In vitro effect of vitellogenin on steroid production by ovarian follicles of greenback flounder Rhombosolea tapirina

Ovarian follicles from vitellogenic greenback flounder (Rhombosolea tapirina) were incubated in L15 medium alone, or containing human chorionic gonadotropin (hCG), dibutyryl cyclic AMP (dbcAMP) or the steroid precursors testosterone (T), 17-hydroxyprogesterone (17P) and androstenedione (A) in the presence of vitellogenin (Vtg) at 0.1-5.0mg mL(-)(1). Medium concentrations of 17beta-estradiol (E(2)) and T were measured by radioimmunoassay. HCG generally stimulated follicular E(2) but not T production, whereas 17P, A and T stimulated production of E(2), T, and E(2) respectively. Treatment of follicles with dbcAMP inhibited follicular E(2) production, but increased follicular T production at high doses. The effect of low concentrations of Vtg on follicular steroid production was variable; however, higher doses of Vtg significantly suppressed basal, hCG-, dbcAMP- and steroid precursor-stimulated follicular E(2) and T production. The results of this study show that high concentrations of Vtg may suppress follicular steroid production by interfering in the steroidogenic pathway. This suggests that Vtg may regulate its own production by limiting the ovarian production of E(2).     

HistoChoice® as an alternative to formalin fixation of undecalcified bone specimens

We compared histochemical and immunohistochemical staining as well as fluorochrome labeling in murine bone specimens that were fixed with 10% neutral buffered formalin to those fixed with HistoChoice®. We showed that sections from undecalcified tibiae fixed for 4 h in HistoChoice® resulted in enhanced toluidine blue and Von Kossa histochemical staining compared to formalin fixation. HistoChoice® produced comparable or improved staining for alkaline phosphatase. Acid phosphatase localization was better in formalin fixed specimens, but osteoclasts were visuralized more easily in HistoChoice® fixed specimens. As expected, immunohistochemical labeling was antibody dependent; some antibodies labeled better in HistoChoice® fixed specimens while others were better in formalin fixed specimens. Toluidine blue, Von Kossa, and alkaline phosphatase staining of sections fixed for 12 h produced sections that were similar to 4 h fixed sections. Fixation for 12 h preserved acid phosphatase activity better. Increasing fixation to 12 h affected immunolocalization differentially. Bone sialoprotein labeling in HistoChoice® fixed specimens was comparable to formalin fixed samples. On the other hand, after 12 h formalin fixation, osteocalcin labeling was comparable to HistoChoice®. For most histochemical applications, fixing murine bone specimens for 4 h with HistoChoice® yielded superior staining compared to formalin fixation. If immunohistochemical localization is desired, however, individual antibodies must be tested to determine which fixation process retains antigenicity better. In addition, there was no detectable difference in the intensity of fluorochrome labeling using either fixative. Finally, fixation duration did not alter the intensity of labeling.     

Histological changes in the gonad, skin, intestine and olfactory epithelium of artificially-matured male American eels, Anguilla rostrata (LeSueur)

Migratory (silver) male American eels were injected weekly for 5 weeks with human chorionic gonadotropin (hCG), salmon pituitary extract, hydrocortisone 21-hemisuccinate (cortisol), or saline to determine the effects of induced maturation on the histology of their gonads, skin, intestines and olfactory epithelia. Treatment with hCG induced full sexual maturity, salmon pituitary evoked only limited spermatogenesis, and neither hydrocortisone nor saline had any effect on the gonad. Eels injected with hCG and salmon pituitary experienced no changes in skin morphology, but the epidermal thickness of saline- and hydrocortisone-treated fish decreased. Intestinal morphology did not change in any of the treatment groups. Both the presumed sensory and non-sensory portions of the olfactory epithelium of hCG- and pituitary-treated males exhibited a decrease in thickness, structural degeneration, and non-significant reductions in mucous cell density. Some of these changes in maturing male eels are similar to those previously observed in maturing females but others are different; it appears likely that maturing Anguilla are sexually dimorphic and that these changes are adaptive and not artifacts of the hormone treatments.     

An ultrastructural study of adherence to buccal epithelial cells of Simonsiella sp.

Simonsiella , an unusual aerobic Gram-negative multicellular, filamentous bacterium present in the oral flora, possesses a marked dorsal-ventral differentiation. The morphology and ultrastructure of the structural appendages on the ventral surface were examined. Two types of fibrils could be distinguished; in vitro adherence of Simonsiella to buccal epithelial cells was by means of the short fibrils.     

Microbiological and chemical properties of soil associated with macropores at different depths in a red-duplex soil in NSW Australia

Some agricultural soils in South Eastern Australia with duplex profiles have subsoils with high bulk density, which may limit root penetration, water uptake and crop yield. In these soils, a large proportion (up to 80%) of plant roots maybe preferentially located within the macropores or in the soil within 1–10 mm of the macropores, a zone defined as the macropore sheath (MPS). The chemical and microbiological properties of MPS soil manually dissected from a 1–3 mm wide region surrounding the macropores was compared with that of adjacent bulk soil (>10 mm from macropores) at 4 soil depths (0–20 cm, 20–40 cm, 40–60 cm and 60–80 cm). Compared to the bulk soil, the MPS soil had higher organic C, total N, bicarbonate-extractable P, Ca⁺, Cu, Fe and Mn and supported higher populations of bacteria, fungi, actinomycetes, Pseudomonas spp., Bacillus spp., cellulolytic bacteria, cellulolytic fungi, nitrifying bacteria and the root pathogen Pythium.In addition, analysis of carbon substrate utilization patterns showed the microbial community associated with the MPS soil to have higher metabolic activity and greater functional diversity than the microbial community associated with the bulk soil at all soil depths. Phospholipid fatty acids associated with bacteria and fungi were also shown to be present in higher relative amounts in the MPS soil compared to the bulk soil. Whilst populations of microbial functional groups in the MPS and the bulk soil declined with increasing soil depth, the differentiation between the two soils in microbiological properties occurred at all soil depths. Soil aggregates (< 0.5 mm diameter) associated with plant roots located within macropores were found to support a microbial community that was quantitatively and functionally different to that in the MPS soil and the bulk soil at all soil depths. The microbial community associated with these soil aggregates thus represented a third recognizable environment for plant roots and microorganisms in the subsoil.     

Crystal structure of a soluble form of the intracellular chloride ion channel CLIC1 (NCC27) at 1.4-A resolution

CLIC1 (NCC27) is a member of the highly conserved class of chloride ion channels that exists in both soluble and integral membrane forms. Purified CLIC1 can integrate into synthetic lipid bilayers forming a chloride channel with similar properties to those observed in vivo. The structure of the soluble form of CLIC1 has been determined at 1.4-A resolution. The protein is monomeric and structurally homologous to the glutathione S-transferase superfamily, and it has a redox-active site resembling glutaredoxin. The structure of the complex of CLIC1 with glutathione shows that glutathione occupies the redox-active site, which is adjacent to an open, elongated slot lined by basic residues. Integration of CLIC1 into the membrane is likely to require a major structural rearrangement, probably of the N-domain (residues 1-90), with the putative transmembrane helix arising from residues in the vicinity of the redox-active site. The structure indicates that CLIC1 is likely to be controlled by redox-dependent processes.     

Evidence that temperature change cues reproductive development in the spiny damselfish, Acanthochromis polyacanthus

The role of temperature in the reproductive development of the spiny damselfish, Acanthochromis polyacanthus, was assessed by measuring changes in ovulatory periodicity and plasma levels of gonadal steroids in response to temperature shifts of 4°C. This was assessed in sexually mature, paired and unpaired fish. The effect of brood presence on reproductive activity was also assessed. Temperature change but not brood presence significantly affected the spawning periodicity of sexually mature, reproductively active pairs. A temperature increase from 24°C to 28°C resulted in a decrease in the ovulatory period from 32 days to 21 days. Incubation temperature was found to dramatically affect survival of the egg clutches with only one of 13 egg clutches surviving to hatching at 24°C, compared with 50 of 51 egg clutches surviving at 28°C. Gonadal steroid production was positively correlated with temperature increase in paired fish. Plasma levels of 17-estradiol and testosterone (T), increased significantly in female fish, whereas there was no change in T and 11-ketotestosterone, in paired male fish exposed to a temperature increase from 24°C to 28°C. In unpaired fish, temperature change had ambiguous affects on plasma steroid levels, suggesting that social context is necessary for stimulation of reproductive development by temperature.