Mieotic micronuclei induced by X-rays in early spermatids of the rat

In mutagenicity studies a rapid detection of chromosomal damage in mammalian germ cells would be very valuable. Encouraged by the usefulness of the bone-marrow micronucleus test, we applied an analogous method to the assay of micronuclei induced during meiotic reduction divisions in the adult male rat by X-irradiation. The micronuclei were observed in early post-meiotic cells which were enriched using a transillumination phase-contrast microscopic method. The frequency of micronuclei was scored at various dose levels and at various time intervals.The results indicate a linear increase in frequency of micronuclei 24 h after X-irradiation with doses of 0, 10, 50, 150, 300 and 600 rad. The highest frequency of micronuclei was observed after 900 rad whereas lower frequencies were found after 1200 rad. The lowest dose giving a statistically significant increase above the control level was 50 rad.The stages of meiosis showed different sensitivities to the chromosome-breaking action of X-rays. The maximal incidence of micronuclei was found 18 h after irradiation which was considered to reflect the great radiosensitivity of diakinesis-metaphase I. The anesthetized group of control animals showed a slightly higher frequency of micronuclei than the non-anesthetized control. Potentials of the new method for mutagen testing are discussed.     

Permeability changes induced by electric impulses in vesicular membranes

Summary Electric impulses were found to cause transient permeability changes in the membranes of vesicles storing biogenic amines. Release of catecholamines induced by electric fields (of the order of 20 kV/cm and decaying exponentially with a decay time of about 150 sec) was studied, using the chromaffin granules of bovine adrenomedullary cells as a vesicular model system. Far-UV-absorption spectroscopy was applied to determine the amount of catecholamines released from suspended vesicles. A polarization mechanism is suggested for the induction of short-lived permeability changes caused by electric fields. Such transient changes in permeability may possibly represent a part of the sequence of events leading to stimulated neurohumoral secretion.     

Permeability changes induced by peroxidation in liposomes prepared from human erythrocyte lipids

The diffusion of [2-(14)C]glucose out of liposomes prepared from extracted human erythrocyte lipids was examined. Increased glucose efflux was observed when the lipids were treated with hydrogen peroxide and CuCl(2) before liposome formation, and this phenomenon required both peroxide and metal. Peroxidation of these lipids also resulted in the destruction of polyunsaturated fatty acids and the generation of conjugated dienes, but neither of these processes appeared to be the sole cause of increased glucose efflux. Thin-layer chromatography and the effects of aqueous washes suggested that surface-active lysophosphatides or other lipid degradation products were responsible for the increased permeability of the treated liposomes. It is suggested that the behavior of this liposome model system may be relevant to the high permeability and fragility of vitamin E-deficient erythrocytes.     

Electrical changes in isolated rat jejunum induced by hypertonicity

Mucosal hypertonicity produces a drop of transepithelial potential difference in isolated rat jejunum with a half time of about 15 s. The same effect is obtained when the osmorality of both bathing solutions is raised simultaneously. Serosal hypertonicity produces an increase of transepithelial potential difference an order of magnitude lower than the drop produced by mucosal hypertonicity. The change in the short circuit current parallels the one in potential difference.When the transepithelial potential difference is varied by adding different concentrations of glucose to the bathing media, the potential drop induced by mucosal hypertonicity is linearly related to the magnitude of the transepithelial potential difference just before the increase in osmolarity.The drop of potential can be explained by a decrease of the electrical resistance of the extracellular shunt pathway due to opening of the tight junctions. The results can be accounted for in terms of an equivalent electrical circuit proposed for small intestine. Using this equivalent circuit model it is possible to obtain estimates of the values of the diffusion potential and the salt gradient across the tight junction.     

Motile components in early rat spermatids

In early rat spermatids, two distinctly different kinds of movements of cell components were detected by video-analysis. The primary flagellum, a typical 9 + 2 axonema, is capable of inducing wave-like movements in three dimensions, unlike late spermatid forms, which display motility of the now thickened flagellum only by repeated bending of its extreme part. Additionally, at the apical regions of spermatids of the same early stage, cytoplasmic protrusions executed rhythmic movements at a frequency of almost three times per second.The two kinds of motility of the different components in the same cell type are thought to be involved in normal orientation and transfer of spermatids in the tubulus seminiferus during their differentiation to sperm.     

Structural changes induced in rat leukemic basophils by immunological stimulus

We have examined morphological events in RBL-2H3 basophilic leukocyte-derived cells following stimulation to secrete with specific antigen. Following stimulation, the cell surface undergoes a rapid and pronounced ruffling concomitant with an overall flattening. Secretory granules which are located in one area before stimulation becomes rapidly dispersed throughout the cytoplasm. Since secretion has been shown to be a relatively slow process in the RBL-2H3 cell line our observations suggest that ruffling and flattening precede the secretion process.     

Sleep changes induced by lipopolysaccharide in the rat are influenced by age

In mammals, aging is associated with immune senescense. To examine whether the sleep changes occurring during immune challenge are affected by age, we assessed sleep alterations induced by the administration of lipopolysaccharide (LPS) in young and middle-aged rats. During vehicle, the middle-aged rats exhibited less pre-rapid eye movement sleep (pre-REMS) as well as REMS, due to a smaller number and shorter duration of REMS episodes, than young rats. LPS elevated body temperature, increased non-REMS, and suppressed both pre-REMS and REMS in the young as well as in the middle-aged rats. However, in the young animals, LPS significantly enhanced slow-wave activity in the electroencephalogram (EEG) within non-REMS, reflecting an increase in sleep intensity. In contrast, LPS attenuated EEG power in most frequency bands in the older animals. This finding indicates age-related changes in the modulation of sleep by LPS.     

Intracellular pH regulation in rat round spermatids

Intracellular pH has been shown to be an important physiological parameter in cell cycle control and differentiation, aspects that are central to the spermatogenic process. However, the pH regulatory mechanisms in spermatogenic cells have not been systematically explored. In this work, measuring intracellular pH (Hi) with a fluorescent probe (BCECF), membrane potential with a fluorescent lipophilic anion (bisoxonol), and net movement of acid using a pH-stat system, we have found that rat round spermatids regulate pHi by means of a V-type H⁺-ATPase, a HCO ^{3 ?} entry pathway, a Na⁺ HCO3^?dependent transport system, and a putative proton conductive pathway. Rat spermatids do not have functional base extruder transport systems. These pH regulatory characteristics seem specially designed to withstand acid challenges, and can generate sustained alkalinization upon acid exit stimulation.     

Permeability changes induced by 130 GHz pulsed radiation on cationic liposomes loaded with carbonic anhydrase

The effects of pulsed 130 GHz radiations on lipid membrane permeability were investigated by using cationic liposomes containing dipalmitoyl phosphatidylcholine (DPPC), cholesterol, and stearylamine. Carbonic anhydrase (CA) was loaded inside the liposomes and the substrate p-nitrophenyl acetate (p-NPA) added in the bulk aqueous phase. Upon permeation across the lipid bilayer, the trapped CA catalyzes the conversion of the p-NPA molecules into products. Because the self-diffusion rate of p-NPA across intact liposomes is very low the CA reaction rate, expressed as Delta A/min, is used to track membrane permeability changes. The effect of 130 GHz radiation pulse-modulated at low frequencies of 5, 7, or 10 Hz, and at time-averaged incident intensity (I(AV)) up to 17 mW/cm(2) was studied at room temperature (22 degrees C), below the phase transition temperature of DPPC liposomes. At all the tested values of I(AV) a significant enhancement of the enzyme reaction rate in CA-loaded liposomes occurred when the pulse repetition rate was 7 Hz. Typically, an increase from Delta A/min = 0.0026 +/- 0.0010 (n = 11) to Delta A/min = 0.0045 +/- 0.0013 (n = 12) (P < 0.0005) resulted at I(AV) = 7.7 mW/cm(2). The effect of 130 GHz pulse-modulated at 7 Hz was also observed on cationic liposomes formed with palmitoyloleoyl phosphatidylcholine (POPC), at room temperature (22 degrees C), above the phase transition temperature of POPC liposomes.     

Biochemical changes in rat testis induced in vitro by reactive oxygen species

We report the effects of reactive oxygen species generated by ultraviolet-A radiation on some biochemical parameters specific for oxidative stress, in rat testis homogenates. Results show an increase in lipid peroxidation products under ultraviolet-A exposure, and suggest that the involved mechanism is typical for a radical-mediated chain reaction. The amount of SH groups also increases during irradiation, probably as a consequence of conformational changes in proteins. Electrophoresis results revealed protein pattern changes mainly in the low molecular weight domain. The catalytic activities of alkaline phosphatase and gamma-glutamil transpeptidase are modified under the oxidative conditions generated by reactive oxygen species. The changes of the enzymatic activities are UVA exposure time-dependent, suggesting that conformational modifications are responsible for enzymatic activities enhancement.     

Fucosylation of glycoproteins in rat spermatocytes and spermatids

Glycoprotein synthesis in pachytene spermatocytes and round spermatids, isolated from rat testes, was studied by analysis of the incorporation of (³H)-fucose. The isolated germ cells were capable of incorporating (³H)-fucose into cell-bound, acid-precipitable components for an incubation period of at least 23 hours (at 32°C). In young spermatids, engaged in the formation of the acrosome, (³H)-fucose was incorporated into more than 16 different glycoproteins within the molecular weight range of 20.000–100,000. A qualitatively similar set of glycoproteins was found to be labeled in spermatocytes. Radioautography showed that after 4 hr most of the incorporated radioactivity was present at one pole in the perinuclear zone of spermatocytes and spermatids, which could reflect incorporation of fucose in the Golgi apparatus. The newly fucosylated glycoproteins were associated with a particulate subcellular fraction (membrane fraction). Trypsin treatment of whole cells after 25 hours of incubation with (³H)-fucose, however, did not cause significant lysis of tritiated glycoproteins. From the results it was concluded that the majority of the newly fucosylated glycoproteins in spermatocytes and spermatids remained associated with an intracellular membrane system, presumably the Golgi apparatus and the vesicles that arise from this structure, to be deposited subsequently in proacrosomic granules and the acrosome. The results also suggest that initiation of the synthesis of spermatidal glycoproteins occurs during the prophase of meiosis in spermatocytes.     

Physicochemical study of platelet aggregation by 3 polylysines]

The action of polylysines of various molecular weight on platelet behaviour is studied with the help of 3 techniques: photometric test, screen filtration pressure on PRP and electrophoretic mobility. Polylysines, which are polybasic substances, produce a platelet aggregation studied by photometry after a short period of latency. Aggregation, depending on the doses of poly-lysine and chiefly on the optic density, shows a "plateau" with the dose of 100 gamma/ml for poly-lysine L, of molecular weight 17000. The screen filtration pressure increases continuously in the presence of the polylysines studies. Finally, the platelet charge decreases. The results depend on the concentration of poly-lysine, and the optimal concentration which involves the more marked alterations is inversely proportional to the molecular weight of the polylysine studied.