---start physio.lec.04.18.97---

the award-winning Dr Sertich...

pregnancy today, parturition next week, lactation after that.
there is a handout, but we didn't get it yet. it is quite comprehensive.

estrus cycle is a constant ongoing event that is an attempt to establish 
pregnancy. never stops until animal gets pregnant. from a farm animal 
standpoint that's kind of delightful; for us, not really, but whatever.

requirements for pregnancy: things that must happen for pregnancy to 
develop

maintenance of corpora lutea, which make progesterone. the CL is what 
stops the estrus cycle.
development of conceptus - relies on endometrial secretions for early 
nourishment
endometrial secretions
placentation - not implantation, but rather placentation, which provides 
connection between conceptus and dam for transfer of nutrients and gas 
exchange.
control of uterine blood flow - uterus gets much larger, blood flow has 
to change and give it more blood.
immunologically priveleged state - 1/2 of conceptus' DNA is different 
from dam's and she has to tolerate that foriegn body in her uterus for 
the whole time.

early events of pregnancy:

follicle develops, matures, releases egg. slide: equine oocyte with 
cumulus cells around it. this sturcture is about 125-135 microns across. 
it is released, the infundibulum accepts it and takes it down into the 
oviduct - three parts of oviduct: isthmus (narrow), ampulla (wide) and 
infundibulum. egg sits in ampulla and waits for fertilization.

if fertilization takes place we have a zygote: fertilized egg. then 
cleaving takes place at about 12 hr intervals. the first 5-7 days, the 
conceptus' total size stays the same, it just cleaves into a larger 
number of cells. so the cells are getting smaller, but more numerous. it 
develops into a morula, the raspberry type thing....in most species, when 
egg drops down into uterus, it is during morula stage. then blastulation 
takes place - it becomes hollow ball of cells filled with fluid (in 
horse, this happens on day 7). in some species eg horse, conceptus stays 
round, but in cattle, sheep, swine, blastocyst kinda hatches from zona 
pellucida, and there is a dramatic elongation of the embryo to increase 
surface area. this is in the handout.

memorize this as a vet:
gestation lengths:

cow 9 mos with some breed variation - 280 days give or take
horse 11 mos - 340 days, with great variation. really, 321-365 days. at 
NBC, they have a 403 day pregnancy right now, though. horse is quite 
variable.
sow is predictable: 3 mos, 3 weeks and 3 days, 114 days
ewe/doe 5 mos, 146 days
dogs/cats 63 days. luteal phase is also 63 days

slide: morula - ball of cells, stage where most embryos enter uterus. 
still surrounded by zona pellucida (ZP). 

in most species, ovulation occurs during estrus/heat - the period of 
receptivity where female will agree to copulate. depending how well 
defined estrus is, may count from beginning or end of estrus...there's a 
chart in the handout about this. but the cow ovulates after estrus, after 
she goes out of heat and is no longer receptive to the male. also, two of 
the common species we deal with are induced ovulators: stimulated to have 
LH peak and ovulation by cervical stimulation from copulation - these are 
the cat and the llama.

slide: 7 day equine embryo. on 7th day, embryo has just come down into 
the uterus and is very spherical and it is interesting that already, we 
can visually see difference between "horse" part and "membrane" part. the 
dark area is inner cell mass, which is the horse. the rest of trophoblast 
cells will be fetal membrane. on 7th day instead of hatching from zp, 
horse embryo zp just disappears and is replaced by a glycoprotein capsule 
unique to equids, which plays a role in keeping embryo spherical. until 
day 16, equine embryo is quite mobile within the uterus. in early 80s, a 
grad student ultrasounded a group of mares every 20 min during the first 
few weeks of gestation. found that after day 10, through day 16, the 
embryo kept moving around the uterus. he found that during that time, 
embryo would move length of 2 football fields (not sure if that's per 
day, or all week). this is embryo signalling mare to prevent luteolysis. 
any embryo has to somehow tell the dam that she's pregnant - this is 
called "maternal recognition of pregnancy". in horse, if embryo isn't 
able to touch all surfaces of endometrium, mare will come back into heat. 
this is just in equids. in cattle/sheep/swine, sometime between day 11 
and 13, blastocyst hatches from zp, rapidly elongates and becomes 
threadlike, and isn't motile in uterus, but contacts uterus via surface 
contact with this long, thin embryo. the elongation is very important. 
the surface contact prevents luteolysis. in swine, uterus is long, 
torturous. need at least four pig embryos to have enough surface contact 
to prevent luteolysis. also, embryos need to be spread through uterus, 
not in one clump.

35 day deer embryo....very small, with long slender stretched out 
membranes which stretch out to contact uterus, as above. 

in dogs, and cats, gestation is 63 days. remember, diestrus is 63 days in 
dogs/cats also. so in dogs/cats, there doesn't seem to be a difference 
between pregnant and nonpregnant progesterone state. the embryos do drop 
into uterus, move around, space themselves apart, causing localized 
bulges to appear. maternal recognition seemingly less imp't here, since 
63 days anyway...

in cow...corpus luteum forms after ovulation, and starts producing 
progesterone. the CL distorts shape of ovary, and enlarges the ovary.

normal luteolysis - the event causing destruction of CL, decreases 
secretion of progesterone, allows estrus to recurr. must be prevented if 
pregnant

cow 15 days
mare 14 
sow 13
ewe 12
dog/cat 63

(so about 2 weeks except in dog/cat)

now, while discussing luteolysis, another interesting phenomenon 
different between species, and important to understand if you want to 
understand how much drug to give to induce luteolysis...
in sheep/cows/sows, there is a very intimate relationship between ovarian 
artery and uterine vein. they are right up against each other. if dam 
isn't pregnant, secretion of PGF2a from uterus occurs. in these species, 
PGF2a is released from uterus, goes down through uterine vein, but 
because the vein is so intimate w/ovarian artery, the artery rapidly 
picks up the PGF2a, and it goes directly to ovary to cause luteolysis. 
what that means is that the CL is seeing a relatively large amt of 
prostoglandin. it gets almost all the prostoglandin the uterus releases. 
practically, this means that when you give exogenous prostoglandin to 
cow/sheep/sow you need to give a high dose. cow would get 30-35 mg. 
contrast this to the horse...

in horse, she doesn't have that anatomy. the uterine vein is quite 
separate from ovarian artery. the prostoglandin has to circulate all the 
way through the heart and back through arterial system to get to the CL. 
so by then, it's quite dilute. but mare CL is very sensitive to 
prostoglandin. relatively low amts of prostoglandin are adequate to cause 
luteolysis. so clinically, you give low amts of prostoglandin to mares. 
the same size mare compared to that cow that needs 35 mg would only need 
10 mg, about 1/3 of dose. 

in ewe, you give 15-20 mg - alot! only weighs like 150 lbs, right, so 
this is proportionally a large dose.


slide - mare ovary. in the horse - well, in all species we've talked 
about how impt the CL is. in some species, the CL may only function for 
part of gestation. in horse, ovary only needed for first 60-100 days. 
this is because the conceptus, or the placenta of conceptus, can also 
produce progesterins, maintaining the function of progesterone. so mare 
doesn't rely on CL all through pregnancy. so in some animals, could 
terminate pregnancy by giving luteolytic prostoglandin, but in mare, you 
couldn't do that, or in other similar species.

Equine chorionic gonadotropin - ECG
in mares, they have ECG elevated between day 40 and 160. is made by 
endometrial cups which only form in the mare, and they make this hormone 
that plays a role in development of secondary ovulations and secondary 
CLs.

in sheep, there is a functional CL for 60 days of gestation, a little 
less than half the full 150 days. after 60 days, the fetal membranes make 
the progesterone. so in ewe, could terminate pregnancy with prostoglandin 
only during first two mos.

but in goats, they are luteal dependent. they need the CL throughout 
gestation. 

in pigs, as far as maternal recognition of pregnancy goes, pig embryos 
make a LOT of estrogen, esp estrone sulfate, and interestingly, there is 
an increase in prostoglandins in the uterus of a sow during pregnancy. 
she has PGF2a and PGE2 elevated during pregnancy. this is quite different 
from those other species, where PGF2a will terminate pregnancy. swine 
embryos also make proteins that play a role in maternal recognition.

dogs/cats not well understood about maternal recognition; probably less 
critical since CL is maintained 63 days anyway.

re: uterine secretions during pregnancy - there are uterine glands in the 
endometrium which make "uterine milk" or "histotroph".  there is a 
variation in species about when placentation takes place. in ewe, it 
occurs at 17 days post ovulation. then, nutrient exchange occurs via 
placenta. in mare, placentation doesn't occur til about 60 days. placenta 
isn't really functioning at all for first two months! embryo relies on 
nutrients from endometrial glands during that time. in mares, there is 
common pathology called periglandular fibrosis, where scar tissue forms 
around glands. this is very highly correlated with embryonic loss. seen 
more often in older mares. these mares can get pregnant, but miscarry 
between 45 and 60 days due to inadequate glandular secretions. in cattle 
and sheep, they get the fibrosis, but it isn't that big a deal, b/c the 
placenta comes into the picture much earlier.

----break----

well, allegedly doug got delayed by the snow (sounds bogus to me) and so 
we don't have our handouts, still.

Stages of gestation: three of them

embryo: from fertilization to start of primitive fetal membrane 
development (short time)
organogenesis: that time during which organs are forming and developing. 
very important time, need to be very careful about exposure to toxins, 
fetus is most vulnerable to teratogens now. in cows: 12-45 days, in horse 
12-60, in ewe: 11-34 days. so for cow and mare, about first 2 mos, for 
sheep the first month - avoid use of exogenous substances. don't 
vaccinate then, don't worm them, etc. even though some drugs are "proven 
safe" for use during pregnancy, many are not, and we should err on side 
of caution, and not use anything unnecessary. human obstetricians say the 
same thing. if not totally necessary, don't use it. can look in PDR to 
see if there are known teratogenic effects, but usually there isn't any 
information except in rats.
fetus: the latter portion of gestation, after completion of 
organogenesis. time of rapid growth of fetus. organ systems are present, 
now fetus is growing. there start to be major nutritional/metabolic 
demands on the dam. first 2/3 of gestation, very minimal demands on dam, 
but now a lot more.

slide: 90 day horse fetus - within amnion, very delicate membrane 
surrounding fetus. can see that already, looks like a tiny horse, all 
organ systems ready, now it just has to grow.

length of gestation: is affected by
dam - younger dams have shorter pregnancies than older dams
fetus - in cattle, study was done - programs using embryo transfer show 
that breed of embryo determines length of gestation. a short-gestation 
embryo in a long gestation cow will have a short gestation.
breed - see above.
environment - seasonal effects...eg horses breed during days of long day 
length, and if they breed in may, will give birth in april, so those 
fetuses scheduled to be born early in year will be carried longer, and 
later in season will be carried shorter - fetuses "want" to be born in 
june :)
male fetuses carried longer than female

Hormones and pregnancy:

progesterone has a lot of positive influences. a happy hormone.
progestogen: any agent capable of producing biological effects similar to 
that of progesterone.
we always talk about progesterone, but that is really a specific steroid 
hormone. sometimes people talk about progestogens - eg the progestogens 
of late pregnancy, made by the fetal membranes. see above definition. 
but:
progestin: a hormone of the corpus luteum.
progesterone is a progestin, but some of the progestogens are not 
progestins.

actions of progesterone: causes dam to:
secrete histotroph (uterine milk)
close cervix - if it were open, embryo would fall out.
increase uterine tone
suppress myometrial activity - prevents contractions

diagram (in handout)
days of gestation on x axis with hormone levels plotted across it.
after mare ovulates, a primary corpus luteum forms and produces luteal 
progesterone - the level of luteal progesterone rises then starts 
dropping at 21 days. then mare has unique phenomenon. will ovulate and 
luteinize more follicles which will secrete more progestorone, so will 
get an even bigger progesterone peak now, peaking at about 90 days. at 60 
days, fetal membrans/placenta start making progestogens so level of 
placental progestogens starts rising then, and luteal hormones drop off, 
and eventually you could ovariectomize mare and she'd stay pregnant.

in cow, early on, most of progseterone is from ovary, and she needs to 
have ovary throughout most of gestation.

in ewe, and women, similar charts shown with species specific 
differences.
clinically, sometimes you might not be able to dx pregnancy by standard 
veterinary methods. usually you palpate uterus per rectum, but now and 
then you can't palpate for some reason, so you would maybe wnat to take a 
blood level. what would you measure? progesterone? no. because it is also 
high during diestrus. so....unless you take the sample when you KNOW 
female should already be back in heat, it's not useful. but...

Estrogen:
all conceptuses produce estrogen. they're pretty good at it, too. some 
groups think that the estrogen plays a role in maternal recognition of 
pregnancy. particularly estrone sulfate is made by conceptus, and it is a 
hormone unique to the conceptus most of the time. when fetus is there, 
estrone sulfate level will be elevated.  in mare, after 60 days, can see 
these estrogens in maternal bloodstream. if you look at embryo culture in 
vitro, can see the estrogens right from beginning. but after 60 days, 
level spills over into mare serum. in mare by 160 days, they even spill 
into urine - could analyze urine for estrone sulfate and pick it up.  so 
if working with zebras that you can't handle or get blood from, you could 
measure estrone sulfate levels in the urine. this hormone is very 
specific for pregnancy. if fetus died and mummified, estrone sulfate 
level would drop off, so this is also good for checking fetal well being. 
urine test is called "cuboni test"

Prostoglandin: usually we don't think about this in connection with 
pregnancy, because PGF2a causes luteolysis and termination of pregnancy. 
but, say, in sheep, PGE is thought to be a maternal recognition signal. 
there are other prostoglandins too, that play a lot of roles eg in blood 
flow regulation, etc. but you don't want to give exogenous 
prostoglandins. often, we abbreviate prostoglandins as PG, but that can 
be confused with progesterone, so don't get confused.

Equine chorionic gonadotropin ECG
pregnant mare serum gonadotropin PMSG is the old name for it. in late 70s 
they changed the name. but in the literature they still use both, so.
women have HCG.
in mare, the development of fetal structures occurs during well defined 
time frame. developing allantoic cavity and regressing yolk sac have 
"chorionic girdle' interface, circular band around embryo dividing these 
areas. cells from the chorionic girdle - fetal cells - invade the uterine 
endometrium. in the horse, those are the only cells going into maternal 
tissue. in humans, rats, mice, fetus "implants" way into the uterus. that 
doesn't happen here. so the endometrial cups are formed by these 
cells...little cuplike structures which are arranged in a circular layer. 
these cups produce the ECG. because those fetal cells have burrowed into 
endometrium, they now come into direct contact with bloodstream of 
mother. she forms an inflammatory rxn against them, eventually killing 
the endometrial cup, so by 140 days there is a pretty much dead cup that 
sloughs off. so ECG is high from day 40-120 to 160, then can no longer be 
detected. so there are thse sacs full of ECG for a while, though.
the ECG peak is on the chart in the handout, too. so, would ECG be a good 
pregnancy test? used to be a lot of tests on the market to detect the 
hormone, was a simple blood test. but what happened was, there was a case 
that came in to NBC w/history of being a young racing filly, with good 
potential, who wasn't running that fast and was getting fat. so they cut 
her feed back. she kept running slower and getting fatter. then someone 
remembered she'd been turned out with colts, and wondered if she was 
pregnant. racetrack vet decided to run blood test on mare b/c he wasn't 
good at palpation. he drew blood and ran the test for ECG - was negative 
- but this was after 150 days. she kept getting bigger and slower. 
finally came to NBC hugely pregnant. the company making the test was 
sued. the test is only good if used at the right time. test was off the 
market for about 10 years, but is back on the market now, mainly due to 
mini horse industry needing it. but if you use it, you ahve to know the 
breeding date. if you use it betwen 40-140 days, it's a good test. but if 
fetus dies during that time, th cups will persist the usual time, so then 
you could be wrong. so an estrone sulfate test is probably the best, but 
can't do it before 60 days.

Placental Lactogen:
sheep, cow, woman. has growth hormone like effects, influences dams 
metabolism to divert nutrients to the fetus, plays role in feeding the 
fetus.
oPL, bPL, hPL.

Placenta composed of:
fetal membranes
endometrium
placenta includes fetal and maternal components working together to 
support the fetus' development. so, it's incorrect to say, though we say 
it all the time, that the mare has passed the placenta. only fetal 
component is passed, unless she prolapses her uterus.

slide: diagram of mare utersu with two muscle layers outside and internal 
endometrium with glands, and in between the glands, the areas of 
endometrium that touch the chorion will develop villi. depending on 
species, will be different morphology to them. in mare, make microvilli. 
cow/sheep make cotyledonary type villi. but will interdigitate with fetal 
portion to form placenta.

layers of fetal membranes:
yolk sac early on, regresses as development occurs
amnion - surrounds fetus, contains fetus and amniotic fluid. floats in 
allantoic fluid.
allanois - carries blood vessels. contains allantoic fluid and the amnion

chorion - outer layer, interdigitates with endometrium. contains 
allantois - in most species, is fused to allantois.

so. embryo has umbilical cord kind of going into allantois. area 
surrounding embryo full of amniotic fluid and then around that the 
amnion...

functions of placenta:
a really coool organ. does it all. has respiratory function, nutritive 
function, endocrine function, excretory function, can HEAR and SEE - ok, 
ok, maybe not see and hear. but, we throw this thing OUT after 
birth...but it is really cool. some people eat it. some species eat it. 
yuck.
functions:
protection
respiration
nutrition
excretion

series of slides to help us understand how layers develop, etc. dr 
ginther gave her these slides. a very nice series. 

-horse conceptus - spherical. this one is 14 days, surrounded by 
glycoprotein coat. can see teeny tiny inner cell mass. this is in a yolk 
sac stage. not only is embryo gaining nutrients from uterine milk, but 
also from yolk sac. kinda like having a powerbar with it. :)

-day 21. can grossly see the embryo itself. still very small relative to 
the rest of the conceptus. can see some blood vessels too. embryo is 
within amnion, but amnion is very intimate with embryo. still a huge amt 
of yolk sac.

-day 24 or 25 - beating heart is present. can see that as a red spot. 
with u/s can see it beating! can see more blood vessels. allantoic cavity 
is starting to form from outpouching of hindgut - can see this paler 
structure surrouding part of embryo. now yolk sac will start to shrink.

-day 30 - embryo proper with much much larger allantoic cavity full of 
blood vessels, and much smaller yolk sac, though embryo still getting 
nutrients from yolk sac. can see chorionic girdle - junction of allantois 
and yolk sac.

- day 42 - mostly allantois, sm amt yolk sac, pronounced chorionic 
girdle. 
-exposed embryo within amnion which is closely adhered to embryo, with 
peeled back allantois. still don't have a chorion. this is about 50 days.


-bovine amniotic vesicle. remember, whole thing elongated, so allantois 
is way far away, but amnion is there as bulging vesicle.

-fetal membranes from mare after parturition. can see umbilical cord, 
extending into amnion. amnion is there. is translucent, a few cells 
thick. cord extends out to allantoic surface, where many many vessels 
branch out through and connect into underlying chorion.

classification of placenta:
shape, tissue layers, tissue shed.
clinically is important to understand these classifications.

shape: in horses, and in llama, placenta is "diffuse" in shape. this 
means that the microvilli are spread all over the chorion, wherever it 
touches endometrium. so whole chorion is covered wtih microvilli. kind of 
looks like velvet some people say. if you immerse it in water can see the 
little fingerlike microvilli sticking out. when you think of microvilli 
or chorion, think of tree branches. because in fact, microvilli do look 
like that. histologically, you can see these little branches coming out, 
increaseing the surface contact between chorion and endometrium.
in cattle, sheep, goats, ruminants: we have cotyledonary type placenta. 
little circumscribed areas develop villi. other areas are called 
intercodyledonary areas, and they don't directly contact the uterus. the 
cotyledons form over the uterine caruncles. when palpating cows, can use 
this to stage pregnancy. on postpartum membrane, can see the cotyledons - 
pretty big. cotyledon + caruncle == placentome. 
cats/dogs: zonary type of placenta. at the widest circumference of lump 
in the uterus, a strip of connection forms between chorion and 
endometrium. just like a wide zone of tape around a ball or something.

by tissue layer:
horse, sow, ruminant: epitheliochorial - 6 layers. so at parturition, 
epithelium of uterus stays intact, and mares can get pregnant again in 6 
days.
dog/cat: endotheliochorial - 5 layers? dogs normally have fair amt of vag 
d/c postpartum
primates/rodents: hemochorial -3 layers - fetus is bathed directly in 
maternal blood. mother can bleed for weeks after birth. there is big 
erosion of uterine lining.

by tissue loss at delivery:
deciduate: rodents primates, kinda modified in dog/cat. some of uterus is 
lost with placenta
indeciduate: mare, sow, ruminant. when placenta is shed, uterus totally 
intact, no damage to uterus. these species can get pregnant sooner after 
parturition.

passive immunity:
in utero: primate, gpig, rabbit - Ig crosses placenta
colostrum: mare, sow, ruminant - Ig doesn't cross placenta - neonates 
NEED colostrum.
both: bitch, queen, rodent - in between.

trivial pursuit question: name this slide: "false tongue" aka "hippomane" 
an accumulation of crap which forms in allantoic cavity and floats 
around. in dystocia, vet can pretend to have pulled a tongue out, but it 
isn't really a tongue :)


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