Content uploaded by Sean Limesand
Author content
All content in this area was uploaded by Sean Limesand on Sep 20, 2018
Content may be subject to copyright.
Available via license: CC BY
Content may be subject to copyright.
Proceedings of the 10th International Ruminant Reproduction Symposium (IRRS 2018); Foz do Iguaçu, PR, Brazil,
September 16th to 20th, 2018.
_________________________________________
1Corresponding author: limesand@email.arizona.edu
Received: April 9, 2018
Accepted: May 16, 2018
Impact of thermal stress on placental function and fetal physiology
Sean W. Limesand1, Leticia E. Camacho, Amy C. Kelly, Andrew T. Antolic
School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson AZ 85719 USA.
Abstract
In ruminants, prolonged exposure to high
ambient temperatures negatively affects placental
development and function. The pursuing limitations in
placental oxygen and nutrient supply between the
mother and fetus slow fetal growth lowering birth
weights and postnatal performance. The pregnant ewe is
a long-standing animal model for the study of maternal-
fetal interactions and is susceptible to naturally
occurring heat stress, which causes fetal growth
restriction. In the pregnant ewe, studies show that the
fetus adapts to hyperthermia-induced placental
insufficiency to preserve placental transport capacity of
oxygen and nutrients. These adaptive responses are at
the expense of normal fetal development and growth.
Enlarged transplacental gradient for oxygen and glucose
facilitates diffusion across the placenta, but develops by
lowering fetal blood oxygen and glucose concentrations.
Fetal hypoxemia and hypoglycemia slow growth and
alter their metabolic and endocrine profiles. Deficits in
amino acids transport across the placenta are present but
are overcome by reduced fetal clearance rates, likely
due to fetal hypoxemia or endocrine responses to
hypoxic stress. Here, we provide an overview of the
performance limitations observed in ruminants exposed
to heat stress during pregnancy, but we focus our
presentation on the sheep fetus in pregnancies
complicated by hyperthermia-induced placental
insufficiency. We define the characteristics of placental
dysfunction observed in the fetus of heat stressed ewes
during pregnancy and present developmental adaptations
in organogenesis, metabolism, and endocrinology that are
proposed to establish maladaptive situations reaching
far beyond the perinatal period.
Keywords: heat stress, intrauterine growth restriction,
placental insufficiency, sheep fetus.
Introduction
Environmental heat stress diminishes revenue
for livestock producers by negatively impacting nutrient
utilization, growth, and reproductive performance.
Consequences of heat stress on early embryonic
survival have been well documented, both scientifically
and economically (Hansen et al., 2001). However,
financial losses from warm environmental conditions
are not limited to embryonic wastage. Maternal
exposure to prolonged high ambient temperatures
during gestation has been associated with lighter birth
weights, greater incidence of morbidity before weaning,
lower survival rates, and less desirable carcass traits
(Shelton, 1964; Monteiro et al., 2016). As we will
explain, these latter complications likely are products of
developmental adaptations to nutrient and oxygen
deprivation caused by hyperthermia-induced placental
insufficiency. During maternal heat stress, fetal growth
restriction may be considered beneficial for the dam, as
a smaller conceptus yields less metabolically active
tissue, greater maternal surface area to mass ratio, and
less nutritional strain on the mother (Wells and Cole,
2002). Although fetal growth restriction is advantageous
for the dam, fetal growth restriction and the
accompanying adaptations to placental restriction are
associated with a myriad of metabolic complications
that negatively affect future performance.
Because offspring from a heat stressed dam are
growth restricted during gestation, we begin with a
closer look at the characteristics of placental
dysfunction that restrict fetal growth and cause
metabolic adaptations. We provide an overview of the
performance limitations observed with maternal heat
stress in ruminants, but focus our presentation on work
conducted in sheep that are experimentally heat stressed
during mid gestation, a time when the placenta is
established and placental growth is at maximum
(Regnault et al., 2002a). The pregnant sheep has been
used extensively over the past 50 years to investigate
placental and fetal physiology due to the ability to
surgically place and maintain catheters in the maternal
and fetal vasculature that allow for repetitive blood
sampling from non-anesthetized ewes (Meschia et al.,
1965; Barry et al., 2008). The substantial groundwork
on maternal-fetal interactions in sheep provides ample
knowledge for normal pregnancy, as well as information
on models of pregnancies complicated by
experimentally or naturally produced placental
restriction, which includes a model of hyperthermia-
induced placental insufficiency.
Pregnant ewes exposed chronically to high
ambient temperatures in the laboratory from early to late
gestation have fetuses that are significantly growth
restricted close to term (Bell et al., 1987; Thureen et al.,
1992). Ultrasonographic measurements indicate that
biometric parameters for determining fetal growth
restriction, for example abdominal circumference begins
to diverge from normal as early as mid-gestation. This is
a developmental point, prior to rapid fetal growth and at
the apex of placental growth (Galan et al., 1999).
Terminal studies indicate that significant reduction of
placental mass precedes declines in fetal weight (Fig.
1A and B). Before 110 days of gestation (term 149
days), placental weights were significantly less in heat
stress ewes than controls (280 ± 32 g vs. 443 ± 32 g).
DOI: 10.21451/1984-3143-AR2018-0056
G e s t at io n a l A g e ( d a y)
F e t a l W e i g h t (k g )
90 100 110 120 130 140 150
0
1
2
3
4
5
G e s t at io n a l A g e ( d a y)
P l a c e n t a (g )
90 100 110 120 130 140 150
0
200
400
600
800
G e s t at io n a l A g e ( d a y)
G l u c o se (m m o l /l)
90 100 110 120 130 140 150
0 .0
0 .5
1 .0
1 .5
2 .0
G e s t at io n a l A g e ( d a y)
O x y g e n C o n t e n t (m m o l /l )
90 100 110 120 130 140 150
0
1
2
3
4
5
A . B .
C . D .
Limesand et al. Heat stress during pregnancy.
Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018 887
However, fetal weights were not affect at this younger
age (0.9 ± 0.2 kg versus 1.0 ± 0.2 kg). After 130 days of
gestation, both fetal and placental weights were
significantly less in heat stressed ewes compared to
controls (49% for fetus and 56% for placenta). These
data indicate that the majority of fetal growth restriction
occurs during the final stages of gestation after placental
growth restriction (de Vrijer et al., 2006; Macko et al.,
2013). Limitations are caused by reduced placental mass
and function, which leads to the development and
progressive decline in fetal glucose (21 to 33% less) and
oxygen (25 to 46%) concentrations over the final third
of gestation, when fetal growth rate is at maximum
(Limesand et al., 2013; Rozance et al., 2018; Fig. 1C
and D). This evidence supports the hypothesis that
placental deficiencies are responsible for fetal growth
restriction in late gestation. Abnormal placental growth,
vascular organization, and angiogenesis were described
as possible causes of placental insufficiency due to
aberrant expression patterns of angiogenic growth
factors and their receptors (Vatnick et al., 1991;
Regnault et al., 2002b; Galan et al., 2005; Hagen et al.,
2005). Together, the pregnant ewe and this model of
hyperthermia-induced placental insufficiency provides a
unique opportunity to investigate fetal adaptive
responses and growth restriction caused by a naturally
induced placental restriction, which negatively effects
their future health and performance. We review the
outcomes in the placenta and fetus that are associated
with adaptive responses to hyperthermia-induced
placental insufficiency and discuss how they relate to
future deficiencies in production.
Figure 1. Progression of hyperthermia-induced placental insufficiency. Fetal weights (A), placental weights (B),
fetal arterial plasma glucose concentrations (C), and fetal arterial blood oxygen contents (D) are presented as means
reported in Ross et al., 1996 (hexagon); Brown et al., 2012 (circle); de Vrijer et al., 2006 (square <100 days);
Limesand et al., 2013 (downward triangle), Macko et al., 2013 (diamond); and Rozance et al., 2018 (square >130
days). Each individual point represents a group mean for thermoneutral control ewes (fill shapes) and heat stress
ewes (open shapes) for that specific report (symbol). At younger gestational ages (<110 days), all group means were
significantly different within a report, except for fetal weight. At older gestational ages (>130 days), group means
within the report were significantly different. Conclusions from these data indicate that advancing gestational age
leads to greater differences in all parameters measured, which includes slower rates of fetal growth.
Effects of heat stress in ruminants during gestation
and lactation
Environmental heat stress imposes significant
limitations on fetal growth and milk production in
several species of ruminants, but some species, usually
those with higher production rates, are more susceptible
to heat stress. For this reason, lactating dairy cows have
a low tolerance to heat stress because as milk
production per cow is increased there is a concurrent
increase in metabolic heat production (Collier et al.,
2006). Similarly, substantial economic consequences
are incurred during gestation and lactation due to high
metabolic heat output (West, 2003; Collier et al., 2006).
Limesand et al. Heat stress during pregnancy.
888 Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018
Mechanisms that explain placental and fetal growth
restriction include the redistribution of blood flow to the
skin and nasal mucosa at the expense of other internal
organs including the pregnant uterus (Reynolds et al.,
1985; McCrabb et al., 1993). Reductions in placental
function, along with maternal factors caused by heat
stress, restrict mammary gland development and lower
the potential yield for the subsequent lactation. In
addition to losses in milk yield and quality following the
immediate pregnancy, heat stress in late pregnancy
effects immune processes associated with poor
transition to lactation including: lower phagocytic
activity and decreased hepatic prolactin signaling (PRL-
R, SOCS-3, and CAV-1 mRNA) during the dry period
(do Amaral et al., 2011). Milk yield and milk protein
are influenced by calving season, as both are lowest
during the warmest months compared to cows that
calved in the winter (Barash et al., 2001). Lactating
sheep are also susceptible to heat stress, which leads to
differences in milk composition, specifically lower fat
and protein content (Abdalla et al., 1993). In addition to
lighter birth weights, calves from heat stress cows
exhibited both immediate and prolonged effects on their
passive immunity, growth, activity patterns, and
thermotolerance (Ahmed et al., 2017; Laporta et al.,
2017). Furthermore, the effects of heat stress on
offspring persist, resulting in lower yearling weights and
less heifers reaching their first lactation. Cows exposed
to heat stress during fetal life produced less milk
compared to cows that received heat abatement
strategies (Monteiro et al., 2016). This evidence shows
that heat stress during gestation affects fetal
development and also creates lasting complications that
lower the future productivity of the offspring, which
may be caused by developmental adaptations to
placental insufficiency.
Maternal heat stress limits placental transport
capacity
The capacity of the placenta to transfer oxygen
and nutrients must increase throughout pregnancy to
meet metabolic demands of the growing fetus. In sheep,
placental transport capacity continues to increase by
expanding the surface area of the maternal-fetal
interface and by thinning of the placental barrier to
promote the exchange and permeability of metabolic
substrates. Amino acids, oxygen, and glucose are
transported across the placenta by active transport,
passive diffusion, and facilitated diffusion (Battaglia
and Meschia, 1978). For diffusion mechanisms, the rate
of transplacental transport is dependent on uterine and
umbilical blood flow, substrate permeability, and
substrate concentration difference across the placenta.
Placental clearance is diminished with heat
stress due to lower permeability for metabolic
substrates. Oxygen and glucose permeability is reduced
by a smaller placenta with less surface area and
transport capacity, which combine to lower uterine
extraction efficiency (Fig. 2). Evidence for this
conclusion is that the transplacental gradients of oxygen
and glucose increase in ewes exposed to environmental
heat stress during pregnancy compared to pregnant ewes
maintained under thermoneutral conditions (Fig. 3).
Unlike placental transfer rates, placental clearance is
independent of concentration gradients, but dependent
on the properties of the exchanger (membrane)
permeability or perfusion. Studies with inert molecules
that have flow-limited placental transport show
equivalent transplacental clearance rates in heat stressed
and thermoneutral ewes. For example, there is no
difference in ethanol clearance across the placenta
between thermoneutral and heat stressed ewes when
expressed relative to placental mass (Bell et al., 1987;
Thureen et al., 1992; Regnault et al., 2007). This
observation excludes shunting or uneven perfusion of
uterine and umbilical blood flow as a cause for
decreased transplacental clearance in heat stressed ewes.
In addition to lower placental permeability, placental
transport capacity of metabolic substrates is hindered by
alterations in uteroplacental consumption of the
substrate itself. In heat stressed ewes, uteroplacental
oxygen consumption normalized to placental weight is
unaffected, and glucose utilization by uteroplacental
tissue is less (Bell et al., 1987; Thureen et al., 1992;
Regnault et al., 2007). Therefore, the enlargement in the
transplacental concentration difference for oxygen and
glucose are due to lower placental permeability, which
for glucose may be explained partially by lower
abundance of facilitated glucose transporters (Limesand
et al., 2004; Wallace et al., 2005).
Transplacental gradients and uterine-umbilical
blood flow ratios adapt under heat stress to preserve the
net umbilical uptake of glucose and oxygen, but as
discussed later, this compensatory mechanism causes
reductions in substrate concentrations in the fetus that
become detrimental to development and growth (Bell et
al., 1987; Thureen et al., 1992; Regnault et al., 2007,
2013). Simple concepts for diffusion indicate that larger
transplacental concentration gradients (Fig. 3A and B)
will increase the net movement of oxygen and glucose
across the placenta to the fetus. Another compensatory
mechanism is greater uterine-to-umbilical blood flow
ratio, which further demonstrates impaired placental
diffusion capacity in heat stressed ewes (Fig. 3C).
Normally the uterine-to-umbilical blood flow ratio is
~2, which is predicted to fulfill the delivery
requirements for the placenta and fetus because
maximum fetal clearance occurs when flows are
equivalent (Wilkening et al., 1982). Increases in the
blood flow ratio lowers uterine arteriovenous
differences for oxygen and aids in expanding the
transplacental gradient to facilitate uterine uptake (Bell
et al., 1987; Regnault et al., 2003). This adaptation may
be advantageous to the fetus because it lowers cardiac
output to the placenta to increase umbilical uptake, but
the increase vascular resistance in the placenta is
postulated to negatively affect the cardiovascular
development (Galan et al., 2005). Together, the
enlarged transplacental concentration gradients of
oxygen and glucose and increased uterine-to-umbilical
blood flow ratio are sufficient to minimize reductions in
net umbilical oxygen and glucose uptake per fetal mass.
However, comparisons for means across several
A . B .
E n v ir o n m e n t a l C o n d it io n
U t e ri n e G lu c o s e E x t r a c ti o n ( % )
C o n t r o l H e a t S tr e s s
0
2
4
6
8
P < 0 . 0 5
E n v i ro n m e n t a l C o n d it i o n
U t e r in e 0 2 E x t ra c t i o n ( % )
C o n t r o l H e a t S tr e s s
0
1 0
2 0
3 0
P < 0 . 0 5
C . D .
P l a c e n ta l M a s s ( k g )
P l a c e n ta l O
2
T r a n s p o rt
C a p a c i t y (m l /m in )
0 . 0 0 . 1 0 . 2 0 . 3 0 . 4 0 . 5
0
200
400
600
800
1000
R
2
= 0 . 7 7
P l a c e n ta l M a s s ( k g )
P l a c e n ta l G l u c o s e T r a n s p o r t
C a p a c i t y (m l /m in )
0 . 0 0 . 1 0 . 2 0 . 3 0 . 4 0 . 5
0
2 0
4 0
6 0
8 0
R2= 0 . 7 5
Limesand et al. Heat stress during pregnancy.
Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018 889
reported cohorts indicate that there were modest but
significant reductions of 9 and 14% in net umbilical
oxygen and glucose uptakes, respectively (Fig. 4).
Placenta delivery of amino acids to the fetus is
also lower in heat stressed ewes even though fetal
plasma amino acid concentrations are not necessarily
reduced (Thorn et al., 2009; Regnault et al., 2013). For
most amino acids, concentrations in fetal circulation are
normally greater than in maternal circulation and
therefore are transported actively across the placenta
against their concentration gradient. In heat stressed
ewes, the absolute placental flux for essential amino
acids from mother to fetus is reduced ~80%, whereas
the flux relative to placental mass is ~40% less (Ross et
al., 1996; Anderson et al., 1997; de Vrijer et al., 2004).
Similar reductions in transplacental uptake of essential
amino acids are seen when expressed relative to fetal
mass, with the exception of lysine (Regnault et al.,
2013). Two important points become evident from these
independent studies on amino acid placental transport in
heat stress ewes. First, fetal uptakes of essential amino
acids are reduced to similar magnitudes. Second, the
impaired transport of amino acids is due to a reduction
in placental size as well as decreased transport capacity
per unit mass. Therefore, amino acid transfer depends
on surface area of the maternal-fetal interface, which is
reduced, as well as on the concentrations of amino acid
transporters (Regnault et al., 2005). Interestingly,
decreased transplacental flux of amino acid does not
always lower their concentration in fetal plasma,
implicating adaptive mechanisms in fetal metabolism or
clearance of amino acids, which were associated with
low oxygen concentrations (Regnault et al., 2013).
Again, these data show that placental insufficiencies
produced by heat stress depend on decreased placental
mass and function, even though compensatory
mechanism by the fetus are in place to assist with the
placental deficiencies.
Figure 2. Placental transport capacity and uterine extraction efficiency for oxygen and glucose. Placental transport
capacity for oxygen (A) and glucose (B) are presented by placental weights for independent group means from
previously reported cohorts of thermoneutral and heat stressed ewes during gestation. Placental transport capacity is
the net umbilical (fetal) uptake (µmol/min) divided by the maternal arterial-fetal arterial plasma concentration
difference (µmol/ml) according to the equation: transport capacity = uptake/concentration difference. Linear
regression analysis shows a positive association for placental transport capacity and placental mass (R2). Uterine
extraction of oxygen was calculated by expressing the whole blood arterial-venous oxygen concentration difference
across the uterine circulation as a percent of the arterial concentration (C). Uterine extraction of glucose was
expressed as the plasma arterial-venous concentration difference as a percent of the arterial glucose concentration
(D). Group means for thermoneutral control (fill symbols) and heat stressed animals (open symbols) were reported
in Bell et al., 1987 (downward triangle); Thureen et al., 1992 (hexagon); Ross et al., 1996 (diamond); Anderson et
al., 1997 (small circle, panel A); Regnault et al., 2003 (large circle, panel A); Limesand et al., 2004 (circle, panel
B); de Vrijer et al., 2004 (triangle); Limesand et al., 2007 (large square, panel B) and Brown et al., 2012 (small
square, panel B). An ANOVA with reported study as the repeated measure identify group (control and heat stress)
differences (P-values) for panels C and D.
E n v i ro n m e n t a l C o n d it io n
M a t e r n a l- F e ta l A rt e r ia l D if fe r e n c e
in O x y g e n ( m M )
C o n t r o l H e a t S tr e s s
0
2
4
6
P < 0 . 0 1
E n v i ro n m e n t a l C o n d it io n
M a t e r n a l- F e ta l A rt e r ia l D if fe r e n c e
in G lu c o s e (m M )
C o n t r o l H e a t S tr e s s
0
1
2
3
4
P < 0 . 0 5
E n v i ro n m e n t a l C o n d it io n
U t e ri n e -t o - U m b i li c a l
B l o o d F lo w R a ti o
C o n t r o l H e a t S tr e s s
0
1
2
3
4
5
P < 0 . 0 5
A .
B .
C .
Limesand et al. Heat stress during pregnancy.
890 Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018
Figure 3. Transplacental gradients and
uterine-umbilical blood flow ratio. Maternal-
fetal arterial difference for oxygen (A) and
glucose (B) concentrations are presented for
pregnant ewes at approximately 90% of
gestation. Uterine-to-umbilical blood flow
rates are presented in panel C. Group means
for thermoneutral control and heat stressed
ewes were reported in Bell et al., 1987;
Thureen et al., 1992; Ross et al., 1996;
Anderson et al., 1997; Regnault et al., 2003;
Limesand et al., 2004, 2007; de Vrijer et al.,
2004. An ANOVA with report as the repeated
measure identifies group (control and heat
stress) differences (P-values).
Fetal responses to hyperthermia-induced placental
insufficiency
Maternal hyperthermia is natural in sheep that,
uncharacteristically, carry pregnancies in summer
months producing a smaller placenta with lower
transport capacity for glucose, amino acids, and oxygen.
Therefore, hyperthermia-induced placental insufficiency
under-nourishes the fetus and leads to asymmetric
intrauterine growth restriction (IUGR) that spares brain
and heart growth relative to overall body weight (Fig.
5). In heat stressed sheep with placental insufficiency,
we and others have characterized fetal adaptations in
metabolism, endocrinology, and selected organ
functions to specify mechanisms responsible for
developmental programming. These studies have
focused on fetal metabolism and endocrinology,
pancreatic insulin secretion, hepatic glucose production,
skeletal muscle growth and metabolism, and cardiac
metabolism. In combination with oxygen and nutrient
deficits, elevations in norepinephrine and epinephrine
impinge on nearly all adaptive fetal responses measured
including growth, glucose metabolism, and insulin
secretion (Davis et al., 2015; Macko et al., 2016). How
these fetal responses allow normal cellular oxidation to
continue, maintain viability at the expense of growth,
but ultimately become maladaptive for future
performance are described herein.
Fetal metabolism and endocrinology
The fetus uses the umbilical uptake of nutrients
to fulfill two major requirements: oxidation to fuel
energy metabolism and accretion for growth and storage
of substrates. Energy metabolism can be estimated from
rates of oxygen consumption, which based on net
umbilical oxygen uptake per fetal mass was only
marginally less in IUGR fetuses compared to control
E n v ir o n m e n t a l C o n d it io n
U m b i lic a l O 2 U p ta k e
(m o l /m in / k g )
C o n t r o l H e a t S tr e s s
200
250
300
350
400
450
P < 0 .0 5
E n v ir o n m e n t a l C o n d it io n
U m b il ic a l G l u c o s e U p t a k e
(
m o l /m in / k g )
C o n t r o l H e a t S tr e s s
0
1 0
2 0
3 0
4 0
P < 0 .0 1
A .
B .
E n v ir o n m e n t a l C o n d it io n
U m b i li c a l L a c ta t e U p t a k e
(
m o l /m in / k g )
C o n t r o l H e a t S tr e s s
-5
0
5
1 0
1 5
2 0
2 5
P < 0 .0 1
C .
Limesand et al. Heat stress during pregnancy.
Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018 891
fetuses (Fig. 4). For metabolic studies, the rate of
oxygen consumption is one of the most useful standards
of reference because the metabolic quotient defines the
quantity of substrate needed to satisfy energy
requirements of the fetus or fetal organs. This allows us
to judge whether the quantity of nutrients being supplied
to the fetus by the placenta is larger or smaller than the
energy demands of the fetus, thus indicating whether the
placental nutrient supply is sufficient to support fetal
growth through accretion. The metabolic quotient for
individual substrates are calculated as the ratio of the
substrate oxygen equivalents to oxygen uptake. Oxygen
equivalents are the quantity of oxygen molecules
required for complete oxidation of that substrate to
carbon dioxide and water. For example, when
calculating the glucose/oxygen quotient, the glucose
concentration difference (mmol/l) is multiplied by six
and then divided by the oxygen concentration difference
(mmol/l). In the fetus, the major sources for oxidative
substrates are glucose, lactate, and amino acids, which
explain the emphasis for studying their placental
transport capacity and the resulting consequences to
fetal growth when their delivery is restricted.
Strikingly, the sum of the nutrient/oxygen quotients for
glucose, lactate, and amino acids in IUGR fetus barely
exceeds the umbilical oxygen quotient, which indicates
that fetal uptake of nutrients is just sufficient to meet
the oxidative requirements with no surplus for
accretion (Regnault et al., 2013). A similar limitation
in substrate availability was identified for the hindlimb
quotients in IUGR fetuses (Rozance et al., 2018).
Numerous studies have been conducted to explain how
IUGR fetuses adapt to placental insufficiency by
altering organ metabolism for these primary substrates
as well as describing the potential endocrine regulation
(Fig. 6).
Figure 4. Umbilical uptakes for oxygen,
glucose, and lactate. Fetal weight-specific net
umbilical uptakes for oxygen (A) and glucose
(B) are presented for pregnant ewes at 90% of
gestation. Group means for thermoneutral
control and heat stressed ewes were reported in
Bell et al., 1987; Thureen et al., 1992; Ross et
al., 1996; Anderson et al., 1997; Regnault et
al., 2003; Limesand et al., 2004, 2007; de
Vrijer et al., 2004; Brown et al., 2012, 2015;
and Thorn et al., 2013. An ANOVA with
reported study as the repeated measure
identifies group (control and heat stress)
differences (P-values) for each of the uptakes.
Although net umbilical (fetal) glucose uptakes
were not significantly lower for all reports individually,
the meta-analysis performed herein for ten reports that
measured umbilical glucose uptake show that glucose
uptake was lower in IUGR fetuses (Fig. 4). Experiments
with glucose tracers to determine rates of fetal glucose
E n v ir o n m e n t a l C o n d it io n
B r a in -t o -F e t u s
(g /k g )
C o n t r o l H e a t S tr e s s
0
1 0
2 0
3 0
P < 0 .0 1
E n v ir o n m e n t a l C o n d it io n
H e a r t - t o F e t u s
(g /k g )
C o n t r o l H e a t S tr e s s
5
6
7
8
9
1 0
E n v ir o n m e n t a l C o n d it io n
L iv e r -t o - F et u s
(g /k g )
C o n t r o l H e a t S tr e s s
2 0
2 5
3 0
3 5
P < 0 .0 5
A .
B .
C .
Limesand et al. Heat stress during pregnancy.
892 Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018
utilization and oxidation show significant alterations in
glucose metabolism in IUGR fetuses with placental
insufficiency (Limesand et al., 2007; Thorn et al., 2013;
Brown et al., 2015). Body-weight specific glucose
utilization rates are not different between IUGR and
control fetuses, despite IUGR fetuses having markedly
lower plasma insulin and glucose concentrations. In
control fetuses, umbilical glucose uptake is normally
equivalent to the glucose utilization rate, which
demonstrates that placental glucose uptake is sufficient
and glucose production is negligible. In IUGR fetuses
with hyperthermia-induced placental insufficiency,
glucose utilization rates exceed umbilical glucose
uptake, thus the IUGR fetus has endogenous glucose
production (Limesand et al., 2007; Thorn et al., 2013).
The fraction of glucose oxidized to carbon dioxide is
also less in IUGR fetuses, which suggests peripheral
tissues may have increased glycolysis to supply lactate
for hepatic glucose production (Limesand et al., 2007;
Brown et al., 2015). Together, these findings identify
important metabolic responses in glucose metabolism
that are predicted to caused alterations in tissues such as
liver and muscle, changes in endocrine factors, or a
combination of both.
Figure 5. Asymmetric growth restriction with
hyperthermia-induced placental insufficiency.
Brain (A), heart (B), and liver (C) weights in
grams are expressed relative to fetal weight (kg)
for fetuses necropsied at 90% of gestation. Group
means for thermoneutral control and heat stressed
ewes were reported in Thureen et al., 1992;
Anderson et al., 1997; de Vrijer et al., 2004;
Brown et al., 2012; Davis et al., 2015; and Barry
et al., 2016. An ANOVA with reported study as
the repeated measure identifies group (control and
heat stress) differences (P-values) for organ to
fetal weight ratios.
For glucose metabolism, three adaptations are
apparent in IUGR fetuses with placental insufficiency.
First, there is greater avidity for glucose uptake and
utilization by fetal tissues in the presence of low glucose
and insulin concentrations, indicating that there is
greater insulin sensitivity in the IUGR fetus.
Explanations for the greater glucose uptake capacity
include a larger proportion of neuronal tissue to body
weight (Fig. 5) and is supported by upregulation of
glucose transporter 1 concentrations in the brain
(Limesand et al., 2007). Furthermore, the glucose
extraction efficiency and glucose uptake into both the
hindlimb and myocardium are similar between control
and IUGR fetuses, despite low glucose and insulin
concentrations in the IUGR fetus (Barry et al., 2016;
Rozance et al., 2018). Because glucose transporter
expression was unaffected in IUGR muscle, adaptations
in proximal insulin signaling were apparent and appear
E n v ir o n m e n t a l C o n d i ti o n
In s u l in (n g / m l )
C o n t r o l H e a t S tr e s s
0 .0
0 .1
0 .2
0 .3
0 .4
0 .5
P < 0 . 0 0 1
E n v ir o n m e n t a l C o n d i ti o n
G l u c a g o n (p g / m l)
C o n t r o l H e a t S tr e s s
0
2 0
4 0
6 0
8 0
100
P < 0 .1 0
E n v ir o n m e n t a l C o n d i ti o n
N o r e p in e p h ri n e (p g /m l)
C o n t r o l H e a t S tr e s s
0
1000
2000
3000
4000
5000
P < 0 .0 1
E n v ir o n m e n t a l C o n d i ti o n
C o r ti s o l ( n g /m l )
C o n t r o l H e a t S tr e s s
0
1 0
2 0
3 0
4 0
P < 0 .0 1
E n v ir o n m e n t a l C o n d i ti o n
IG F -I (n g / m l )
C o n t r o l H e a t S tr e s s
0
5 0
100
150
P < 0 .0 1
A . B .
C . D .
E .
Limesand et al. Heat stress during pregnancy.
Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018 893
to enhance insulin sensitivity due to increased insulin
receptor concentrations and decreased phosphoinositide-
3 kinase (p85) with no change in the p110 catalytic
subunit (Limesand et al., 2007; Thorn et al., 2009).
Second, IUGR fetuses exhibit hepatic glucose
production, which is normally uncommon, but
augmented by significant increases in gluconeogenic
enzymes, PEPCK and glucose-6-phosphatase, perhaps
in response to cAMP-response element-binding protein
activation due to increases in cortisol, glucagon, and
norepinephrine (Fig. 6; Limesand et al., 2007; Thorn et
al., 2013). Third, enzymes that regulate the tricarboxylic
acid cycle are altered in skeletal muscle and liver of
IUGR fetuses. For example, pyruvate dehydrogenase
kinase 4 mRNA expression is increased 5-fold in IUGR
skeletal muscle. Pyruvate dehydrogenase kinase 4, when
phosphorylated, inhibits pyruvate dehydrogenase, which
converts pyruvate to acetyl CoA for use in the
tricarboxylic acid cycle (Brown et al., 2015). Pyruvate
carboxylase and lactate dehydrogenases expression also
were depressed in the skeletal muscle from IUGR
fetuses, and the former may also play a role in sparing
pyruvate via glycolysis from oxidative metabolism.
Interestingly, lactate output from the hindlimb was not
increased in IUGR fetuses. However, the lactate/oxygen
quotient was greater, which shows greater lactate output
per mole of oxygen consumed by the hindlimb of the
IUGR fetus (Rozance et al., 2018). The sum of glucose
and lactate quotient was similar between control and
IUGR fetuses, and the amino acid/oxygen quotient was
lower. This indicates that alterations in substrate
utilization are more dependent on amino acid
metabolism and protein synthesis and growth are
expendable.
Figure 6. Endocrine profile in fetuses with hyperthermia-induced placental insufficiency. Mean plasma insulin (A),
glucagon (B), norepinephrine (C), cortisol (D), and insulin-like growth factor I (IGF-I, E) concentrations are
presented for fetuses at 90% of gestation. Group means for thermoneutral control and heat stressed ewes were
reported in Limesand et al., 2006; Brown et al., 2012, 2016; Thorn et al., 2013; Macko et al., 2016; and Rozance et
al., 2018. An ANOVA with reported study as the repeated measure identifies group (control and heat stress)
differences (P-values) for hormone concentrations.
Limesand et al. Heat stress during pregnancy.
894 Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018
We have followed IUGR lambs from heat
stressed ewes and identified persistent augmentations in
insulin sensitivity at least until two weeks of age
(Camacho et al., 2017). These experiments demonstrate
that in utero adaptations could have negative
consequences during the postnatal growth period, and
ultimately progress into insulin resistance because
IUGR lambs with less muscle mass are adapting to their
postnatal environment while also undergoing age related
declines in insulin sensitivity (Gatford et al., 2004).
Low birth weight and less muscle mass may cause
metabolic limitation that decrease subsequent
performance (Wu et al., 2006). As postnatal growth is
largely dependent upon fetal growth and development, it
is important to determine how management decisions
could impact the growth trajectory of the fetus;
however, it is not always feasible to prevent IUGR in
production settings. Thus, identifying the mechanism
that regulate skeletal muscle metabolism in IUGR
during pre- and postnatal development is necessary.
Pancreatic insulin secretion
Pancreatic β-cells secrete insulin in response to
elevated plasma glucose concentrations to stimulate
glucose uptake into fetal tissues and promote growth. In
fetal sheep, β-cells become responsive to glucose after
mid-gestation (Aldoretta et al., 1998). Since insulin
cannot cross the placenta, insulin secretion from fetal
pancreatic β-cells is critical for coordinating fetal
growth rates with the placental glucose transport.
Insulin secretion parallels changes in glucose
concentrations, and both glucose and insulin
concentrations are lower in IUGR fetuses (Boehmer et
al., 2017; Fig. 6). In IUGR fetuses, glucose
concentrations are chronically low (Fig. 1), which
negatively affects β-cell mass and insulin secretory
capacity. The IUGR fetus has impaired insulin secretion
responsiveness due to two primary deficits: less β-cell
mass as a consequence of slower rates of β-cell
proliferation and less insulin content per β-cell
(Limesand et al., 2005, 2006).
Another factor contributing to the suppression
of insulin secretion in IUGR fetuses is persistent
elevations of plasma norepinephrine and epinephrine,
which inhibit insulin secretion through α2-adrenergic
receptors on β-cells (Jackson et al., 2000). In the IUGR
fetus, glucose-stimulated insulin concentrations during
an adrenergic receptor blockade are equivalent to
maximal insulin concentrations in control fetuses (Leos et
al., 2010; Macko et al., 2013). This response occurs
despite IUGR fetuses having fewer β-cells that contain
less insulin. Therefore, chronic adrenergic stimulation
inhibits insulin secretion from fetal β-cells, but the
chronic suppression in IUGR fetuses causes
developmental changes in β-cells insulin secretion
responsiveness. Following chronically high
norepinephrine fetal infusions, a subsequent hyper-
secretory response of insulin has been confirmed in
normally grown fetuses (Chen et al., 2014). Strikingly, in
fetuses with a chronic norepinephrine infusion, the
enhanced insulin secretion responsiveness to glucose and
arginine persisted for five days after termination of the
infusion. This is consistent with the observation of
insulin hyper-secretion in week old IUGR lambs
because norepinephrine concentrations are high during
gestation but expected to decrease after birth when
oxygen and nutrients are sufficient (Camacho et al.,
2017; Chen et al., 2017; Limesand and Rozance, 2017).
Surgical ablation of the fetal adrenal medulla prevents
acute hypoxia-induced norepinephrine secretion and
partially explains the lower glucose stimulated insulin
concentrations in IUGR fetuses (Macko et al., 2016).
While mechanisms for adrenergic inhibition of insulin
secretion include distal steps in exocytosis, acute
adrenergic stimulation also inhibits oxidative metabolism
in β-cells and islets, which supports a role for
norepinephrine to lower oxidation rates of glucose and to
inhibit islet metabolism in IUGR fetuses (Kelly et al.,
2018).
When islets from IUGR and control fetuses
were analyzed for molecular changes using high
throughput RNA sequencing (RNAseq), more than 1000
genes were differentially expressed and explained
decreased cell proliferation (Kelly et al., 2017). This
unbiased approach also revealed novel mechanisms
underlying IUGR islet dysfunction including down
regulation of immune function, suppressed Wnt
signaling, adaptive stress responses, and impaired
proteolysis. These transcriptional changes define
adaptive responses of β-cells during IUGR and may
provide the framework for understanding programming
mechanisms that lead to metabolic complications in
later life following hyperthermia-induced placental
insufficiency.
Skeletal muscle growth and metabolism
Lambs with fetal growth restriction are lighter
at birth and grow less efficiently, yielding carcasses
with insufficient muscle growth (Greenwood et al.,
1998, 2000). In sheep, the formation of new fibers
(myogenesis) is complete around 110 days of gestation
after which the myofibers grow by hypertrophy, which
is when declines in fiber size are detected in fetuses
with placental insufficiency-induced IUGR (Maier et
al., 1992; Wilson et al., 1992; Hay et al., 2016).
Conserved myonuclear domains during early muscle
growth increase protein synthesis, such that myonuclear
accumulation drives growth in young animals (Pavlath
et al., 1989). Fetal myoblast incorporation (via
differentiation) into myofibers is required to increase
nuclei content because nuclei within these myofibers are
post-mitotic (Allen et al., 1979). Myoblasts proliferate
and differentiate in response to an activation signal,
followed by a cascade of regulatory transcription factors
(e.g. Pax7, MyoD, myogenin, and others). IUGR fetuses
from hyperthermia-induced placental insufficiency have
similar numbers of myoblasts compared to controls but
have impaired myoblast differentiation due to slower
rates of myoblast proliferation. (Yates et al., 2014,
2016; Brown et al., 2017). When myoblasts are isolated
and cultured from IUGR skeletal muscle, they replicate
slower than controls. This is likely an intrinsic defect in
Limesand et al. Heat stress during pregnancy.
Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018 895
the IUGR myoblast because it occurs in culture and
independent of nutrient availability (Yates et al., 2014).
Furthermore, IUGR fetuses have decreased fiber size
regardless of the fiber type (Yates et al., 2016). These
complications explain smaller muscle fibers, as well as
identify programing effects that could limit growth later
in life (Brown, 2014).
Skeletal muscle is a primary target for
metabolic complications because it represents
approximately 40% of total body weight and greater
than 50% of energy expenditure (Brown, 2014).
Metabolic energy requirements in skeletal muscle are
met through oxidative phosphorylation. Thus, the
number and efficiency of mitochondria determines
metabolic capacity of muscle fibers. Traditionally,
fibers are classified by oxidative capacity including
Type Ia (slow oxidative), Type IIb (fast glycolytic), and
Type IIa and IIx (fast oxidative; Dunlop et al., 2015). In
the developing sheep the percentage of slow oxidative
fibers (Type I) increases in number from less than 10%
at 110 days gestation to greater than 25% near term and
continues to increase postnatally (Maier et al., 1992).
Myofiber area for both Type I and Type IIa fibers are
decreased in hindlimb muscles from IUGR fetuses
(Yates et al., 2016; Rozance et al., 2018). While there
are less oxidative fibers in hindlimb muscles from
IUGR fetuses, the proportion of glycolytic fibers is
similar to control fetuses (Yates et al., 2016). We expect
this developmental adaptation will lower oxidative-to-
glycolytic fiber ratios and explains impaired glucose
oxidation rates in the IUGR fetus (Limesand et al.,
2007; Brown et al., 2015).
Long-term exposure to elevated
catecholamines down regulates adrenergic receptor
concentrations, lowers sensitivity, and impairs skeletal
muscle metabolism (Yates et al., 2014). Chronic
adrenergic stimulation is associated with adaptive
programming responses in fetal metabolic tissues:
pancreatic islets (Chen et al., 2014; Camacho et al.,
2017), skeletal muscle (Yates et al., 2012), and adipose
(Chen et al., 2010). Moreover, adaptations to chronic
adrenergic stimulation persist after the insult, creating the
potential for life-long metabolic programming.
Specifically, adrenergic receptor β2 mRNA
concentrations are reduced >60% in IUGR fetuses and
lambs but adrenergic receptors β1 and β3 are not different
(Chen et al., 2010). Therefore, potential desensitization of
adrenergic receptor β2, but not other β-adrenergic
receptors, might impair insulin responsiveness in
skeletal muscle because it persists in lambs after birth.
Cardiac metabolism
The effects of maternal heat stress on fetal
development and maturation are also evident in the
heart. Blood flow to essential fetal organs (the brain,
heart, and adrenal glands) is increased preferentially in
response to acute bouts of hypoxia. As a result, blood
flow decreases to the gastrointestinal, renal, and
peripheral vasculature. This pattern for redistribution of
cardiac output is also maintained during chronic periods
of hypoxia such as those found in hyperthermia-induced
placental insufficiency, which may result in the
asymmetric fetal growth (Fig. 5).
IUGR fetal myocardium responds through
unique metabolic and cardiovascular adaptations that
support myocardial growth and function. The reduced
circulating anabolic factors (Fig. 6) present in IUGR
would be expected to suppress cardiac growth, as the
fetal heart is sensitive to insulin and IGF-1, and reliant
on glucose and lactate as major carbon sources for
metabolism (Bartelds et al., 1998, 2000). The
myocardium of the IUGR fetus responds by increasing
plasma membrane concentrations of the insulin-
stimulated glucose transporter 4 and insulin receptor β
protein (Barry et al., 2006). In contrast, myocardial
membrane protein concentrations of glucose transporter
1 are unchanged in the IUGR fetus. Additionally, blood
flow to, and glucose delivery/uptake by, the left
ventricle is significantly increased by insulin stimulation
(Barry et al., 2016). These adaptations appear to
promote myocardial energy supply and utilization by
increasing its sensitivity to insulin, which supports
cardiac growth despite the significant nutrient
deprivation.
Fetal cardiac function appears to be relatively
unaffected by placental insufficiency because heart rates
are similar between IUGR and control fetuses (Galan et
al., 2005; Barry et al., 2016). However, IUGR fetuses
have increased indices of umbilical artery resistance and
a significant reduction in umbilical blood flow, which
did not always cause greater mean aortic blood pressure
(Galan et al., 2005; Barry et al., 2016). The increased
placental vascular resistance might reflect a mechanism
by which the fetus is able to increase extraction of
nutrients from the placenta as discussed above. A similar
model that induces IUGR by removing the majority of
the uterine caruncles in the sheep also found no
difference in fetal blood pressure under baseline
conditions; however, there was a greater hypotensive
effect in IUGR fetuses following administration of
phentolamine, an α-adrenergic antagonist, and captopril,
an angiotensin-converting enzyme inhibitor (Edwards et
al., 1999; Danielson et al., 2005). These findings indicate
that the α-adrenergic and renin-angiotensin systems have
a greater role in blood pressure maintenance in IUGR
fetuses, which are likely involved in mediating some of
the organ sparing phenomena present in this model
(McMillen et al., 2001; Danielson et al., 2005).
Conclusions
We have presented evidence for prolonged
exposure to heat stress causing placental insufficiency in
ruminants. Maladaptive responses during development,
which include fetal growth restriction, persist as lifelong
deficiencies lowering the performance and health of the
animal. We discuss how the enlarged transplacental
gradient for oxygen and glucose facilitates umbilical
uptakes but results in low blood oxygen and plasma
glucose concentration in the fetus. These conditions
slow growth by altering glucose metabolism, decreasing
amino acid clearance, and decreasing anabolic
hormones while increasing catabolic hormones.
Limesand et al. Heat stress during pregnancy.
896 Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018
Acknowledgements
Funding for this work was from the National
Institutes of Health R01 DK084842 (S.W. Limesand,
Principal Investigator). L.E. Camacho was supported by
Award 2015-03545 (L.E. Camacho, Principal
Investigator) from the National Institute of Food and
Agriculture, USDA. A.T. Antolic was supported by T32
HL7249 (J. Burt, Principal Investigator). We thank all
our colleagues who contributed to the seminal studies
reported in this review. The content is solely the
responsibility of the authors and does not necessarily
represent the official views of the funding agencies.
References
Abdalla EB, Kotby EA, Johnson HD. 1993.
Physiological responses to heat-induced hyperthermia
of pregnant and lactating ewes Small Rumin Res,
11:125-134.
Ahmed BMS, Younas U, Asar TO, Dikmen S Hansen
PJ, Dahl GE. 2017. Cows exposed to heat stress during
fetal life exhibit improved thermal tolerance. J Anim
Sci, 95:3497-3503.
Aldoretta PW, Carver TD, Hay WW Jr. 1998.
Maturation of glucose-stimulated insulin secretion in
fetal sheep. Biol Neonate, 73:375-386.
Allen RE, Merkel RA, Young RB. 1979. Cellular
aspects of muscle growth: myogenic cell proliferation. J
Anim Sci, 49:115-127.
Anderson AH, Fennessey PV, Meschia G, Wilkening
RB, Battaglia FC. 1997. Placental transport of
threonine and its utilization in the normal and growth-
restricted fetus. Am J Physiol, 272:E892-E900.
Barash H, Silanikove N, Shamay A, Ezra E. 2001.
Interrelationships among ambient temperature, day
length, and milk yield in dairy cows under a
Mediterranean climate. J Dairy Sci, 84:2314-2320.
Barry JS, Davidsen ML, Limesand SW, Galan HL,
Friedman JE, Regnault TR, Hay WW Jr. 2006.
Developmental changes in ovine myocardial glucose
transporters and insulin signaling following
hyperthermia-induced intrauterine fetal growth
restriction. Exp Biol Med (Maywood), 231:566-575.
Barry JS, Rozance PJ, Anthony RV. 2008. An animal
model of placental insufficiency-induced intrauterine
growth restriction. Semin Perinatol, 32:225-230.
Barry JS, Rozance PJ, Brown LD, Anthony RV,
Thornburg KL, Hay WW Jr. 2016. Increased fetal
myocardial sensitivity to insulin-stimulated glucose
metabolism during ovine fetal growth restriction. Exp
Biol Med (Maywood), 241:839-847.
Bartelds B, Gratama JW, Knoester H, Takens J,
Smid GB, Aarnoudse JG, Heymans HS, Kuipers JR.
1998. Perinatal changes in myocardial supply and flux
of fatty acids, carbohydrates, and ketone bodies in
lambs. Am J Physiol, 4(6, pt. 2):H1962-1969.
Bartelds B, Knoester H, Smid GB, Takens J, Visser
GH, Penninga L, van der Leij FR, Beaufort-Krol
GC, Zijlstra WG, Heymans HS, Kuipers JR. 2000.
Perinatal changes in myocardial metabolism in lambs.
Circulation, 102:926-931.
Battaglia FC, Meschia G. 1978. Principal substrates of
fetal metabolism. Physiol Rev, 58:499-527.
Bell AW, Wilkening RB, Meschia G. 1987. Some
aspects of placental function in chronically heat-stressed
ewes. J Dev Physiol, 9:17-29.
Boehmer BH, Limesand SW, Rozance PJ. 2017. The
impact of IUGR on pancreatic islet development and
beta-cell function. J Endocrinol, 235:R63-R76.
Brown LD, Rozance PJ, Thorn SR, Friedman JE,
Hay WW Jr. 2012. Acute supplementation of amino
acids increases net protein accretion in IUGR fetal
sheep. Am J Physiol Endocrinol Metab, 303:E352-64.
Brown LD. 2014. Endocrine regulation of fetal skeletal
muscle growth: impact on future metabolic health. J.
Endocrinol, 221:R13-R29.
Brown LD, Rozance PJ, Bruce JL, Friedman JE,
Hay WW Jr, Wesolowski SR. 2015. Limited capacity
for glucose oxidation in fetal sheep with intrauterine
growth restriction. Am J Physiol Regul Integr Comp
Physiol, 309:R920-R928.
Brown LD, Davis M, Wai S, Wesolowski SR, Hay
WW Jr, Limesand SW, Rozance PJ. 2016.
Chronically Increased Amino Acids Improve Insulin
Secretion, Pancreatic Vascularity, and Islet Size in
Growth-Restricted Fetal Sheep. Endocrinology,
157:3788-3799.
Brown LD, Kohn, JR, Rozance PJ, Hay WW Jr,
Wesolowski SR. 2017. Exogenous amino acids
suppress glucose oxidation and potentiate hepatic
glucose production in late gestation fetal sheep. Am J
Physiol Regul Integr Comp Physiol, 312:R654-R663.
Camacho LE, Chen X, Hay WW Jr, Limesand SW.
2017. Enhanced insulin secretion and insulin sensitivity
in young lambs with placental insufficiency-induced
intrauterine growth restriction. Am J Physiol Regul
Integr Comp Physiol, 313:R101-R109.
Chen X, Fahy AL, Green AS, Anderson MJ, Rhoads
RP, Limesand SW. 2010. beta2-Adrenergic receptor
desensitization in perirenal adipose tissue in fetuses and
lambs with placental insufficiency-induced intrauterine
growth restriction. J Physiol, 588:3539-3549.
Chen X, Green AS, Macko AR, Yates DT, Kelly AC,
Limesand SW. 2014. Enhanced insulin secretion
responsiveness and islet adrenergic desensitization after
chronic norepinephrine suppression is discontinued in fetal
sheep. Am J Physiol Endocrinol Metab, 306:E58-E64.
Chen X, Kelly AC, Yates DT, Macko AR, Lynch
RM, Limesand SW. 2017. Islet adaptations in fetal
sheep persist following chronic exposure to high
norepinephrine. J Endocrinol, 232:285-295.
Collier RJ, Dahl GE, VanBaale MJ. 2006. Major
advances associated with environmental effects on dairy
cattle. J Dairy Sci, 89:1244-1253.
Danielson L, McMillen IC, Dyer JL, Morrison JL.
2005. Restriction of placental growth results in greater
hypotensive response to alpha-adrenergic blockade in
fetal sheep during late gestation. J Physiol, 563:611-620.
Davis MA, Macko AR, Steyn LV, Anderson MJ,
Limesand SW. 2015. Fetal adrenal demedullation
lowers circulating norepinephrine and attenuates growth
restriction but not reduction of endocrine cell mass in an
ovine model of intrauterine growth restriction.
Nutrients, 7:500-516.
de Vrijer B, Regnault TR, Wilkening RB, Meschia
Limesand et al. Heat stress during pregnancy.
Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018 897
G, Battaglia FC. 2004. Placental uptake and transport
of ACP, a neutral nonmetabolizable amino acid, in an
ovine model of fetal growth restriction. Am J Physiol
Endocrinol Metab, 287:E1114-1124
de Vrijer B, Davidsen ML, Wilkening RB, Anthony
RV, Regnault TR. 2006. Altered placental and fetal
expression of IGFs and IGF-binding proteins associated
with intrauterine growth restriction in fetal sheep during
early and mid-pregnancy. Pediatr Res, 60:507-512.
do Amaral BC, Connor EE, Tao S, Hayen MJ,
Bubolz JW, Dahl GE. 2011. Heat stress abatement
during the dry period influences metabolic gene
expression and improves immune status in the transition
period of dairy cows. J Dairy Sci, 94:86-96.
Dunlop K, Cedrone M, Staples JF, Regnault TR.
2015. Altered fetal skeletal muscle nutrient metabolism
following an adverse in utero environment and the
modulation of later life insulin sensitivity. Nutrients,
7:1202-1216.
Edwards LJ, Simonetta G, Owens JA, Robinson JS,
McMillen IC. 1999. Restriction of placental and fetal
growth in sheep alters fetal blood pressure responses to
angiotensin II and captopril. J Physiol, 515:897-904.
Galan HL, Hussey MJ, Barbera A, Ferrazzi E,
Chung M, Hobbins JC, Battaglia FC. 1999.
Relationship of fetal growth to duration of heat stress in
an ovine model of placental insufficiency. Am J Obstet
Gynecol, 180:1278-1282.
Galan HL, Anthony RV, Rigano S, Parker TA, de
Vrijer B, Ferrazzi E Wilkening RB, Regnault TR.
2005. Fetal hypertension and abnormal Doppler
velocimetry in an ovine model of intrauterine growth
restriction. Am J Obstet Gynecol, 192:272-279.
Gatford KL, De Blasio MJ, Thavaneswaran P,
Robinson JS, McMillen IC, Owens JA. 2004.
Postnatal ontogeny of glucose homeostasis and insulin
action in sheep. Am J Physiol Endocrinol Metab,
286:E1050-E1059.
Greenwood PL, Hunt AS, Hermanson JW, Bell AW.
1998. Effects of birth weight and postnatal nutrition on
neonatal sheep: I. Body growth and composition, and
some aspects of energetic efficiency. J Anim Sci,
76:2354-2367.
Greenwood PL, Hunt AS, Hermanson JW, Bell AW.
2000. Effects of birth weight and postnatal nutrition on
neonatal sheep: II. Skeletal muscle growth and
development. J Anim Sci, 78:50-61.
Hagen AS, Orbus RJ, Wilkening RB, Regnault TR,
Anthony RV. 2005. Placental expression of
angiopoietin-1, angiopoietin-2 and tie-2 during placental
development in an ovine model of placental
insufficiency-fetal growth restriction. Pediatr Res,
58:1228-1232.
Hansen PJ, Drost M, Rivera RM, Paula-Lopes FF,
al-Katanani YM, Krininger CE, 3rd Chase CC Jr.
2001. Adverse impact of heat stress on embryo
production: causes and strategies for mitigation.
Theriogenology, 55:91-103.
Hay WW Jr, Brown LD, Rozance PJ, Wesolowski
SR, Limesand SW. 2016. Challenges in Nourishing the
IUGR fetus-lessons learned from studies in the IUGR
fetal sheep. Acta Paediatr, 105:881-889.
Jackson BT, Piasecki GJ, Cohn HE, Cohen WR.
2000. Control of fetal insulin secretion. Am J Physiol
Regul Integr Comp Physiol, 279:R2179-R2188.
Kelly AC, Bidwell CA, McCarthy FM, Taska DJ,
Anderson MJ, Camacho LE, Limesand SW. 2017.
RNA Sequencing exposes adaptive and immune
responses to intrauterine growth restriction in fetal
sheep islets. Endocrinology, 158:743-755.
Kelly AC, Camacho LE, Pendarvis K, Davenport
HM, Steffens NR, Smith KE, Weber CS, Lynch RM,
Papas KK, Limesand SW. 2018. Adrenergic receptor
stimulation suppresses oxidative metabolism in isolated
rat islets and Min6 cells. Mol Cell Endocrinol. doi:
10.1016/j.mce.2018.01.012.
Laporta J, Fabris TF, Skibiel AL, Powell JL, Hayen
MJ, Horvath K, Miller-Cushon EK, Dahl GE. 2017.
In utero exposure to heat stress during late gestation has
prolonged effects on the activity patterns and growth of
dairy calves. J Dairy Sci, 100:2976-2984.
Leos RA, Anderson MJ, Chen X, Pugmire J,
Anderson KA, Limesand SW. 2010. Chronic exposure
to elevated norepinephrine suppresses insulin secretion
in fetal sheep with placental insufficiency and
intrauterine growth restriction. Am J Physiol Endocrinol
Metab, 298:E770-E778.
Limesand SW, Regnault TR, Hay WW Jr. 2004.
Characterization of glucose transporter 8 (GLUT8) in
the ovine placenta of normal and growth restricted
fetuses. Placenta 25, 70-77.
Limesand SW, Jensen J, Hutton JC, Hay WW Jr.
2005. Diminished b-cell replication contributes to
reduced b-cell mass in fetal sheep with intrauterine
growth restriction. Am J Physiol Regul Integr Comp
Physiol, 288, R1297-R1305.
Limesand SW, Rozance PJ, Zerbe GO, Hutton JC,
Hay WW Jr. 2006. Attenuated insulin release and
storage in fetal sheep pancreatic islets with intrauterine
growth restriction. Endocrinology, 147:1488-1497.
Limesand SW, Rozance PJ, Smith, D, Hay WW Jr.
2007. Increased insulin sensitivity and maintenance of
glucose utilization rates in fetal sheep with placental
insufficiency and intrauterine growth restriction. Am J
Physiol Endocrinol Metab, 293:E1716-E1725.
Limesand SW, Rozance PJ, Macko AR, Anderson
MJ, Kelly AC, Hay WW Jr. 2013. Reductions in
insulin concentrations and beta-cell mass precede
growth restriction in sheep fetuses with placental
insufficiency. Am J Physiol Endocrinol Metab,
304:E516-E523.
Limesand SW, Rozance PJ. 2017. Fetal adaptations in
insulin secretion result from high catecholamines during
placental insufficiency. J Physiol 595, 5103-5113.
Macko AR, Yates DT, Chen X, Green AS, Kelly AC,
Brown LD, Limesand SW. 2013. Elevated plasma
norepinephrine inhibits insulin secretion, but adrenergic
blockade reveals enhanced b-cell responsiveness in an
ovine model of placental insufficiency at 0.7 of
gestation. J Dev Orig Health Dis, 4:402-410.
Macko AR, Yates DT, Chen X, Shelton LA, Kelly
AC, Davis MA, Camacho LE, Anderson MJ,
Limesand SW. 2016. Adrenal demedullation and
oxygen supplementation independently increase
glucose- stimulated insulin concentrations in fetal sheep
with intrauterine growth restriction. Endocrinology,
Limesand et al. Heat stress during pregnancy.
898 Anim. Reprod., v.15, (Suppl.1), p.886-898. 2018
157:2104-2115.
Maier A, McEwan JC, Dodds KG, Fischman DA,
Fitzsimons RB, Harris AJ. 1992. Myosin heavy chain
composition of single fibres and their origins and
distribution in developing fascicles of sheep tibialis
cranialis muscles. J Muscle Res Cell Motil, 13:551-572.
McCrabb GJ, McDonald BJ, Hennoste LM. 1993.
Lamb birthweight in sheep differently acclimatized to a
hot environment. Aust J Agric Res, 44:933-943.
McMillen IC, Adams MB, Ross JT, Coulter CL,
Simonetta G, Owens JA, Robinson JS, Edwards LJ.
2001. Fetal growth restriction: adaptations and
consequences. Reproduction, 122:195-204.
Meschia G, Cotter JR, Breathnach CS, Barron DH.
1965. The diffusibility of oxygen across the sheep
placenta. Q J Exp Physiol Cogn Med Sci, 50:466-480.
Monteiro APA, Tao S, Thompson IMT, Dahl GE.
2016. In utero heat stress decreases calf survival and
performance through the first lactation. J Dairy Sci,
99:8443-8450.
Pavlath GK, Rich K, Webster SG, Blau HM. 1989.
Localization of muscle gene products in nuclear
domains. Nature, 337:570-573.
Regnault TR, Galan HL, Parker TA, Anthony RV.
2002a. Placental development in normal and
compromised pregnancies. Placenta, 23(suppl.
A):S119-S129.
Regnault TR, Orbus RJ, de Vrijer B, Davidsen ML,
Galan HL, Wilkening RB, Anthony RV. 2002b.
Placental expression of VEGF, PlGF and their receptors
in a model of placental insufficiency-intrauterine growth
restriction (PI-IUGR). Placenta, 23:132-144.
Regnault TR, de Vrijer B, Galan HL, Davidsen ML,
Trembler KA, Battaglia FC, Wilkening RB, Anthony
RV. 2003. The relationship between transplacental O2
diffusion and placental expression of PlGF, VEGF and
their receptors in a placental insufficiency model of fetal
growth restriction. J Physiol, 550:641-656.
Regnault TR, Friedman JE, Wilkening RB, Anthony
RV, Hay WW Jr. 2005. Fetoplacental transport and
utilization of amino acids in IUGR--a review. Placenta,
26(suppl. A):S52-62.
Regnault TR, de Vrijer B, Galan HL, Wilkening RB,
Battaglia FC, Meschia G. 2007. Development and
mechanisms of fetal hypoxia in severe fetal growth
restriction. Placenta, 28:714-723.
Regnault TR, de Vrijer B, Galan HL, Wilkening RB,
Battaglia FC, Meschia G. 2013. Umbilical uptakes and
transplacental concentration ratios of amino acids in
severe fetal growth restriction. Pediatr Res, 73:602-611.
Reynolds LP, Ferrell CL, Nienaber JA, Ford SP.
1985. Effects of chronic enviromental heat stress on
blood flow and nutrient uptake of the gravid bovine
uterus and foetus. J Agric Sci, 104:289-297.
Ross JC, Fennessey PV, Wilkening RB, Battaglia
FC, Meschia G. 1996. Placental transport and fetal
utilization of leucine in a model of fetal growth
retardation. Am J Physiol, 270:E491-E503.
Rozance PJ, Zastoupil L, Wesolowski SR,
Goldstrohm DA, Strahan B, Cree-Green M,
Sheffield-Moore M, Meschia G, Hay WW Jr,
Wilkening RB, Brown LD. 2018. Skeletal muscle
protein accretion rates and hindlimb growth are reduced
in late gestation intrauterine growth-restricted fetal
sheep. J Physiol, 596:67-82.
Shelton M. 1964. Relation of birth weight to death
losses and to certain productive characters of fall-born
lambs. J Anim Sci, 23:355-359.
Thorn SR, Regnault TR, Brown LD, Rozance PJ,
Keng J, Roper M, Wilkening RB, Hay WW Jr,
Friedman JE. 2009. Intrauterine growth restriction
increases fetal hepatic gluconeogenic capacity and
reduces messenger ribonucleic acid translation initiation
and nutrient sensing in fetal liver and skeletal muscle.
Endocrinology, 150:3021-3030.
Thorn SR, Brown LD, Rozance PJ, Hay WW Jr,
Friedman JE. 2013. Increased hepatic glucose
production in fetal sheep with intrauterine growth
restriction is not suppressed by insulin. Diabetes, 62:65-
73.
Thureen PJ, Trembler KA, Meschia G, Makowski
EL, Wilkening RB. 1992. Placental glucose transport
in heat-induced fetal growth retardation. Am J Physiol,
263:R578-R585.
Vatnick I, Ignotz G, McBride BW, Bell AW. 1991.
Effect of heat stress on ovine placental growth in early
pregnancy. J Dev Physiol, 16:163-166.
Wallace JM, Regnault TR, Limesand SW, Hay WW
Jr, Anthony RV. 2005. Investigating the causes of low
birth weight in contrasting ovine paradigms. J Physiol,
565:19-26.
Wells JC, Cole TJ. 2002. Birth weight and
environmental heat load: a between-population analysis.
Am J Phys. Anthropol, 119:276-282.
West JW. 2003. Effects of heat-stress on production in
dairy cattle. J Dairy Sci, 86:2131-2144.
Wilkening RB, Anderson S, Martensson L, Meschia
G. 1982. Placental transfer as a function of uterine
blood flow. Am J Physiol, 242:H429-H436.
Wilson SJ, McEwan JC, Sheard PW, Harris AJ.
1992. Early stages of myogenesis in a large mammal:
formation of successive generations of myotubes in
sheep tibialis cranialis muscle. J Muscle Res Cell Motil,
13:534-550.
Wu G, Bazer FW, Wallace JM, Spencer TE. 2006.
Board-invited review: intrauterine growth retardation:
implications for the animal sciences. J Anim Sci,
84:2316-2337.
Yates DT, Macko AR, Nearing M, Chen X, Rhoads
RP, Limesand SW. 2012. Developmental programming
in response to intrauterine growth restriction impairs
myoblast function and skeletal muscle metabolism. J
Pregnancy, 2012:631038. doi: 10.1155/2012/631038.
Yates DT, Clarke DS, Macko AR, Anderson MJ,
Shelton LA, Nearing M, Allen RE, Rhoads RP,
Limesand SW. 2014. Myoblasts from intrauterine
growth-restricted sheep fetuses exhibit intrinsic
deficiencies in proliferation that contribute to smaller
semitendinosus myofibres. J Physiol, 592:3113-3125.
Yates DT, Cadaret CN, Beede KA, Riley HE, Macko
AR, Anderson MJ, Camacho LE, Limesand SW.
2016. Intrauterine growth-restricted sheep fetuses
exhibit smaller hindlimb muscle fibers and lower
proportions of insulin-sensitive Type I fibers near term.
Am J Physiol Regul Integr Comp Physiol, 310:R1020-
R1029.
