----start---- repro 9/18 turner mares alternate between estrus and diestrus. this process is referred to as 'cycling' [slide of mare on bicycle] Today we're going to discuss the estrus cycle. NOT in as much depth as we did in physiology. Also we'll go into how to manipulate the cycle with various hormones. this is something we do a lot to try and improve breeding efficiency. this lecture will be sort of an overview, prior to the species specific lectures. ok? you have to learn the other stuff for each species, separately. So, you have the estrus cycle. all species cycle b/w estrus and diestrus, though the pattern varies. Males do not cycle. this is due to the Y chromosome. early in embryogenesis the Y chromosome does something to the fetus to prevent this cycling. male hormones are more stable. So there are phases of the estrus cycle that are described based on ovarian structure or hormones. So, you have follicular/luteal phases - follicular is when the follicle is dominant in the ovary (litter bearing species have multiple dominant follicles at once) and estrogen predominates, a corpus luteum is absent. During the luteal phase, the corpus luteum (CL) is dominant. Progesterone is dominant. There are some follicles present during diestrus too, but the animal isn't in heat. The mature CL will always override any follicular effects. another way to describe the estrus cycle is based on the hormonal environment - proestrus, estrus, metestrus, diestrus, and anestrus. proestrus: early follicular growth. CL is regressing, progesterone levels declining, estrogen levels rising, follicles starting to grow. in domestic animals, proestrus is best demonstrated by the bitch, who has a long proestrus, and obvious behavior - considering a date with male, but far from commitment. may flag tail a bit, flirt, etc, but ultimately refuses the male. estrus: slide of mini-horses (Kewpie Doll and Rowdy Commander) mating. a female in estrus is in heat, shows heat behavior, will permit breeding. dominant hormone is estrogen, progesterone levels are low. metestrus: not well defined. estrogen is declining, following ovulation, progesterone is rising as CL develops. in cow we may see metestral bleeding due to diapedesis of RBCs across uterine lining in response to something or other i didn't hear. when you see this, cow has already ovulated and it is too late to breed. diestrus: CL is predominating. Progesterone is high. estrogen is low. she will actively reject the male. anestrus: this is when all hormones are at baseline levels - there is no cycling. not all species have this. some do. mare will only cycle during long days - spring and summer. in winter, all ovarian activity shuts down and she's in anestrus. also in queen, ewe, doe. sometimes seen with lactation esp in beef cattle and sometimes you see pathologic anestrus in thin or sick animals. patterns of cyclicity: pigs are polyestrus. that's what we'll discuss first polyestrus: cycle year round. no periods of anestrus unless pregnant or lactating. see this pattern in sow and cow. estrus, diestrus, estrus, diestrus... seasonal polyestrus: mare, ewe, doe - cycle as above during certain seasons. usually evolutionary adaptation to ensure young are born in the spring. so mares cycle during long days, foal 11 mos later in spring. sheep/goat, cycle in fall/winter due to shorter gestation, so they also have young in spring. sheep/goat anestrus during spring and summer. monestrus: bitch. one single estrus cycle and then anestrus after each cycle. period of anestrus varies - about 6 mos, give or take a lot. induced ovulators: effecient breeders. cat, llama. these animals recruit waves of follicles, but do not ovulate or enter diestrus unless they are bred by a male. diestrus is much longer than estrus. if you haven't been bred, no need to waste time in diestrus, right? So they bring along wave after wave of follicles - acting like they are always in heat - they seem to be in heat all the time - only ovulate if bred - then enter diestrus. now, cat is a seasonal induced ovulator- only cycles in long days (spring/summer). nonseasonal induced ovulators like llama, lab rabbit, will do this year round. you would not expect to find a CL on ovary of induced ovulator unless she had been bred. Follicular maturation: slide of clam egg and clam sperm on it (wow.) The follicle contains the egg within the follicular fluid. The normal fate of most follicles is atresia. only about 1 in 400 actually get to ovulate. The rest end up just regressing. Only the ones that come along in the right hormonal environment get to actually ovulate. Mechanisms of choosing which ones do what aren't fully understood. Atresia is marked by pyknosis of follicular cells, decreased FSH and LH receptors on the surface of the follicular wall. mature follicles also make a lot of estrogen; atretic ones do not. also there are changes in microvasculature of follicle. not sure if this is cause or effect of atresia. but clinically, we look at size of follicle and if CL is present. if CL is there, follicle will not ovulate. if no CL and really big follicle, that follicle will likely ovulate. slide: cow looking at us :) in induced ovulators no CL forms until breeding, so follicles keep coming and going. in others, follicles still keep coming but don't get that big when progesterone is high - won't ovulate til CL goes away. it's believed that inhibin, made by preovulatory follicles, feeds back to reduce FSH production so other follicles will regress. u/s of mare ovary: the dense white structure is the CL. the clear black areas are antral follicles that will not ovulate. this is a diestrus ovary, animal not in heat. slide:mare's preovulatory follicle on U/S. no CL, big giant black follicle about 5.2 cm across. very large. this mare would have high estrogen, low progesterone, would be in heat. review of basic estrus cycle: using a sheep model - the ewe has most standard estrus cycle 1. proestrus - early growing follicle, not mature. Progesterone levels are in decline - progesterone inhibits FSH and LH release, so as progesterone declines, you start to get more FSH and slowly more LH. in response to FSH, follicles start to grow. 2. FSH binds receptors on follicle and follicle grows, antrum gets bigger, fluid increases, and steroids increase - estrogen levels rise. now you have preovulatory follicle. progesterone levels are low, LH increases. this is estrus. early on, estrogen kind of holds LH in check so it doesn't really surge, but when estrogen gets really high, it becomes a positive feedback signal and there is an LH surge 3. LH surge signals follicle to release oocyte - this is ovulation. follicle ruptures, oocyte enters oviduct, passes down into ampulla and hopes to meet a sperm. 4. after follicle rupture, the follicular wall cells change from granulosa and theca cells into luteal cells. under influence of LH, they stop making estradiol and start making progesterone. the cavity fills in with blood. they call this the corpus hemorrhagicum, or CH. it's like a jelly-blood-clot on the ovary, you can kind of palpate this. it's painful on the mare's ovary, and indicates ovulation w/in past few hours. now estrogen is declining and progesterone rises. metestrus is the phase. 5. now, you have the CL. a large structure on ovary, producing progesterone. this is diestrus. FSH and LH levels are baseline b/c progesterone represses their secretion. if animal is pregnant she will maintain high progesterone - embryo takes care of that. if not, she needs to go back to estrus. in most spp but not bitch and queen, after a few weeks, if nont pregnant, uterus sends signal to ovary to tell it to get rid of CL- signal is PGF2a - luteolytic hormone. this lyses CL, progesterone levels decline 6. regressing CL - FSH and LH start to increase, follicles start to grow, we start over. The Perils of Priscilla Pig: facts about pigs: a young female swine that has never been bred is a gilt. Gilts are raised in a large group with other gilts and enter early puberty and have enhanced heat behavior (standing still) b/c of this. a sow not in heat will scream and yell if boar tries to mount her. boars have corkscrew penis and will secrete smelly boar saliva and aroma when they approach sow. this also enhances heat behavior of sow. you can buy boar aroma in a can. things to enhance early onset of puberty in gilts is raising them in a group, and stressing them out. boars are also usually raised in groups for the same reason. ok, so once upon a time was a pig named priscilla who turned 6 mos old (puberty age) and all her friends were boar crazy...but priscilla was a loner, had a gilt-complex, had been raised alone and didn't want to seek a boar, was very sad and alone. that is part one of the story. more later. Slide: big horse and little horse. one was the biggest draft horse ever. the other is a tiny little mini horse. they just wanted to take pictures of these stallions next to each other. but the mini stallion wanted to kill the big one :) ok, we now enter a very boring part of the lecture....but it is review from physiology so you may know it. [as if.] you need to understand follicle physiology to know how to manipulate these things. you start with small antral follicle in ovary, with granulosa and theca cells in the wall. there are FSH receptors on it, but just a few. inside is a tiny little egg, surrounded by cumulous cells, attached to wall by a stigma (stalk). so, if hormones are right, FSH levels are rising, and FSH binds receptors, causing follicular cells to divide, and multiply, and to start making low levels of estrogen. the estrogen acts on the follicle and increases FSH receptors, and starts to induce LH receptors. so in repsonse to low levels of estradiol and FSH you upregulate LH and FSH receptors. now LH and FSH bind their receptors. in response to this binding, esp of LH, you get big increase of estrogen production, and big growth of follicle. estradiol now reaches threshold level causing LH surge and ovulation. key points: youneed time to develop receptors before follicle can respond to FSH or LH. if you randomly inject FSH or LH, you won't do anythig b/c the receptors aren't there, unless you did it at the right time. FSH works with estrogen to increase LH receptors. binding of LH receptors drives steroidogenesis and follicle maturation, and the estrogen finally causes LH surge and ovulation. GnRH from hypothalamus tells pituitary to make FSH and LH which cause estrogen, progesterone and inhibin production. inhibin feeds back to stop FSH and LH production. also i think inhibits GnRH? sorry, I missed that. egg is in prophase of meiosis I. remember meiosis starts in embryo and then stays in prophase of meiosis I until it's in the follicle - first meiotic division occurs just prior to ovulation - ovulated oocyte is in meiosis II except in the bitch. bitch ovulates a primary oocyte and needs a little time for that to mature. meiosis II isn't completed until after sperm binds egg. actual mechanism of ovulation- not well known. LH surge is key, but also prostaglandins are involved. follicle slowly collapses, egg is trapped by infundibulum, goes down oviduct, waits. may sit there a while, or eventually pass through to uterus, depends. after ovulation, empty follicle fills with blood, forms CH, cells luteinize, form CL, make progesterone, diestrus occurs. so when you think of this stuff, both LH and FSH are required. FSH has early differentiating function by causing early steroid production and upregulating LH receptors. LH sustains final growth, really causes estrogen production, and surge causes ovulation. there is a lot more to it but this is the basic stuff. The oocyte in meiosis I - we think the granulosa and theca cells release cAMP which holds oocyte frozen in meiosis I, and then when egg ruptures off of stigma, cAMP levels fall off and meiosis starts up again. so it is when egg is starting to be released that this happens. effects of steroids fertilization - sperm meets egg, takes of acrosomal cap, gets hyperactivated, and gloms onto the egg. Priscilla the pig part II Oscar Meyer the boar lived near her. he grew up in a big family. he saw priscilla and went hog wild. he knew young sows would forget her gilty feelings if taken for a ride in a truck....so he borrowed dad's truck... ---break--- p 5 of handout effects of major steroids - you think of an anabolic effect, building up muscle mass, etc, when you think of steroids, but we'll see. estrogen (mainly 17 estradiol) - theca and granulosa interact to make estradiol17b. theca make androstenedione and sometimes testosterone, and these go to granulosa which finish it into estrogen. mainly it acts on CNS to induce estrus behavior in the abscence of progesterone. it also does physiological things to prepare for reciept of sperm - cervix relaxes, vulva relaxes. in sow, you see vulvar relaxation - pink, swollen, long vulva is seen. also increases myometrial tone, stimulates endometrial gland development, increases uterine circulation and primes uterine immune system. these effects are all generally blocked by progesterone. progesterone - progestational hormone. prepares tract to nurture embryo. causes rejection of male, diestrus behavior. overrides effects of estrogen when both hormones are present. quiets the myometrium, makes it tolerant of strange things. creates immunotolerant environment. kicks up growth of endometrial glands another notch. closes cervix and keeps it closed. decreases immunocompetence of uterus. stimulates mammary gland developement. ways to manipulate estrus cycle with hormones: there are a limited number of pharmacologic agents used by repro people. you use them a lot though. 1. prostoglandin F2a (PGF2a) aka Lutalyse, prosta, etc. this acts to lyse the CL, causing decline in progesterone, and onset of another estrus. HOWEVER it only works when CL is mature - it must have PGF2a receptors. this is after day 5 post ovulation in mare, or day 7 post ovulation in cow. if you give it to a cow 3 days after ovulation, nothing happens. if you give it at day 7, you will lyse CL, progesterone levels will drop, you short cycle her and bring her back into heat soon. in bitch, works after about day 5; usually requires multiple doses though. not effective til about day 12 in the sow and since her cycle isn't really much longer than that anyway, we tend not to use this in pigs. when do you use PGF2a? To terminate pregnancy early in species dependent on CL for gestation. Doe, bitch, queen - pregnancy depends on CL and progesterone, so you can end pregnancy this way. tx of choice to end pregnancy in bitch. people used to use estrogen for this - delays tubal transport of egg - so if you give animal estrogen it takes too long for egg to get to uterus - but there were too many side effects. in large animals, PGF2a is used most often for "short cycling", early return to estrus- about 3 days after shot, animal will come back into heat. this won't work in a bitch. if she's in diestrus and you give her PGF2a, you'll lose the CL and drop progesterone level, but then she'll enter anestrus, not go back into estrus - because of that monestrus thing she does. there is always anestrus after estrus. you can use it to synchronize cycles for AI or embryo transfer herds so they are all in heat at the same time. usually you do this with a two dose regimen. you take the animals and about 70% have a CL at that time. give all of them a shot. those that have a CL will respond to the shot; the ones that don't will not. then you wait 10 days in cow, longer in mare. the responders will now have another mature CL; the nonresponders will now have a CL. so now they all have a mature CL and you give a shot, and you synchronize everyone, and they all come into heat at about the same time. in mare, you wait about 12 days in between b/c estrus is longer. they do this in dairy operations too. also, to tx pyometra in spp in which CL is required for definition of pyometra. to correct that often if you lyse the CL, progesterone drops, uterine immune system improves, tone improves, and animal can clear the infection. or you can use it with abx. can use this in the bitch and the cow. what if you do not feel a CL but cow has pus in uterus? Some think the PGF2a will increase uterine tone and maybe help some, and it probably doesn't hurt, so it might be worth a try. 2. LH luteinizing hormone. hard to get in pure form, so we use stuff with LH like activity - "Follutein" - HCG, human chorionic gonadotropin. You could also use GnRH which stimulates pituitary to make LH and FSH. Both GnRH and HCG act like LH, causing final maturation and ovulation of mature follicles. so you can use these to induce ovulation in animal with mature follicles. mare - we use it a lot to time ovulation. mares have a long estrus, about 5-7 days of heat. that is a long time. we'd like to know when exactly she will ovulate - if you are using shipped semen, and FedEx brings it Tuesday, you'd like her to ovulate at the right time. so if she's in heat, and has a follicle that is relatively mature and has some LH receptors, and you induce LH surge with HCG, you can make her ovulate within 36 hrs of injection. repeat u/s at 36 hrs shows a CL present. Also, in fall, when she's getting ready to enter anestrus, you can induce ovulation of a reluctant follicle cow- commonly gets follicular cyst that never ovulates. she can't get pregnant b/c follicle won't ovulate so to induce luteinization you can give GnRH or HCG to cause it and then you can short cycle her and bring her back into heat. queen - if you have one in heat anddon't wanna breed or spay her and she's going nuts, you can give a shot of HCG or GnRH to simulate the LH surge and cause ovulation. tricks her into thinking she's been bred. be careful though, b/c you cause a rise in progesterone and risk pyometra. infusions of it can be used to induce ovulation in anestrus animals. also you can use it to improve conception rates in "problem breeder cows." back to priscilla and oscar - oscar was wearing his BoarMate cologne and had the truck. priscilla was nervous. oscar took out a bottle of wine andwhipped out his corkscrew. she wouldn't stand for it. He asked if she'd go for a ride. she jumped in the car. he drove really fast. she got stressed, released cortisol, smelled his cologne, lost gilty feelings, rooted him on to go faster and faster as she squealed with delight, having fallen whole hog in love ;) 3. FSH: recruits follicles and induces early follicular development. given when progesterone is declining, in proestrus, you can induce many follicles to develop. pure FSH is expensive, so we use FSH like hormones like PMSG (pregnant mare serum gonadotropin)(hard to get in US except as PG600 for swine) - it causes follicular development in some spp - not mare, but works in cow =- can induce superovulation using it. mainly for purposes of embryo transfer. wno't work in anestrus cow, or cow in middle of diestrus. have to do it in the right part of the cycle. results vary - a few follicles to 20 or more. and you only do it a few times a year, not every cycle. PG600 - Born to be Bred - tattoo on sow on motorcycle in the PG600 ad :) using this in sows will induce earlier onset of estrus in postweaning sows, and may increase ovulation rates as well. also we use this in sheep, esp as people start using AI in sheep. we use PGF2a to synchronize estrus, then use PG600 to superovulate them into ovulating 2,3, or 4 follicles instead of 1. btw, remember you have to give stuff at the right time of cycle! People like to give HCG to fillies - any time a filly misbehaves they say she is in heat and give her a big dose of HCG. but most of the time she wasn't even in heat and the HCG is just being wasted. 4. Progesterone: slide of bovine ovary with mature CL secreting progesterone. we also see an older regressed CL, and a CA - corpus albicans - an old scar left on ovary from an old CL. If you think about the effects of progesterone - think about why you might use it clinically. there are many forms- injectable that you give daily; Regu-mate, an oral progestogen with progesterone like activity; there used to be repositol forms that lasted a month, no longer available; implants available used in cattle - like a capsule you put under the skin of the ear. you can use it to synchronize estrus - it acts on hypothalamus to suppress FSH and LH secretion. so if you give progesterone for 2 wks, you suppress FSH and LH the whole time, and when you release the inhibition there are FSH/LH surges, and all animals get synchronized - more tightly than with prostoglandin - but more expensive and takes longer. also is used to maintain pregnancy esp in mare, bitch. not proven, though. some think if you have mare or bitch that miscarries and you don't knwo why, supplemental progesterone might help. used commonly in mares. if it works, is a subject of controversy. we use it sometimes also in mares that come into high risk pregnancy program at NBC - mares with a lot of problems. here is one with pituitary adenoma - thin, weak, thick haircoat - abd u/s indicated fetus wasn't doing so well. we put these animals on progesterone because we don't know what else to do, and b/c it might work. also, if mare is severely stressed due to surgery, or has been exposed to endotoxin, progesterone supplements DO definitely help. also use for synchronization for embryo transfer b/w donor and recipient - better than PGF. and, to suppress behavioral estrus in performance mares (and performance sows :)). like, if your horse is going to Devon as a show hunter, you might put her on a progestogen to block this behavior. 5. Estrogen - estradiol - in many forms. will induce behavioral estrus in the absence of progesterone. will block tubal transport of ova, slowing it down drastically. also acts on pituitary to suppress FSH, LH release. stimulates uterine immune response, causes relaxation of cervix, increased uterine tone, and in some spp like cow is luteolytic, but luteotropic in sow. remember it will not induce a fertile estrus! in animal with low progesterone will make her act like she's in heat but she won't develop a follicle. it will stimulate uterine immune defense so used to be used to tx uterine infections but doesn't work clinically and leaves milk residues. used to use as mismate shot in bitch but not anymore b/c of risk of pyometra - causes increased progesterone receptors in myometrium, and ultimately suppresses immune response and closes cervix really tightly. again- behavioral estrus but not fertile estrus. used in jump mares. these mares have no ovaries. but you can give them estrogen and they act like they are in heat. weight gain in feedlot cattle - it is an anabolic steroid. prolonged use of estradiol in cattle can cause cystic ovaries, in reality we do not use much estradiol in the clinic these days. there are more hormones but these are the main ones. how to manipulate estrus w/o drugs - -contact with a male, esp in sheep and pigs- introducing a male can synchronize ovulation. when ewes come out of summer anestrus and it is early fall, they are getting ready to ovulate, if you introduce a ram, they all ovulate at about the same time. also in peri-pubertal gilts, they are preparing to ovulate, if you introduce a boar it synchronizes them. slide: pig testicles - these are really big. slide: the quad. contact with females can synchronize estrus cycles. this has been shown in women (who do not have estrus, but whatever). we think it is a pheromone effect. housing boars with boars or gilts with gilts can induce earlier onset of puberty. light is another way - seasonally polyestrus animals can be tricked into cycling at different times of the year by changing the light cycle. mares don't cycle during short day seasons like winter. in spring, melatonin starts to decline, mare starts to cycle. so you can put them under light for longer time than the real sun, and they will cycle. sheep/goats, need shorter day lengths. you need to lock them into dark so it is harder to trick them. but you can feed them melatonin and trick them into thinking the day is short. Priscilla pig - the conclusion. they lived happily ever after. she ovulated 15 ova on her 2ndheat. was bred twice with high quality semen with oscar. she farrowed a litter of 12 and wasn't sent for sausage. moral of the story: is that a corkscrew in your pocket, or are you just happy to see me? ----end-----