Doctoral Dissertations

Date of Award

6-1987

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Zoology

Major Professor

John R. Kennedy

Committee Members

Leaf huang, John Koontz, Roland Bagby

Abstract

The large granulosa lutein cells of the corpus luteum of the ovary in the pig synthesize and secrete progesterone. Receptors that bind luteinizing hormone (LH) and others that bind low density lipoproteins (LDL) located on the outer surface of this cell type play important roles in the regulation of the secretory event. Both types of receptors when bound with their specific ligand have been demonstrated to be internalized by endocytosis. New receptors replace the lost receptors and the process is repeated many times for the life of the cell. Neither the source of new receptors nor the mechanism for the replacement process has been identified.

Earlier studies by the author (Gonadotropin Receptors of the Ovarian Granulosa Lutein Cell of the Pregnant Pig, Master's Thesis, University of Tennessee, June 1982) revealed that a pool of unbound gonadotropin receptors was located within the cell and that subcellular fractions enriched with Golgi body membranes or endoplasmic reticulum possessed gonadotropin receptors, especially the Golgi body fractions. A presupposition was made that the intracellular receptors located in these organelles were the source of new receptors, but the mechanism of transport of these receptors from within the cell to the outer surface was unknown.

Examination of this cell type by electron microscopy revealed that coated vesicles, known membrane transporters within cells, were located near the vicinity of the Golgi body as well as near the surface of the cell. A hypothesis was presented that the coated vesicles seen at or near the Golgi bodies were responsible for the transport of newly synthesized or recycled receptors in the Golgi to the outer cell surface.

To test the hypothesis, coated vesicles were isolated from these cells using differential and density gradient centrifugation techniques and their receptor content was examined. A fraction of coated vesicles estimated to be greater than 97% pure was obtained. Analysis of the coated vesicle fraction using radioreceptor assays revealed that it possessed receptors for both LH and LDL and thus is consistent with our hypothesis.

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