Vitellogenin transport and yolk formation in the quail ovary.

Morphological and biochemical investigations were made on the yolk formation in ovaries of the quail Coturnix japonica. Morphologically, two ways of nutrient uptake were observed in follicles. In small oocytes of white follicles, vitellogenin (VTG) was taken up through fluid-phase endocytosis which was assisted by follicular lining bodies. The lining bodies were produced in follicle cells. They adhered to the lateral cell membrane, moved along the membrane in the direction of the enclosed oocyte and were posted to the tips of the microvilli. These tips, now with lining bodies, were pinched off from the main cell body, engulfed by indented cell membranes of the oocyte, and transported to yolk spheres. In large oocytes of yellow follicles, VTG and very-low-density lipoproteins (VLDL) were taken up through receptor-mediated endocytosis. The VTG and VLDL particles diffused through the huge interspaces between follicle cells, and once in oocytes were transported to yolk spheres via coated vesicles. Immunohistochemistry showed that the VTG resides on or near the surface of the follicle cell membrane at the zona radiata whereas the cathepsin D resides at or near the oocytic cell membranes. Tubular and round vesicles in the cortical cytoplasm of oocytes were also stained with both antisera, suggesting that these vesicles are the sites where the VTG is enzymatically processed by cathepsin D. Upon analysis by SDS-PAGE, a profile similar to that of yolk-granule proteins was produced by incubating VTG with a quail cathepsin D of 40 kD.

Dose-response relationships and pharmacokinetics of vitellogenin in rainbow trout after intravascular administration of 17alpha-ethynylestradiol.

A commonly used endpoint in bioassays testing the estrogenicity of chemicals is the induction of the egg yolk precursor vitellogenin (VTG) in male fish. However, relatively little is known about the kinetics of induction and elimination of VTG in fish exposed to xenoestrogens. In this study, we administered graded intra-arterial doses (0.001, 0.1, 1.0 and 10.0 mg/kg) of 17alpha-ethynylestradiol (EE(2)) to male rainbow trout via a dorsal aortic cannula which allowed repetitive blood sampling from individual fish for up to 48 days after injection. The plasma concentrations of VTG was quantified using an enzyme-linked immunosorbent assay procedure and the simultaneous concentrations of EE(2) were determined by gas chromatography-mass spectrometry. The pattern of VTG induction was similar for all doses of EE(2), with a 12-h lag-time before increase from basal levels (0.006-0.008 microg/ml), then increasing sharply to maximum levels within 7-9 days (C(max)=0.05, 711, 1521 and 2547 microg/ml VTG for the 0.001, 0.1, 1.0 and 10.0 mg/kg doses, respectively). After induction by EE(2), VTG declined mono-exponentially with an elimination half-life of 42-49 h. The half-life of VTG increased to 145 h in the 10 mg/kg treated fish. The pharmacokinetics of EE(2) were distinctly nonlinear with substantial increases in the elimination half-life with increasing dose. The plasma concentration-time profiles of EE(2) were influenced by enterohepatic recirculation that caused multiple or secondary peaks in the profiles. In a separate experiment, the pharmacokinetics of purified VTG was characterized after intra-arterial injection in trout. After direct injection of VTG, plasma levels declined tri-exponentially with an apparent steady-state volume of distribution of 837 ml/kg; total body clearance was 31.1 ml/h per kg, and the elimination half-life was 43.7 h.

Synchronized endocytosis studied in the oocyte of a temperature-sensitive mutant of Drosophila melanogaster.

This study demonstrates that endocytosis in the oocyte of Drosophila melanogaster is reversibly blocked at the stage of pit formation by the temperature-sensitive, single-gene mutant, shibirts1. Uptake of horeradish peroxidase conjugated with wheat-germ agglutinin was observed to be normal in mutant oocytes at 19 degrees C, but was blocked at 29 degrees C. After 10 min at 29 degrees C, there was a build-up of coated pits along invaginations of the plasma membrane. Also, the endosomal compartment consisting of tubules, bulbs, and small yolk spheres, disappeared. Lowering the temperature to 19 degrees C after 10 min at 29 degrees C released a synchronized wave of endocytosis into a cytoplasm cleared of uptake-related organelles. By observing this synchronized wave after exposure to 19 degrees C for varying durations, we determined that endocytosis proceeds as follows: coated pits/vesicles----tubules----small yolk spheres----mature yolk spheres. The observations suggest that these organelles transform one into another within this sequence.