Utilization of the Relaxed Symphysis Pubis of Guinea Pigs for Clues to the Mechanism of Ovulation

This ultrastructural study reveals that the collagen of the relaxed symphysis pubis is decomposed by some substance in multivesicular structures of symphyseal fibroblasts. The multivesicular structures appear identical to those found in fibroblasts of the ovarian follicle. The data support the hypothesis that these multivesicular structures are instrumental in preovulatory decomposition of mammalian follicles. (Endocrinology 93: 1441, 1973) HE symphysis pubis of the guinea pig is predominantly dense collagenous tissue. Under physiological conditions this tissue decomposes in response to relaxin (1,2). Likewise, the tunica albuginea and theca externa in the Graafian follicle of the mammalian ovary consist of dense collagenous tissue. In the hours preceding ovulation, this thecal framework undergoes physiological deterioration which results in a marked decrease in the tensile strength of the follicle wall (3,4). The specific cause(s) of the preovulatory decomposition are uncertain; however, recent ultrastructural studies of the rabbit ovary have revealed that follicular fibroblasts produce unusual multivesicular structures which protrude from the cell surface and appear to digest the adjacent extracellular collagen (5, 6). The general appearance of these structures Received November 17, 1972. 1 Current address: Department of Physiology, Louisiana State University Medical Center, New Orleans, Louisiana. suggests that they could have a major role in the mechanism of ovulation. To evaluate this hypothesis it seemed worthwhile to determine if similar multivesicular structures could be detected in another collagenous tissue that is undergoing rapid physiological decomposition. This report reveals that fibroblasts in the relaxing symphysis pubis of the guinea pig produce multivesicular structures which appear to decompose the surrounding collagenous tissue and are morphologically similar to the structures observed in the thecal layers of ovulating Graafian follicles. Materials and Methods Seven mature virgin female guinea pigs were injected with 1 mg of 17P-estradiol in sesame oil each day for 4 days. Relaxin® (Releasin W1164, Lot #48E-2103a from Wamer-Lambert, courtesy Dr. Bernard Steinetz) was dissolved in 1% benzopurpurin-4B and injected (0.002 mg relaxin/g body wt) 24 hr after the last dose of estrogen (7). Relaxation was confirmed by palpation of the pubic symphysis 6-8 hr after relaxin treatment (8). If 1442 NOTES AND COMMENTS Endo Vol 93 1973 No 6 the palpation test was positive, the animal was anesthetized by an injection (ip) of sodium pentobarbitol, and tissue was surgically removed from the interpubic ligament. The tissue was placed in 9% glutaraldehyde solution in a petri dish and cut into small sections suitable for rapid fixation. The sections were fixed for 2 hr (at 4 C) with 9% glutaraldehyde in Millonig's phosphate buffer, and postfixed in 2% osmium tetroxide for 1 hr. The tissue was dehydrated with ethanol, rinsed in propylene oxide, and embedded in Spurr's Epon mixture. Control tissue from untreated animals was fixed each time experimental tissue was prepared. The embedded material was sectioned on a PorterBlum MT-2 ultramicrotome and placed on 75-300 mesh copper grids. The tissue was stained with 2% uranyl acetate in 50% aqueous ethanol for 45 min, followed by Reynolds' stain (lead citrate) for 15 min. Micrographs were made with a Hitachi HS-8 electron microscope.