Reproduction in Eutherian Mammals

Colin A Finn, University of Liverpool, Liverpool, UK

The eutherians make up the vast majority of extant mammals. They are all viviparous but vary in details of their ovarian cycle, behaviour, placentation and length of gestation.

Introductory article
Article Contents
. Sexual Differentiation . Hormones of Reproduction . Gametogenesis . The Ovarian Cycle . Fertilization . Pregnancy . Parturition . Lactation and Maternal Care

Sexual Differentiation
All mammals breed sexually and all are unisexual. Sex is determined at fertilization by the sex chromosomes. Unlike the autosomal chromosomes the pair of sex chromosomes are not matched. They are unequal in size; the Y being short, while the X is similar in size to the other chromosomes. The nucleated somatic cells of normal females have a pair of X-chromosomes while those of males have an X and Y. Nuclei containing pairs of chromosomes are diploid. Reproduction takes place through the germ cells, which are set aside from the somatic cells very early in the development of the embryo. Germ cells are the only cells in the body that can undergo meiosis, a form of cell division in which the pairs of chromosomes are separated, with each daughter cell receiving one of each pair of chromosomes, giving a haploid cell. Germ cells can rst be seen in the yolk sac, although they are actually set aside earlier in the embryonic disc. As development of the embryo proceeds and organs are formed, the germ cells migrate by amoebic movement through the gut wall to an area of mesoderm called the genital ridge (the indierent gonad). Here, if a Ychromosome is present, the indierent gonad develops into a testis. In the absence of a Y-chromosome an ovary develops. The genital tract is formed from the mesonephric (Wolan) duct in the male and the paramesonephric (Mu llerian) duct in the female. The Wolan duct forms the epididymis and vas deferens while the Mu llerian duct forms the oviduct, uterus, cervix and part of the vagina. Which duct system develops and which degenerates depends on whether a testis is present. The secretions of the testis, testosterone and Mu llerian inhibitory hormone, cause the Wolan system to develop and the Mu llerian to degenerate. The absence of these secretions leads to the development of the female reproductive tract (the default condition is the female). The male sex hormone is also responsible for sex changes in the nervous system, which occur soon after birth (e.g. behaviour).

. Ageing and Reproduction

Hormones of Reproduction
The main hormones involved in reproduction are polypeptides, steroids and prostaglandins. The hypothalamus synthesizes short-chain polypeptides, which either pass down the nerve bres to the posterior pituitary for storage and secretion (oxytocin) or pass down the hypothalamic pituitary portal bloodstream to the pituitary (gonadotrophin-stimulating hormone). Here they stimulate secretion of protein hormones. In both sexes the main protein hormones are luteinizing hormone (LH), follicle-stimulating hormone (FSH) and prolactin. These have a major control over the ovary and the testis, which in response, secrete steroid hormones, testosterone in the male, oestradiol and progesterone in the female. The ovary and testis also secrete polypeptides, such as inhibin and activin. The uterus secretes prostaglandins.

Gametogenesis
The germ cells, having colonized the indierent gonad, are transformed into gametes ready for fertilization. Morphologically, the gametes of the two sexes are very dierent. Female gametes, ova, are large round cells with a large central nucleus, whilst male gametes, sperm, take on a tadpole-like structure. The sperm of each species has a characteristically shaped head, largely made up of the condensed DNA of the nucleus. Passing from the head is a middle piece (containing mitochondria) and a tail (for movement). The process of forming the gametes (gametogenesis) is also dierent between the two sexes. All germ cells initially undergo mitosis but then, at some stage, undergo meiosis and become haploid. The formation of male gametes, spermatogenesis, is a continuous process from puberty to death. Within the testis the somatic cells form continuous tubules lined by specialized cells called Sertoli cells. Situated inside these seminiferous tubules and between the Sertoli cells, the germ cells (spermatogonia) multiply by
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a set number of mitotic divisions (specic for a species) and then undergo two meiotic divisions to give four round spermatocytes. Some of the spermatogonia do not progress to meiosis but return to mitosis, thus ensuring a continuous supply of gametes. After the two meiotic divisions the spermatocytes are transformed into spermatozoa (a process known as spermiogenesis). Some way up from the basement membrane the Sertoli cells are joined together by specialized junctions. This divides the space between them into a basal compartment and an adluminal compartment. The spermatogonia, whilst undergoing mitosis, are contained within the basal compartment but move up into the adluminal compartment with the start of meiosis. The junction between the Sertoli cells restricts the transfer of substances into the adluminal compartment. This barrier, together with the secretions of the Sertoli cells, creates the specialized uid in which spermiogenesis can take place. Fully formed spermatozoa pass out of the seminiferous tubules into the epididymis and then to the vas deferens. During passage down the vas deferens, secretions are added from the seminal vesicles and prostate (the seminal plasma). Situated between the seminiferous tubules are the interstitial cells of Leydig which secrete male sex hormone, testosterone. This has an important role in the control of spermatogenesis, in the growth and development of the male reproductive organs and in sexual behaviour. In the ovary the germ cells, oogonia, undergo numerous mitotic divisions during the prenatal period but before the animal is born all the oogonia enter the rst stage of meiosis, so that at birth they are all oocytes and no further mitosis takes place. A single layer of somatic cells surrounds each oocyte to form a primary follicle. Each oocyte is capable of undergoing two maturation divisions to become a haploid ovum. Most, however, do not do so but are removed by atresia (programmed cell death). Very few actually become ova. After puberty a few of the oocytes will, at regular intervals, progress to become ova. The primary follicle enlarges by multiplication of the somatic cells, which form into two layers, the granulosa and thecal layers, with a membrane between them. The oocyte grows rapidly in size and becomes encased in a protein covering, the zona pellucida. The cells in the thecal layer, in response to LH, synthesize androgens, which, in response to FSH, are aromatized to oestradiol in the granulosa cells. Levels of oestradiol gradually increase, and by a feedback cascade reaction cause the follicle to grow rapidly. The number of granulosa cells increases and a uid-lled cavity forms between the cells, to become the graaan follicle. When oestradiol levels reach their peak, a surge of LH is released from the anterior pituitary which causes the follicle to burst, releasing the ovum into the oviduct (ovulation). After ovulation, the granulosa and theca cells, in response to the high levels of LH and sometimes prolactin, enlarge to form the corpus luteum, which secretes progesterone.
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The rst meiotic division of the oocyte occurs at ovulation and the second at fertilization. Both of the meiotic divisions are unequal, with most of the cytoplasm going into one of the cells, the other half of the cell forming a small polar body, which is discarded. Each oocyte, therefore, gives rise to only one ovum.

The Ovarian Cycle

The ovary regularly undergoes cycles of follicle development, ovulation and corpus luteum formation, which results in the sequential secretion of oestradiol and progesterone. At the height of oestradiol secretion, mammals (except women) show sexual behaviour (oestrus or heat); this accompanies ovulation, and occurs at intervals specic for each species. Following ovulation the female is prepared for pregnancy by progesterone. Thus the cyclic changes in the ovary are followed by a cycle of changes throughout the body, brought about by the changing levels of ovarian hormones. The stages of the oestrous cycle are prooestrus, when the follicle is developing, oestrus, near the time of ovulation, and dioestrous, when the corpus luteum is formed. Similar changes occur in the ovary of women, but as they do not show oestrous behaviour but menstruate, it is called the menstrual cycle. The aim of the oestrous cycle is to encourage pregnancy, but if mating does not take place the corpus luteum is removed (luteolysis) and another ovulation takes place. Luteolysis is controlled in some species by the secretion of prostaglandin F2a by the uterus. Other species have evolved dierent mechanisms to control the switch between return to oestrus and maintenance of the corpus luteum for pregnancy. In women it is the secretion by the embryo of chorionic gonadotrophin, a hormone with LHlike activity, which controls the switch (the presence of this hormone in the urine is used as an indication of pregnancy). A few animals, such as rodents, do not form a functional corpus luteum unless copulation has occurred, or some other mechanism has been used to stimulate the nerve endings in the vagina and cervix. There are also a few animals (e.g. cats, rabbits and camels) that do not ovulate unless copulation takes place. They are known as induced ovulators to distinguish them from the majority of species, which are spontaneous ovulators. In a few species (dogs) the corpus luteum of the oestrous cycle lasts as long as pregnancy, so no mechanism is necessary for its maintenance.

Fertilization
Fertilization takes place in the upper part of the oviduct. Sperm are deposited by the male, during coitus, into the

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Reproduction in Eutherian Mammals

vagina or sometimes directly into the uterus. They pass along the uterus, mostly by the churning action of the uterine muscles on the uterine uid, to reach the lower part of the oviduct. The ejaculate contains millions of sperm, but only hundreds reach the oviduct. During their passage through the uterus and oviduct, capacitation, which removes a layer (partly cholesterol) from the surface of the sperm, takes place. If an ovum is in the oviduct a few sperm pass towards it by the whipping action of their tails and undergo the acrosome reaction. This involves changes in the sperm head so that the sperm can pass through the zona pellucida of the ovum and fuse with it. After fusion of the sperm and ovum, a block to polyspermy is initiated. This involves electrical changes in the vitelline membrane around the ovum and changes to the zona pellucida, which together prevent the attachment of another sperm to the ovum. Following fertilization the nucleus of the sperm decondenses and joins up with the nucleus of the ovum to form the diploid zygote. The zygote divides many times as it passes along the oviduct. When about 32 to 63 cells have been formed, the zygote is a bunch of cells that looks like a raspberry (morula). The outer cells join together with tight junctions to form a seal. A uid-lled cavity then forms in the morula to give the blastocyst. The cells destined to be the embryo are separated o, as the embryonic disc, and the outer layer becomes the trophoblast, by which the blastocyst attaches to the wall of the uterus. As the embryonic disc grows and dierentiates into the embryo, a membranous sac forms around it (the amnion), and two sacs grow out from the embryo into the blastocyst cavity. These are the allantois and the yolk sac. They are important in the formation of the placenta.

Pregnancy
The passage of the zygote along the oviduct to the uterus takes a characteristic time for each species. During this time the uterus is transformed from the open, uid-lled vessel necessary for sperm transport and capacitation, to the more cellular structure with a closed lumen in preparation for pregnancy. This transformation involves division and dierentiation of the cells of the endometrium (the inner lining of the uterus). The three layers of the endometrium are luminal epithelium, glandular epithelium and connective tissue stroma. Cells divide and dierentiate in sequence, in response to the ovarian hormones, to prepare the surface of the luminal epithelium for attachment of the blastocyst, the glands for the secretion of histotroph (taken up as nutrient by the early embryo), and the stroma for the invasion of the embryo into the wall of the uterus. Invasion does not happen in all species but is most pronounced in women. In anticipation of this, the stromal broblasts enlarge, become polyploid and bi-

nuclear, and due to the presence of tight junctions form a continuous sheet of tissue. This is the decidual cell reaction and although decidual cells are formed in other mammals (rodents for example), they only dierentiate if the animal has been mated and a blastocyst is present. It is this very advanced preparation of the endometrium in anticipation of a blastocyst in women that accounts for the loss of tissue and blood during menstruation. After arrival in the uterus the blastocyst attaches to the wall by apposition of the surface of the trophoblast to the luminal surface of the uterine epithelium. This is the rst stage in the formation of the placenta. What follows depends on the species. In many species the trophoblast increases greatly in size so that a very large area of contact is attained. In the embryo, blood vessels grow out into the allantois to vascularize the trophoblast. This allows the passage of nutrients, gases and waste products between the mother and the embryo. In the simplest form of placenta six layers of cells separate the bloodstreams of the mother and embryo and this is as far as placentation goes in some species, e.g. pig and horse. In many species, however, the two bloodstreams have been brought closer together by removal of some of the maternal layers (rodents) or invasion of the blastocyst into the wall of the uterus (human). Where the bloodstream of the embryo has become closer to that of the mother the expansion of the trophoblast is not so extensive and the placenta does not occupy all the surface of the endometrium. This gives each species a characteristically shaped placenta. In ruminants, special areas of the endometrium, caruncles, are dierentiated for the attachment of the trophoblast. After attachment they are called cotyledons. During pregnancy the cells of the embryonic disc proliferate and dierentiate into all the organ systems of the body. When they are fully formed the embryo is called a fetus. In response to the growth of the embryo the placenta and uterus increase in size and become extremely vascular. To allow for this growth and to ensure that the muscles (myometrium) do not contract, the uterus must be under the inuence of progesterone. This is provided in most species by the corpus luteum, which is maintained for the duration of pregnancy. In a few species, including the human, the placenta at a certain stage of development synthesizes progesterone, making the animal independent of the corpus luteum.

Parturition
Gestation lasts for a set length of time depending on the species. The time of birth is determined by the maturation of the fetus, in particular of the fetal hypothalamic pituitaryadrenal system. At a certain stage of develop3

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Reproduction in Eutherian Mammals

ment the synthesis of glucocorticoids from the fetal adrenal gland increases. This has several important eects on both the fetus and the mother, all connected with birth. It stimulates secretion of surfactant, important for expansion of the fetal lungs after birth, increases the production of brown fat, important for the maintenance of body temperature and sets o a chain of reactions that bring about parturition. For parturition to take place it is necessary for the cervix to soften and for the myometrium to contract. These changes are brought about by changes in the hormone levels in the maternal bloodstream, most importantly a shift in the balance of progesterone and oestradiol. During pregnancy the level of progesterone is very high compared to that of oestradiol. Under these conditions the cervix consists of a hard collagen matrix which together with thick mucus provides a seal to the entrance to the uterus. When the level of oestradiol exceeds that of progesterone, the cervix softens due to a change in the collagen matrix. High levels of progesterone also aect the cells of the myometrium by reducing their inclination to form junctions between neighbouring cells. When the level of progesterone falls, gap junctions are formed between myometrial cells facilitating a coordinated contraction. For a full contraction other hormones (prostaglandin F2a and oxytocin) act on the oestrogen-dominated uterus. The changes in hormone balance in the mother are brought about by the action of fetal glucocorticoids on the placenta. Depending on the species, this involves either switching the synthesis of progesterone towards the synthesis of oestradiol, or initiating the synthesis of prostaglandin F2a, which then passes to the maternal ovary causing breakdown of the corpus luteum and reduction in progesterone. Once the change in steroid hormones has been eected, the prostaglandin can stimulate the myometrium to contract. This causes the fetus to be forced against the cervix, stimulating nerve endings and causing a nervous message to be transmitted to the hypothalamus. In response, oxytocin is released from the posterior pituitary and passes to the uterus, where it causes further contraction of the myometrium. This is an example of a neurohumoral reex and is responsible for a positive feedback eect by which parturition is brought to a rapid conclusion, followed by ejection of the placenta.

alveoli (the sac-like terminations to the ducts that synthesize and secrete milk) progesterone is also required. During gestation there is massive growth of the glands due largely to the high levels of progesterone, assisted by lower but still signicant levels of oestrogen. The hormones of the anterior pituitary gland control the secretion of milk (lactogenesis). Prolactin is important in most species, although growth hormone and other metabolic hormones are very important in some (especially ruminants). Whilst the animal is pregnant the high levels of progesterone inhibit the enzymes involved in lactogenesis so that synthesis and secretion of milk is inhibited. At parturition, when levels of progesterone fall, milk synthesis proceeds rapidly, assisted by the greatly increased secretion of prolactin that occurs when the ospring starts to suckle. During suckling stimulation of nerve endings in the teat leads to two neurohumoral reexes, one resulting in the release of prolactin from the anterior pituitary, the other causing the release of oxytocin from the posterior pituitary. The latter is involved in the removal of milk. Having been synthesized in the alveolar cells, the milk passes into the alveoli and from there down the ducts to the teat. To facilitate removal of milk, special contractile cells, called myoepithelial cells, surround the alveoli and small ducts. These contract in response to oxytocin and are important in the ejection of milk. Apart from being provided with nutrients the young must be protected and cared for. Mammals vary considerably in the mechanisms that have been evolved. Some young are born in a very dependent state and require complete protection. These altricial species (rodents and dogs for example) usually build nests in which the young are cared for. Young of other mammals are born needing only a small amount of protection (precocious) and can take their place among the other members of the herd. However, it is still essential that the mother protects and feeds them. A bond has to be established between mother and ospring. This is particularly important in animals like sheep in which it has been shown that the bond is dependent on the release of oxytocin, in response to the passage of the young through the birth canal during parturition. Oxytocin has thus evolved as a hormone of the three main facets of birth, parturition, milk provision and maternal care. Other hormones that have been shown to be involved in maternal care are oestradiol and prolactin.

Lactation and Maternal Care

Before puberty mammogenesis (the growth of the mammary glands) proceeds at the same rate as the rest of the body (isometric growth). After puberty, following the secretion of oestrogens and progesterone, there is increased growth of the mammary glands (allometric growth). In most species oestrogens are responsible for growth of the mammary ducts, while for the development of the
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Ageing and Reproduction

From what has been discussed it is clear that in mammals the germ cells are completely dependent on the somatic organs for their development and successful union with gametes of the opposite sex. As the somatic organs concerned in reproduction get older they will be less ecient in their role in reproduction. There is, however, a

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big dierence between the sexes in the role played by the somatic organs in reproduction. While both sexes rely on the somatic cells of the gonads and the endocrine organs for gametogenesis, their dependence after release of the gametes from the gonads is very dierent. In males, the spermatozoa, after addition of secretions from the accessory glands (prostate, seminal vesicles), are passed to the exterior through ducts, a fairly short and simple process. Females have the more onerous task of carrying the ospring and providing it with nutrients while it is growing and developing. This dierence in burden between the sexes is reected in the eect of ageing. Males continue producing sperm and reproducing until they reach a very advanced age and stop only when the mental and physical eort available is no longer compatible with that required for successful mating. Females, on the other hand, stop breeding well before the end of their life. In the case of rodents, where most investigations have been carried out, the size of litters produced gets smaller as the mother gets older and breeding ceases about half to two-thirds of the way through their expected lifespan. The cause of the declining litter size and premature cessation of breeding is failure of the uterus to carry all or any of the zygotes to term. Mating and fertilization take place but changes in the uterus necessary for pregnancy are impaired. Thus the uterus appears to be the crucial organ in determining reproductive ageing in female mammals. There is, however, one exception to this women. In women it is not the uterus which fails but the ovary. All female mammals are born with a nite store of oocytes, which are gradually lost from the time of birth, or even before. In most mammals this has no eect on reproductive output because somatic ageing, especially of the uterus, causes the animal to stop reproducing and die well before the stock of oocytes is exhausted. Rodents, for example, continue having oestrous cycles after they have stopped having litters and oocytes can be found in the ovary at autopsy. Cycles do, however, become very irregular, indicating that somatic ageing is taking place in the organs concerned in regulating the cycle, although as the eects follow the ageing of the uterus they are not critical in inuencing reproductive output. In women, however, this is not the case. Women run out of oocytes when only about halfway through their expected lifespan. Thus women have a menopause, in which not only do they not breed but also they do not have ovarian cycles. Cycles depend on the presence in the ovary of follicles on which the pituitary hormones can act.

The crucial question is why do women run out of oocytes while other mammals do not? The answer is almost certainly related to the very extended lifespan of humans. It is well known that there is, in general, a relationship between body size and longevity in mammals. Very small rodents live only a matter of months whereas elephants and other large mammals live for many years. A graph of longevity against bodyweight is approximately a straight line, with the notable exception of humans, who live considerably longer than any mammal of equivalent size. The reason for the relationship is thought to be the geometrical relationship between mass and surface area. The smaller the body the greater the surface area. Warmblooded animals have to expend energy keeping warm and as most heat is lost through the body surface, smaller animals will have to use up more energy keeping warm. During the metabolic processes associated with this they will produce free radical oxygen. A major theory of ageing suggests that the accumulated damage caused by free radical oxygen is responsible for ageing of somatic organs. Why humans should be immune from this relationship is unknown. It may have something to do with the fact that humans rely to a far less extent than other mammals on metabolic energy to keep warm, although it is unlikely that this is the whole story. The only other mammals with an unexpectedly long lifespan are bats. Again the reason is unknown. Whatever the reason, it appears that women have a menopause due to two evolutionary developments. One, common to all mammals, the entry of germ cells into meiosis before birth thus limiting the supply of oocytes, and the other, common to all humans, male and female, extended longevity. In view of the dual origin of the menopause, neither specic to the human female, it is dicult to see how it can be an adapted response to a human need such as grandmothering, as suggested by some anthropologists.

Further Reading
Austin CR and Short RV (1980) Reproduction in Mammals. Cambridge: Cambridge University Press. Hunter RHF (1982) Reproduction of Farm Animals. London: Longmans. Johnson MH and Everitt BJ (1995) Essential Reproduction, 4th edn. Oxford: Blackwell Scientic. McDonald LE and Pineda MH (eds) (1989) Veterinary Endocrinology and Reproduction, 4th edn. Philadelphia: Lea and Febiger.

ENCYCLOPEDIA OF LIFE SCIENCES / & 2001 Nature Publishing Group / www.els.net