Placental
amino
acid transport
AARON J. MOE
The Edward
Mallinkrodt
Department
of Pediatrics,
Children’s
Washington
University
School of Medicine, St. Louis, Missouri
Hospital,
63110
Moe, Aaron
J. Placental amino acid transport.
Am. J. Physiol. 268
(CeZZ PhysioZ. 37): C1321-C1331,
1995. -Normal
fetal growth and development depend on a continuous
supply of amino acids from the mother to
the fetus. The placenta is responsible
for the transfer
of amino acids
between the two circulations.
The human placenta is hemomonochorial,
meaning that the maternal and fetal circulations
are separated by a single
layer of polarized
epithelium
called the syncytiotrophoblast,
which is in
direct contact with maternal
blood. Transport
proteins
located in the
microvillous
and basal membranes
of the syncytiotrophoblast
are the
principal
mechanism
for transfer
from maternal
blood to fetal blood.
Knowledge
of the function and regulation
of syncytiotrophoblast
amino
acid transporters
is of great importance
in understanding
the mechanism
of placental
transport
and potentially
improving
fetal and newborn
outcomes. The development
of methods for the isolation of microvillous
and basal membrane
vesicles from human placenta over the past two
decades has contributed
greatly to this understanding.
Now a primary
cultured trophoblast
model is available to study amino acid transport
and
regulation
as the cells differentiate.
The types of amino acid transporters
and their distribution
between the syncytiotrophoblast
microvillous
and
basal membranes
are somewhat
unique compared with other polarized
epithelia. These differences may reflect the unusual circumstance
of this
epithelium
that is exposed to blood on both sides. The current state of
knowledge as to the types of transport systems present in syncytiotrophoblast, their regulation,
and the effects of maternal consumption
of drugs
on transport are discussed.
syncytiotrop
epithelium
hoblast;
microvillous
membrane;
THE PLACENTA IS A specialized organ of fetal origin that
performs numerous functions critical to normal growth
and development of the fetus (24). The nature and
importance of each function change with the intrauterine development of the fetus. The placental function of
nutrient. transport from maternal blood to the fetal
circulation is important throughout pregnancy and particularly in the last trimester as fetal size increases
rapidly. Amino acids are an essential nutrient class,
since they provide the building blocks for protein synthesis for the rapidly growing fetus. Normal fetal growth
and development depend on a continuous supply of
amino acids from maternal blood, involving uptake and
transfer across the syncytiotrophoblast (87).
The maternal and fetal circulations in the human are
separated by a polarized epithelium known as the
syncytiotrophoblast (Fig. 1). Although resembling other
epithelia including that of intestine, placental epithelium is a true syncytium, lacking lateral cell membranes.
Undifferentiated stem cells (cellular trophoblast or cytotrophoblast) underlying this epithelium give rise to it by
fusion (24). The apical membrane of the syncytiotropho0363-6143/95
$3.00
Copyright
o
basal membrane;
polarized
blast is in direct contact with maternal blood (hemochorial) and is composed of a microvillous structure resembling morphologically
the apical membrane of the
enterocyte. The basal membrane faces the fetal circulation and lacks the high degree of organization of the
apical membrane. The trophoblast layer is the most
metabolically active tissue of the placenta, even though
it comprises only 13% of placental weight (88). The
syncytiotrophoblast
is the anatomic portion of the human placenta that mediates the exchange of nutrients
and waste products between maternal and fetal circulations.
TRANSPLACENTAL
PATHWAYS
Pathways of transsyncytial flux may be divided into
either transcellular or paracellular mechanisms. Transcellular mechanisms may include simple diffusion down
a concentration gradient or transfer by specific membrane-bound transport proteins (usually described as
transport systems). The primary mechanism for the
movement of amino acids from mother to fetus is
1995 the American
Physiological
Society
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REVIEW
MicroviIIous
Fig. 1. Simplified
drawing
of the human placental
villous structure.
ST, syncytiotrophoblast;
CT, cytotrophoblast;
FV, fetal vessel.
thought to be a result of the functioning of specific
amino acid transporters located in the microvillous and
basal membranes of the syncytiotrophoblast (i.e., mediated transport). Maintenance of electrochemical gradients of sodium, potassium, chloride, and other ions
provides the driving force for the operation of many of
these transporters, whereas others operate by exchange
mechanisms.
The paracellular pathway in the human placenta may
represent sparsely distributed but relatively wide pores
that transverse the syncytiotrophoblast
(124, 125).
Transepithelial pores that potentially represent the
morphological structure responsible for paracellular
transport have been described in the hemomonochorial
placenta of the degu (Octodon degus) and of humans
(33, 66). Paracellular transport can occur by diffusion
from areas of higher concentration to areas of lower
concentration or by bulk volume flow (126). Paracellular
pathways predominate for transplacental flux of the
inert hydrophilic solutes, mannitol, Cr-EDTA, and inulin (126). It is likely that paracellular pathways account
for a negligible proportion of flux by small molecules in
relatively low plasma concentrations such as amino
acids.
The hypothesis that specific transport proteins in the
two membranes of the syncytiotrophoblast are primarily responsible for the maternal-to-fetal
transfer of
amino acids is based partially on the fact that most
amino acids are in higher concentration in fetal blood
compared with maternal blood and in even higher
concentrations in the placenta (23, 25, 77, 128, 133).
Therefore the transepithelial flux of most amino acids is
against a concentration gradient that would necessitate
a concentrative mechanism of transfer rather than
simple diffusion. As an example, the mean fetal plasma
lysine concentration for the five published studies (23,
25,77,128,133) was threefold higher than the maternal
plasma lysine concentration (92). For alanine, the concentration in fetal blood has been estimated to be twice
that in maternal blood. Syncytial alanine concentration
is sevenfold higher compared with maternal blood (92).
There are a few exceptions, like glutamate and aspartate, which are found in roughly equal concentrations in
each circulation (92).
TECHNIQUES
TO STUDY
PLACENTAL
TRANSPORT
As the understanding of placental amino acid transport processes has grown, it has become apparent that,
for the most part, the transporters present are not
unique to the placenta but are types common to other
cells and tissues. The distribution of these transporters
between the apical and basal membranes of the syncytiotrophoblast may be different from other epithelia, and
the role these transporters play in placental function is
unique. Knowledge of these processes has come with the
development of in vitro techniques to study the placental villous structure and specifically the two plasma
membranes of the syncytiotrophoblast.
Placental amino
acid transport has been investigated using villous fragments or slices (35,113-115), perfused placental cotyledon (48, 105-108), isolated plasma membrane vesicles
from the microvillous (43, 58, 82, 85, 100) and basal
membranes (40, 49, 50, 70, 71), and primary cultured
trophoblast cells (42, 60) as well as choriocarcinoma cell
lines (42, 73, 84, 97). Chronically catheterized sheep
have proved to be a very important model for whole
animal studies (45,51, 74,80,85,86). It should be noted
that the sheep placenta is a cotyledonary epitheliochorial (the trophoblast is not in direct contact with maternal blood) type placenta, whereas the human placenta is
discoidal and hemomonochorial (meaning the tropho-
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blast is in direct contact with maternal
blood) (63).
Several studies have been published using the perfused
guinea pig placenta model (31, 131, 134). Although the
guinea pig placenta does not resemble the human placenta, it is similar, in that they are both of the hemochorial type (63). The study of placental transport
necessitates the use of different
models, and they have all
contributed
to our understanding
of placental amino
acid transport.
Investigators
should consider
species
differences
in placental structure
and morphology
as
well as the fact that length of gestation,
fetal growth
rates, and fetal composition vary among species (6).
A significant achievement in the development of appropriate placental models was the isolation of relatively
pure membrane vesicles from the syncytiotrophoblast
microvillous
(10, 117, 119) membrane
and later from
the basal membrane (54, 64) of human placenta. Membrane vesicles allow investigators
to study the plasma
membrane transporters
more directly, free from cellular
metabolism,
and in membranes
specific to the syncytiotrophoblast
of the human placenta. Because transporter
distribution
is polarized, it is helpful to investigate each
membrane domain separately to determine which transport systems are present for a given substrate.
Using
these established procedures
(10, 64) as well as a later
method (54), investigators
routinely isolate paired microvillous and basal membranes
from the same placenta.
Subsequent
transport
experiments
can then directly
compare the two membranes.
The amino acid transporters
found in the microvillous
and basal membranes
of the syncytiotrophoblast
have
largely been identified
by the classical technique
of
functional
characterization
as originally
described by
Christiansen
and co-workers
(16-21).
For the two decades since the first isolation of placental microvillous
membrane vesicles, a good deal of effort has been applied
to identification
of amino acid transport
systems. Although this work continues, in recent years a new phase
has begun with the development
of primary
cultured
trophoblasts
(67) as a model to investigate
amino acid
transport
(42, 60). The trophoblast
model is attractive
because it allows investigation
of regulatory
factors and
changes in transporter
distributions
that occur in conjunction with cellular differentiation
in vitro. The cultured trophoblast
model has added another powerful
tool for the study of placental amino acid transport.
Recently, there have been advances in applying modern molecular biology techniques
to the task of transporter discovery and subsequent
study of function and
regulatory
mechanisms.
Thus far, most transporters
have been cloned in tissues other than placenta, but
these techniques
are now being applied to problems in
placental amino acid transport
as well. It will be possible, for the first time, to study specific placental amino
acid transport
proteins
in pure form, independent
of
other related transporters.
It should also be possible to
make inferences about transporter
function and regulation directly from transport
protein structure.
In this review the location, type, and functional
characteristics
of amino acid transport
systems
are
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discussed with respect to the classical divisions based on
preferred
substrates.
Regulatory
factors
and recent
advances using molecular biology techniques are considered. Finally, the effects of factors in pregnancy
(i.e.,
alcohol, smoking, drugs) on placental amino acid transport are examined.
NEUTRAL
AMINO
ACID
TRANSPORT
SYSTEMS
The neutral amino acid transport
systems identified
in the placental syncytiotrophoblast
are common to
many cell types. In this respect, the syncytiotrophoblast
differs from intestine and kidney, which possess specialized epithelial transport
systems
(122). The known
syncytial amino acid transporters
can be found listed in
Table 1, according to the nomenclature
originally developed by Christiansen.
Throughout
this article the convention of designating
sodium-dependent
transporters
with upper case letters and sodium-independent
transporters by lower case letters is followed.
There are at least two sodium-dependent
and two
sodium-independent
neutral amino acid systems found
in the microvillous
membrane.
These are the sodiumdependent system A and system N and the sodiumindependent system 1 (formerly
referred to as system L)
system, which pre(58). A set on d sodium-independent
fers alanine, has also been proposed but is not well
characterized
(58). The A system resembles that found
in other cell types, in that it has broad substrate
specificity and is sensitive to N-methylated
amino acids
(58) such as a-(methylamino)isobutyric
acid (MeAIB),
which is considered to be a specific system A analogue.
System N transports
histidine
and glutamine
and is
insensitive
to methylated
amino acids (62). System N is
sodium dependent,
electrogenic,
and transstimulated.
The sodium-independent
system 1 was identified by its
sensitivity
to the amino acid analogue
2-amino-2norbornane
carboxylic
acid and by its high affinity
(apparent Michaelis constant 20 PM) for leucine uptake
(58) .
Table 1. Amino acid transport
in syncytiotrophoblast
System
systems
Substrates
Excludes
A
Neutral
AA,
MeAIB
ASC
Ala, Ser, Cys,
anionic AA
His, Gln
Tau
Asp, Glu
Leu, BCH
LYS, At-g
LYS, kg
LYS, f%T
Anionic
and
cationic AA,
BCH, Leu
MeAIB,
cationic
AA, Pro
MeAIB,
Cys
(X-AA
Nonanionic
AA
MeAIB,
Pro
Anionic
AA
Anionic
AA
Anionic
AA
N
P
FG
Y+
bo, +
YfL
found
Membrane
M, B
B
M
M
M, B
M, B
M, B
B
M
Reference
41,49
41
53
70
42,72
41,49
27,33
33
27
Dicke et al. (29) have suggested
the presence
of system
GLY in
microvillous
membrane
vesicles
but this has not been confirmed.
System t has been excluded
from microvillous
(M) membrane
(33) but
may be present
in basal (B) membrane
(58). An uncharacterized
sodium-independent
neutral
amino acid transporter
has been reported
in microvillous
membrane
(47). MeAIB,
ar-(methylamino)isobutyric
acid; BCH, 2-amino-2-norbornane
carboxylic
acid. AA, amino acids.
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INVITED
The microvillous
membrane apparently lacks system
ASC (42,58,71)
or system t for tyrosine uptake (43,72).
Sodium-dependent
cysteine transport
(i.e., suggestive of
ASC transport)
was reported in an investigation
of the
importance of storage conditions on the results of amino
acid transport
studies with placental microvillous
membrane vesicles. Furesz et al. (42) did not observe ASC
system transport
in freshly prepared microvillous
membrane vesicles as measured by MeAIB-resistant
sodiumdependent alanine uptake. In the same report, measurable ASC activity was lost from cultured trophoblast
cells and BeWo cells after differentiation
and syncytial
formation
(42). These data suggest that ASC system
activity decreases with syncytial formation and is consistent with the absence of the ASC system from the
microvillous
membrane. A portion of glycine uptake by
microvillous
membranes is MeAIB resistant, which suggests some non-system
A uptake that perhaps represents system GLY activity (29).
The basal membrane of the syncytiotrophoblast
possesses system A and system 1 transporters
and, in
addition, the sodium-dependent
system ASC (50). System ASC is characterized
by insensitivity
to the analogue MeAIB and has somewhat
narrower
substrate
specificity compared with system A. Alanine, serine, and
cysteine are preferred
substrates
and thus give this
system its name. There has been a suggestion but not
confirmation
that the basal membrane
also possesses
the sodium-independent
system t for tyrosine transport
(71).
ANIONIC
AMINO
ACIDS
Placental transport
of anionic amino acids is unique
because, unlike other amino acids, glutamate and aspartate are not transferred
across the placenta (76, 105,
121). Net uptake of glutamate from the fetal circulation
has been observed in some studies (51, 76, 81). The
ovine fetal liver is a net producer of glutamate (81). The
placental tissue concentration
of glutamate is orders of
magnitude
higher than concentrations
in either the
maternal or fetal circulation
(92). Blood concentrations
are roughly equal on both sides of the syncytiotrophoblast. The elevation of glutamate concentration
within
the placenta is accomplished by sodium-dependent
potassium-coupled
transporters
located in the microvillous
and basal membranes (49, 53,SS). In functional studies,
this transporter
has been identified as the fairly ubiquitous Xio system (49,85). A distinguishing
characteristic
of the Xio system is that it is stereoselective
for the
stereoisomers
of glutamate
but does not discriminate
between L- and D-aspartate. The normal orientation of
sodium and potassium gradients in the syncytiotrophoblast of the placenta should drive concentrative uptake
of anionic amino acids into the placenta. There the
amino acids would be disposed of via metabolism, since
an effective means of egress has not been found. Recent
studies indicate an extremely high glutamate clearance
rate from the fetal circulation, which is the result of
glutamate disposal via oxidation to COZ (87). The main
route of glutamate metabolism in human placental
trophoblast also seems to be oxidation to COZ (13). The
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unique placental handling of glutamate may offer the
developing fetal brain a protective mechanism from a
potential neurotoxin. Consistent with this hypothesis is
the recent observation in my laboratory that the human
placental trophoblast expresses both of the brain type
sodium-dependent glutamate transporters. In addition,
glutamate oxidation may provide an important energy
source to the placenta (13, 87) or provide a source of
NADPH for placental lipid synthesis (86). The full-term
placenta is an active producer of fatty acids and steroid
hormones but has a low level of pentose phosphate
pathway activity (83). Oxidation of glutamate via glutamate dehydrogenase (83) could provide the needed
NADPH for fatty acid and cholesterol sysnthesis.
CATIONIC
AMINO
ACID
TRANSPORTERS
The cationic amino acids, like most of the neutral
amino acids, are concentrated in the fetal circulation
relative to concentrations in maternal circulation. Several types of cationic transporters seem to be present in
the two membranes of the syncytiotrophoblast.
The
most widely known is system y+, which is found in many
cell types and in both membranes of the syncytiotrophoblast (34, 40, 41). This transporter is sodium independent, has a relatively low affinity and high capacity, and
is specific for cationic amino acids, with the exception
that lysine uptake can be inhibited by certain neutral
amino acids in the presence of sodium (40). Recent work
in this laboratory suggests that the y+ systems of the
placental microvillous and basal membranes may differ
functionally. Lysine uptake by system y+ in the basal
membrane is completely inhibited by neutral amino
acids, including leucine, in the presence of sodium,
whereas lysine uptake in the microvillous membrane is
partially inhibited by neutral amino acids other than
leucine (41). Thus there may be different subtypes of
system y+ in the syncytiotrophoblast.
Given its higher
maximum velocity compared with other cationic amino
acid transporters, the y+ system has usually been assigned the function of the major cationic amino acid
transporter in the placenta.
Other cationic transporters are present in both syncytial membranes. A high-affinity low-capacity system has
been described in the basal membrane that has initially
been ascribed to system b”T+ (40). The boy+system was
originally characterized in mouse blastocytes, is distinguished by a higher lysine affinity and lower capacity
than system y+, and is inhibited by leucine in the
absence of sodium (40, 130). A second cationic system
has been described in microvillous membranes and
given the name y+L . This transporter was originally
described in human erythrocytes (27) and subsequently
by the same laboratory in the human placental microvillous membrane (34). Smith and co-workers (41) have
been able to discriminate two systems in the microvillous membrane kinetically and by inhibition of system
y+ with N-ethylmaleimide
(NEM). The high-affinity
system (which may be y+L) is temperature sensitive,
with activity observable only at temperatures > 25°C
(41). The system y+L as described by Deves et al. (27)
has a much higher affinity for lysine and neutral amino
INVITED
acids than system y+. In addition, the y+L system is
insensitive
to treatment
with NEM (26) and less sensitive to changes in membrane
potential
(34). These
investigators
(34) have hypothesized
that the difference
in sensitivity
to membrane potential may play an important role in transepithelial
flux of lysine (34).
The maternal-to-fetal
transfer of cationic amino acids
in the human placenta is a complex process involving
multiple transport
systems whose identities are still an
area of controversy.
Eleno et al. (34) report that a
broad-spectrum
transporter
accounts for > 90% of lysine uptake in the basal membrane. These authors also
question the interpretation
that the high-affinity
system
reported earlier in the basal membrane (40) represents
system boy+. However,
the evidence suggests that the
high-affinity
transporter
of the basal membrane differs
from the comparable
microvillous
system. The basal
transporter,
unlike the microvillous
transporter,
is measurable at room temperature
and inhibitable by leucine
in the absence of sodium (41). Thus the high-affinity
cationic transporter
described originally by Furesz et al.
(40) as boy+ does not appear to resemble y+L, although
y+L activity
in the basal membrane
has not been
excluded.
fMMIN0
ACIDS
Taurine is also concentrated
in the fetal blood and is
in the highest concentration
of any amino acid in
placental tissue (92). Placental transfer
has an obligatory role in supplying this important
amino acid to meet
fetal demands. Taurine is taken up at the microvillous
membrane by a P-amino acid-specific
transporter
(61,
73, 82). The transporter
is sodium and chloride dependent and is energized by gradients of these ions (82), and
uptake is selectively
impaired by cyclosporin
A in a
placental cell line (JAR) (97). Recently, the microvillous
taurine transporter
has been solubilized and reconstituted in proteoliposomes
in functional
form (95). This
same group has cloned the transporter
from a human
placental cDNA library and used in situ hybridization
to
localize the 6.9-kilobase
transcript
to human chromosome 3 p24-p26 (98). The mechanism of transfer across
the basal membrane
is unknown.
There is some evidence for channel-mediated
taurine efflux from villous
fragments
(111).
PEPTIDE
TRANSPORT
Placental peptide transporters
have received much
less attention
than amino acid transporters.
Plasma
contains large amounts of small peptides in the di- and
tripeptide size, and it is conceivable that the fetus may
derive a significant
amount of its necessary
supply of
amino acids in this form. Ganapathy
et al. (44) have
demonstrated
that placental
microvillous
membrane
vesicles transport
glcylsarcosine
intact into an osmotically sensitive space. Transport
was inhibited by other
peptides and not by amino acids (44). The transporter
is
sodium and proton independent. There are no reports of
peptide transport
in the basal membrane.
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TRANSPORT
REGULATION
Regulation of amino acid transepithelial
flux has great
potential importance to normal fetal growth and development, since the placental syncytiotrophoblast
fulfills an
obligatory role in providing amino acids to the developing fetus. Amino acid transport
regulation
occurs at
several levels, and clearly the transport
proteins in the
two membranes must work in concert to bring about the
concentration
of amino acids in the fetal blood. The
polarization
of the two membrane domains with respect
to transport
proteins most likely accounts for a share of
this coordination. For example, sodium-dependent
transporters
in the microvillous
membrane
can drive the
uptake of neutral amino acids against a concentration
gradient and thereby raise intracellular
concentrations
above that in fetal blood. Transport
to the fetal circulation could then occur “downhill”
by diffusion or exchange type mechanisms.
The actual situation
in the
placenta is more complicated,
since the sodium-dependent A system is found in both the microvillous
and
basal membranes
and system ASC is found only in the
basal membrane.
The sodium-independent
system 1 is
located in both membranes.
Another
weakness
of the
“pump/leak”
hypothesis
is that it does not allow for
fetal regulation
of transport.
Regulation
of transport
most likely involves the transport
systems of either or
both of the membrane domains.
Lack of understanding
of the underlying
mechanisms
responsible
for asymmetrical
flux has complicated the
investigation
of transport
regulation.
Hence less is
currently
known
about regulation
of transsyncytial
amino acid flux than about the presence and characteristics of individual transport
systems. Nevertheless,
several regulatory
factors have been identified and associated with
certain
transport
systems.
Substrate
availability
can influence system A transport
by a protein synthesis-dependent
mechanism
(115) and by
transinhibition
(120). Transinhibition
is apparently specific to system A and does not regulate system 1 or
system ASC activity (120). Transstimulation
of system 1
does occur (58, 120). System 1 in choriocarcinoma
cells
(JAR) is stimulated
by an acidic extracellular
pH (11)
and phorbol esters and calmodulin
antagonists
(96).
System 1 activity in JAR cells is modified by calmodulin
as is the pH sensitivity
(11). Leucine transport
in JAR
cells is also competitively
inhibited by thyroid hormone
3,5,3’-triiodothyronine
transport,
although these two
substrates
are apparently transported
by separate transport systems
(94). The taurine
transport
system is
under the regulation of protein kinase C (12, 72) and is
inhibited by calcium (73).
Recent evidence indicates
hormonal
regulation
of
placental amino acid transport.
Cultured
trophoblast
uptake of cx-aminoisobutyrate
(AIB) is stimulated
by
insulin, insulin-like
growth factor I (IGF-I), and epiderma1 growth factor (EGF) (9,60). Bloxam et al. (9) used a
two-sided
culture technique so that the apical and basal
surfaces were reportedly
treated independently.
These
authors did not provide evidence as to the quality of
their preparation
in terms of maintaining
two separate
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INVITED
aqueous phases, and therefore
it is difficult to judge
whether
the hormonal
treatments
were limited to a
specific membrane as stated. Nevertheless,
they report
that EGF or IGF-I on the apical side reduced the rate of
unidirectional
(apical-to-basal)
transfer
and increased
cellular retention. EGF on the basal side increased AIB
flux in both directions,
whereas
IGF-I induced an increased flux into the basal media (9). AIB transport
is
potentially
upregulated
by IGF-II, dibutyryl
adenosine
3’,5’-cyclic monophosphate,
and phorbol esters, and the
insulin and IGF-I effects can be abolished by ethanol
exposure (P. Karl, personal communication).
The sodium-independent
systems and system ASC do not seem
to be regulated by any of the above hormonal factors.
Much work needs to be done to understand
the regulation of placental amino acid transport
at the cellular
level and its role in transsyncytial
flux.
MOLECULAR
BIOLOGY
TRANSPORTERS
OF AMINO
ACID
Recently, a number of amino acid transporters have
been cloned in various mammalian tissues. With the
exception of the taurine transporter (98), none have
been cloned directly from placenta, although many are
transporters that are found in the placenta. Two laboratories have independently cloned an ASC-like transporter based on its belonging to the same gene family as
the previously cloned glutamate transporter (4, 109).
Transporter message was found in placenta by both
laboratories but at much lower levels than in brain,
skeletal muscle, and pancreas. The identity of this
transporter was investigated by expression in HeLa cells
or expression in Xenopus oocytes. Both laboratories
observed uptake of neutral amino acids, which was
insensitive to the A system substrate MeAIB. However,
more extensive characterization in HeLa cells revealed
certain discrepancies between the cloned activity and
known ASC system characteristics (i.e., a lack of mutual
inhibition between serine and cysteine and a lack of
transstimulation) (109). As more transport proteins are
cloned, a major focus will be the characterization
of
these proteins with respect to function and comparison
with the classically defined transport systems in the
placenta and other organs.
System A is located in both the microvillous and basal
membranes of the syncytiotrophoblast. A putative clone
of system A (subsequently, doubts have been raised as to
whether this is system A) from a kidney cell line has
been reported (68). The authors concluded that this
transporter that they designated SAATl is closely related to the sodium-glucose transporter (SGLUT- 1).
As mentioned previously, the sodium-dependent glutamate transporter has been cloned. Two separate transporters have been cloned from the brain (93, 123) and a
third from rabbit small intestine (59). In collaboration
with Danbolt’s laboratory, we have obtained preliminary evidence that antibodies to at least two of these
transporters (the brain type) react with microvillous
and basal membrane vesicles from human placenta
(A. J. Moe, N. C. Danbolt, M. Landt, J. Storm-Mathisen,
and C. H. Smith, unpublished observations). We have
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also established the presence of message for a transporter similar to the human brain (110) and rat brain
transporter (123) by reverse transcriptase-polymerase
chain reaction from trophoblast total RNA. The presence of a GLT-l-like
transporter was established by
screening a placental cDNA library with probes to the
rat brain transporter (N. C. Danbolt, unpublished observations).
Several cationic amino acid transporters have recently
been cloned from the tissues of various species (3, 22,
65). These transporters apparently represent subtypes
of system y+, but functional differences between the
subtypes have yet to be reported. As already described in
this review, our laboratory has reported on a functional
difference between system y+ in the microvillous and
basal membranes of the syncytiotrophoblast.
Perhaps
these differences between membranes may correlate
with the molecular biology of cationic amino acid transporters. The p-amino acid transporter for taurine uptake has been cloned from rat brain (1 IS), thyroid cells
(57), and a kidney cell line (127).
One of the difficulties that has been encountered with
attempts to clone mammalian amino acid transporters
has been the isolation of transporter activators rather
than the transporters themselves (129). Many of the
transporters of interest are endogenous to the expression systems (the most common beingxenopus oocytes),
and it is difficult to rule out stimulation of an endogenous transporter in these experiments (129). It is clear
from the current state of knowledge that many potential
variants exist for the classical systems. The rapid pace of
cloning and increased knowledge about the structure of
these transport proteins is resulting in a major rethinking about the number and categorization of transport
systems. Through cloning it may finally be possible to
study the functional characteristics and regulation of
individual amino acid transport proteins rather than a
collection of different transporters.
INTRAUTERINE
GROWTH
ACID TRANSPORT
RETARDATION
AND AMINO
Impairment of fetal/neonatal growth and development results in intrauterine
growth retardation or
small-for-gestational age (SGA) babies (55). This condition can result from multiple causes, including many
that are related to placental function (56, 90). Placental
amino acid transport is of potential importance, since
the developing fetus is totally dependent on the supply
of amino acids from maternal blood for protein synthesis for accumulation of fetal mass. The umbilical venoarterial concentration differences of most essential amino
acids is lower for intrauterine growth-retarded fetuses
(15). The cord blood of growth-retarded neonates and
fetuses also shows an increase in the glycine-to-valine
ratio (15, 78). Recently, it has been reported that system
A transport is defective in microvillous membrane
vesicles isolated from syncytiotrophoblasts of placentas
of SGA babies (80). This is the first report linking a
defect in a specific transport system of the syncytiotrophoblast to intrauterine growth retardation (80). It has
now been established that factors that disrupt or inhibit
INVITED
the supply of amino acids across the placenta can lead to
SGA babies (28).
Several conditions, including the ingestion of drugs by
the pregnant woman, can cause decreased amino acid
transport
by the placenta and thus a decreased supply of
amino acids to the fetus. Maternal ethanol consumption
by pregnant rats caused a 30-40% reduction in valine
uptake by placental villous fragments
(47) and reductions of a similar magnitude when measured in vivo in
the rat (91). Chronic
ethanol consumption
30 days
before and throughout
gestation reduced placental valine uptake by 44% (47). Both short- and long-term
ethanol exposure
decreased fetal valine uptake
(91).
Other studies have found an effect of ethanol administration on AIB uptake in rodent placenta (39, 76). Amino
acid transport
in the human and nonhuman
primate
placentas has proven to be relatively resistant
to acute
ethanol treatment
(36, 38, 39, 103). Chronic alcohol
administration
has depressed uptake of some amino
acids in nonhuman primates (37).
Another
factor that depresses fetal growth
and is
often found in conjunction
with ethanol is tobacco
smoke (38). Use of ethanol concurrent
with tobacco
smoking may be more harmful
then either substance
used independently
(38). Babies born to mothers who
smoke are on average 200 g lighter than those born to
nonsmokers
(46, 90, 132). One possible cause of this
lower birth weight is depressed amino acid transport
across the placenta (101). Smoking increases nicotine
concentrations
in placenta, amniotic
fluid, and fetal
serum above concentrations
found in maternal serum
(79). Uptake of amino acids by isolated human placental
villi is reduced by nicotine and several components
of
tobacco smoke (5, 99, 101, 102). The mechanism
of
nicotineor ethanol-induced
depression
of placental
amino acid uptake
is still unknown.
Studies using
membrane vesicles have failed to show a direct effect of
nicotine (104) or ethanol (103) on amino acid transporters.
Another
major problem today is the ingestion
of
cocaine by pregnant women. Cocaine use can lead to
numerous
neonatal complications,
with intrauterine
growth retardation
among them (112). Cocaine is known
to inhibit placental uptake of amino acids (5) and binds
to a high-affinity
binding protein in human placenta (2,
30). Recent studies using membrane vesicles suggest a
potential direct effect of cocaine on amino acid transport
(30). The exact mechanism of this inhibition is unknown
but seems to be limited to sodium-dependent
transport
systems (30).
Maternal diabetes and preeclampsia are complications
of pregnancy that can also lead to alterations
in placental amino acid transport
and serious perinatal implications. A large or macrosomic
fetus can’ result from
excessive intrauterine
growth, which is a frequent outcome of diabetes mellitus-complicated
pregnancies
(7).
Sodium-dependent
MeAIB uptake was reduced 49% in
microvillous
membrane vesicles isolated from placentas
of diabetic pregnancies
resulting
in macrosomic
babies
(74). As with an early report from another laboratory
(28), these authors failed to demonstrate
an effect of
Cl327
REVIEW
diabetic pregnancy without
macrosomia
on MeAIB uptake (74). Intuitively,
a lower system A transport
activity associated with macrosomia
seems surprising
but is
consistent
with results from the rat where streptozototin-induced
diabetes resulted
in lower fetal plasma
amino acid concentrations
(1).
ASYMMETRICAL
FLUX
OF AMINO
ACIDS
The cellular basis for the asymmetrical maternal-tofetal transfer of amino acids across the placenta has yet
to be established. Tran .sepithelial amino acid flux across
the intestine has been explained by ion-driven concentrative uptake at the microvillous membrane and diffusion
across the basolateral membrane (122). The same pump/
leak mechanism has been used to explain asymmetrical
flux across the syncytiotrophoblast (116). As mentioned
previously, a weakness of this theory is that it does not
allow for fetal regulation of placental transport or the
presence of sodium-dependent transporters in the basal
membrane. In addition, there are alternative explanations for the possible role of the relatively high syncytial
amino acid concentrations (92) in cellular functions
other than transsyncytial flux. The nonprotein amino
acid taurine is the amino acid fou nd in the highest
concentration (10.4 mM) in the placenta (92). In many
cell types, taurine plays a role in maintaining cell
volume (52), and Shennan et al. (111) have provided
eviden .ce that taurine may Play an osmoregulatory role
in the placenta. The amino acid next highest in concentration in the placenta is glutamate (92), which we know
is oxidized by the placenta (13,86) and is not transferred
to the fetus. A recent report has also implicated amino
acid-induced hyperosmolarity in activation of maternalto-fetal Cl- transfer (8).
Maternal-to-fetal
transfer of amino acids requires
movement across two membrane barriers. As discussed
in this review, these membranes contain specific transport proteins for the translocation of all important
amino acids and some small peptides. Because of this,
the transport paradigm has always included the specific
transport systems, even though it has not been possible
to explain how the operation of these systems results in
maternal-to-fetal transfer.
Some of the key aspects of maternal-to-fetal transfer
that have received less attention and perhaps play
significant roles in this process include intrasyncytial
factors such as amino acid metabolism and membrane
efflux pathways. Placental amino acid metabolism has
received little attention (especially in humans), yet is
known to impact placental handling of amino acids (13,
14, 85, 86). Differential efflux across the microvillous
and basal membranes may play an important role in
maternal-to-fetal transfer (32). Uptake from the blood
across each membrane could be roughly equivalent,
provided that efflux across th .e basal membrane was
more rapid relative to the microvillous membrane.
Differential rates of efflux for the two membranes could
very well be determined by the distribution of specific
transporters in the two membranes. With our present
ability to differentiate between transporter subtypes
and to identify and characterize specific transport pro-
Cl328
INVITED
REVIEW
teins, we hopefully
will be able to understand
the
mechanisms
responsible for asymmetric
amino acid flux
across the placental syncytiotrophoblast.
A great deal is yet to be learned about placental amino
acid transporter
structure,
function, and regulation as
well as the interrelationship
of drugs and other substances.
The application
of molecular
biology techniques, along with the development
of improved
cell
culture models, suggests the possibility
of major advances in this important
area of investigation.
I thank
the investigators
Thanks
to Dr. Carl H. Smith
manuscript.
This review
was supported
and Human
Development
Grant
Address for reprint
requests:
Medicine,
One Children’s
Place,
who
for
shared
reading
their
and
unpublished
commenting
16.
17.
18.
19.
work.
on the
20.
by National
Institute
HD-27258.
A. J’. Moe, Washington
St. Louis, MO 63110.
of Child
Univ.
Health
School
of
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