Gynecology Past Paper PDF (Suez Canal University 2019-2020)

Suez Canal University Faculty of Veterinary Medicine Department of Theriogenology Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan https://mohamedmedan.wordpress.com Email: [email protected] Email: [email protected] Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Contents Terminology...................................................................................................................................................................... 1 NEUROENDOCRINE CONTROL OF ESTRUS AND OVULATION...................................................................... 3 Pattern of hormone secretion:......................................................................................................................................... 4 Reproductive hormones................................................................................................................................................... 4 Hypothalamus:.................................................................................................................................................................. 4 Gonadotropin Releasing Hormone (GnRH):.................................................................................................................. 4 Kisspeptin and its effect on GnRH:................................................................................................................................ 5 The pituitary gland:.......................................................................................................................................................... 6 Follicle stimulating hormone (FSH):.............................................................................................................................. 6 Luteinizing hormone (LH):............................................................................................................................................. 7 Oxytocin:........................................................................................................................................................................ 8 Ovarian Steroids:.............................................................................................................................................................. 9 Estrogens:....................................................................................................................................................................... 9 Progesterone:................................................................................................................................................................ 10 Endometrial hormones:................................................................................................................................................. 11 Prostaglandins:............................................................................................................................................................. 11 Other hormones:............................................................................................................................................................. 12 Puberty............................................................................................................................................................................ 14 Factors affecting the age of puberty:............................................................................................................................ 14 Endocrinology of Puberty:............................................................................................................................................ 15 Ovarian follicular dynamics during estrous cycle....................................................................................................... 16 Stages of follicle growth:.............................................................................................................................................. 17 Follicular waves during bovine estrous cycle:.............................................................................................................. 18 HORMONAL CHANGES DURING ESTROUS CYCLE IN CATTLE................................................................... 20 FEMALE REPRODUCTIVE CYCLE......................................................................................................................... 21 Estrous Cycle:............................................................................................................................................................... 21 Periods of estrous cycle:............................................................................................................................................... 21 Heat (estrus) Detection................................................................................................................................................... 23 Estrus detection aids:.................................................................................................................................................... 23 SYNCHRONIZATION OF ESTRUS........................................................................................................................... 27 Why do we synchronize estrus?................................................................................................................................... 27 How do we synchronize estrus:.................................................................................................................................... 27 Protocols of estrus synchronization by PGF2α............................................................................................................ 27 Interval from PGF2α administration to estrus:............................................................................................................. 28 Estrous synchronization using PGF2α and a Progestin:............................................................................................... 28 Estrous synchronization using GnRH–PGF2α–GnRH:................................................................................................ 29 ESTROUS CYCLE OF THE MARE............................................................................................................................ 31 Seasonality:................................................................................................................................................................... 31 Estrus and ovulation:.................................................................................................................................................... 31 ESTROUS CYCLE IN SHEEP AND GOATS............................................................................................................. 34 Detection of estrus in ewes:.......................................................................................................................................... 34 Estrous cycle in goats (doe or nanny):.......................................................................................................................... 34 a Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) FOLLICULAR WAVE (estrous cycle) IN SHE CAMEL........................................................................................... 35 What causes ovulation in she camel?........................................................................................................................... 35 Signs of estrus in she camel:......................................................................................................................................... 37 ESTROUS CYCLE OF THE BITCH........................................................................................................................... 38 Hormonal changes during estrous cycle in bitches and LH surge................................................................................ 39 Canine Vaginal Cytology (smear):............................................................................................................................... 40 Pseudopregnancy (Pseudocyesis) in bitches:................................................................................................................ 41 ESTROUS CYCLE OF THE QUEEN (FEMALE CAT)............................................................................................ 43 Induction of ovulation without copulation:.................................................................................................................. 44 UNDERSTANDING ULTRASOUND TECHNOLOGY............................................................................................ 46 Important notes:............................................................................................................................................................ 46 WHY ULTRASOUND IS BETTER?........................................................................................................................... 47 How are ultrasound waves produced?.......................................................................................................................... 47 Physics and fundamental of diagnostic ultrasound:...................................................................................................... 48 Attenuation of ultrasound beam:.................................................................................................................................. 48 Resolution..................................................................................................................................................................... 49 ULTRASOUND EQUIPMENTS.................................................................................................................................. 51 Ultrasound scan gels:.................................................................................................................................................... 53 Control knobs in the ultrasound machine:.................................................................................................................... 53 SCANNNG MODES....................................................................................................................................................... 55 How can you read an ultrasound image?...................................................................................................................... 56 TECHNIQUES OF EXAMINATIONS........................................................................................................................ 57 ULTRASOUND APPLICATIONS IN VETERINARY REPRODUCTION............................................................ 60 Normal ovarian structures:........................................................................................................................................... 60 Ovarian pathology:....................................................................................................................................................... 61 Cyclic changes of the uterus:........................................................................................................................................ 62 Delayed uterine clearance:............................................................................................................................................ 62 Normal pregnancy:....................................................................................................................................................... 63 Imaging of the Reproductive Tract of Bitches:............................................................................................................ 65 ULTRASOUND APPLICATIONS IN MALES........................................................................................................... 68 Testicular ultrasound:................................................................................................................................................... 68 Ultrasonography of male accessory sex glands:........................................................................................................... 68 ARTEFACTS.................................................................................................................................................................. 70 Biologic effect of ultrasound:......................................................................................................................................... 74 b Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Terminology Theriogenology: is the branch of veterinary medicine concerned with reproduction, including the physiology and pathology of male and female reproductive systems of animals and the clinical practice of veterinary obstetrics, gynecology, andrology and artificial insemination. Theriogenologists are veterinarians with advanced training in animal reproduction and obstetrics. Gynecology: Gynec. means female,..ology means science. It is the science, which deals with the study of the female during childhood and from puerperium till conception. Or it deals with the study of fertility and infertility of the female. Andrology: Andro. means male..ology means science. It is the science, which deals with the study of male genitalia and any deviation from normal physiological condition. Artificial Insemination: Artificial insemination (A.I.) involves collection of semen from a male and transfer of the semen to the reproductive tract of sexually receptive female at the time of ovulation in order to result in fertilization. Females can be inseminated with either fresh semen or with frozen semen. Obstetrics: is the branch of veterinary medicine that deals with medical and surgical care together with manipulations of the female during gestation, labor (parturition) and puerperium. Fertility: It is the complete ability of the female to give a viable ovum ready for fertilization by a viable sperm to give a viable offspring. Infertility: It is a temporal inability of the female either to produce viable ovum or keep the zygote, embryo, and fetus till parturition. Sterility: Absolute infertility or complete inability of the female to reproduce. Estrus period (heat): It is the period in which the female accepts the male and mating occurs. It is characterized by restlessness, flow of clear mucus secretion from the vulva, hyperemia and redness of the vulva, frequent urination and mounting other animals. Ovulation occurs during this phase of the cycle in all domestic species with the exception of the cow, where it occurs about 12 hours after the end of estrus. Ovulation is a spontaneous process in all domestic species with the exception of the cat, rabbit and camel, in which it is induced by the act of coitus. Silent heat: The animal doesn‘t show the typical external sings of estrus although the normal cyclic changes occurring in the ovaries. Anestrum (absence of estrus): It is the period of sexual rest in the female with typical breeding season. It may be physiological (pregnancy, after parturition and outside the breeding season in seasonal breeders) and pathological (the genital system is mainly quiescent). Time of breeding: It is that time during the estrus (heat) at which insemination or natural mating results in a high conception rate. Cow Ewe Mare Bitch (*) Sow nd nd from the middle to 2 day of Mount each Optimal breeding day is Late 2 day or early the end of heat heat 2 days 2 days after ovulation. 3rd day of heat ------------------------------------------ --------------------------------------- 1 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) *Bitch breeding management: The average bitch ovulates on the second day of estrus. If only one breeding is possible or if artificial insemination is to be performed, optimal breeding day is 2 days after ovulation. In bitches, ovulation day is best determined by measurement of serum progesterone concentration; serum progesterone concentration 2 days before ovulation is 2-3 ng/ml and on ovulation day is 5-8 ng/ml. Age of senility: It is the age at which the ovaries fail to produce viable ova after the full life of breeding. Sex ratio: It is the number of the male to the number of the female, theoretically it is 1:1. Conception rate: The ratio between the numbers of the females, which don‘t return to heat 21 days after insemination to the total number of the females inseminated. Pregnancy rate: It is the ratio between the numbers of the pregnant females to the total number of females inseminated. Calving rate: It is the ratio between the females, which give a live offspring to the number of females inseminated. Repeat breeder syndrome: It is a cow which has normal estrous cycle and bred for more than 2 times by a good fertile bull but failed to conceive. The cause may be failure of fertilization or early embryonic death. Diagnostic ultrasound: uses very high frequency sound waves that are pulsed into the body, and the returning echoes are then analyzed by computer to yield high resolution images of organs, tissues, and blood flow (Ultrasound depends on pulse echo principle). N.B: High-quality ultrasound studies require a firm understanding of the important physical principles of diagnostic ultrasound. Echogenic = echoic: has echoes and depends on the density of the tissue. If the density of the tissue increased, it produces more echoes (hyperechoic) and the density of the tissue decreased it produces less echoes (Hypoechoic). Anechogenic = anechoic: without echoes or echo free as urine, fetal fluids, follicular fluids). Artefact: A feature appearing in an ultrasound image that does not correspond to actual normal or pathologic structure in the body, For example as shadow, mirror, enhancement ….. etc. ------------------------------------------ --------------------------------------- 2 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) NEUROENDOCRINE CONTROL OF ESTRUS AND OVULATION - Nervous system and endocrine system control reproduction. The eyes (optic nerve), nose (olfactory nerve) and tactile senses (sensory nerves) transmit stimuli (light, sex odor and tactile stimuli) from the environment to the brain. - The neuroendocrine control of the estrous cycle involves the integration of multiple regulatory signals. Secretion of gonadotropin-releasing hormone (GnRH) into the hypothalamic–adenohypophyseal portal circulation is considered the primary endocrinological mechanism regulating gonadotropin synthesis and release. Neuroendocrine control of estrus and ovulation - Under stimulation by GnRH, gonadotropes in the pituitary gland synthesize and release luteinizing hormone (LH) and follicle-stimulating hormone (FSH). These two gonadotropins reach the gonads through the blood circulation and act in the ovary to stimulate follicle growth and maturation, ovulation of the preovulatory follicle, and synthesis of gonadal steroid. Gonadal hormones, in turn, exert feedback actions at the central and adenohypophyseal levels to control the release of GnRH and gonadotropins. ------------------------------------------ --------------------------------------- 3 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Pattern of hormone secretion: - A pulsatile pattern of secretion is seen for all hormones, with variations in pulse characteristics. - Differential secretion of gonadotropin-releasing hormone (GnRH) with respect to pulse frequency and amplitude has profound effects on reproductive function in both the female and male. In the female, a slow GnRH pulse frequency favors (tonic secretion) increased secretion of FSH, resulting in the development of the ovarian follicle during the follicular phase of the estrous cycle. Hypothalamic Secretion of GnRH - The onset of high frequency and amplitude GnRH pulses (Surge secretion) results in a sudden increase in LH leading to ovulation, marking the onset of the luteal phase. Conversely, constant GnRH secretion has been shown to suppress pituitary LH and FSH secretion which is only restored once pulsatile secretion recommences. Reproductive hormones Hypothalamus: - The hypothalamus is a part of the brain that lies just above the pituitary gland. It releases polypeptide hormones that start and stop the release of pituitary hormones. The hypothalamus controls hormone production in the pituitary gland through several "releasing" hormones including gonadotropin- releasing hormone (GnRH). - GnRH stimulates pituitary gland to make luteinizing hormone (LH) and follicle- stimulating hormone (FSH), which are important for normal puberty and reproduction. - The hypothalamus also secretes oxytocin which travel to the posterior pituitary gland. Gonadotropin Releasing Hormone (GnRH): - Before puberty, secretion of GnRH occurs infrequently and the amplitude is also low. This means that corresponding effects on secretion of FSH and LH from the anterior pituitary are also reduced. However, as ovarian development progresses, secretion of estradiol from waves of growing follicles increases. ------------------------------------------ --------------------------------------- 4 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) - The frequency and amplitude of GnRH secretion from the tonic center in the hypothalamus are increased. This produces more FSH and LH which leads to more follicular activity. - However, ovulation needs a preovulatory LH surge. This requires activation of the surge center. The prepubertal female is characterized by having insufficient ovarian- derived estradiol to stimulate the surge center. As she matures, her hypothalamus becomes progressively more sensitive to estradiol. Kisspeptin and its effect on GnRH: - Gonadotropin-releasing hormone neurons do not express progesterone, androgen or estradiol receptors involved in the feedback control of GnRH. This means that gonadal steroids must exert their effect on GnRH indirectly via steroid-responsive neurons. - Kisspeptin is secreted from neurons in the forebrain and play a critical role in the control of GnRH secreting cells. Because kisspeptin neurons have receptors for steroid hormones, they can respond to steroid (e.g., estrogen) signals. High levels of estrogen stimulate kisspeptin neurons to deliver excitatory signals to the GnRH-secreting cells to increase their secretion of GnRH. The increased GnRH secretion, stimulates a huge output of LH (the LH surge) that triggers ovulation. Endocrine control of cyclical reproductive activity. Solid line, Hypothalamic-Pituitary Ovarian (HPO) axis; dashed line, extra HPO axis inputs; GH, growth hormone; IGFs, insulin-like growth factors; IGFBPs, insulin-like growth factor binding proteins; PGF2alpha, prostaglandin F2alpha. ------------------------------------------ --------------------------------------- 5 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Drugs available for GnRH: 1- Receptal (Contains Buserelin 4 μg/ml = 0.004 mg /ml and the dose is 20 μg (5 ml) for cows and 40 μg (10 ml) for mares. 2- Fertagyl (Gonadorelin 0.1 mg/ml and the dose is 2 ml). Uses and dose rate: 1. Treatment of cystic ovaries – 20 μg IM (Receptal). Repeat after 8-14 days if required. 2. Improvement of conception rate – 10 μg IM before or at the time of insemination. 3- Delayed ovulation and anovulation. 4- Anestrum – 20 μg IM. Repeat after 8-22 days if required. N.B: Injection (IM) of GnRH will induce a surge of LH within 2 to 4 hours and ovulation of a viable dominant follicle (≥ 10 mm) will occur within 24 to 36 hours. The pituitary gland: - The pituitary gland is located at the base of the brain near the hypothalamus. The pituitary gland is considered the master gland because it secretes many hormones that control various endocrine glands. The anterior pituitary gland: - The anterior pituitary gland secretes several hormones, all large molecular weight proteins that control various functions in the body. These include: Follicle stimulating hormone (FSH): - FSH is a glycoprotein consists of two protein fractions, the a-subunit and the b- subunit. This hormone may be regarded as the initiator of ovarian activity since it directly promotes growth of ovarian follicles. - Equine Chorionic Gonadotropin (eCG) previously termed Pregnant Mare Serum Gonadotropin (PMSG) having predominantly FSH like activity and little LH like activity. PMSG obtained from endometrial cups in pregnant mares from day 40 to 120 of gestation. Drugs available: Folligon (Intervet): Injection: Powder for reconstitution. Each vial contains Pregnant Mare Serum Gonadotrophin (PMSG) as a white freeze dried crystalline powder (1000 I.U.). Uses: - Anestrum - Superovulation. - Increase fertility rate. ------------------------------------------ --------------------------------------- 6 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Dose rate: - Cows: 1500 - 3000 I.U. S/C or I/M. Repeat after 10-14 days. - Sheep – goat: 500 – 600 IU. - Sow: 1000 IU. - Bitch: 50 – 200 IU. Folltropin-V: is a purified follitropin extract obtained from carefully selected porcine pituitary glands. It is lyophilized to maintain potency under normal storage conditions. Each 20 mL vial contains FSH equivalent to 400 mg. For superovulation in cows: Administer 2.5 mL (50 mg) of FOLLTROPIN-V intramuscularly, twice daily, for four days. Administer prostaglandin F2α or a prostaglandin F2α analogue to cause luteolysis, in conjunction with the 6th dose of FOLLTROPIN-V. Luteinizing hormone (LH): - LH is a glycoprotein hormone. The function of LH is to stimulate the maturation and ovulation of the antral follicle and secondly to stimulate the formation and maintenance of the corpus luteum. - LH concentrations are lowest during the mid-luteal phase of estrous cycle, rising only a few days before the onset of estrus to a peak usually on the day of ovulation, to then drop to previous levels over the next few days. - Human chorionic gonadotrophin (hCG), obtained from the urine of pregnant women, which has mainly an ‗LH-like‘ effect but with some ‗FSH-like‘ activity. Drugs available: Chorulon, Injection: Powder for reconstitution. Human chorionic gonadotropin, 1500 IU/vial Pregnyl, Human chorionic gonadotropin, 1500 IU/vial. Uses: Induction of ovulation. Delayed ovulation. Prolonged estrus. Anovulation - 1000-1500 IU - at the time of insemination, IV or IM. Cystic ovaries - 3000-5000 IU I/M or IV. ------------------------------------------ --------------------------------------- 7 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Dose rate - Cattle: 1500–3000 IU i.v. or i.m. - Mare: 1500–3000 IU i.v. or i.m. - Sow: 500–1000 IU i.m. or s.c. - Sheep and goat: 100–500 IU i.v. or i.m. - Bitch, 100–500 IU i.m. - Cat: 100–200 IU i.m. N.B: FSH and LH are protein in nature (so, they cannot be given orally, digested by proteolytic enzymes). Also, repeated uses lead to antibody formation. The posterior pituitary gland: - The posterior pituitary gland secretes two peptide hormones, oxytocin and vasopressin. These hormones are synthesized in the paraventricular and supraoptic nuclei of the hypothalamus and droplets pass down the axons of specialized secretory neurons into the posterior pituitary gland from where they are secreted into the systemic circulation. Oxytocin: - It is secreted by hypothalamus and stored in posterior pituitary. IT causes contractions of the smooth muscles of the uterus and myoepithelial cells around the alveoli of the udder. So, it is used in uterine inertia and let down of milk. Oxytocin is secreted under cervical stimulation during coitus and parturition. This leads to suction of the sperm to the site of fertilization and expulsion of the fetus. - The straining forces the fetus through the cervix and anterior vagina, initiate Ferguson‘s reflex which leads to release of large amount of oxytocin from poasterior pituitary causing further contractions of Ferguson the myometrium. reflex Milk let down Drugs available and dose rate: -Syntocinon: Small animals (5-10 IU); large animals (20-30 IU). N.B: The myometrium is very sensitive to the effects of oxytocin and high dose rates can cause spasm rather than synchronized contractions. The myometrium will also become refractory to oxytocin effect. So, oxytocin will be most effective when used in an intravenous drip in saline. ------------------------------------------ --------------------------------------- 8 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Uses: 1- Uterine inertia. 4-Enhance ovulation 2-Pyometra 5-Retained placenta 3-Milk letdown 6-Hasten uterine involution Contra-indications of oxytocin: - When the product is used as an aid to parturition, cervical dilation must be confirmed prior to administration to prevent the risk of fetal death and possible uterine rupture. - The product is contra-indicated in any form of obstructive dystocia. - Excessive doses of the product may delay parturition by producing in coordinated uterine contractions which interfere with the progress of the fetus especially in multiple pregnancies. - Adrenaline at physiological levels markedly reduces the effect of oxytocin on the uterus or mammary gland. For this reason the animal should not be frightened when complete oxytocin effect is desired to cause either milk 'let down' or uterine contractions. Ovarian Steroids: Estrogens: - Estrogens are secreted by theca interna of the Graafian follicle. Commercially it is synthesized from coal tar. (Folone 1 or 5 mg). - There are several forms of estrogen. In the non-pregnant animal estradiol is the most active; in the pregnant animal there are several, but the most important is estrone sulphate. Actions of estradiol: - Growth and development of reproductive organs. - Development of the secondary female sexual characteristics. - Development and growth of mammary ducts. - Estradiol is associated with the behavioral displays common to estrus (acts on CNS). - Estradiol also causes the relaxation of the cervix during estrus. - Estrogen presence has a negative impact on FSH, however it has a positive impact on LH. - Estrogen causes the retention of fluid in the cells of the genital tract thus the tract feels more turgid on palpation through the rectum. - Estrogen also increases blood flow to the reproductive tract and stimulates the production of mucus by the vagina, promoting the conditions for copulation. - In the uterus, estrogen facilitates the passage of larger cells, specifically white blood cells, into the lumen, increasing the resistance of the uterus to any infection which may be introduced at insemination. - Makes uterus more responsive to oxytocin. ------------------------------------------ --------------------------------------- 9 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) - Metabolic effect - enhances water retention, favors protein build-up, decreases blood cholesterol, increases body fat. - Anabolic effect due to nitrogen retention (used in fattening). N.B: repeated use of estrogen may leads to cessation of milk production and cystic ovaries. Also may lead to carcinogenesis. Therapeutic uses: - Uterine infections (pyometra; opening of cervix). - Uterine atony. - Poor uterine drainage. - Feed additives and implants for improved weight gain. - Induction of heat symptoms. Sources of estrogen: a. Natural - plants, mammalian tissues (ovary, testicle, adrenal cortex, placenta, urine, blood). b. Synthetic - diethylstilbesterol, estradiol, and others Dose: Small animals ► 5-10 mg Large animals ► 20-30 mg. Progesterone: - Progesterone is secreted by the corpora lutea, suppresses the further development of follicles and the secretion of estrogen. - High levels of progesterone and low levels of estrogen prevent a cow from coming into heat. - Administration of progesterone to cattle prevents the expression of estrus and subsequent ovulation as long as cattle are under its influence. Progestins: Sources: mammalian tissue (ovary, testicle, adrenal cortex, placenta, blood) and synthetic. Physiologic actions: - The primary role of high progesterone concentrations during the luteal phase is to prepare the uterus for reception of the embryo. - Increased secretion of uterine glands (uterine milk), reduced motility of uterus less sensitive to oxytocin. - Maintenance of pregnancy (formation of cervical plug). - Development of secretory system of mammary gland. - Inhibits hypothalamus and anterior pituitary (negative feedback). - Chronic use may favor pyometra-increased secretions, closed cervix, decreased uterine immune response. ------------------------------------------ --------------------------------------- 10 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Therapeutic uses: 1. Estrus suppression (Negative feedback on GnRH and thus anterior pituitary). 2-Estrous synchronization (with or without PGF2α, will be explained later). Endometrial hormones: Prostaglandins: - Prostaglandins are synthesized in the body from arachidonic acid. Pulsatile secretion of Prostaglandin F2α by the lining of the uterus (endometrium) commences if pregnancy is not detected at about day 14 post-ovulation. - It is well known that oxytocin can induce luteolysis in the cow by inducing the release of PGF2a. Specific oxytocin receptors are present on the outer membranes of endometrial cells in the uterus. Binding of oxytocin to the receptor stimulates the conversion of arachidonic acid to PGF2 α within the endometrial cell. The concentration of oxytocin receptors is low during the early part of the cycle but increases as the cycle progresses, stimulated by estradiol secretion from waves of follicle growth during the luteal phase. - The presence of PGF2α causes the "lysis" of the CL and an almost immediate drop in circulating levels of progesterone, which permits the animal to start displaying estrus. PGF2α also causes a contraction of smooth muscle - which includes the uterus. - In cow, it appears that the close proximity of the ovarian artery and uteroovarian vein is important, particularly since at their points of approximation the walls of the two vessels are thinnest; there is no anastomosis. This allows the leakage of the luteolytic substance (PGF2a) from the uterine vein into the ovarian artery and thus to the ovary, by a form of counter-current exchange through the walls of the vessels. Drugs available: 1) Dinoprost (Lutalyse; dose is 5 ml). 2) Cloprostinol (Estrumate; dose is 2 ml). Uses: ▪ Causes lysis of mature CL ---> progesterone drops off ----> the animal returns to heat. ▪ As abortifacient in feed lot heifers. ▪ Pyometra, ▪ Expulsion of mummified fetus. ▪ Estrus synchronization of a herd basis - two injections 10 days apart should find a functional CL present at the second injection. ▪ Cystic ovaries (luteinized). ▪ Solution to estrous detection problem. ▪ Metritis. ▪ Induce parturition. ------------------------------------------ --------------------------------------- 11 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) N.B: PGF2α is secreted by the endometrium and cause lysis of the CL. However, the presence of a fetus secretes PGE2 which has luteotrophic effect. The half-life of PG2α is very short, so it cannot be measured in the blood. We measure its metabolite. Other hormones: Relaxin: - It is secreted from the CL and placenta in the last stage of pregnancy. It leads to relaxation of the pelvic ligaments in the last stage of pregnancy. Leptin: - Leptin has an important role in not only regulating food intake in domestic animals, but also in controlling reproduction and in particular the time of onset of puberty and fertility. Leptin is a 16-kDa protein (of 140 amino acids) that is synthesized by the white fat cells of adipose tissue and acts not only primarily on the hypothalamus, but also the anterior pituitary and reproductive tract because leptin receptors have also been identified in these structures. The production of leptin is influenced by body condition/weight and thus informs the brain of the level of the body‘s energy stores. The interaction between food intake and the hypothalamic–pituitary axis has been shown by examining the effect of acute fasting and chronic feed restriction on both serum leptin and LH levels. In the cow and ewe there was a marked decrease in serum LH release coinciding with reduced leptin levels. The current evidence indicates that leptin is likely to exert its effects on LH secretion by modulating kisspeptin. Insulin-Like Growth Factor System: - It is clear that the ‗insulin-like growth factor (IGF) system‘ plays a fundamental role in the growth and selection of follicles, as well as luteal development in most of the domestic species. It is likely that the IGF system integrates the metabolic status of the animal with reproductive performance. The circulatory levels of IGF1 parallel the ------------------------------------------ --------------------------------------- 12 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) energy status of that animal in several species; for example, plasma IGF1 levels decline during the period of negative energy balance postpartum in dairy cows. How does the insulin-like growth factor system function? IGFs stimulate follicular growth by stimulating granulosa cell proliferation, as well as the emergence of a dominant follicle, by sensitizing follicular granulosa cells to the effects of FSH and LH. In the latter stages of follicular development, IGF1 is more involved with follicular maturation and differentiation. Inhibin: - Inhibin is a protein hormone secreted by granulosa cells (female) and Sertoli cells (male) and its major action is the negative feedback control of pituitary FSH secretion.. It is found in blood plasma and in great quantities in seminal plasma and follicular fluid. While inhibins are dimers of alpha and beta subunits, dimers of beta-beta subunits form another hormone, activin. Activin is a general growth-stimulating factor, and appears to have local effects on many cell types. Additionally, activin stimulates FSH secretion, thus antagonizing the main biological action of inhibin. Inhibin acts localy on ovary and systemically on pituitary glands - Inhibin (secreted from the dominant follicles) acts locally on the ovary to make atresia of the subordinate growing follicles and on the pituitary gland to decrease FSH secretion. - Immunization against inhibin resulted in increased plasma FSH concentration (presumably by suppressing the negative feedback of inhibin on pituitary FSH secretion), the development of more follicles and an increased ovulation rate. ------------------------------------------ --------------------------------------- 13 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Puberty Puberty can be defined as the time whereby animals become capable of reproducing. By another way: Puberty in the Female: Puberty may be defined as  Age at first estrus (heat)  Age at first ovulation  Age at which the female can support pregnancy Also, puberty can be defined as the first day that serum progesterone (determined in blood samples collected at weekly intervals) exceeds 1 ng/mL. N.B: If animals are bred at puberty, a high percentage will have difficulty at parturition (dystocia). Age of breeding (sexual maturity): It is the age at which the female reproductive tract become more suitable for pregnancy and parturition, where bony and somatic structure reaches their maximum growth. The age of puberty and sexual maturity in different species: Species Puberty Maturity Cow 9 months 18 months Buffalo 2 years 3 years Mare 2 years 3 years Sheep & goats 5-7 months 12 months Swine 4-7 months 12 months Dog 9-12 months 12 months Cat 6-12 months 12-15 months She camel 3 years 4-5-years Factors affecting the age of puberty: 1-Animal species: smaller species usually reach puberty before larger species 2-Breed: some breeds within a species reach puberty earlier than other breeds. In cattle, Hereford and Angus reach puberty before Brahma and Zebu. 3-Season: many breeds and species of animals are seasonal breeders. Long-day breeders begin to cycle and breed during increasing day length. Horses are long-day breeders. Also cats are long day breeder. Short-day breeders begin to cycle and breed and are most fertile during decreasing day length. Sheep and goats are examples of species who are normally fertile and actively breeding in the fall of the year during decreasing day length. 4-Climate: animal generally have an earlier onset of puberty in warmer climates, however excessive heat may actually delay the onset of puberty. 5-Nutrition: an adequate, well-balanced diet is necessary for animals to reach puberty at an optimal time. Severely malnourished animals will have a delayed onset of puberty. 6-Sex: the female of the species or breed usually reaches puberty earlier than male animal. ------------------------------------------ --------------------------------------- 14 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) 7-Management system: the type of housing, pen arrangement, lighting, and the presence or absence of mature animals will affect the onset of puberty. The sight, smell and sound of the opposite sex enhances the onset of puberty 8-Stress: recent studies indicate that low levels of stress results in early onset of puberty, excess stress, however, may delay the onset of puberty. 9- Proximity of the male: exposure to the male will advance the timing of the onset of puberty. This so-called ‗male effect‘ is probably mediated by pheromonal and other sensory cues influencing hypothalamic GnRH secretion. Endocrinology of Puberty: - There is one hypothesis (called gonadostat theory) can be used to demonstrate the control of puberty at least in some species. Under this theory all components of the endocrine system become potentially functional shortly after birth. LH is released from the anterior pituitary gland and stimulates the production of estradiol-17b from ovarian follicles. However, the hypothalamo-pituitary unit is excessively sensitive to the negative feedback effect of estradiol and therefore LH secretion is inhibited. Eventually this excessive sensitivity is reduced, allowing gonadotropin secretion to rise, thereby stimulating follicular development and eventual ovulation.. Physiological and endocrinological changes during the peripubertal period in heifers ------------------------------------------ --------------------------------------- 15 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Ovarian follicular dynamics during estrous cycle Folliculogenesis: - The current evidence indicates that the formation of follicles is completed during fetal life, such that at birth, there are approximately a half to 1 million follicles. Each follicle will contain an oocyte that is arrested in meiosis I. The majority of these follicles will remain at the primordial stages until puberty when, in cohorts, they will undergo a ‗committed‘ transition into primary follicles. The primary follicle cohort will then progress into preantral (secondary) follicles once it has multiple granulosa cell layers alongside the recruitment of the vascularized theca layer. These preantral follicles grow in size, and at 0.5 mm will form a fluid-filled antrum and become antral follicles. Antral follicles are also known as tertiary or Graafian follicles. These early stages of folliculogenesis are controlled by intraovarian growth factors such as insulin-like growth factors (IGF), members of the transforming growth factor beta (TGFB) superfamily, and vascular endothelial growth factor (VEGF). Schematic representation of development of the primordial follicle to the pre- ovulatory Graafian follicle. Key: BL basement lamina, O oocyte, GC granulosa cell, ZP zona pellucida, TC theca cell, FF follicular fluid, BV blood vessel/capillary, CT connective tissue (loose) ------------------------------------------ --------------------------------------- 16 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Stages of follicle growth: Recruitment : gonadotrophin stimulation of a pool of rapidly growing follicles. Selection : a process whereby one or more of the recruited follicles are selected to develop further. Growth Dominance : the mechanism whereby one (the dominant follicle), or several follicles, undergo rapid development in an environment where the growth and development of other follicles is suppressed. Atresia : This process is to an extent controlled by the production, from the dominant follicle, of inhibin, which acts at the local level in limiting the responsiveness of other follicles, and at the pituitary level by limiting the release of FSH. - Ovarian follicular growth and development in ruminants is characterized by two or three consecutive follicular waves per estrous cycle. - Emergence of a new follicular wave is supported by a transient (1–2 days) Follicle development occurs as a wave-like increase in follicle-stimulating pattern consisting of ―Recruitment‖ , hormone (FSH). Peak FSH ―Selection‖ , ―Growth‖ , ―Dominance‖ , and concentrations occur just before wave ―Regression‖ phases emergence, and then subsequently decline over several days. - A follicular wave in cattle is characterized by the emergence of a group of small follicles that grow at a similar rate for about 2–3 days, at which time one follicle is selected to continue growth (dominant follicle) FSH precedes recruitment of follicles while the rest become atretic and (causes follicles to start growing) regress (subordinate follicles). Follicle maturation and ovulation: - LH stimulates follicle growth and a growing follicle produces high levels of estrogen. - High levels of estrogen, in turn, cause estrus and surge release of LH that triggers ovulation. ------------------------------------------ --------------------------------------- 17 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Follicular waves during bovine estrous cycle: - Several follicular waves occur during the estrous cycle as shown in the following figures. During each wave, some follicles are recruited, some of these are selected, and some become dominant. Most eventually regress or become atretic. Only follicles recruited during the third wave or after luteolysis of the corpus luteum produced in the previous cycle will become eligible for ovulation. Several follicular waves occur during the estrous cycle. The small filled circles represent gonadotropin-sensitive follicles. During each wave, some follicles are recruited (R), some of these are selected (S), and some become dominant (D). Most eventually regress or become atretic (A). Only follicles recruited during the third wave or after luteolysis of the corpus luteum produced in the previous cycle will become eligible for ovulation. ------------------------------------------ --------------------------------------- 18 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Ovulation: - The preovulatory LH surge triggers a series of cellular and molecular changes in the follicle that culminate in ovulation. Ovulation is a complex process, involving the measured destruction of the follicle enabling the release of the oocyte and conversion of the follicle into the CL. The three key events involved in ovulation are: (1) elevated local blood flow (hyperaemia) and increased vascular permeability stimulated by prostaglandin E2 and vascular endothelial growth factor (VEGF); (2) the breakdown of the connective tissue within the tunica albuginea and the basement membrane of the ovulatory follicle by the proteolytic enzymes (e.g., collagenase, plasmin). Collectively, this will increase follicular fluid volume and weaken the apex of the follicle, leading it to protrude and form a stigma; and (3) the contractions of the smooth muscle within the theca externa layer of the follicle. These are stimulated by PGF2α and may increase pressure locally to force the stigma to protrude more and eventually open up to release the oocyte and follicular fluid (N.B: there is a dramatic preovulatory increase in the follicular fluid concentration of both PGE2 and PGF2 alpha in cattle, and suggest that an increased concentration of PGF2alpha is more essential for ovulation to occur and suppression of PGF2 alpha rise with indomethacin will inhibit ovulation. Formation of the Corpus Luteum: - The CL is rapidly formed from the Graafian follicle after ovulation, primarily from the luteinisation of the granulosa and thecal cells. Although the CL develops as a result of ovulation, in some species, notably the bitch, there are early signs of luteinisation of the follicle before it has ovulated. - The stimulus for the formation and maintenance of the CL probably varies within species. The hormones that are most likely to be involved are prolactin and LH, with some evidence that they act in conjunction with each other. The treatment of ewes, cows and pigs with LH antiserum causes a dramatic decline in luteal weight and progesterone content, and therefore prolactin is unlikely to be essential for normal luteal function. However, prolactin plays a critical role in maintaining the CL in rodents and the dog. ------------------------------------------ --------------------------------------- 19 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) HORMONAL CHANGES DURING ESTROUS CYCLE IN CATTLE As mentioned before, there are 2, 3 or more follicular waves during estrous cycle in cattle. The next figure shows variation in hormonal level during estrous cycle. Peak concentrations of follicle stimulating hormone (FSH) are detected 1to 2 days before the recruitment of each follicular wave. During diestrus, when elevated concentrations of progesterone are secreted by the corpus luteum, pulses of luteinizing hormone (LH) are sufficient to allow selection of dominant follicles but insufficient to allow maturation of dominant follicles until progesterone returns to basal concentrations after spontaneous luteolysis (due to increased level of prostaglandin F2α secreted by the uterus) or in response to exogenous injections of PGF2 α. After luteolysis, during proestrus, estradiol (Estrogen) increases sufficiently to induce behavioral estrus and LH surge necessary for ovulation. Changes in hormonal levels during estrous cycle in cattle ------------------------------------------ --------------------------------------- 20 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) FEMALE REPRODUCTIVE CYCLE Domestic animals ► Estrous cycle Primates ► Menstrual cycle Estrous Cycle: - After puberty the female enters a period of reproductive cyclicity. - An estrous cycle is the period of time from one estrus to the next or it is a definite physiologic functional rhythm of reproductive system. - Cattle comes in heat (estrus) approximately every 21 days, with the normal range being 17 to 24 days. This periodic pattern of sexual receptivity is the result of an organized and complex series of changes that occur in the reproductive system of cattle. Classification of animals according to occurrence of estrus: 1- Monoestrous: the animals that have one estrous cycle per year as wild animals. 2- Seasonal monoestrous: The animals that have one estrous cycle per season as bitch (spring and fall). 3- Seasonal polyestrous: Animals that have multiple estrous cycles during certain periods of the year as mare (late spring and early summer) and sheep and goat (late summer and early autumn). 4-Polyestrous: the animals show estrus throughout the year as cow and swine. Phases of estrous cycle: A. Follicular Phase: - Period from the regression of the corpus luteum to ovulation. - Relatively short in length. - Growth and maturation of ovulatory follicles. - Rise in blood estrogen and LH concentrations. - Low concentration of blood progesterone. B. Luteal Phase - The period from ovulation until CL regression. - Longer in length compared to the follicular phase. - Growth and maturation of the corpora lutea. - Rise in blood progesterone from the CL. Periods of estrous cycle: -The estrous cycle can be divided into four periods: - Proestrus, estrus, metestrus, and diestrus. - An additional period of anestrus (no estrus) occurs in some animals, and is located between diestrus or parturition and proestrus. Because females can only become pregnant and produce offspring during a short interval within the estrous cycle, it is very important to understand the cycle and be able to identify animals at critical stages of the cycle. ------------------------------------------ --------------------------------------- 21 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) 1. Proestrus (building-up period): - Begins with regression of the CL and drop in progesterone. - Rapid follicle growth occurs during proestrus due to increasing FSH levels. - Estrogen levels begin to increase also. - At the end of proestrus, the animal begins to show signs of estrus. - Estrogens absorbed from the follicles into the blood stimulate increases in vascularity and cell growth of the tubular genitalia in preparation for estrus and pregnancy. Late in proestrus the vaginal wall thickness, and the external genitalia may increase in vascularity. 2. Estrus (heat) (period of sexual receptivity): - Definition-mad desire (Greek). It is the period at which the female accept the male (time of coitus or breeding). ―Heat,‖ or estrus, is the period of time that occurs, on the average, every three weeks (18–24 days) in sexually mature, non-pregnant female cattle, when they are receptive to mounting or riding actively by a bull or other cows. - Proestrus and estrus together comprise the follicular phase of the estrous cycle. 3. Metestrus: - Period of formation of the corpus luteum (CL). - Ovulation occurs in some species (cow). - Progesterone secretion by CL begins. 4. Diestrus - Period in the estrous cycle when the corpus luteum is fully functional. The purpose of CL is to produce progesterone. Progesterone prepares the uterus for pregnancy and supports pregnancy. - If the ovum is not fertilized, the CL will regress, progesterone levels will drop and diestrus will cease. Length of estrous cycle: Species Estrous cycle Estrus phase Ovulation Cow 21 (17-24) days 12 – 18 hours 12 hours after the end of estrus Ewe 17 (13-19) days 24-36 hours 24-36 hours after the onset of estrus Mare 21 (15-25) days 6 days 24-48 hours before the end of estrus Sow 21 (17-25) days 48-72 hours Near the end of estrus ------------------------------------------ --------------------------------------- 22 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Heat (estrus) Detection - Heat detection is an important livestock management tool - missed heats (estrous cycles in which an animal remains "open" or is not bred) result in: - Increased parturition intervals, decreased numbers of offspring produced per year or lifespan of the female. - Longer dry periods and consequently decreased milk production per year or lifespan of a dairy cow. - Reduced profits due to the costs of upkeep for animals that are not producing at their optimal efficiency. - Currently, the best means of heat detection is observation of the animals. Observation should be done during the animals' normal activities. Observation is usually done during morning hours, when the animals are normally more active. (With cattle, observation must be done twice daily, or estrus may be missed due to the short 18 hour length of estrus). Behavioral signs of estrus: ○Restlessness and nervousness. ○Isolation from the rest of the herd or flock. ○Tail elevated, ears perked (cattle & horses). ○Mounting of other members of the herd or flock and stand to be mounted. Physical signs of estrus: ○Thick, clear, stringy mucus from vulva. ○Reddened, swollen vulva. ○Reduced milk production. The best way of detecting estrus is through observation of the animals and their interactions Estrus detection aids: Records: - All heats, including those observed in the early postpartum period, must be recorded. Record systems should be used on a daily basis. ------------------------------------------ --------------------------------------- 23 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Heat Watch: - The system uses an electronic sensing device placed in a small pouch that is glued to the tail- head of the cow. The HeatWatch system uses radiotelemetry to send a signal to a computer every time a cow is mounted and the pressure sensitive transmitter is activated. KaMar Pressure-Sensitive Mount Detectors: - These devices are glued on the topline of the rump. Sustained pressure for several seconds by the sternum of the mounting cow will expel red fluid from a small storage chamber into a larger visible plastic chamber. The detectors should be placed further forward on small cows to avoid false activation when large cows attempt to mount them. The devices can be used for various groups of cattle. To help detect early postpartum heats, they can be applied to cows at 30 days postpartum Tailhead Markings: - Marking the tailhead with chalk, paint, or crayon and observing for evidence of rubbed off or smeared markings is less expensive than Kamar detectors and has gained popularity in larger herds. Markings 10 to 12 inches long and 2 to 3 inches wide are made across the tail head with a livestock marking crayon or heat detector paint. Tail head paint is less convenient to use than crayon but lasts longer (up to three weeks). Electronic Mount Detectors: - Mount detectors are being developed which detect and record mounts. Each detector is coded with the cow‘s identification number, and the information is transmitted to a computer to be stored. At regular intervals during the day, the herd manager can access the information to determine which cows were mounted at a particular time. Various prototypes are currently being tested. Videotape: - This system has been used extensively in research to continuously monitor estrous behavior. The cattle must be clearly identifiable from a distance and must be allowed to interact in a loose-housing arrangement. The video camera(s) should monitor a ------------------------------------------ --------------------------------------- 24 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) large proportion of the housing area. Several cameras may have to be strategically positioned, and the video equipment should be protected from moisture. This system will be effective only if the videotapes are reviewed daily, especially after the herd has been monitored in the evening. The disadvantages are the initial investment for purchase and installation of the equipment and the time required to review the tapes. If used properly, however, videotaping is a very efficient and accurate estrous detection system. Vasectomized or Surgically Altered Bull: - A vasectomized bull or a bull with a surgically altered penis can be an effective heat detector. These animals can be most effective if they are equipped with a chin-ball or ballpoint marker harness which marks the loin and rump of cows that were mounted with a bright-colored marking solution. Surgically altering the penis of a bull to prevent intromission may be more costly than vasectomy, but this method is preferred since vasectomized bulls can copulate with cows and possibly spread disease. bull attempting to mount female in estrus. The penis has been altered so he cannot successfully mate Chin-ball marking device positioned under with the female. bull's chin. The major disadvantage is the danger of handling a bull and allowing him to interact with farm employees. There also is the veterinary cost of performing the vasectomy or surgical alteration and general veterinary costs of maintaining the bull. Testosterone-Treated Female: - Testosterone, a male hormone, causes increased sexual aggressiveness when injected or implanted into cows or heifers. Potential nonlactating cull cows or heifers, even freemartin heifers, are candidates for masculinization. Most people who have used this system prefer cows that have completed a lactation. However, there are several reports of success with freemartin or virgin heifers. A typical treatment regime consists of administering 200 mg of testosterone propionate intramuscularly every other day for three weeks. Some veterinarians have used larger doses (500 to 600 mg) once weekly or a single injection of two grams of testosterone given in three or four locations. When mounting activity is increased following these initial injections, the cow is given a maintenance injection of about 500 mg testosterone propionate every 10 to 14 days. ------------------------------------------ --------------------------------------- 25 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Vaginal electrical resistance: - In early research done in Europe, the electrical resistance (ER) of vaginal fluids decreased during proestrus and through the estrous period. Numerous studies have validated this concept. Several probes that measure the ER of vaginal fluids are now commercially available. - This tool is labor intensive since cattle must be probed frequently to detect significant changes in ER. The probe also is expensive. It must be washed in disinfectant, thoroughly rinsed, and dried before it is used in another cow. Without proper sanitation, the device could spread disease among cows Activity monitors as pedometer: - It is well documented that cattle are more active during estrus and thus spend more time walking and standing than resting. A study showed that activity increased approximately 400 percent during estrus for cows housed in a free-stall barn, and 275 percent for cows in comfort stalls. If pedometers are used properly and the equipment remains functional, this method can be effective in identifying some silent heat cows which fail to show other Pedometer obvious signs of estrus. - This heat detection aid can be effective if used in a herd environment where the devices are checked twice daily, excessive time is not devoted to cleaning mud and manure from them to observe the readings, and cattle are not agitated or moved excessively. Changes in management activities that prompt excessive cow activity on certain days can cause inaccurate readings. ------------------------------------------ --------------------------------------- 26 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) SYNCHRONIZATION OF ESTRUS (Artificial control of the estrous cycle) Why do we synchronize estrus? 1) Facilitate the use of artificial insemination (AI) and allowing the greater uses of genetically superior sires.. 2) Greatly reduces or eliminates labor and time required for estrus detection. 3) Greatly reduces days needed to artificially inseminate the cattle. 4) Allow batch (group) management of inseminations and calvings. Concentration of labor for calving and breeding seasons. Shortens the duration of calving season and subsequent breeding season. 5) May offer some advantages in shortening the calving to conception interval, and thus the calving interval. How do we synchronize estrus: The approved drugs for estrus synchronization are: 1- Artificial induction of luteolysis using luteolytic agents such as prostaglandin F2α. This will obviously only be effective in cycling cows with an active corpus luteum. 2- Using GnRH to manipulate follicular and luteal function. This procedure could potentially be used for the induction of ovulation. 3- Simulation of corpus luteum function, by administration of progesterone (or one of its synthetic derivatives) for a number of days, followed by abrupt withdrawal. This procedure is also effective for the induction of ovulation in acyclic cows. Protocols of estrus synchronization by PGF2α One Injection of PGF2α : - Inject all cows, check heat, and inseminate all cows 12 hours after detection of standing heat. With a single injection of PGF2α, ~70% of the cyclic cows would be expected to display heat (those on Day 6 or greater of the cycle at injection) during the next four to five days. Two Injection PGF2α Programs: - The two injection programs for synchronization with PGF2α are designed to increase the proportion of females with a CL that is responsive to regression with PGF2α. Traditionally, the recommendation was that two injections of PGF2α be administered 11 days apart, but recent data suggest that a 14-day interval is more effective. - With the first PGF2α injection, ~70% of the cyclic cows would be expected to display heat (those on Day 6 or greater of the cycle at injection) during the next four to five days. - Those animals that were not responsive to the first injection (i.e., on Days 1 to 5 of the cycle at the first injection) would respond to the second injection. - Also, cows in heat after the first injection would be on Day 6 or greater of their next estrous cycle and would be expected to show heat a second time, following the second PGF2α injection. ------------------------------------------ --------------------------------------- 27 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) - With all systems that use PGF2α, another PGF2α injection could be added to any protocol (11 to 14 days later) in cows not inseminated to provide another opportunity for A.I. - Remember that, if cows did not initially respond because they were anestrus, an additional PGF2α injection will be of no benefit. Interval from PGF2α administration to estrus: (a) Animals receiving PGF2α following selection of a dominant follicle are in estrus within 2–3 days because the dominant follicle ovulates soon after induced luteolysis. (b) Animals receiving PGF2α at emergence of a follicular wave will require 4–5 days for a small follicle to reach preovulatory diameters and hence have a longer interval to onset of estrus. Estrous synchronization using PGF2α and a Progestin: - Progestins mimic the natural progesterone produced by the corpus luteum (CL) after ovulation. - The progesterone from the CL prepares the uterus for pregnancy and prevents the heifer from coming back into heat. - Melengestrol Acetate (MGA), Controlled Intravaginal Drug Release devices (CIDR), Progestogen-impregnated intravaginal sponge or tampon and Progesterone-releasing intravaginal device (PRID). Progestogen-impregnated Progesterone-releasing intravaginal sponge or Controlled internal drug intravaginal device (PRID) tampon (s) with speculum release (CIDR) device (p) with speculum/applicator and introducer. ------------------------------------------ --------------------------------------- 28 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Melengestrol Acetate (MGA) and PGF2α (14 days treatment): - In this system, MGA is fed at 0.5 mg/head/day for 14 days. Feeding MGA for 14 days prevents cyclic females from showing heat even if their CL regresses, until the MGA is removed from their feed. - Essentially all cyclic females and some anestrous females will exhibit estrus within a week after withdrawal of the MGA. This is a subfertile heat, with many females ovulating a persistent follicle. They should not be inseminated at this estrus. - A single injection of PGF2α , administered 17 to 19 days after the MGA has been withdrawn, will regress the CL that developed following the infertile heat. Most females will show estrus 48 to 72 hours after PGF2α and can be inseminated 12 hours after detection of estrus. CIDR and PGF2α (7 days treatment): - A second progestin-PGF2α system involves the use of progesterone, delivered by means of a CIDR, and PGF2α. - With this system, the CIDR is inserted into the vagina of the female for seven days. - An injection of PGF2α is given either one day before or on the day of CIDR withdrawal. - Females are inseminated based upon detection of estrus during the three- to five-day period following treatment. Estrous synchronization using GnRH–PGF2α–GnRH: - The protocol involves an injection of GnRH at a random stage of the estrous cycle, followed by an injection of PGF2α 7 days later. - A second injection of GnRH follows the PGF2α injection by 48 hours. - The first injection of GnRH causes either ovulation or luteinization of all dominant or large growing follicles (This means, the 1st GnRH injection eliminates the current follicular wave). As a result, a new follicular wave is initiated in all cows by approximately 3 days after the injection. - Therefore, all the females in the group have growing follicles of about the same stage of development. In addition, GnRH stimulates development of luteal tissue from the cells that were previously the dominant follicle. ------------------------------------------ --------------------------------------- 29 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) - PGF2α injection lyses the CL resulting from the initial GnRH injection. Furthermore, the final GnRH injection serves to increase the synchrony of ovulation within a group of cows. The effect of GnRH and PGF2α in Estrus synchronization protocols Addition of progesterone in GnRH–PGF2α : - Progesterone supplementation between the first GnRH and PGF2α for 7 days in Ov- synch and CO-Synch protocols enhances estrus/ovulation synchrony and conception in cows and heifers. - These protocols are advantageous in anestrus cows and heifers and in cows with cystic ovaries. ------------------------------------------ --------------------------------------- 30 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) ESTROUS CYCLE OF THE MARE - Filly reaches puberty between 10 and 24 months of age, with 18 months being the average. At this time she can become pregnant. However, most horse breeders believe that a 2-year-old mare is too immature to be bred. After she is 1 year old, separate fillies from stallions to prevent accidental pregnancy. Seasonality: - The pineal gland plays a critical role in controlling reproductive activity in seasonal breeders. Melatonin is produced by the pineal gland during periods of darkness, with increasing melatonin secretion driving the reproductive response in ‗shortday‘ breeders such as goats and sheep. In ‗long-day‘ breeders such as the horse, the increasing daylight length will ‗switch-on‘ the hypothalamus–pituitary–ovarian–axis. - Reproductive seasonality in the mare refers to morphological and physiological changes of the reproductive organs primarily as a result of the changing ratio of light- hours to dark-hours - Mare is a seasonal breeder, she will only breed or show sexual activity during the spring, summer and autumn, this is termed the breeding season. Winter is the non- breeding season or anestrus. On average the breeding season lasts from April until October in the northern hemisphere and October to May in the southern Hemisphere. During this time, the mare will have an estrous cycle of 21 days (seasonal polyestrous). Mare breeding season in northern hemisphere Estrus and ovulation: - The duration of estrus varies. Early in the year it lasts 6-8 days, but by midsummer it decreases to about 4 days. - A mare in standing heat displays a typical estrus behavior in the presence of a stallion. She presents her hindquarters to the stallion and, if separated by a partition, leans back against it. The receptive attitude includes a squatting posture, raised tail, flexed pelvis, urination, and spasmodic ―winking‖ of the labia with presentation of the clitoris (winking means rhythmic opening of the ventral vulva lips to expose the clitoris). - Ovulation: occurs at the last 48 hours of estrus at ovulation fossa. -Time of breeding in mares: ► Best results without palpation: - Obtained by multiple breedings starting on the third day and repeating at 48-hours intervals until the mare is no longer in estrus. ► When only one breeding is desired: ------------------------------------------ --------------------------------------- 31 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) - It is recommended that the mare be palpated and bred when she has a 35mm follicle. She should be palpated 2 days later to see if ovulation occurred and if not, she should be rebred. Or inject LH at the time of breeding. ► When two large follicles are detected: - by palpation or ultrasonography, mares should not be bred, since pregnancies involving twins are usually terminated by abortion. Uterine edema in mares during estrus: - Ultrasonography of the uterus is an important part of a reproductive examination. Mares in estrus usually have edema present in the endometrium. Edema is caused by the presence of elevated estrogen and low progesterone levels. The pattern of edema changes over the course of estrus. Edema scores typically increase with growth of the dominant follicle, peak 1 to 2 days prior to ovulation, and decrease the day prior to ovulation. In contrast, the uterus of a mare in diestrus appears tubular and homogeneous without edema. Care in interpretation of uterine edema is necessary as inflammatory changes can also cause edema. A small volume of clear (non- echogenic) fluid may be present in the uterus of a normal mare in estrus. However, an increased volume of echogenic fluid is suggestive of inflammation and infection. Uterine edema Ultrasonographic image Endometrial folds easily observed in a light ‘spoke- wheel‘ pattern; edema may be more evident in uterine horns than uterine body. Typical of early estrus as dominant follicle is developing or late estrus prior to or at time of ovulation Endometrial folds increased in thickness; edema pattern obvious throughout uterus. Typical of mid- estrus and usually represents the peak estrogen effect noted 1 to 2 days prior to ovulation ------------------------------------------ --------------------------------------- 32 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Large distended endometrial folds; exaggerated degree of edema; not typical of a normal mare in estrus. May be associated with uterine inflammation Teasing: - The teaser stallion is a stallion, often of low value, used under controlled conditions to detect whether a mare is in estrous or not, but he is not allowed to cover her. - Once the teaser stallion has confirmed that the mare is in estrus she can then be prepared and covered by the stallion of your A mare showing typical signs of estrus, standing still in choice. the presence of the stallion and exposing her clitoris (lower part of the vulva), an event known as ‗winking‘. The postpartum estrus or foal heat: Should you breed or not at foal heat? - Foal heat is generally around at 9 days postpartum. - Foal heat breeding is good in that you can easily detect the estrus, whereas subsequent heats may be harder to detect. - Foal heat breeding also keeps the mare foaling earlier in the year. Yes if the mare had a normal foaling and postpartum period. - The pregnancy rates on foal heat breeding are dependent on when a mare ovulates postpartum. This is because the mares that ovulate early do not have complete uterine involution, so conception rate is lower. - Conception rates are definitely lower if fluid is observed in uterus on ultrasound examination. Do not breed if fluid is observed. - The fertility of the next heat cycle is not adversely affected if the mare does not become pregnant. - Mares should be bred at this time only if they have been given a careful examination to determine if there has been adequate recovery since parturition. ------------------------------------------ --------------------------------------- 33 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) ESTROUS CYCLE IN SHEEP AND GOATS - The sexual (breeding) season of most breeds of sheep is from October to February, (seasonal polyestrous). During the breeding season, there are 8 to 10 recurrent estrous cycles.. The stimulus for the annual onset of sexual activity is declining length of daylight (short-day breeder). The extent of the breeding season diminishes with increase of latitude; thus at the equator ewes may breed at any time of year. - In sheep, estrous cycle length ranged from 15–19 days (average 17 days) and the estrus duration ranged from 24 to 36 hours. Detection of estrus in ewes: - It is better to use ram to detect estrus in sheep. The ram nuzzle the genital region of the ewe and showing a particular form of behavior known as the ‗flehmen response‘. This behavior enables the vomeronasal organ, located in the roof of the mouth, to examine pheromones. The ‗flehmen response‘ is characterized by the ram elevating its head and curling its upper lip; this closes the nostrils and enables fluids to be aspirated into the vomeronasal organ, via the nasopalatine duct, where they can be analyzed by the organ‘s sensory neurons. Similar activities are found with the bull and stallion and, less commonly, with the boar. Estrous cycle in goats (doe or nanny): - The breeding season in northern hemisphere is from August to February with the greatest activity in the months of October, November and December (decreasing day length). Near the equator, there is no evidence of seasonality but continuous cyclic activity. The doe is polyestrous, with an interestrus interval of 20–21 days, although it is rather irregular at the beginning of the breeding season. The duration of estrus is 36–48 hours, with ovulation occurring 12–36 hours after the onset. - The detection of heat in a doe is difficult in the absence of a male goat. The vulva shows some evidence of edema and hyperemia; the tail is wagged from side to side and up and down. The doe is restless and more vocal, has a reduced appetite and milk yield, and shows mounting behavior. The presence of the pheromones from the male goat will often intensify the signs. Signs of estrus in goats: Cervical mucus discharge, which causes hairs to – red colored vulva stick together ------------------------------------------ --------------------------------------- 34 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) FOLLICULAR WAVE (estrous cycle) IN SHE CAMEL - The camel is a most valuable animal which contributes effectively to the welfare of people in harsh and difficult environments. - Although reproduction is the key to improve livestock performance, in camel, reproduction is not as well understood as in other species of domestic animals. - The camel is a non-spontaneous ovulating animal (induced ovulatory) and in this respect it is incorrect to apply the term "estrous cycle". - Camel is a seasonal breeder (In northern hemisphere from November to April, while in southern hemisphere from May to October), - The ovarian activity during the "cycle" is follicular. Graafian follicle(s) will develop in one or both ovaries reaching a mature size. If no copulation occurs it will regress and then another follicle will start developing. Usually the left and right ovaries function equally and keep alternating in activity throughout the breeding season. It is more appropriate to refer to such pattern of activity in the ovary as follicular wave. This could be divided into four phases: a- The growing follicular stage, equivalent to prooestrus. b- Mature follicular stage, equivalent to estrus or heat. c- Atretic follicular stage if mating did not occur. d- The non-follicular stage. Duration of the follicular wave (estrous cycle): - The duration of the follicular wave (the estrous cycle) varies slightly in relation to geographical locations. In India the mean length of the cycle was 23.4 days. In Egypt it was 24.2 days and in Sudan it was 28 days. Estrus lasts about 4 to 6 days in general. The prooestrous phase lasts about 6 days and the atretic phase lasts 7 to 10 days. - In the bactrian camel the duration of the cycle averages 19 days (range 14 to 24 days), the prooestrous or developing phase lasts 5 days. What causes ovulation in she camel? - Ovulation in the camel occurs normally after the coitus and in this respect it resembles the cat and the rabbit. It is known that in these animals the neuroendocrine reflex involving the initiation of luteinizing hormone release is delayed until coitus occurs. - Ovulation occurs 32 to 40 hours after copulation under the influence of luteinizing hormone (LH). - Manual stimulation of the cervix for 15 minutes in the camel did not induce ovulation but cause only partial luteinisation of the Graafian follicle. - Bactrian camel revealed that ovulation is induced by a factor or factors in seminal plasma. Furthermore, ovulation could be stimulated by either intrauterine or intramuscular injection of seminal plasma. The intervals from the intramuscular injection to the subsequent luteinizing hormone (LH) peak in peripheral blood, and to ovulation, were the same as those that follow natural mating. It is concluded that there is a gonadotrophin-releasing (GnRH)-like hormone in seminal plasma. ------------------------------------------ --------------------------------------- 35 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) - Intramuscular administration or luteinizing hormone (LH), human chorionic gonadotrophone (hCG) and Gonadotropin-releasing hormone (GnRH) produced ovulation in a similar manner to that of seminal plasma. Proposed model hypothesis for kisspeptin (Kp) neurons involvement in the β nerve growth factor (β-NGF). initiation of the preovulatory luteinizing hormone (LH) surge and induced ovulation following mating in camelids. The figure shows how Kp neurons may act as central processors for relaying β-NGF/matting signals to gonadotropin-releasing hormone (GnRH) neurons activation. Following coitus, the β- NGF of the seminal plasma is absorbed through the genital mucosa within the general circulation and conveyed toward the central nervous system. β-NGF crosses the blood–brain barrier (BBB) to reach the hypothalamus at the level of preoptic area (POA) and arcuate nucleus (ARC) where it binds the tyrosine kinase receptor (TrkA) receptor expressed by Kp neurons. By activating Kp neurons, β-NGF induces a ------------------------------------------ --------------------------------------- 36 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) sequence of neuroendocrine events leading to the ovulation: Kp released at axonal ends binds the G-protein coupled receptors (GPR54) located on the GnRH neurons and stimulate the release of GnRH. The GnRH on the other hand stimulates the release of a pulse of LH, which leads to the rupture of the dominant follicle and thus induced the ovulation. In such seasonal breeders, Kp would be also responsible, as for other species, for the seasonal stimulating of the neuroendocrine reproductive axis. The known steroids positive feedback responsible of ovulation in spontaneous ovulators is lacking in induced (reflex) ovulators. Estradiol would be responsible of a tonic effect while the β-NGF exerts a surge effect. Signs of estrus in she camel: - The signs of estrus are restlessness, bleating, vulval swelling and mucous vaginal discharge. The female camel urinates and moves its tail up and down in rapid succession on the approach of the male, or when hearing the gurgling voice of the rutting male. - At range, the rutting male pursues the estrous female, presses his head on her neck and induces her to sit down. The male then mounts in a squatting posture. Copulation lasts from 8 to 15 minutes; it is accompanied by much male oral frothing and gurgling, with intermittent Protrusion of the Squatting posture protrusion of the soft palate, and soft palate by female bleating. N.B: in Saudi Arabia, well-fed camels will cycle and breed throughout the year, but in case of food shortage there is a decrease in breeding activity. ------------------------------------------ --------------------------------------- 37 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) ESTROUS CYCLE OF THE BITCH - Bitch is mono-estrous (come into heat once, then exhibit an anestrus period). i.e. All bitches have a prolonged period of anestrus or sexual quiescence between successive heats irrespective of whether they have been pregnant or not; this pattern has been described as monocyclic The estrous cycle is traditionally divided into four phases. Proestrus: - The bitch has a true proestrus (9 days) characterized by the presence of vulval edema, swelling and a sanguineous discharge. Some bitches show no evidence of discharge as they are continually cleaning the perineum. The bitch is attractive to males but will not accept the male. Estrus and fertilization period: - The bitch will accept the male (estrus phase = 9 days). The vulva becomes less edematous and the vulval discharge becomes clearer, less sanguineous and less copious. The duration of proestrus and estrus combined as mentioned is about 18 days, i.e. 9 days each. However, this can be very variable, some bitches showing very little sign of proestrus before they will accept and stand for the dog producing a copious sanguineous discharge during true estrus. Ovulation usually occurs 1 or 2 days after the onset of estrus or 2 days after LH surge. In the bitch the ova are not mature and capable of being fertilized until an additional 2 days after ovulation. The mature ova live another 1 to 3 days. Counting the LH surge as day 0, the bitch's peak fertile period is days 4, 5 and 6 post the LH surge. N.B: Crystallization of mucus collected from the anterior vagina has been described in the bitch, occurring after the peak in plasma estrogen concentrations. Assessment of the mucus, which originates from cervical glandular tissue may be useful when combined with vaginal cytology for determining the optimal mating time. Photomicrograph of the cervicovaginal fluid collected from a bitch during estrus. Crystallization of the mucus has resulted in the formation of a fern-like pattern. Metestrus: - This stage starts when the bitch ceases to accept the dog; however, there is dispute about its duration. Some consider that it ends when the corpora lutea have regressed at 70–80 days whilst others measure it in relation to the time taken for repair of the endometrium, 130–140 days. Anestrus: - At the end of metestrus the bitch passes into a period of anestrus without any external signs. The same is also true after parturition following a normal pregnancy. This phase lasts about 3 months before the bitch returns to proestrus. ------------------------------------------ --------------------------------------- 38 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Copulation in bitch: - Bitch stands in the mating position with her tail slightly raised or held to one side. She remains still while the male mounts and copulates. - In the later stages of the copulatory tie, which occupies from 15 to 25 minutes, she becomes restless and irritable and her attempt to free herself may cause the male considerable physical embarrassment. - After the first 2 days of estrus, sexual desire gradually decreases, but with the continued persuasion of the male she will accept coitus until the end of the period. - Bleeding, although reduced in amount, generally continues well into estrus. More often, however, the discharge becomes yellow as estrus proceeds. During the course of estrus the enlarged labia become softer in consistency. Some labial swelling continues into the first part of the metestrus phase. First stage coitus The turn Second stage coitus (Copulatory tie) Hormonal changes during estrous cycle in bitches and LH surge - In bitches, estradiol-17beta has a positive feedback effect on the occurrence of LH surge as other animal species. However in bitches, the start of the pre-ovulatory LH surge is associated with an increase in the plasma progesterone concentration which is different from other species. The source of progesterone is luteinization of follicles before ovulation. Progesterone level in bitches is used to expect ovulation timing. Ovulation timing in bitches: - Determining the actual day of the LH surge requires many blood testing which many be not practical. The LH surge causes the follicular cells to convert from secreting estrogen to secreting progesterone. The time of the LH surge can be estimated by monitoring progesterone serum levels. - In the bitch progesterone levels are baseline prior to the LH surge (typically < 1.0 ng/ml). Corresponding to the beginning of the LH surge serum progesterone levels will increase (initial rise) to a typical range of 1.5 to 2.0 ng/ml. After the LH surge, progesterone will continue to rise, it usually rises to a level > 5 ng/ml by day 2 to day 4 post the LH surge and continue to rise to >10-15 ng/ml by the end of the peak fertile period. The initial rise in progesterone correlates closely to the LH surge and can be used reliably to expect ovulation and plan breedings during the bitch's peak fertile period. ------------------------------------------ --------------------------------------- 39 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) Canine Vaginal Cytology (smear): Technique: The sample should be collected from the cranial vagina because cells from the clitoral fossa, vestibule or urethral papilla are not as indicative of the stage of the cycle and provide confusing results. Moisten a cotton swab with few drops sterile saline. Open the vulvar lips, insert the swab dorsally into anterior vagina. If you do not pass the swab far enough, you will get vestibular cells and result in false cornification. If you pass the swab too ventral, you may enter the bladder and get a falsely non-cornified smear. The swab is gently rolled across a microscope slide in two to three rows. The slide is dried and then stained with a modified Wright-Giemsa stain. You may also use new methylene blue stain. Read the slide under low power first to establish the trend of cellularity and cell types. Move to a higher power to establish the cell types. View several fields to get an overall visual idea of the percentage of cornified cells. Technique of preparing a vaginal smear in bitches How do I interpret vaginal cytology in bitches? - The healthy epithelial cells that line the vagina at all times are termed non–cornified and include the parabasal and intermediate cells. - Under the influence of estrogen, these cells are stimulated to divide. As the cells divide and the vaginal lining thickens, the cells nearest the lumen become nonviable and lose the characteristic appearance of the healthy cell. The misshapen, clumped cells are termed cornified. ------------------------------------------ --------------------------------------- 40 Prof. Dr. Mohamed Sabry Medan https://msmedan.wixsite.com/mohamedmedan Gynecology: 4th year (2019-2020) (Hormones-Estrus and Ultrasound) - During proestrus, the cell population changes from completely non-cornified in early proestrus to completely cornified in late proestrus. Red blood cells (RBCs) may be present throughout. - Estrus is also called standing heat. During this stage, the epithelium is at maximum thickness, with a prominent layer of keratinized cells on the luminal surface. Estrus is defined cytologically as 100% cornified cells with greater than 50% being anuclear squame cells. - Anestrus is characterized by presence of few cells, all of which are non-cornified. Illustration of the hormonal events during the estrous cycle of the bitch. The vaginal epithelial cells seen during proestrus are also shown. £2, Estradiol; LH, luteinizing hormone; P4, progesterone; Ovut, ovulation. Pseudopregnancy (Pseudocyesis) in bitches: - During estrus, the surge in LH that results in ovulation also stimulates the ruptured follicles, from which the ova are released, to secrete progesterone. At this point the follicular cells become the corpus luteum. If the bitch becomes pregn

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