Collection, staining and examination of exfoliated vaginal epithelial cells has proven to be an effective method for monitoring different stages of the estrous cycle.1 The Mirage Dolphin Habitat monitored vaginal cytology on two adult bottlenose dolphins (Tursiops truncatus) for six months.
Hormones affect the vaginal epithelium, and estrogen in particular has a strong influence; high estrogen causes cells to become "cornified" and can show when ovulation is likely. Changes can be noted in the cellular structure of exfoliated vaginal cells throughout the female's cycle. During proestrus, estrogen concentrations increase, and the mucosa of the vagina thickens. During this transition, surface cells become larger, irregular in shape, and flat nucleated (intermediate). They continue to change and become anuclear cornified cells (superficial). As more surface cells become cornified, proestrus is coming to an end and estrus begins. When 80-100% of the cells are cornified, optimal breeding time has arrived. Following peak cornification, estrogen levels begin to decrease and progesterone levels increase, causing vaginal epithelial cells to slough off. The cell layer decreases, and the appearance of deeper, parabasal and intermediate cells indicates the onset of metestrus and diestrus. At this point, ovulation has already taken place.2
Two bottlenose dolphins were trained to allow manipulation of the genital slit and insertion of a swab into the vagina. Sterile swabs were used to collect vaginal cells, and slides were prepared using a Jogensen lab dip quick stain. Cells were evaluated microscopically and cell types identified according to percentage. This process showed clear changes in the cellular structure of vaginal cells according to the estrous cycle phase.
Currently, the use of tricolor Papanicolaou stain is being tested for accuracy with bottlenose dolphins. This method has been proven to identify circulating estrogen levels in the giant panda.1 If also successful with dolphins, the Papanicolaou stain could be a simple method for obtaining detailed information about the estrous cycle. This staining process reveals the same classification of vaginal cells as the Jogensen stain. Additionally, it shows three distinct color phases: basophilic cells are blue, acidophilic cells are pink, and keratinized cells are yellow or orange.1 The Mirage has been conducting this study since November 2003.
The authors are grateful to Dr. Dennis Arn for his expertise. We also thank Dr. Barbara Durrant and the Center for Reproduction of Endangered Species (CRES) staff. None of this would be possible without the hard work and dedication of The Mirage Dolphin Trainers.
1. Durrant B, N Czekala, M Olson, A Anderson, D Amodeo, R Campos-Morales, F Gual-Sill, DJ Ramos-Garza. 2002. Papanicolaou Staining of Exfoliated Vaginal Epithelial Cells Facilitates the Prediction of Ovulation in the Giant Panda. Theriogenology. Apr 15; 57(7). Pp. 1855-64.
2. England G, PW Concannon. 2002. Determination of the Optimal Breeding Time in the Bitch: Basic Considerations. Recent Advances in Small Animal Reproduction. International Veterinary Information Service, Ithaca NY. A 1231.0602.