The posterior pituitary is an extension of the hypothalamus and contains the axon terminals of magnocellular neurons located in the supraoptic and paraventricular nuclei (see Figures 2–2
). These neurons generate and propagate action potentials, producing membrane depolarization and exocytosis of the contents of their secretory granules. The neuropeptides produced by the magnocellular neurons, and consequently released from the posterior pituitary, are oxytocin and AVP. As the axons leave the supraoptic and paraventricular nuclei, they give rise to collaterals, some of which terminate in the median eminence.
Oxytocin and AVP are closely related peptides consisting of 9 amino acids (nonapeptides) with ring structures (Figure 2–4). They are synthesized as part of a larger precursor protein, consisting of a signal peptide, the hormone, a peptide called neurophysin 2, and a glycopeptide called copeptin. Following cleavage of the signal peptide in the endoplasmic reticulum, the remaining precursor folds, dimerizes, exits from the Golgi apparatus, and moves down the neurohypophyseal axons packaged within neurosecretory vesicles (see Figure 2–4).
Within the neurosecretory vesicles, as they migrate down the axons, the precursor protein undergoes post-translational processing, producing the peptides AVP, oxytocin, neurophysins (defined below), and copeptin all of which are stored in the vesicles. The release of the contents of the vesicles is triggered by the neuronal influx of calcium ions through voltage-gated calcium channels, which open as the wave of depolarization reaches the axon terminals. The increase in intracellular calcium triggers the movement and docking of the secretory vesicles on the plasma membrane, resulting in exocytosis of the vesicle contents into the extracellular space. These neuropeptides enter the systemic circulation through venous drainage of the posterior pituitary into the intercavernous sinus and internal jugular vein. In the systemic circulation, oxytocin and AVP circulate unbound. They are rapidly cleared from the circulation by the kidney and, to a lesser extent, by the liver and brain. Their half-life is short and is estimated to range between 1 and 5 minutes.
Neurophysins are by-products of post-translational prohormone processing in the secretory vesicles. The release of AVP and oxytocin is accompanied by the release of neurophysins from the secretory granules. Although the exact function of these by-products is not clear, it appears that neurophysins play an important role in AVP release. This role has become more evident since the identification of the inherited disease of familial neurogenic diabetes insipidus (DI). This disease is characterized by AVP deficiency caused by mutations in neurophysins and improper targeting of the hormone to neurosecretory granules. Neurophysins thus have an important role in the transport of AVP from the cell bodies of magnocellular neurons to their final release from the posterior pituitary. Impairment in hormone targeting leads to retention of the mutated neuropeptide precursor in the endoplasmic reticulum of the magnocellular neurons, and these cells progress to programmed cell death (apoptosis).
The neuropeptide oxytocin is synthesized by magnocellular neurons in the supraoptic and paraventricular nuclei of the hypothalamus and is released from the posterior pituitary into the peripheral circulation. The release of oxytocin is stimulated by sucking during breast-feeding (lactation) and stretch of the cervix during childbirth (parturition) (Figure 2–5).
Physiologic effects and regulation oxytocin release. The release of oxytocin is stimulated by the distention of the cervix toward the term of pregnancy as well as by the contraction of the uterus during parturition. The signals are transmitted to the paraventricular (PVN) and supraoptic (SON) nuclei of the hypothalamus where they provide a positive feedback regulation of oxytocin release. The increased number in oxytocin receptors, the increase number in gap junctions between smooth muscle cells, and the increased synthesis of prostaglandins enhance the responsiveness of the uterine muscle. The suckling of the nipple of the lactating breast also stimulates oxytocin release. The afferent sensory signals elicit an increase in oxytocin release into the circulation.
Physiologic Effects of Oxytocin
The 2 main target organs for oxytocin’s physiologic effects are the lactating breast and the uterus during pregnancy (see Figure 2–5). In the lactating breast, oxytocin stimulates milk ejection by producing contraction of the myoepithelial cells that line the alveoli and ducts in the mammary gland. In the pregnant uterus, oxytocin produces rhythmic smooth muscle contractions to help induce labor and to promote regression of the uterus following delivery (see Chapter 9). Oxytocin analogs may be used clinically during labor and delivery to promote uterine contractions and during the postpartum period to help decrease bleeding and return the uterus to its normal size (uterine involution) (Table 2–2).
Table 2–2. Key Aspects of Posterior Pituitary Hormones ||Download (.pdf)
Table 2–2. Key Aspects of Posterior Pituitary Hormones
|Receptor||Gq/11 protein-coupled||G protein-coupled (V1R, Gq/11, V2R, Gs)|
|Second messenger||Phospholipase C ↑ [Ca2+]i|
(V1R) Phospholipase C ↑ [Ca2+]i
(V2R) Adenylate cyclase cAMP
|Target organ or cells|
- Mammary myoepithelial cells
V2R; kidney collecting ducts
V1R; smooth muscle cells
- Uterine contraction
- Milk ejection
- Increased H2O permeability
The physiologic effects of oxytocin in the pregnant uterus are augmented by a dramatic increase in sensitivity to the hormone during the onset of labor. This increased sensitivity to oxytocin is caused by an increased density (upregulation) of oxytocin receptors in the uterine muscle. Receptor levels can be 200 times greater at the onset of labor than in the nonpregnant uterus. Because of this increase, levels of oxytocin that would normally not be effective can produce forceful uterine contractions toward the end of pregnancy. The increased density of oxytocin receptors is mediated by steroid hormone regulation of oxytocin-receptor synthesis. Responsiveness of the uterus is also enhanced by increased gap-junction formation between smooth muscle cells, facilitating conduction of action potentials between one cell and the next; and by increased synthesis of prostaglandin, a known stimulator of uterine contraction toward the end of gestation. All of these factors enhance myometrial contractile activity in response to oxytocin at term (see Chapter 9). Additional secondary effects that have been attributed to oxytocin are potentiation of the release of ACTH by CRH, interaction with the AVP receptor to produce vasoconstriction, stimulation of prolactin release, and an influence on maternal behavior and amnesia.
Control of Oxytocin Release
The principal stimulus for oxytocin release is mechanical stimulation of the uterine cervix by the fetus near the end of gestation. Oxytocin release is also stimulated by the forceful contractions of the uterus during the fetal expulsion reflex. Hence, the contractile activity of the uterus acts through positive feedback mechanisms during parturition to stimulate oxytocin neurons, and this further increases the secretion of oxytocin. Before the oxytocin neurons can secrete oxytocin, they must be released from inhibition by other neurons containing endogenous opioids, nitric oxide, and γ-aminobutyric acid. This modulation of oxytocin release is partly caused by the declining blood levels of progesterone and increasing levels of estrogen during late pregnancy. The neurotransmitters involved in stimulating oxytocin release are thought to be acetylcholine and dopamine.
Oxytocin release is also triggered by stimulation of tactile receptors in the nipples of the lactating breast by suckling (see Figure 2–5). Breast-feeding generates sensory impulses that are transmitted to the spinal cord and then to the oxytocin-producing neurons in the hypothalamus. The information transmitted by these sensory afferents produces intermittent synchronized burst-firing of oxytocin neurons, resulting in pulsatile release of oxytocin and increases in blood oxytocin concentrations.
In addition to its secretion from neurohypophysial terminals in the posterior pituitary, oxytocin is released within the hypothalamic supraoptic and paraventricular nuclei. The function of this intrahypothalamic release of oxytocin is to control the activity of oxytocin neurons in an autocrine fashion by a positive feedback mechanism, increasing the neurohypophysial release of oxytocin. The release of oxytocin is inhibited by severe pain, increased body temperature, and loud noise. Note how the environmental, hormonal, and neural mechanisms of hypothalamic hormone regulation are in play to regulate oxytocin release at the appropriate time of gestation and in response to the relevant stimuli. The role of oxytocin in males is not clear, although recent studies have suggested that it may participate in ejaculation.
The physiologic effects of oxytocin are achieved by binding to cell membrane Gq/11 protein–coupled oxytocin receptors expressed in the uterus, mammary glands, and brain (Figure 2–6). Binding of oxytocin to the receptor activates phospholipase C, producing an increase in inositol trisphosphate and 1,2-diacylglycerol, which in turn results in an increase in cytosolic calcium concentrations leading to smooth muscle contraction.
Receptor-mediated oxytocin effects. Oxytocin binds to cell membrane Gαq/11 protein–coupled receptors expressed in the uterus, mammary glands, and brain. Binding of oxytocin to the receptor activates phospholipase C (PLC), producing an increase in inositol trisphosphate (IP3) and 1,2-diacylglycerol, which in turn results in an increase in cytosolic calcium concentrations. Calcium binds to calmodulin (CaM) and the Ca2+-CaM complex activates myosin light chain kinase (MLCK), which in turn phosphorylates (activating) myosin regulatory chains. The phosphorylated myosin filament combines with the actin filament leading to smooth muscle contraction. ATPase, adenosine triphosphatase; MLC, myosin light chain.
Disorders of Oxytocin Production
Diseases resulting from oxytocin excess have not been described. Although oxytocin deficiency causes difficulty with nursing because of impaired milk ejection, it is not associated with altered fertility or delivery. Normal oxytocin levels have been detected in women with DI (AVP deficiency).
AVP, also known as antidiuretic hormone (ADH), is the other neuropeptide produced by magnocellular neurons of the hypothalamus and released from the posterior pituitary. The principal effect of AVP is to increase water reabsorption by enhancing permeability to water in the distal convoluted tubules and the medullary collecting ducts in the kidney. In addition, AVP increases vascular resistance. This function of AVP may be important during periods of severe lack of responsiveness to other vasoconstrictors, as may occur during severe blood loss (hemorrhagic shock) or systemic infection (sepsis). The circulating concentrations of AVP range from 1.5 to 6 ng/L.
Arginine Vasopressin Receptors
The cellular effects of AVP are mediated by binding to G protein–coupled membrane receptors. Three AVP receptors have been characterized thus far, which differ in terms of where they are expressed as well as in the specific G proteins to which they are coupled and, thus, in the second-messenger systems that they activate.
V1R (also known as V1a) is coupled to Gq/11 and is specific for AVP. It is found in the liver, smooth muscle, brain, and adrenal glands. It activates phospholipases C, D, and A2 and stimulates the hydrolysis of phosphatidylinositol, resulting in an increase in intracellular calcium concentrations. The vasopressor effects of AVP are mediated through the V1R.
V2R is coupled to Gs and is expressed in the kidney. Binding of AVP to the V2R receptor activates adenylate cyclase and increases cyclic 3′,5′-adenosine monophosphate (cAMP) formation and aquaporin 2 (AQP2) phosphorylation and insertion into the luminal membrane. The water reabsorptive effects of AVP are mediated through the V2R.
V3R (also known as V1b) is coupled to Gq/11 and is expressed in the majority of anterior pituitary corticotroph cells and in several tissues, including the kidney, thymus, heart, lung, spleen, uterus, and breast. Binding of AVP to the V3R receptor stimulates the activity of phospholipase C, resulting in an increase in intracellular calcium.
Physiologic Effects of Arginine Vasopressin
The main target site of ADH is the collecting duct in the kidney (Figure 2–7
). Water permeability in the collecting duct is relatively low compared with that in the proximal tubule and the thin descending limb of Henle’s loop. In the proximal tubule and the descending limb of Henle’s loop, the water channel protein aquaporin 1 (AQP1) is constitutively expressed both in the apical (luminal) and basolateral (interstitial) membranes of epithelial cells (Table 2–3
). The proximal tubule is responsible for reabsorbing approximately 90% of filtered water. Reabsorption of the remaining 10% of filtered water in the distal collecting ducts is under tight control by AVP. Although it may seem that only a small fraction of the total filtered water reabsorption is under AVP control, water permeability of the collecting duct can be dramatically increased (within a few minutes) through the production of cAMP following AVP binding to V2
receptors in the basolateral membrane of the principal cells (see Figure 2–7
Cellular mechanism of arginine vasopressin (AVP) water conservation. The principal function of AVP is to increase water reabsorption and to conserve water. AVP binds to the V2 G protein–coupled receptor (V2R) in the principal cells of the distal tubule. This triggers the activation of adenylate cyclase and the formation of cyclic 3′,5′-adenosine monophosphate (cAMP), leading to activation of protein kinase A (PKA). PKA phosphorylates the water channel aquaporin 2 (AQP2), leading to the insertion of AQP2 into the luminal cell membrane. The insertion of water channels into the membrane increases the permeability to water. Water reabsorbed through these water channels leaves the cell through aquaporin 3 (AQP3) and aquaporin 4 (AQP4), which are constitutively expressed in the basolateral membrane of the principal cells. GPCR, G protein–coupled receptor.
Table 2–3. Key Features of Aquaporins ||Download (.pdf)
Table 2–3. Key Features of Aquaporins
|AQP1||Constitutively expressed in apical and basolateral membranes of epithelial cells of proximal tubules and descending limb of Henle’s loop. Involved in 90% of water reabsorption.|
|AQP2||Exclusively expressed in the collecting ducts. The only aquaporin directly regulated by ADH. Binding to the V2 AVP receptor stimulates insertion into the luminal membrane.|
|AQP3, AQP4||Constitutively expressed in the basolateral membranes of epithelial cells in the collecting ducts. Enhance water reabsorption following AQP2 insertion into the luminal membrane.|
The importance of AVP is better understood in terms of the total amount of urine that would be excreted in its absence. For example, in a healthy individual an average of 180 L of glomerular filtrate is formed per day. Thus, without AVP-mediated reabsorption of 10% of filtered water in the distal collecting ducts, urine output would be close to 18 L/d. This is 10-fold higher than the volume of urine output (1.5–2 L/d) under normal conditions.
The increase in cAMP that is stimulated by AVP binding to the receptor located in the basolateral membrane activates protein kinase A and subsequently the phosphorylation of AQP2, another protein. Phosphorylation of AQP2 is essential for its movement from cytoplasmic pools and its insertion in the luminal (apical) epithelial cell membrane of the collecting duct cells. The result is an increase in the number of functional water channels in the luminal membrane, making it more permeable to water. Thus, AVP-mediated insertion of AQP2 into the luminal membrane results in water conservation and urine concentration. This event is a short-term regulation of water permeability in response to an increase in circulating levels of AVP. In addition, AVP is thought to regulate water permeability over hours to days as a result of an increase in the total cellular amount of AQP2 caused by increased protein synthesis.
AQP2, one of several members of the aquaporin family, is exclusively expressed in the collecting ducts of the kidney. It is the only aquaporin that is directly regulated by ADH via the V2 AVP receptor. Water that diffuses into the cells through AQP2 exits through the basolateral side through aquaporin 3 (AQP3) and 4 (AQP4) eventually entering the vasculature. AQP3 and AQP4 are constitutively expressed in the basolateral membranes of the collecting ducts.
Water reabsorption through this mechanism is driven by the hydroosmotic gradient generated by a countercurrent mechanism in the renal medulla. The result is an increase in the concentration and reduction of urine volume, which minimizes urinary water loss. This antidiuretic mechanism may increase urine osmolarity to approximately 1200 mmol/L and reduce urine flow to approximately 0.5 mL/min. Without AVP-mediated effects, the principal cells of the collecting duct are impermeable to water, resulting in large volumes of diluted urine entering the collecting tubules from the ascending limb of the loop of Henle. As mentioned above, this could translate into excessive urine output and reduced urine osmolarity.
AVP also binds the V1 receptor, expressed in vascular smooth muscle, producing contraction and increasing peripheral vascular resistance (Figure 2–8). The hormone is known as vasopressin because of these vasoconstrictor effects. In particular, renal medullary blood flow has been demonstrated to be under AVP regulation. AVP circulates unbound and is distributed in a volume approximately equal to that of the extracellular space. Because of its relatively low molecular weight, AVP permeates peripheral and glomerular capillaries readily, so the urinary excretion rate of AVP is extraordinarily high.
Cellular mechanism of AVP vasoconstrictor effects. AVP also known as vasopressin binds the V1 receptor, expressed in vascular smooth muscle, producing contraction and increasing peripheral vascular resistance. DG, diacylglycerol; IP3, inositol trisphosphate; MLC, myosin light chain; PCR, protein-coupled receptor; PKC, protein kinase C.
Control of Arginine Vasopressin Release
AVP is released into the circulation following either an increase in plasma osmolarity or a decrease in blood volume (Figure 2–9
). Under physiologic conditions, the most important stimulus for AVP release is the “effective” plasma osmolarity. The changes in osmotic pressure are detected by special osmoreceptor neurons located in the hypothalamus and in 3 structures associated with the lamina terminalis: the subfornical organ, the median preoptic nucleus, and the organum vasculosum lamina terminalis. Dehydration produces loss of intracellular water from the osmoreceptors, resulting in cell shrinkage, which signals the AVP magnocellular neurons to stimulate AVP release. Release of AVP occurs even before the sensation of thirst. Therefore, on a hot day, an increase in AVP has already been initiated, and AVP is at work conserving fluid even before one feels the desire to drink water. In contrast, hypotonic stimuli, such as excess fluid intake or intravenous fluid administration in a hospital setting, result in cell swelling and stretching and hyperpolarization of the magnocellular neurons, which result in decreased depolarization and firing and consequently decreased release of AVP. The sensitivity of this system is quite high. That is, very small changes in plasma osmolarity (as little as 1% change) above the osmotic threshold of 280–284 mOsm/L produce significant increases in AVP release.
Integration of signals that trigger arginine vasopressin (AVP) release. Release of AVP is stimulated by an increase in plasma osmolarity and by a decrease in blood volume. Small changes in plasma osmolarity above a threshold of 280–284 mOsm/L produce an increase in antidiuretic hormone (ADH) release before the stimulation of thirst. A decrease in blood volume sensitizes the system and increases the responsiveness to small changes in plasma osmolarity. Blood loss and a decrease in mean arterial blood pressure (MABP) greater than 10% signal the hypothalamus to increase the release of AVP. The afferent signals are transmitted by the 9th and 10th cranial nerves. These signals increase sympathetic tone, therefore decreasing magnocellular neuron inhibition and stimulating AVP release.
AVP secretion is also stimulated by a decrease in blood pressure of greater than 10% (see Figure 2–9). Decreases in blood volume or blood pressure are detected by pressure-sensitive receptors in the cardiac atria, aorta, and carotid sinus. Factors that reduce cardiac output, such as a decrease in blood volume, orthostatic hypotension, and positive-pressure breathing, are all potent stimuli for AVP release. A decrease in blood pressure decreases the stretch of the baroreceptors and decreases their firing rate. These signals are transmitted to the central nervous system by neurons of the vagus and glossopharyngeal nerves. The reduced stimulation produces a decrease in tonic inhibition of AVP release, leading to an increase in AVP release from the magnocellular neurosecretory neurons. In addition to signaling the brain to stimulate the release of AVP, the decrease in blood pressure is also perceived by the macula densa in the kidney. This results in stimulation of renin release from the juxtaglomerular apparatus in the kidney. Renin catalyzes the conversion of angiotensinogen produced in the liver to angiotensin I, which is then converted to angiotensin II, by angiotensin-converting enzyme. The resulting rise in circulating levels of angiotensin II sensitizes the osmoreceptors, leading to enhanced AVP release. This is another example of hormonal regulation of hypothalamic neuropeptide release.
The volume-induced sensitization of AVP release results in a more accentuated AVP response to changes in plasma osmolarity. However, AVP secretion is far more sensitive to small changes in plasma osmolarity than to changes in blood volume. AVP is barely detectable below a certain plasma osmolarity (287 mOsm/L) threshold. Above this threshold, the plasma AVP concentration increases steeply in direct proportion to plasma osmolarity. An increase in osmolarity of only 1% will change AVP by an average of 1 pg/mL, an amount sufficient to significantly alter urine concentration and output. Circulating levels of AVP may reach 15–20 pg/mL under a strong osmotic stress. Because of this extraordinary sensitivity, the osmoreceptor plays the primary role in mediating the antidiuretic response to changes in water balance. In contrast, the response to pressure-volume changes is exponential. Reductions in plasma volume by 5%–10% usually have little effect on AVP levels, whereas decreases of 20%–30% result in intense hormone secretion, bringing AVP concentrations to a level many times higher than that required to produce maximum antidiuresis (up to 50–100 pg/mL). In other words, small changes in plasma osmolarity are more effective than small changes in blood pressure and volume in stimulating AVP release. Release of AVP can be modulated by estrogen and progesterone, opiates, nicotine, alcoholic beverages, and atrial natriuretic factor.
Disorders of Arginine Vasopressin Production
Either deficiency or excess of AVP can result in clinical disease. Deficiency of ADH release results in DI, a clinical syndrome resulting from the inability to form concentrated urine. Excess AVP release is known as syndrome of inappropriate antidiuretic hormone secretion
(SIADH). The concentrations of AVP may be altered in various chronic pathophysiologic conditions, including congestive heart failure, liver cirrhosis, and nephritic syndrome.
DI is characterized by the excretion of abnormally large volumes (30 mL/kg of body weight per day for adult subjects) of dilute (<250 mmol/kg) urine and excessive thirst. Three basic defects have been identified in the etiology. Only the first 2 pertain to alterations related to components of the AVP system itself. The third, excess water intake does not involve alterations in ADH release.
Decreased Arginine Vasopressin Release
Neurogenic (central or hypothalamic) DI is the most common defect and is caused by a decrease in AVP release resulting from diseases affecting the hypothalamic-neurohypophysial axis. Three causes can be identified: traumatic, inflammatory or infectious, and cancer related.
Decreased Renal Responsiveness to Arginine Vasopressin
Renal (nephrogenic) DI results from renal insensitivity to the antidiuretic effect of AVP. AVP production and release are not affected, but responsiveness at the distal tubule is impaired. Nephrogenic DI can be inherited or acquired and is characterized by an inability to concentrate urine despite normal or elevated plasma concentrations of AVP. Approximately 90% of cases are males with the X-linked recessive form of the disease who have mutations in the AVP type-2 receptor gene. A small number of cases of inherited nephrogenic DI are caused by mutations in the AQP2 water channel gene. Acquired nephrogenic DI can result from lithium treatment, hypokalemia, and postobstructive polyuria.
The differential diagnosis of DI relies on an understanding of the physiologic regulation of AVP release and its effects on the kidney. Plasma levels of AVP are interpreted together with the indirect assessment of antidiuretic activity triggered by a dehydration test. This test determines the ability of the body to increase the production and release of AVP during water deprivation. Fluids are withheld from the individual, and the rise in urine osmolarity, which indicates the response of the body to conserve fluid, is measured. Normal function consists of an increase in urine osmolarity and a decrease in urine output during water deprivation alone.
Another way of testing the system is by a challenge test with synthetic AVP. Individuals with normal pituitary function do not exhibit a further increase in urine osmolarity following the administration of a synthetic AVP analog (desmopressin). Individuals with central DI have a greater than 9% increase in urine osmolarity following desmopressin administration, indicating that the body is not capable of producing maximal release of AVP and consequently does a better job when a synthetic analog of AVP is administered. Another approach to diagnosing central DI is also based on the physiologic regulation of AVP release and uses the response to osmotic stimulation produced by the intravenous infusion of hypertonic (5%) saline solution. These examples illustrate that understanding the etiology of the disease requires an understanding of the normal physiologic regulation of the endocrine system in question.
Syndrome of Inappropriate Antidiuretic Hormone Secretion
An increase or excess in the release of ADH, in the absence of a physiologic stimuli for its release (thus the name inappropriate) is known as SIADH. This may be the result of brain injury or tumor production of AVP. The tumor can be located in the brain, but malignancies of other organs, such as the lung, have also been shown to produce high levels of AVP. The excess production of AVP results in the production of very small volumes of concentrated urine and dilutional hyponatremia. Management of this condition entails fluid restriction and, in some cases, the use of saline solutions to restore adequate plasma sodium levels.