Clinical use of hormone status determination in canine and feline reproduction Suppawiwat Ponglowhapan, DVM (Hons), M.S., M.Sc, PhD Department of Obstetrics Gynaecology and Reproduction Faculty of Veterinary Science, Chulalongkorn University

Introduction Hormone  Hormaein (ภาษากรีก) = Impetus (a moving force, stimulus) แปลว่า กระตุ้น (excite), ก่อกวน (stir up) หรือทำให้เกิดการเปลี่ยนแปลง (to set in motion) Endocrine The endocrine system is a group of glands that work together and secrete many types of different hormones that regulate the body.

Introduction Major endocrine glands. (Male on the left, female on the right.) 1. Pineal gland 2. Pituitary gland 3. Thyroid gland 4. Thymus 5. Adrenal gland 6. Pancreas 7. Ovary 8. Testes

Introduction การควบคุมการทำงาน Autocrine Paracrine Endocrine

Introduction Hormone feedback regulatory system Negative feedback Positive feedback Physiological mechanisms of endocrine Pathologic mechanisms of endocrine Hormone excess (tumor, exogenous route) Hormone deficiency (autoimmune, surgery, inflammation) Hormone resistant (genetic factor)

Introduction สามารถแบ่งฮอร์โมนออกเป็น 5 ชนิด Amino acid derivatives; dopamine, catecholamines, thyroid hormone Small neuropeptides; GnRH, TRH, somatostatin, vasopression Large proteins; LH, FSH, insulin, parathyroid hormone Steroid hormones; oestrogen, progesterone, testosterone, cortisol Vitamin derivatives; retinoids

Introduction หน้าที่หลักของฮอร์โมน Growth and differentiation Maintenance of homeostasis Reproduction

Introduction สามารถแบ่งฮอร์โมนออกเป็น 5 ชนิด Amino acid derivatives; dopamine, catecholamines, thyroid hormone Small neuropeptides; GnRH, TRH, somatostatin, vasopression Large proteins; LH, FSH, thyroid hormone, insulin Steroid hormones; oestrogen, progesterone, testosterone,cortisol Vitamin derivatives; retinoids

Reproduction Progesterone Oestrogen Testosterone Gonadotrophins: LH & FSH Thyroid hormone

Progesterone Definitions 1. A female hormone and the principal progestational hormone that is made mainly by the corpus luteum in the ovary and by the placenta. Progesterone prepares the lining (endometrium) of the uterus to receive and sustain the fertilized egg and so permits pregnancy. 2. The principal progestational hormone liberated by the corpus luteum, adrenal cortex, and placenta, whose function is to prepare the uterus for the reception and development of the fertilized oocyte by inducing transformation of the endometrium from the proliferative to the secretory stage.

Progesterone A sex steroid hormone CL, adrenal gland, placenta (cat) Pregnancy, mammary gland development

Progesterone ประโยชน์ของการตรวจโปรเจสเตอโรนในสุนัข เพื่อกำหนดวันผสม (optimal breeding time) เพิ่มโอกาสผสมติด เพิ่มจำนวนลูกต่อครอก เพื่อตรวจความผิดปกติของการตั้งท้อง เพื่อทำนายวันคลอด

Determination of optimal breeding time in the dog

Luteinization of the follicle in the dog

Guidelines for interpretation of serum progesterone concentration in the estrous bitch measured by RIA Serum Progesterone (ng/mL) Estimated day of ovulation Best breeding days Estimated parturition from ovulation days 1.0-1.9 + 2 days + 4 days (3-6) + 63 days (62-64) 2.0-3.9 + 1 days + 3 days (2-5) 4.0-10.0 0 days + 2 days (1-4) For natural mating or AI using fresh semen, inseminate the day after ovulation For AI using chilled or frozen semen, inseminate 2-4 days after ovulation

Predicted time of ovulation based on serum progesterone (n = 50; > 20 breeds) Key points Traditional or classical dates for canine breeding: No one-for-all rule Importance of determining serum progesterone level for canine breeding

Determination of optimal breeding time in the dog

Determination of optimal breeding time in the dog LH and ovulation? Why LH check is less likely to be recommended in practice? Specificity of LH test Lab Availability :Thailand?

Determination of optimal breeding time in the dog Is there any in-house LH test kit? YES

Determination of optimal breeding time in the dog Some considerations on the clinical use of LH test kit Advantages Price ($230 per box/ 6 tests) The number of cases Frequency of the use/ cycle

Determination of optimal breeding time in the dog Unlike other mammals, dogs ovulate when the oestrogen level is declining. Oestrogen levels can only give us a general idea of when a dog will come into cycle.

Vaginal cytology in the cat

Determination of optimal breeding time in the dog Oestrogen and vaginal cytology? basal/ parabasal cell intermediate cell superficial cell

Determination of optimal breeding time in the dog The use of vaginal cytology for canine breeding management To ensure a dog is in cycle When to start vaginal swab? How often? When to start collecting blood sample for progesterone assay?

(previous oestrus, first day of vaginal bleeding: Day0) Physical exam Recommended workup for determination of optimal breeding time in the bitch History taking (previous oestrus, first day of vaginal bleeding: Day0) Physical exam (General and reproductive exam; vaginal discharge, vulvar swelling) Estrous behaviors Vaginal smear to confirm the bitch is in estrous stage Blood collection for progesterone measurement

Guidelines for interpretation of serum progesterone concentration in the estrous bitch Serum Progesterone (ng/mL) Estimated day of ovulation Best breeding days Estimated parturition from ovulation days 1.0-1.9 + 2 days + 4 days (3-6) + 63 days (62-64) 2.0-3.9 + 1 days + 3 days (2-5) 4.0-10.0 0 days + 2 days (1-4) Key Notes Stress and progesterone hypersecretion Combination of behaviour, vaginal cytology and progesterone Delivery of live puppies can not be guaranteed.

Progesterone ประโยชน์ของการตรวจโปรเจสเตอโรนในสุนัข เพื่อกำหนดวันผสม (optimal breeding time) เพิ่มโอกาสผสมติด เพิ่มจำนวนลูกต่อครอก เพื่อตรวจความผิดปกติของการตั้งท้อง เพื่อทำนายวันคลอด

Pregnancy monitoring

Hypoluteoidism in the bitch Hypoluteoidism = luteal insufficiency Progesterone levels > 2.0-2.5 ng/mL are necessary to support pregnancy. Incidence? Cause? Insufficient relaxin low prolactin low progesterone Endometrial infection/inflammation PGF2 alpha luteolysis

Hypoluteoidism in the bitch Treatment Medroxyprogesterone acetate (MPA) 0.1 mg/kg PO sid Altrenogest 0.088 mg/kg PO sid Progesterone in oil 1-2 mg/kg IM every other day NOTE Discontinued 2-3 days prior to the expected due date

Progesterone ประโยชน์ของการตรวจโปรเจสเตอโรนในสุนัข เพื่อกำหนดวันผสม (optimal breeding time) เพิ่มโอกาสผสมติด เพิ่มจำนวนลูกต่อครอก เพื่อตรวจความผิดปกติของการตั้งท้อง เพื่อทำนายวันคลอด

Prediction of parturition date

กำไร 3 Oct 2009 Golden Retriever, 2 yrs 54 days of pregnancy Severe bloody vaginal discharge General health status is good No signs of parturition X-ray: 3 fetal skeletons (not yet full-term) U/S: one fetus with slow heat beat (< 180 bpm) Predicted parturition date by U/S (biparietal diameters) = -7 days

กำไร 6 Oct 2009 Abortion: one fetus Progesterone level 1.9 ng/mL 7 Oct 2009 at night Deliver 2 living puppies

Oestrogen and testosterone Research purpose Clinical use is for diagnosis of the presence of gonadal tissue by GnRH stimulating test Ovarian remnant syndrome oestrogen Bilateral abdominal cryptorchidism testosterone

Ovarian remnant syndrome How to diagnose wait till the bitch/queen shows oestrus signs GnRH stimulation test Adverse effects Owner Animal

Bilateral abdominal cryptorchidism How to diagnose Dog: GnRH stimulation test Cat: penile spine Adverse effects Testicular tumour (high incidence) Before castration After castration Testicular tumour

Ovarian remnant syndrome Exner et al., 2008. Theriogenology. Estradiol measurement after GnRH stimulation as a method to diagnose the presence of ovaries in the female domestic cats GnRH analogue buserelin (Receptal, Intervet) 0.4 microgram/kg IM