil samples taken worldwide revealed a more or less uniform distribution of strains which synthesize specific metabolites. Therefore these moderately thermophilic Myxobacteria, which grow 2-3 times faster, have the potential to replace the slow-growing isolates and provide a means for fast and cost-saving production of myxobacterial metabolites in the future.     

Morphological characterization of a human glioma cell l ine

A human malignant continuous cell line, named NG97, was recently established in our laboratory. This cell line has been serially subcultured over 100 times in standard culture media presenting no sign of cell senescence. The NG97 cell line has a doubling time of about 24 h. Immunocytochemical analysis of glial markers demonstrated that cells are positive for glial fibrillary acidic protein (GFAP) and S-100 protein, and negative for vimentin. Under phase-contrast microscope, cultures of NG97 showed cells with variable morphological features, such as small rounded cells, fusiform cells (fibroblastic-like cells), and dendritic-like cells. However, at confluence just small rounded and fusiform cells can be observed. At scanning electron microscopy (SEM) small rounded cells showed heterogeneous microextentions, including blebs and filopodia. Dendritic-like cells were flat and presented extensive prolongations, making several contacts with small rounded cells, while fusiform cells presented their surfaces dominated by microvilli.     

Nitrogen binding to deoxyhemoglobin at high pressures and its relation to changes in hemoglobin-oxygen affinity

The stoichiometric and thermodynamic properties of nitrogen (N2) binding to human deoxyhemoglobin (Hb) at N2 saturation pressures up to 400 atm were derived from measured N2 solubilities in protein-free buffers (pH 7.1) and in corresponding buffer + Hb (6.5% w/w) solutions at 20.0, 25.0, and 37.0 degrees C. At each temperature, approximately 3 N2 molecules bind per Hb tetramer at N2 pressures of 100 atm, while about 7 N2 molecules bind per tetramer at 400 atm N2 pressure, where available binding sites are still not fully saturated. Calculated N2-Hb binding isotherms are well described by a simple binding model with 12 independent and equivalent binding sites/Hb tetramer. N2 binding at each of the sites is hydrophobic, exhibiting the typical increase of the dissociation enthalpy with temperature. Enthalpies of dissociation are slightly more negative, while corresponding unitary entropies are somewhat less negative than those for the transfer of N2 from olive oil to water. Calculated partial molar volumes of N2 in Hb are positive but less than the corresponding partial molar volumes of N2 in buffer. Results indicate that N2 binding to Hb is accompanied by only small protein conformational changes which entail slight structural destabilization and loss of free volume in the protein that partially accommodates the volume of the N2 ligand. Results are related to previously reported effects of high pressure and high-pressure N2 on HbO2 affinity, illuminating essential features of the molecular mechanisms for these effects.     

Insights into the biodiversity of the Succulent Karoo hotspot of South Africa: the population genetics of a rare and endemic halictid bee,Patellapis doleritica

Population genetic analyses are especially relevant for species considered threatened or highly endemic and for which other forms of biological information are lacking. Patellapis doleritica is a recently described communally nesting halictid bee of conservation concern because it is rare and endemic to the Succulent Karoo of South Africa. Moreover, its dispersal is considered to be restricted by its specialised nesting requirements and inclement weather conditions during its limited annual flight period, traits which may be common to other bee species of the region. We hypothesised that gene flow in P. doleritica was low, leading to marked genetic differentiation. Using 7 microsatellites, we investigated its mating and population genetic structure in 258 individuals (171 females and 87 males) from 7 populations spanning most of its known range. Deviation from Hardy–Weinberg equilibrium (F_IS = + 0.254) suggested P. doleritica to be inbred, as in many other communal nesting bee species. Global F_ST (0.028) and global G′_ST (0.216) revealed modest but significant differentiation between most populations, even across the very limited range of the species (ca. 25 km), with one genetically extreme outlier population. Despite inbreeding, we detected a surprisingly low frequency of diploid males (2 %). Patellapis doleritica nevertheless deserves special conservation attention since it is an endemic species with a low overall abundance and therefore possibly prone to environmental change and local extinction.