David L. Olive
Steven F. Palter
• The female reproductive process involves the central nervous system (primarily hypothalamus), the pituitary gland, the ovary, and the uterus (endometrium). All must function appropriately for normal reproduction to occur.
• Hypothalamic gonadotropin-releasing hormone (GnRH) simultaneously regulates both luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in the pituitary by being secreted in a pulsatile manner. The pulse frequency determines the relative amounts of LH and FSH secretion.
• The ovary responds to FSH and LH in a defined, sequential manner to produce follicular growth, ovulation, and corpus luteum formation. The cycle is designed to produce an optimal environment for pregnancy; if pregnancy does not occur, the cycle begins again.
• In the early menstrual cycle the ovary produces estrogen, which is responsible for endometrial growth. Following ovulation, progesterone is also produced in significant quantities, which transforms the endometrium into a form ideal for implantation of the embryo. If no pregnancy occurs, the ovary ceases to produce estrogen and progesterone, the endometrium is sloughed, and the cycle begins again.
The reproductive process in women is a complex and highly evolved interaction of many components. The carefully orchestrated series of events that contributes to a normal ovulatory menstrual cycle requires precise timing and regulation of hormonal input from the central nervous system, the pituitary gland, and the ovary. This delicately balanced process can be disrupted easily and result in reproductive failure, which is a major clinical issue confronting gynecologists. To manage effectively such conditions, it is critical that gynecologists understand the normal physiology of the menstrual cycle. The anatomic structures, hormonal components, and interactions between the two play a vital role in the function of the reproductive system. Fitting together the various pieces of this intricate puzzle will provide “the big picture”: an overview of how the reproductive system of women is designed to function.
Neuroendocrinology
Neuroendocrinology represents facets of two traditional fields of medicine: endocrinology, which is the study of hormones (i.e., substances secreted into the bloodstream that have diverse actions at sites remote from the point of secretion), and neuroscience, which is the study of the action of neurons. The discovery of neurons that transmit impulses and secrete their products into the vascular system to function as hormones, a process known as neurosecretion, demonstrates that the two systems are intimately linked. For instance, the menstrual cycle is regulated through the feedback of hormones on the neural tissue of the central nervous system (CNS).
Anatomy
Hypothalamus
The hypothalamus is a small neural structure situated at the base of the brain above the optic chiasm and below the third ventricle (Fig. 7.1). It is connected directly to the pituitary gland and is the part of the brain that is the source of many pituitary secretions. Anatomically, the hypothalamus is divided into three zones: periventricular (adjacent to the third ventricle), medial (primarily cell bodies), and lateral (primarily axonal). Each zone is further subdivided into structures known as nuclei, which represent locations of concentrations of similar types of neuronal cell bodies (Fig. 7.2).
Figure 7.1 The hypothalamus and its neurologic connections to the pituitary.
Figure 7.2 The neuronal cell bodies of the hypothalamus.
The hypothalamus is not an isolated structure within the CNS; instead, it has multiple interconnections with other regions in the brain. In addition to the well-known pathways of hypothalamic output to the pituitary, there are numerous less well-characterized pathways of output to diverse regions of the brain, including the limbic system (amygdala and hippocampus), the thalamus, and the pons (1). Many of these pathways form feedback loops to areas supplying neural input to the hypothalamus.
Several levels of feedback to the hypothalamus exist and are known as the long, short, and ultrashort feedback loops. The long feedback loop is composed of endocrine input from circulating hormones, just as feedback of androgens and estrogens onto steroid receptors is present in the hypothalamus (2,3). Similarly, pituitary hormones may feed back to the hypothalamus and serve important regulatory functions in short-loop feedback. Finally, hypothalamic secretions may directly feed back to the hypothalamus itself in an ultrashort feedback loop.
The major secretory products of the hypothalamus are the pituitary-releasing factors (Fig. 7.3):
Figure 7.3 The hypothalamic secretory products function as pituitary-releasing factors that control the endocrine function of the ovaries, the thyroid, and the adrenal glands.
1. Gonadotropin-releasing hormone (GnRH), which controls the secretion of luteinizing hormone (LH) and follicle-stimulating hormone (FSH)
2. Corticotropin-releasing hormone (CRH), which controls the release of adrenocorticotrophic hormone (ACTH)
3. Growth hormone–releasing hormone (GHRH), which regulates the release of growth hormone (GH)
4. Thyrotropin-releasing hormone (TRH), which regulates the secretion of thyroid-stimulating hormone (TSH)
The hypothalamus is the source of all neurohypophyseal hormone production. The neural posterior pituitary can be viewed as a direct extension of the hypothalamus connected by the fingerlike infundibular stalk. The capillaries in the median eminence differ from those in other regions of the brain. Unlike the usual tight junctions that exist between adjacent capillary endothelial lining cells, the capillaries in this region are fenestrated in the same manner as capillaries outside the CNS. As a result, there is no blood–brain barrier in the median eminence.
Pituitary
The pituitary is divided into three regions or lobes: anterior, intermediate, and posterior. The anterior pituitary (adenohypophysis) is quite different structurally from the posterior neural pituitary(neurohypophysis), which is a direct physical extension of the hypothalamus. The adenohypophysis is derived embryologically from epidermal ectoderm from an infolding of Rathke’s pouch. Therefore, it is not composed of neural tissue, as is the posterior pituitary, and does not have direct neural connections to the hypothalamus. Instead, a unique anatomic relationship exists that combines elements of neural production and endocrine secretion. The adenohypophysis itself has no direct arterial blood supply. Its major source of blood flow is also its source of hypothalamic input—the portal vessels. Blood flow in these portal vessels is primarily from the hypothalamus to the pituitary. Blood is supplied to the posterior pituitary via the superior, middle, and inferior hypophyseal arteries. In contrast, the anterior pituitary has no direct arterial blood supply. Instead, it receives blood via a rich capillary plexus of the portal vessels that originates in the median eminence of the hypothalamus and descends along the pituitary stalk. This pattern is not absolute, however, and retrograde blood flow has occurred (4). This blood flow, combined with the location of the median eminence outside the blood–brain barrier, permits bidirectional feedback control between the two structures.
The specific secretory cells of the anterior pituitary are classified based on their hematoxylin- and eosin-staining patterns. Acidophilic-staining cells primarily secrete GH and prolactin and, to a variable degree, ACTH(5). The gonadotropins are secreted by basophilic cells, and TSH is secreted by the neutral-staining chromophobes.
Reproductive Hormones
Hypothalamus
Gonadotropin-Releasing Hormone
GnRH (also called luteinizing hormone–releasing hormone, or LHRH) is the controlling factor for gonadotropin secretion (6). It is a decapeptide produced by neurons with cell bodies primarily in the arcuate nucleus of the hypothalamus (7–9) (Fig. 7.4). Embryologically, these neurons originate in the olfactory pit and then migrate to their adult locations (10). These GnRH-secreting neurons project axons that terminate on the portal vessels at the median eminence where GnRH is secreted for delivery to the anterior pituitary. Less clear in function are multiple other secondary projections of GnRH neurons to locations within the CNS.
Figure 7.4 Gonadotropin-releasing hormone is a decapeptide.
The gene that encodes GnRH produces a 92 amino acid precursor protein, which contains the GnRH decapeptide and a 56 amino acid peptide known as GnRH-associated peptide (GAP). The GAP is a potent inhibitor of prolactin secretion and a stimulator of gonadotropin release.
Pulsatile Secretion
GnRH is unique among releasing hormones in that it simultaneously regulates the secretion of two hormones—FSH and LH. It also is unique among the body’s hormones because it must be secreted in a pulsatile fashion to be effective, and the pulsatile release of GnRH influences the release of the two gonadotropins (11–13). Using animals that had undergone electrical destruction of the arcuate nucleus and had no detectable levels of gonadotropins, a series of experiments were performed with varying dosages and intervals of GnRH infusion (13,14). Continual infusions did not result in gonadotropin secretion, whereas a pulsatile pattern led to physiologic secretion patterns and follicular growth. Continual exposure of the pituitary gonadotroph to GnRH results in a phenomenon called down-regulation, through which the number of gonadotroph cell surface GnRH receptors is decreased (15). Similarly, intermittent exposure to GnRH will “up-regulate” or “autoprime” the gonadotroph to increase its number of GnRH receptors (16). This allows the cell to have a greater response to subsequent GnRH exposure. Similar to the intrinsic electrical pacemaker cells of the heart, this action most likely represents an intrinsic property of the GnRH-secreting neuron, although it is subject to modulation by various neuronal and hormonal inputs to the hypothalamus.
The continual pulsatile secretion of GnRH is necessary because GnRH has an extremely short half-life (only 2–4 minutes) as a result of rapid proteolytic cleavage. The pulsatile secretion of GnRH varies in both frequency and amplitude throughout the menstrual cycle and is tightly regulated (17,18) (Fig. 7.5). The follicular phase is characterized by frequent, small-amplitude pulses of GnRH secretion. In the late follicular phase, there is an increase in both frequency and amplitude of pulses. During the luteal phase, however, there is a progressive lengthening of the interval between pulses. The amplitude in the luteal phase is higher than that in the follicular phase, but it declines progressively over the 2 weeks. This variation in pulse frequency allows for variation in both LH and FSH throughout the menstrual cycle. For example, decreasing the pulse frequency of GnRH decreases LH secretion but increases FSH, an important aspect of enhancing FSH availability in the late luteal phase. The pulse frequency is not the sole determinant of pituitary response; additional hormonal influences, such as those exerted by ovarian peptides and sex steroids, can modulate the GnRH effect.
Figure 7.5 The pulsatile secretion of gonadotropin-releasing hormone in the follicular and luteal phases of the cycle.
Although GnRH is primarily involved in endocrine regulation of gonadotropin secretion from the pituitary, it is apparent that this molecule has autocrine and paracrine functions throughout the body. The decapeptide is found in both neural and nonneural tissues; receptors are present in many extrapituitary structures, including the ovary and placenta. Data suggest that GnRH may be involved in regulating human chorionic gonadotropin (hCG) secretion and implantation, as well as in decreasing cell proliferation and mediating apoptosis in tumor cells (19). The role of GnRH in the extrapituitary sites remains to be fully elucidated.
Gonadotropin-Releasing Hormone Agonists
Mechanism of Action
Used clinically, GnRH agonists are modifications of the native molecule to either increase receptor affinity or decrease degradation (20). Their use leads to a persistent activation of GnRH receptors, as if continuous GnRH exposure existed. As would be predicted by the constant GnRH infusion experiments, this leads to suppression of gonadotropin secretion. An initial release of gonadotropins is followed by a profound suppression of secretion. The initial release of gonadotropins represents the secretion of pituitary stores in response to receptor binding and activation. With continued activation of the gonadotroph GnRH receptor, however, there is a down-regulation effect and a decrease in the concentration of GnRH receptors. As a result, gonadotropin secretion decreases and sex steroid production falls to castrate levels (21).
Additional modification of the GnRH molecule results in an analogue that has no intrinsic activity but competes with GnRH for the same receptor site (22). These GnRH antagonists produce a competitive blockade of GnRH receptors, preventing stimulation by endogenous GnRH and causing an immediate fall in gonadotropin and sex steroid secretion (23). The clinical effect is observed within 24 to 72 hours. Moreover, antagonists may not function solely as competitive inhibitors; evidence suggests they may also produce down-regulation of GnRH receptors, further contributing to the loss of gonadotropin activity (24).
Structure—Agonists and Antagonists
As a peptide hormone, GnRH is degraded by enzymatic cleavage of bonds between its amino acids. Pharmacologic alterations of the structure of GnRH led to the creation of agonists and antagonists (Fig. 7.4). The primary sites of enzymatic cleavage are between amino acids 5 and 6, 6 and 7, and 9 and 10. Substitution of the position-6 amino acid glycine with large bulky amino acid analogues makes degradation more difficult and creates a form of GnRH with a relatively long half-life. Substitution at the carboxyl terminus produces a form of GnRH with increased receptor affinity. The resulting high affinity and slow degradation produces a molecule that mimics continuous exposure to native GnRH (20). Thus, as with constant GnRH exposure, down-regulation occurs. GnRH agonists are widely used to treat disorders that are dependent on ovarian hormones (21). They are used to control ovulation induction cycles and to treat precocious puberty, ovarian hyperandrogenism, leiomyomas, endometriosis, and hormonally dependent cancers. The development of GnRH antagonists proved more difficult because a molecule was needed that maintained the binding and degradation resistance of agonists but failed to activate the receptor. Early attempts involved modification of amino acids 1 and 2, as well as those previously utilized for agonists. Commercial antagonists have structural modifications at amino acids 1, 2, 3, 6, 8, and 10. The treatment spectrum is expected to be similar to that of GnRH agonists, but with more rapid onset of action.
Nonpeptide, small molecule structures with high affinity for the GnRH receptor were developed (25). These compounds demonstrated the ability to suppress the reproductive axis in a dose-related manner via oral administration, unlike the parenteral approach required with traditional peptide analogues (26). Investigation may elucidate an expanded therapeutic role for these antagonists.
Endogenous Opioids and Effects on GnRH
The endogenous opioids are three related families of naturally occurring substances produced in the CNS that represent the natural ligands for the opioid receptors (27–29). There are three major classes of endogenous opioids, each derived from precursor molecules:
1. Endorphins are named for their endogenous morphinelike activity. These substances are produced in the hypothalamus from the precursor proopiomelanocortin (POMC) and have diverse activities, including regulation of temperature, appetite, mood, and behavior (30).
2. Enkephalins are the most widely distributed opioid peptides in the brain, and they function primarily in regulation of the autonomic nervous system. Proenkephalin A is the precursor for the two enkephalins of primary importance: methionine–enkephalin and leucine–enkephalin.
3. Dynorphins are endogenous opioids produced from the precursor proenkephalin B that serve a function similar to that of the endorphins, producing behavioral effects and exhibiting a high analgesic potency.
The endogenous opioids play a significant role in the regulation of hypothalamic–pituitary function. Endorphins appear to inhibit GnRH release within the hypothalamus, resulting in inhibition of gonadotropin secretion (31). Ovarian sex steroids can increase the secretion of central endorphins, further depressing gonadotropin levels (32).
Endorphin levels vary significantly throughout the menstrual cycle, with peak levels in the luteal phase and a nadir during menses (33). This inherent variability, although helping to regulate gonadotropin levels, may contribute to cycle-specific symptoms experienced by ovulatory women. For example, the dysphoria experienced by some women in the premenstrual phase of the cycle may be related to a withdrawal of endogenous opiates (34).
Pituitary Hormone Secretion
Anterior Pituitary
The anterior pituitary is responsible for the secretion of the major hormone-releasing factors—FSH, LH, TSH, and ACTH—as well as GH and prolactin. Each hormone is released by a specific pituitary cell type.
Gonadotropins
The gonadotropins FSH and LH are produced by the anterior pituitary gonadotroph cells and are responsible for ovarian follicular stimulation. Structurally, there is great similarity between FSH and LH (Fig. 7.6). They are both glycoproteins that share identical α subunits and differ only in the structure of their β subunits, which confer receptor specificity (35,36). The synthesis of the β subunits is the rate-regulating step in gonadotropin biosynthesis (37). Thyroid-stimulating hormone and placental hCG also share identical α subunits with the gonadotropins. There are several forms of each gonadotropin, which differ in carbohydrate content as a result of posttranslation modification. The degree of modification varies with steroid levels and is an important regulator of gonadotropin bioactivity.
Figure 7.6 The structural similarity between follicle-stimulating hormone (FSH), luteinizing hormone (LH), and thyroid-stimulating hormone (TSH). The α subunits are identical, and the β subunits differ.
Prolactin
Prolactin, a 198–amino acid polypeptide secreted by the anterior pituitary lactotroph, is the primary trophic factor responsible for the synthesis of milk by the breast (38). Several forms of this hormone, which are named according to their size and bioactivity, are normally secreted (39). Prolactin gene transcription is principally stimulated by estrogen; other hormones promoting transcription are TRH and a variety of growth factors.
Prolactin secretion is under tonic inhibitory control by the hypothalamic secretion of dopamine (40). Therefore, disease states characterized by decreased dopamine secretion or any condition that interrupts transport of dopamine down the infundibular stalk to the pituitary gland will result in increased synthesis of prolactin. In this respect, prolactin is unique in comparison with all other pituitary hormones: It is predominantly under tonic inhibition, and release of control produces an increase in secretion. Clinically, increased prolactin levels are associated with amenorrhea and galactorrhea, and hyperprolactinemia should be suspected in any individual with symptoms of either of these conditions.
Although prolactin appears to be primarily under inhibitory control, many stimuli can elicit its release, including breast manipulation, drugs, stress, exercise, and certain foods. Hormones that may stimulate prolactin release include TRH, vasopressin, γ-aminobutyric acid (GABA), dopamine, β-endorphin, vasoactive intestinal peptide (VIP), epidermal growth factor, angiotensin II, and possibly GnRH (41–43). The relative contributions of these substances under normal conditions remain to be determined.
Thyroid-Stimulating Hormone, Adrenocorticotropic Hormone, and Growth Hormone
The other hormones produced by the anterior pituitary are TSH, ACTH, and GH. Thyroid-stimulating hormone is secreted by the pituitary thyrotrophs in response to TRH. As with GnRH, TRH is synthesized primarily in the arcuate nucleus of the hypothalamus and is secreted into the portal circulation for transport to the pituitary. In addition to stimulating TSH release, TRH is a major stimulus for the release of prolactin. Thyroid-stimulating hormone stimulates release of T3 and T4 from the thyroid gland, which in turn has a negative feedback effect on pituitary TSH secretion. Abnormalities of thyroid secretion (both hyper- and hypothyroidism) are frequently associated with ovulatory dysfunction as a result of diverse actions on the hypothalamic–pituitary–ovarian axis (44).
Adrenocorticotrophic hormone is secreted by the anterior pituitary in response to another hypothalamic-releasing factor, CRH, and stimulates the release of adrenal glucocorticoids. Unlike the other anterior pituitary products, ACTH secretion has a diurnal variation with an early morning peak and a late evening nadir. As with the other pituitary hormones, ACTH secretion is negatively regulated by feedback from its primary end product, which in this case is cortisol.
The anterior pituitary hormone that is secreted in the greatest absolute amount is GH. It is secreted in response to the hypothalamic-releasing factor, GHRH, and by thyroid hormone and glucocorticoids. This hormone is secreted in a pulsatile fashion but with peak release occurring during sleep. In addition to its vital role in the stimulation of linear growth, GH plays a diverse role in physiologic hemostasis. The hormone plays a role in bone mitogenesis, CNS function (improved memory, cognition, and mood), body composition, breast development, and cardiovascular function. It also affects insulin regulation and acts anabolically. Growth hormone appears to have a role in the regulation of ovarian function, although the degree to which it serves this role in normal physiology is unclear (45).
Posterior Pituitary
Structure and Function
The posterior pituitary (neurohypophysis) is composed exclusively of neural tissue and is a direct extension of the hypothalamus. It lies directly adjacent to the adenohypophysis but is embryologically distinct, derived from an invagination of neuroectodermal tissue in the third ventricle. Axons in the posterior pituitary originate from neurons with cell bodies in two distinct regions of the hypothalamus, the supraoptic and paraventricular nuclei, named for their anatomic relationship to the optic chiasm and the third ventricle. Together these two nuclei compose the hypothalamic magnocellular system. These neurons can secrete their synthetic products directly from axonal boutons into the general circulation to act as hormones. This is the mechanism of secretion of the hormones of the posterior pituitary, oxytocin and arginine vasopressin (AVP). Although this is the primary mode of release for these hormones, numerous other secondary pathways were identified, including secretion into the portal circulation, intrahypothalamic secretion, and secretion into other regions of the CNS (46).
Figure 7.7 Oxytocin and arginine-vasopressin (AVP) are 9–amino acid peptides produced by the hypothalamus. They differ in only two amino acids.
In addition to the established functions of oxytocin and vasopressin, several other diverse roles were suggested in animal models. These functions include modulation of sexual activity and appetite, learning and memory consolidation, temperature regulation, and regulation of maternal behaviors (47). In the human, these neuropeptides were linked to social attachment (48–50). Receptor variants for these two molecules were linked to the spectrum of autistic disorders, suggesting that proper function of these two neuropeptides with their receptors is required for positive group interactive behavior. This relationship is strengthened by a strong association between altruistic behavior and the length of the AVP-1a receptor promoter region (51). It is both surprising and humbling that complex human behaviors may be partially explained by such a relatively simple neuropeptide system. Continuing investigation should help elucidate this physiology and potential therapeutic interventions.
Oxytocin
Oxytocin is a 9–amino acid peptide primarily produced by the paraventricular nucleus of the hypothalamus (Fig. 7.7). The primary function of this hormone in humans is the stimulation of two specific types of muscular contractions (Fig. 7.8). The first type, uterine muscular contraction, occurs during parturition. The second type of muscular contraction regulated by oxytocin is breast lactiferous duct myoepithelial contractions, which occur during the milk letdown reflex. Oxytocin release may be stimulated by suckling, triggered by a signal from nipple stimulation transmitted via thoracic nerves to the spinal cord and then to the hypothalamus, where oxytocin is released in an episodic fashion (45). Oxytocin release also may be triggered by olfactory, auditory, and visual clues, and it may play a role in the conditioned reflex in nursing animals. Stimulation of the cervix and vagina can cause significant release of oxytocin, which may trigger reflex ovulation (the Ferguson reflex) in some species.
Figure 7.8 Oxytocin stimulates muscular contractions of the uterus during parturition and the breast lactiferous duct during the milk letdown reflex. Arginine-vasopressin (AVP) regulates circulating blood volume, pressure, and osmolality.
Figure 7.9 The menstrual cycle. The top panel shows the cyclic changes of follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), and progesterone (P) relative to the time of ovulation. The bottom panel correlates the ovarian cycle in the follicular and luteal phases and the endometrial cycle in the proliferative and secretory phases.
Arginine Vasopressin
Also known as antidiuretic hormone (ADH), AVP is the second major secretory product of the posterior pituitary (Fig. 7.7). It is synthesized primarily by neurons with cell bodies in the supraoptic nuclei (Fig. 7.8). Its major function is the regulation of circulating blood volume, pressure, and osmolality (52). Specific receptors throughout the body can trigger the release of AVP. Osmoreceptors located in the hypothalamus sense changes in blood osmolality from a mean of 285 mOSM/kg. Baroceptors sense changes in blood pressure caused by alterations in blood volume and are peripherally located in the walls of the left atrium, carotid sinus, and aortic arch (53). These receptors can respond to changes in blood volume of more than 10%. In response to decreases in blood pressure or volume, AVP is released and causes arteriolar vasoconstriction and renal free-water conservation. This in turn leads to a decrease in blood osmolality and an increase in blood pressure. Activation of the renal renin–angiotensin system can also activate AVP release.
Menstrual Cycle Physiology
In the normal menstrual cycle, orderly cyclic hormone production and parallel proliferation of the uterine lining prepare for implantation of the embryo. Disorders of the menstrual cycle and, likewise, disorders of menstrual physiology, may lead to various pathologic states, including infertility, recurrent miscarriage, and malignancy.
Normal Menstrual Cycle
The normal human menstrual cycle can be divided into two segments: the ovarian cycle and the uterine cycle, based on the organ under examination. The ovarian cycle may be further divided into follicular and luteal phases, whereas the uterine cycle is divided into corresponding proliferative and secretory phases (Fig. 7.9). The phases of the ovarian cycle are characterized as follows:
1. Follicular phase—hormonal feedback promotes the orderly development of a single dominant follicle, which should be mature at midcycle and prepared for ovulation. The average length of the human follicular phase ranges from 10 to 14 days, and variability in this length is responsible for most variations in total cycle length.
2. Luteal phase—the time from ovulation to the onset of menses has an average length of 14 days.
A normal menstrual cycle lasts from 21 to 35 days, with 2 to 6 days of flow and an average blood loss of 20 to 60 mL. However, studies of large numbers of women with normal menstrual cycles showed that only approximately two-thirds of adult women have cycles lasting 21 to 35 days (54). The extremes of reproductive life (after menarche and perimenopause) are characterized by a higher percentage of anovulatory or irregularly timed cycles (55,56).
Hormonal Variations
The relative pattern of ovarian, uterine, and hormonal variation along the normal menstrual cycle is shown in Fig. 7.9.
1. At the beginning of each monthly menstrual cycle, levels of gonadal steroids are low and have been decreasing since the end of the luteal phase of the previous cycle.
2. With the demise of the corpus luteum, FSH levels begin to rise, and a cohort of growing follicles is recruited. These follicles each secrete increasing levels of estrogen as they grow in the follicular phase. The increase in estrogen, in turn, is the stimulus for uterine endometrial proliferation.
3. Rising estrogen levels provide negative feedback on pituitary FSH secretion, which begins to wane by the midpoint of the follicular phase. In addition, the growing follicles produce inhibin-B, which suppresses FSH secretion by the pituitary. Conversely, LH initially decreases in response to rising estradiol levels, but late in the follicular phase the LH level is increased dramatically (biphasic response).
4. At the end of the follicular phase (just before ovulation), FSH-induced LH receptors are present on granulosa cells and, with LH stimulation, modulate the secretion of progesterone.
5. After a sufficient degree of estrogenic stimulation, the pituitary LH surge is triggered, which is the proximate cause of ovulation that occurs 24 to 36 hours later. Ovulation heralds the transition to the luteal–secretory phase.
6. The estrogen level decreases through the early luteal phase from just before ovulation until the midluteal phase, when it begins to rise again as a result of corpus luteum secretion. Similarly, inhibin-A is secreted by the corpus luteum.
7. Progesterone levels rise precipitously after ovulation and can be used as a presumptive sign that ovulation has occurred.
8. Progesterone, estrogen, and inhibin-A act centrally to suppress gonadotropin secretion and new follicular growth. These hormones remain elevated through the lifespan of the corpus luteum and then wane with its demise, thereby setting the stage for the next cycle.
Uterus
Cyclic Changes of the Endometrium
In 1950, Noyes, Hertig, and Rock described the cyclic histologic changes in the adult human endometrium (57) (Fig. 7.10). These changes proceed in an orderly fashion in response to cyclic hormonal production by the ovaries (Fig. 7.9). Histologic cycling of the endometrium can best be viewed in two parts: the endometrial glands and the surrounding stroma. The superficial two-thirds of the endometrium is the zone that proliferates and is ultimately shed with each cycle if pregnancy does not occur. This cycling portion of the endometrium is known as the decidua functionalis and is composed of a deeply situated intermediate zone (stratum spongiosum) and a superficial compact zone (stratum compactum). The decidua basalis is the deepest region of the endometrium. It does not undergo significant monthly proliferation but, instead, is the source of endometrial regeneration after each menses (58).
Figure 7.10 The number of oocytes in the ovary before and after birth and through menopause.
The existence of endometrial stem cells was assumed but difficult to document. Researchers found a small population of human epithelial and stromal cells that possess clonogenicity, suggesting that they represent the putative endometrial stem cells (59). Further evidence of the existence of such cells, and their source, was provided by another study that showed endometrial glandular epithelial cells obtained from endometrial biopsies of women undergoing bone marrow transplants, express the HLA type of the donor bone marrow (60). This finding suggests that endometrial stem cells exist, and that they reside in bone marrow and migrate to the basalis of the endometrium. Furthermore, the timing of the appearance of these cells following the transplant was as long as several years. This fact may prove to be of clinical importance in patients with Asherman syndrome who experienced a loss of functional endometrium; repair of the uterine anatomy may eventually result in a functioning endometrial cavity.
Proliferative Phase
By convention, the first day of vaginal bleeding is called day 1 of the menstrual cycle. After menses, the decidua basalis is composed of primordial glands and dense scant stroma in its location adjacent to the myometrium. The proliferative phase is characterized by progressive mitotic growth of the decidua functionalis in preparation for implantation of the embryo in response to rising circulating levels of estrogen (61). At the beginning of the proliferative phase, the endometrium is relatively thin (1--2 mm). The predominant change seen during this time is evolution of the initially straight, narrow, and short endometrial glands into longer, tortuous structures (62). Histologically, these proliferating glands have multiple mitotic cells, and their organization changes from a low columnar pattern in the early proliferative period to a pseudostratified pattern before ovulation. Throughout this time, the stroma is a dense compact layer, and vascular structures are infrequently seen.
Secretory Phase
In the typical 28-day cycle, ovulation occurs on cycle day 14. Within 48 to 72 hours following ovulation, the onset of progesterone secretion produces a shift in histologic appearance of the endometrium to the secretory phase, so named for the clear presence of eosinophilic protein-rich secretory products in the glandular lumen. In contrast to the proliferative phase, the secretory phase of the menstrual cycle is characterized by the cellular effects of progesterone in addition to estrogen. In general, progesterone’s effects are antagonistic to those of estrogen, and there is a progressive decrease in the endometrial cell’s estrogen receptor concentration. As a result, during the latter half of the cycle, estrogen-induced DNA synthesis and cellular mitosis are antagonized (61).
During the secretory phase, the endometrial glands form characteristic periodic acid–Schiff positive–staining, glycogen-containing vacuoles. These vacuoles initially appear subnuclearly and then progress toward the glandular lumen (57) (Fig. 7.10). The nuclei can be seen in the midportion of the cells and ultimately undergo apocrine secretion into the glandular lumen, often by cycle day 19 or 20. At postovulatory day 6 or 7, secretory activity of the glands is generally maximal, and the endometrium is optimally prepared for implantation of the blastocyst.
The stroma of the secretory phase remains unchanged histologically until approximately the seventh postovulatory day, when there is a progressive increase in edema. Coincident with maximal stromal edema in the late secretory phase, the spiral arteries become clearly visible and then progressively lengthen and coil during the remainder of the secretory phase. By around day 24, an eosinophilic-staining pattern, known as cuffing, is visible in the perivascular stroma. Eosinophilia then progresses to form islands in the stroma followed by areas of confluence. This staining pattern of the edematous stroma is termed pseudodecidual because of its similarity to the pattern that occurs in pregnancy. Approximately 2 days before menses, there is a dramatic increase in the number of polymorphonuclear lymphocytes that migrate from the vascular system. This leukocytic infiltration heralds the collapse of the endometrial stroma and the onset of the menstrual flow.
Menses
In the absence of implantation, glandular secretion ceases and an irregular breakdown of the decidua functionalis occurs. The resultant shedding of this layer of the endometrium is termed menses. The destruction of the corpus luteum and its production of estrogen and progesterone is the presumed cause of the shedding. With withdrawal of sex steroids, there is a profound spiral artery vascular spasm that ultimately leads to endometrial ischemia. Simultaneously, there is a breakdown of lysosomes and a release of proteolytic enzymes, which further promote local tissue destruction. This layer of endometrium is then shed, leaving the decidua basalis as the source of subsequent endometrial growth. Prostaglandins are produced throughout the menstrual cycle and are at their highest concentration during menses (60). Prostaglandin F2α (PGF2α) is a potent vasoconstrictor, causing further arteriolar vasospasm and endometrial ischemia. PGF2α produces myometrial contractions that decrease local uterine wall blood flow and may serve to expel physically the sloughing endometrial tissue from the uterus.
Dating the Endometrium
The changes seen in secretory endometrium relative to the LH surge were thought to allow the assessment of the “normalcy” of endometrial development. Since 1950, it was felt that by knowing when a patient ovulated, it was possible to obtain a sample of endometrium by endometrial biopsy and determine whether the state of the endometrium corresponds to the appropriate time of the cycle. Traditional thinking held that any discrepancy of more than 2 days between chronologic and histologic date indicated a pathologic condition termed luteal phase defect; this abnormality was linked to both infertility (via implantation failure) and early pregnancy loss (63).
Evidence suggests a lack of utility for the endometrial biopsy as a diagnostic test for either infertility or early pregnancy loss (56). In a randomized, observational study of regularly cycling, fertile women, it was found that endometrial dating is far less accurate and precise than originally claimed and does not provide a valid method for the diagnosis of luteal phase defect (64). Furthermore, a large prospective, multicenter trial sponsored by the National Institutes of Health showed that histologic dating of the endometrium does not discriminate between fertile and infertile women (65). Thus, after half a century of using this test in the evaluation of the subfertile couple, it became clear that the endometrial biopsy has no role in the routine evaluation of infertility or early pregnancy loss.
Ovarian Follicular Development
The number of oocytes peaks in the fetus at 6 to 7 million by 20 weeks of gestation (66) (Fig. 7.10). Simultaneously (and peaking at the 5th month of gestation), atresia of the oogonia occurs, rapidly followed by follicular atresia. At birth, only 1 to 2 million oocytes remain in the ovaries, and at puberty, only 300,000 of the original 6 to 7 million oocytes are available for ovulation (66,67). Of these, only 400 to 500 will ultimately be released during ovulation. By the time of menopause, the ovary will be composed primarily of dense stromal tissue with only rare interspersed oocytes remaining.
A central dogma of reproductive biology is that in mammalian females there is no capacity for oocyte production postnatally. Because oocytes enter the diplotene resting stage of meiosis in the fetus and persist in this stage until ovulation, much of the DNA, proteins, and messenger RNA (mRNA) necessary for development of the preimplantation embryo is synthesized by this stage. At the diplotene stage, a single layer of 8 to 10 granulosa cells surround the oogonia to form the primordial follicle. The oogonia that fail to become properly surrounded by granulosa cells undergo atresia (68). The remainder proceeds with follicular development. Thus, most oocytes are lost during fetal development, and the remaining follicles are steadily “used up” throughout the intervening years until menopause.
Evidence has begun to challenge this theory. Studies in the mouse showed that production of oocytes and corresponding folliculogenesis can occur well into adult life (69). The reservoir of germline stem cells responsible for this oocyte development appears to reside in the bone marrow (70). It is not clear whether such stem cells exist in adult humans, and if they do, what clinical function they might provide.
Meiotic Arrest of Oocyte and Resumption
Meiosis (the germ cell process of reduction division) is divided into four phases: prophase, metaphase, anaphase, and telophase. The prophase of meiosis I is further divided into five stages: leptotene, zygotene, pachytene, diplotene, and diakinesis.
Oogonia differ from spermatogonia in that only one final daughter cell (oocyte) forms from each precursor cell, with the excess genetic material discarded in three polar bodies. When the developing oogonia begin to enter meiotic prophase I, they are known as primary oocytes (71). This process begins at roughly 8 weeks of gestation. Only those oogonia that enter meiosis will survive the wave of atresia that sweeps the fetal ovary before birth. The oocytes arrested in prophase (in the late diplotene or “dictyate” stage) will remain so until the time of ovulation, when the process of meiosis resumes. The mechanism for this mitotic stasis is believed to be an oocyte maturation inhibitor (OMI) produced by granulosa cells (72). This inhibitor gains access to the oocyte via gap junctions connecting the oocyte and its surrounding cumulus of granulosa. With the midcycle LH surge, the gap junctions are disrupted, granulosa cells are no longer connected to the oocyte, and meiosis I is allowed to resume.
Follicular Development
Follicular development is a dynamic process that continues from menarche until menopause. The process is designed to allow the monthly recruitment of a cohort of follicles and, ultimately, to release a single, mature, dominant follicle during ovulation each month.
Primordial Follicles
The initial recruitment and growth of the primordial follicles is gonadotropin independent and affects a cohort over several months (73). The stimuli responsible for the recruitment of a specific cohort of follicles in each cycle are unknown. At the primordial follicle stage, shortly after initial recruitment, FSH assumes control of follicular differentiation and growth and allows a cohort of follicles to continue differentiation. This process signals the shift from gonadotropin-independent to gonadotropin-dependent growth. The first changes seen are growth of the oocyte and expansion of the single layer of follicular granulosa cells into a multilayer of cuboidal cells. The decline in luteal phase estrogen, progesterone, and inhibin-A production by the now-fading corpus luteum from the previous cycle allows the increase in FSH that stimulates this follicular growth (74).
Preantral Follicle
During the several days following the breakdown of the corpus luteum, growth of the cohort of follicles continues, driven by the stimulus of FSH. The enlarging oocyte secretes a glycoprotein-rich substance, the zona pellucida, which separates it from the surrounding granulosa cells (except for the aforementioned gap junction). With transformation from a primordial to a preantral follicle, there is continued mitotic proliferation of the encompassing granulosa cells. Simultaneously, theca cells in the stroma bordering the granulosa cells proliferate. Both cell types function synergistically to produce estrogens that are secreted into the systemic circulation. At this stage of development, each of the seemingly identical cohort members must either be selected for dominance or undergo atresia. It is likely that the follicle destined to ovulate was selected before this point, although the mechanism for selection remains obscure.
Figure 7.11 The two-cell, two-gonadotropin theory of follicular development in which there is compartmentalization of steroid hormone synthesis in the developing follicle. LH, luteinizing hormone; FSH, follicle-stimulating hormone.
Two-Cell, Two-Gonadotropin Theory
The fundamental tenet of follicular development is the two-cell, two-gonadotropin theory (73,75,76) (Fig. 7.11). This theory states that there is a subdivision and compartmentalization of steroid hormone synthesis activity in the developing follicle. Most aromatase activity (for estrogen production) is in the granulosa cells (77). Aromatase activity is enhanced by FSH stimulation of specific receptors on these cells (78,79). Granulosa cells lack several enzymes that occur earlier in the steroidogenic pathway and require androgens as a substrate for aromatization. Androgens, in turn, are synthesized primarily in response to stimulation by LH, and the theca cells possess most of the LH receptors at this stage (78,79). Therefore, a synergistic relationship must exist: LH stimulates the theca cells to produce androgens (primarily androstenedione), which in turn are transferred to the granulosa cells for FSH-stimulated aromatization into estrogens. These locally produced estrogens create a microenvironment within the follicle that is favorable for continued growth and nutrition (80). Both FSH and local estrogens serve to further stimulate estrogen production, FSH receptor synthesis and expression, and granulosa cell proliferation and differentiation.
Androgens have two positive regulatory roles in follicular development. Within the ovary, androgens promote granulose cell proliferation, aromatase activity, and inhibit programmed death of these cells (81).
As the peripheral estrogen level rises, it negatively feeds back on the pituitary and hypothalamus to decrease circulating FSH levels (82). Increased ovarian production of inhibin-B further decreases FSH production at this point.
The falling FSH level that occurs with the progression of the follicular phase represents a threat to continued follicular growth. The resulting adverse environment can be withstood only by follicles with a selective advantage for binding the diminishing FSH molecules; that is, those with the greatest number of FSH receptors. The dominant follicle, therefore, can be perceived as the one with a richly estrogenic microenvironment and the most FSH receptors (83). As it grows and develops, the follicle continues to produce estrogen, which results in further lowering of the circulating FSH and creates a more adverse environment for competing follicles. This process continues until all members of the initial cohort, with the exception of the single dominant follicle, have suffered atresia. The stage is then set for ovulation.
Chronic elevation of androgens suppresses hypothalamic–pituitary secretion of FSH, a detriment to the development and maturation of a dominant follicle (81). Clinically, androgen excess results in chronic anovulation, as is seen in polycystic ovarian syndrome.
Preovulatory Follicle
Preovulatory follicles are characterized by a fluid-filled antrum that is composed of plasma with granulosa cell secretions. The granulosa cells at this point have further differentiated into a heterogenous population. The oocyte remains connected to the follicle by a stalk of specialized granulosa known as the cumulus oophorus.
Rising estrogen levels have a negative feedback effect on FSH secretion. Conversely, LH undergoes biphasic regulation by circulating estrogens. At lower concentrations, estrogens inhibit LH secretion. At higher levels, estrogen enhances LH release. This stimulation requires a sustained high level of estrogen (200 pg/mL) for more than 48 hours (84). Once the rising estrogen level produces positive feedback, a substantial surge in LH secretion occurs. Concomitant to these events, the local estrogen–FSH interactions in the dominant follicle induce LH receptors on the granulosa cells. Exposure to high levels of LH results in a specific response by the dominant follicle—the result is luteinization of the granulosa cells, production of progesterone, and initiation of ovulation. Ovulation will occur in the single mature, or Graafian, follicle 10 to 12 hours after the LH peak or 34 to 36 hours after the initial rise in midcycle LH (85–87).
As suggested previously, the sex steroids are not the only gonadotropin regulators of follicular development. Two related granulosa cell–derived peptides were identified that play opposing roles in pituitary feedback (88). The first of these peptides, inhibin, is secreted in two forms: inhibin-A and inhibin-B. Inhibin-B is secreted primarily in the follicular phase and is stimulated by FSH, whereas inhibin-A is mainly active in the luteal phase (89). Both forms of inhibin act to inhibit FSH synthesis and release (90,91). The second peptide, activin, stimulates FSH release from the pituitary gland and potentiates its action in the ovary (92,93). It is likely that there are numerous other intraovarian regulators similar to inhibin and activin, each of which may play a key role in promoting the normal ovulatory process (94). Some of these include follistatin, insulinlike growth factor-1 (ILGF-1), epidermal growth factor (EGF)/transforming growth factor-α (TGF-α), TGF-β1, fibroblast growth factor-β (FGF-β), interleukin-1, tissue necrosis factor-α, OMI, and renin–angiotensin.
Ovulation
The midcycle LH surge is responsible for a dramatic increase in local concentrations of prostaglandins and proteolytic enzymes in the follicular wall (95). These substances progressively weaken the follicular wall and ultimately allow a perforation to form. Ovulation most likely represents a slow extrusion of the oocyte through this opening in the follicle rather than a rupture of the follicular structure (96). Direct measurements of intrafollicular pressures were recorded and failed to demonstrate an explosive event.
Luteal Phase
Structure of Corpus Luteum
After ovulation, the remaining follicular shell is transformed into the primary regulator of the luteal phase: the corpus luteum. Membranous granulosa cells remaining in the follicle begin to take up lipids and the characteristic yellow lutein pigment for which the structure is named. These cells are active secretory structures that produce progesterone, which supports the endometrium of the luteal phase. In addition, estrogen and inhibin-A are produced in significant quantities. Unlike the process that occurs in the developing follicle, the basement membrane of the corpus luteum degenerates to allow proliferating blood vessels to invade the granulosa-luteal cells in response to secretion of angiogenic factors such as vascular endothelial growth factor (97). This angiogenic response allows large amounts of luteal hormones to enter the systemic circulation.
Hormonal Function and Regulation
The hormonal changes of the luteal phase are characterized by a series of negative feedback interactions designed to lead to regression of the corpus luteum if pregnancy does not occur. Corpus luteum steroids (estradiol and progesterone) provide negative central feedback and cause a decrease in FSH and LH secretion. Continued secretion of both steroids will decrease the stimuli for subsequent follicular recruitment. Similarly, luteal secretion of inhibin also potentiates FSH withdrawal. In the ovary, local production of progesterone inhibits the further development and recruitment of additional follicles.
Continued corpus luteum function depends on continued LH production. In the absence of this stimulation, the corpus luteum will invariably regress after 12 to 16 days and form the scarlike corpora albicans (98). The exact mechanism of luteolysis is unclear and most likely involves local paracrine factors. In the absence of pregnancy, the corpus luteum regresses, and estrogen and progesterone levels wane. This, in turn, removes central inhibition on gonadotropin secretion and allows FSH and LH levels to again rise and recruit another cohort of follicles.
If pregnancy does occur, placental hCG will mimic LH action and continually stimulate the corpus luteum to secrete progesterone. Successful implantation results in hormonal support to allow continued maintenance of the corpus luteum and the endometrium. Evidence from patients undergoing oocyte donation cycles demonstrated that continued luteal function is essential to continuation of the pregnancy until approximately 5 weeks of gestation, when sufficient progesterone is produced by the developing placenta (99). This switch in the source of regulatory progesterone production is referred to as the luteal–placental shift.
Summary of Menstrual Cycle Regulation
Following is a summary of the regulation of the menstrual cycle:
1. GnRH is produced in the arcuate nucleus of the hypothalamus and secreted in a pulsatile fashion into the portal circulation, where it travels to the anterior pituitary.
2. Ovarian follicular development moves from a period of gonadotropin independence to a phase of FSH dependence.
3. As the corpus luteum of the previous cycle fades, luteal production of progesterone and inhibin-A decreases, allowing FSH levels to rise.
4. In response to FSH stimulus, the follicles grow, differentiate, and secrete increasing amounts of estrogen and inhibin-B.
5. Estrogen stimulates growth and differentiation of the functional layer of the endometrium, which prepares for implantation. Estrogens work with FSH in stimulating follicular development.
6. The two-cell, two-gonadotropin theory dictates that with LH stimulation, the ovarian theca cells will produce androgens that are converted by the granulosa cells into estrogens under the stimulus of FSH.
7. Rising estrogen and inhibin levels negatively feed back on the pituitary gland and hypothalamus and decrease the secretion of FSH.
8. The one follicle destined to ovulate each cycle is called the dominant follicle. It has relatively more FSH receptors and produces a larger concentration of estrogens than the follicles that will undergo atresia. It is able to continue to grow despite falling FSH levels.
9. Sustained high estrogen levels cause a surge in pituitary LH secretion that triggers ovulation, progesterone production, and the shift to the secretory, or luteal, phase.
10. Luteal function is dependent on the presence of LH. The corpus luteum secretes estrogen, progesterone, and inhibin-A, which serve to maintain gonadotropin suppression. Without continued LH secretion, the corpus luteum will regress after 12 to 16 days. The resulting loss of progesterone secretion results in menstruation.
11. If pregnancy occurs, the embryo secretes hCG, which mimics the action of LH by sustaining the corpus luteum. The corpus luteum continues to secrete progesterone and supports the secretory endometrium, allowing the pregnancy to continue to develop.
References
1. Bloom FE. Neuroendocrine mechanisms: cells and systems. In: Yen SCC, Jaffe RB, eds. Reproductive endocrinology. Philadelphia, PA: Saunders, 1991:2–24.
Simerly RB, Chang C, Muramatsu M, et al. Distribution of androgen and estrogen receptor mRNA-containing cells in the rat brain: an in situ hybridization study. J Comp Neurol 1990;294:76–95.
3. Brown TJ, Hochberg RB, Naftolin F. Pubertal development of estrogen receptors in the rat brain. Mol Cell Neurosci 1994;5:475–483.
4. Bergland RM, Page RB. Can the pituitary secrete directly to the brain? Affirmative anatomic evidence. Endocrinology 1978;102:1325–1338.
5. Duello TM, Halmi NS. Ultrastructural-immunocytochemical localization of growth hormone and prolactin in human pituitaries. J Clin Endocrinol Metab 1979;49:189–196.
Blackwell RE, Amoss M Jr, Vale W, et al. Concomitant release of FSH and LH induced by native and synthetic LRF. Am J Physiol 1973;224:170–175.
7. Krey LC, Butler WR, Knobil E. Surgical disconnection of the medial basal hypothalamus and pituitary function in the rhesus monkey. I. Gonadotropin secretion. Endocrinology 1975;96:1073–1087.
Plant TM, Krey LC, Moossy J, et al. The arcuate nucleus and the control of the gonadotropin and prolactin secretion in the female rhesus monkey (Macaca mulatta). Endocrinology 1978;102:52–62.
Amoss M, Burgus R, Blackwell RE, et al. Purification, amino acid composition, and N-terminus of the hypothalamic luteinizing hormone releasing factor (LRF) of ovine origin. Biochem Biophys Res Commun 1971;44:205–210.
10. Schwanzel-Fukuda M, Pfaff DW. Origin of luteinizing hormone releasing hormone neurons. Nature 1989;338:161–164.
Dierschke DJ, Bhattacharya AN, Atkinson LE, et al. Circhoral oscillations of plasma LH levels in the ovariectomized rhesus monkey. Endocrinology 1970;87:850–853.
12. Knobil E. Neuroendocrine control of the menstrual cycle. Recent Prog Horm Res 1980;36:53–88.
Belchetz PE, Plant TM, Nakai Y, et al. Hypophyseal responses to continuous and intermittent delivery of hypothalamic gonadotropin-releasing hormone. Science 1978;202:631–633.
Nakai Y, Plant TM, Hess DL, et al. On the sites of the negative and positive feedback actions of estradiol and the control of gonadotropin secretion in the rhesus monkey. Endocrinology 1978;102:1008–1014.
15. Rabin D, McNeil LW. Pituitary and gonadal desensitization after continuous luteinizing hormone–releasing hormone infusion in normal females. J Clin Endocrinol Metab1980,51:873–876.
16. Hoff JD, Lasley BL, Yen SSC. Functional relationship between priming and releasing actions of luteinizing hormone–releasing hormone. J Clin Endocrinol Metab 1979;49:8–11.
Soules MR, Steiner RA, Cohen NL, et al. Nocturnal slowing of pulsatile luteinizing hormone secretion in women during the follicular phase of the menstrual cycle. J Clin Endocrinol Metab 1985;61:43–49.
Filicori M, Santoro N, Marriam GR, et al. Characterization of the physiological pattern of episodic gonadotropin secretion throughout the human menstrual cycle. J Clin Endocrinol Metab 1986;62:1136–1144.
19. Yu B, Ruman J, Christman G. The role of peripheral gonadotropin-releasing hormone receptors in female reproduction. Fertil Steril 2011;95:465–473.
20. Karten MJ, Rivier JE. Gonadotropin-releasing hormone analog design. Structure function studies towards the development of agonists and antagonists: rationale and perspective. Endocr Rev 1986;7:44–66.
21. Conn PM, Crowley WF Jr. Gonadotropin-releasing hormone and its analogs. Annu Rev Med 1994;45:391–405.
22. Loy RA. The pharmacology and potential applications of GnRH antagonists. Curr Opin Obstet Gynecol 1994;6:262–268.
23. Schally AV. LH-RH analogues. I. Their impact on reproductive medicine. Gynecol Endocrinol 1999;13:401–409.
Halmos G, Schally AV, Pinski J, et al. Down-regulation of pituitary receptors for luteinizing hormone–releasing hormone (LH-RH) in rats by LH-RH antagonist Cetrorelix. Proc Natl Acad Sci U S A 1996;93:2398–2402.
25. Betz SF, Zhu YF, Chen C, Struthers RS. Non-peptide gonadotropin-releasing hormone receptor antagonists. J Med Chem 2008;51:3331–3348.
Struthers RS, Nicholls AJ, Grundy J, et al. Suppression of gonadotropins and estradiol in premenopausal women by oral administration of the nonpeptide gonadotropin-releasing hormone antagonist elagolix. J Clin Endocrinol Metab 2009;94:545–551.
Hughes J, Smith TW, Kosterlitz LH, et al. Identification of two related pentapeptides from the brain with potent opiate agonist activity. Nature 1975;258:577–580.
28. Howlett TA, Rees LH. Endogenous opioid peptide and hypothalamo-pituitary function. Annu Rev Physiol 1986;48:527–536.
29. Facchinetti F, Petraglia F, Genazzani AR. Localization and expression of the three opioid systems. Semin Reprod Endocrinol 1987;5:103.
30. Goldstein A. Endorphins: physiology and clinical implications. Ann N Y Acad Sci 1978;311:49–58.
31. Grossman A. Opioid peptides and reproductive function. Semin Reprod Endocrinol 1987;5:115–124.
Reid Rl, Hoff JD, Yen SSC, et al. Effects of exogenous β-endorphin on pituitary hormone secretion and its disappearance rate in normal human subjects. J Clin Endocrinol Metab 1981;52:1179–1184.
33. Gindoff PR, Ferin M. Brain opioid peptides and menstrual cyclicity. Semin Reprod Endocrinol 1987;5:125–133.
34. Halbreich U, Endicott J. Possible involvement of endorphin withdrawal or imbalance in specific premenstrual syndromes and postpartum depression. Med Hypotheses 1981;7:1045–1058.
35. Fiddes JC, Talmadge K. Structure, expression and evolution of the genes for human glycoprotein hormones. Recent Prog Horm Res 1984;40:43–78.
Vaitukaitis JL, Ross JT, Bourstein GD, et al. Gonadotropins and their subunits: basic and clinical studies. Recent Prog Horm Res 1976;32:289–331.
37. Lalloz MRA, Detta A, Clayton RN. GnRH desensitization preferentially inhibits expression of the LH β-subunit gene in vivo. Endocrinology 1988;122:1689–1694.
Brun del Re R, del Pozo E, de Grandi P, et al. Prolactin inhibition and suppression of puerperal lactation by a Br-ergocriptine (CB 154): a comparison with estrogen. Obstet Gynecol 1973;41:884–890.
39. Suh HK, Frantz AG. Size heterogeneity of human prolactin in plasma and pituitary extracts. J Clin Endocrinol Metab 1974:39:928–935.
40. MacLeod RM. Influence of norepinephrine and catecholamine depletion agents synthesis in release of prolactin growth hormone. Endocrinology 1969;85:916–923.
Vale W, Blackwell RE, Grant G, et al. TRF and thyroid hormones on prolactin secretion by rat pituitary cell in vitro. Endocrinology 1973;93:26–33.
Matsushita N, Kato Y, Shimatsu A, et al. Effects of VIP, TRH, GABA and dopamine on prolactin release from superfused rat anterior pituitary cells. Life Sci 1983;32:1263–1269.
Dufy-Barbe L, Rodriguez F, Arsaut J, et al. Angiotensin-II stimulates prolactin release in the rhesus monkey. Neuroendocrinology 1982;35:242–247.
44. Burrow GN. The thyroid gland and reproduction. In: Yen SCC, Jaffe RB, eds. Reproductive endocrinology. Philadelphia, PA: Saunders, 1991:555–575.
45. Katz E, Ricciarelli E, Adashi EY. The potential relevance of growth hormone to female reproductive physiology and pathophysiology. Fertil Steril 1993;59:8–34.
46. Yen SCC. The hypothalamic control of pituitary hormone secretion. In: Yen SCC, Jaffe RB, eds. Reproductive endocrinology. Philadelphia, PA: Saunders, 1991:65–104.
47. Insel TR. Oxytocin and the neuroendocrine basis of affiliation. In: Schulkin J, ed. Hormonally induced changes in mind and brain. New York: Academic Press, 1993:225–251.
48. Stein DJ. Oxytocin and vasopressin: social neuropeptides. CNS Spectr 2009;14:602–606.
49. Donaldson ZR, Young LJ. Oxytocin, vasopressin, and the neurogenetics of sociality. Science 2008;322:900–904.
Yamasue H, Kuwabara H, Kawakubo Y, et al. Oxytocin, sexually dimorphic features of the social brain, and autism. Psychiatry Clin Neurosci 2009;63:129–140.
Israel S, Lerer E, Shalev I, et al. Molecular genetic studies of the arginine vasopressin 1a receptor (AVPR1a) and the oxytocin receptor (OXTR) in human behaviour: from autism to altruism with some notes in between. Prog Brain Res 2008;170:435–449.
McNeilly AS, Robinson IC, Houston MJ, et al. Release of oxytocin and PRL in response to suckling. BMJ 1983;286:257–259.
Dunn FL, Brennan TJ, Nelson AE, et al. The role of blood osmolality and volume in regulating vasopressin secretion in the rat. J Clin Invest 1973;52:3212–3219.
54. Vollman RF. The menstrual cycle. In: Friedman E, ed. Major problems in obstetrics and gynecology. Philadelphia, PA: Saunders, 1977:1–193.
Treloar AE, Boynton RE, Borghild GB, et al. Variation of the human menstrual cycle through reproductive life. Int J Fertil 1967;12:77–126.
56. Collett ME, Wertenberger GE, Fiske VM. The effects of age upon the pattern of the menstrual cycle. Fertil Steril 1954;5:437–448.
57. Noyes RW, Hertig AW, Rock J. Dating the endometrial biopsy. Fertil Steril 1950;1:3–25.
58. Flowers CE Jr, Wilbron WH. Cellular mechanisms for endometrial conservation during menstrual bleeding. Semin Reprod Endocrinol 1984;2:307–341.
59. Chan RW, Schwab KE, Gargett CE. Clonogenicity of human endometrial epithelial and stromal cells. Biol Reprod 2004;70:1738–1750.
60. Taylor HS. Endometrial cells derived from donor stem cells in bone marrow transplant recipients. JAMA 2004;292:81–85.
61. Ferenczy A, Bertrand G, Gelfand MM. Proliferation kinetics of human endometrium during the normal menstrual cycle. Am J Obstet Gynecol 1979;133:859–867.
62. Schwarz BE. The production and biologic effects of uterine prostaglandins. Semin Reprod Endocrinol 1983;1:189.
63. Olive DL. The prevalence and epidemiology of luteal-phase deficiency in normal and infertile women. Clin Obstet Gynecol 1991;34:157–166.
Murray MJ, Meyer WR, Zaino RJ, et al. A critical analysis of the accuracy, reproducibility, and clinical utility of histologic endometrial dating in infertile women. Fertil Steril 2004;81:1333–1343.
Coutifaris C, Myers ER, Guzick DS, et al. Histologic dating of timed endometrial biopsy tissue is not related to fertility status. Fertil Steril 2004;82:1264–1272.
66. Peters H, Byskov AG, Grinsted J. Follicular growth in fetal and prepubertal ovaries in humans and other primates. J Clin Endocrinol Metab 1978;7:469–485.
Himelstein-Braw R, Byskov AG, Peters H, et al. Follicular atresia in the infant human ovary. J Reprod Fertil 1976;46:55–59.
68. Wassarman PM, Albertini DF. The mammalian ovum. In: Knobil E, Neill JD, eds. The physiology of reproduction. New York: Raven Press, 1994:240–244.
Johnson J, Canning J, Kaneko T, et al. Germline stem cells and follicular renewal in the postnatal mammalian ovary. Nature 2004;428:145–150.
Johnson J, Bagley J, Skaznik-Wikiel M, et al. Oocyte generation in adult mammalian ovaries by putative germ cells in bone marrow and peripheral blood. Cell 2005;122:303–315.
71. Gondos B, Bhiraleus P, Hobel CJ. Ultrastructural observations on germ cells in human fetal ovaries. Am J Obstet Gynecol 1971;110:644–652.
72. Tsafriri A, Dekel N, Bar-Ami S. A role of oocyte maturation inhibitor in follicular regulation of oocyte maturation. J Reprod Fertil 1982;64:541–551.
Halpin DMG, Jones A, Fink G, et al. Post-natal ovarian follicle development in hypogonadal (HPG) and normal mice and associated changes in the hypothalamic-pituitary axis. J Reprod Fertil 1986;77:287–296.
74. Vermesh M, Kletzky OA. Longitudinal evaluation of the luteal phase and its transition into the follicular phase. J Clin Endocrinol Metab 1987;65:653–658.
Erickson GF, Magoffin DA, Dyer CA, et al. Ovarian androgen producing cells: a review of structure/function relationships. Endocr Rev 1985;6:371–399.
76. Erickson GF. An analysis of follicle development and ovum maturation. Semin Reprod Endocrinol 1986;46:55–59.
77. Ryan KJ, Petro Z. Steroid biosynthesis of human ovarian granulosa and thecal cells. J Clin Endocrinol Metab 1966;26:46–52.
78. Kobayashi M, Nakano R, Ooshima A. Immunohistochemical localization of pituitary gonadotropin and gonadal steroids confirms the two cells two gonadotropins hypothesis of steroidogenesis in the human ovary. J Endocrinol 1990;126:483–488.
79. Yamoto M, Shima K, Nakano R. Gonadotropin receptors in human ovarian follicles and corpora lutea throughout the menstrual cycle. Horm Res 1992;37[Suppl 1]:5–11.
Hseuh AJ, Adashi EY, Jones PB, et al. Hormonal regulation of the differentiation of cultured ovarian granulosa cells. Endocr Rev 1984;5:76–127.
Weil SJ, Vendola K, Zhou J, et al. Androgen receptor gene expression in the primate ovary: cellular localization, regulation, and functional correlations. J Clin Endocrinol Metab 1998;83:2479–2485.
Chappel SC, Resko JA, Norman RL, et al. Studies on rhesus monkeys on the site where estrogen inhibits gonadotropins: delivery of 17 β-estradiol to the hypothalamus and pituitary gland. J Clin Endocrinol Metab 1981;52:1–8.
Chabab A, Hedon B, Arnal F, et al. Follicular steroids in relation to oocyte development in human ovarian stimulation protocols. Hum Reprod 1986;1:449–454.
84. Young SR, Jaffe RB. Strength-duration characteristics of estrogen effects on gonadotropin response to gonadotropin-releasing hormone in women: II. Effects of varying concentrations of estradiol. J Clin Endocrinol Metab 1976;42:432–442.
Pauerstein CJ, Eddy CA, Croxatto HD, et al. Temporal relationship of estrogen, progesterone, luteinizing hormone levels to ovulation in women and infra-human primates. Am J Obstet Gynecol 1978;130:876–886.
86. World Health Organization Task Force Investigators. Temporal relationship between ovulation and defined changes in the concentration of plasma estradiol-17β luteinizing hormone, follicle stimulating hormone and progesterone. Am J Obstet Gynecol 1980;138:383.
87. Hoff JD, Quigley NE, Yen SSC. Hormonal dynamics in mid-cycle: a re-evaluation. J Clin Endocrinol Metab 1983;57:792–796.
Demura R, Suzuki T, Tajima S, et al. Human plasma free activin and inhibin levels during the menstrual cycle. J Clin Endocrinol Metab 1993;76:1080–1082.
Groome NP, Illingworth PG, O'Brien M, et al. Measurement of dimeric inhibin B throughout the human menstrual cycle. J Clin Endocrinol Metab 1996;81:1401–1405.
McLachlan RI, Robertson DM, Healy DL, et al. Circulating immunoreactive inhibin levels during the normal human menstrual cycle. J Clin Endocrinol Metab 1987;65:954–961.
91. Buckler HM, Healy DL, Burger HG. Purified FSH stimulates inhibin production from the human ovary. J Endocrinol 1989;122:279–285.
Ling N, Ying S, Ueno N, et al. Pituitary FSH is released by heterodimer of the β-subunits from the two forms of inhibin. Nature 1986;321:779–782.
93. Braden TD, Conn PM. Activin-A stimulates the synthesis of gonadotropin-releasing hormone receptors. Endocrinology 1992;130:2101–2105.
94. Adashi EY. Putative intraovarian regulators. Semin Reprod Endocrinol 1988;7:1–100.
Yoshimura Y, Santulli R, Atlas SJ, et al. The effects of proteolytic enzymes on in vitro ovulation in the rabbit. Am J Obstet Gynecol 1987;157:468–475.
96. Yoshimura Y, Wallach EE. Studies on the mechanism(s) of mammalian ovulation. Fertil Steril 1987;47:22–34.
Anasti JN, Kalantaridou SN, Kimzey LM, et al. Human follicle fluid vascular endothelial growth factor concentrations are correlated with luteinization in spontaneously developing follicles. Hum Reprod 1998;13:1144–1147.
98. Lenton EA, Landgren B, Sexton L. Normal variation in the length of the luteal phase of the menstrual cycle: identification of the short luteal phase. Br J Obstet Gynaecol 1994;91:685.
Scott R, Navot D, Hung-Ching L, et al. A human in vivo model for the luteal placental shift. Fertil Steril 1991;56:481–484.