Which of the following best explains how treatment with a drug that stimulates the production of insulin receptors on target cells will affect the insulin signaling pathway?

V Physiology of insulin secretion in vivo

Insulin secretionin vivo has also been extensively studied. As predictable from the studies of single beta cells described above, the most important regulators of insulin secretion are circulating nutrients, in particular, glucose. In the fasting state, insulin secretion is maintained at levels that provide sufficient insulin to constrain hepatic glucose release at rates that match glucose utilization (∼2 mg/kg/min) and so the plasma glucose concentration is maintained at normal levels of ∼90 mg/dl (∼5 mmol/liter). After meal ingestion, glucose concentrations in the circulation rise and stimulate insulin secretion (Fig. 7). Increased delivery of insulin into the circulation causes further suppression of hepatic glucose release (to ∼0.5 mg/kg/min) and increased stimulation of glucose uptake by insulin-sensitive tissues such as muscle to restore normoglycemia. Therefore, the simplest model to describe insulin secretion in vivo would have two components: a constant basal rate of insulin secretion superimposed on which are meal-related increments. Although this model (concept of basal-bolus insulin therapy) is commonly employed by physicians attempting to replace insulin in patients who secrete insufficient insulin, it is an oversimplification of a very complex dynamic neuroendocrine secretory system.

Even basal insulin secretion in the fasting state can be resolved into several complex secretory patterns. First, there is a circadian rhythm of insulin release with decreased insulin secretion during the night. Second, insulin secretion can also be resolved into an ultradian rhythm with an ∼40 min oscillatory period that might reflect the feedback loop between insulin secretion and insulin action through the intermediary of the prevailing plasma glucose concentration. Third, insulin secretion can also be resolved into high-frequency discrete insulin secretory bursts, so-called pulsatile insulin secretion (Fig. 8). These pulses occur approximately once every 4 min and yield an insulin concentration profile in the portal vein that shows dramatic peaks and troughs of as much as 5000 pmol/liter in amplitude. Virtually all insulin is secreted in these discrete 4 min insulin secretory bursts, indicating that regulation of insulin secretion is accomplished through changes in either insulin pulse frequency or pulse mass. In fact, almost all regulation of insulin secretion is accomplished by the modulation of pulse mass (vide infra). To understand the mechanisms that result in pulsatile insulin secretion, it is necessary to contemplate both a pacemaker and a pulse coordinator (between islets).

Individual or perfused islets also secrete insulin in ∼4 min discrete insulin pulses, indicating that the pacemaker responsible for generation of pulsatile insulin release is present in each islet. Two hypotheses are currently being investigated: one suggests that the pacemaker might arise at the level of the membrane due to intermittent depolarization of the membrane; the other hypothesis is that oscillations in glycolysis drive an oscillatory production of ATP. Perhaps both hypotheses are correct and an intermittent membrane depolarization temporarily decreases ATP concentrations (ATP is consumed, restoring the resting membrane potential). The dip in ATP would be expected to elicit a surge in glycolysis and entrain the natural glycolytic oscillations generated at the level of the reversible interconversion of fructose-1-phosphate and fructose-1,6-diphosphate. Even if the individual islets are potentially independent pacemakers, the question remains how approximately one million islets, scattered throughout the pancreas, are synchronized to secrete insulin in coordinate insulin secretory bursts? The best available evidence suggests that this is mediated through the intrapancreatic neural network. The denervated, isolated-perfused pancreas retains pulsatile insulin secretion, indicating that intrinsic innervation is not required for this process. Islets that are transplanted into the portal vein secrete insulin in a noncoordinated manner until they become reinnervated and coordinate pulsatile insulin secretion is reestablished. Regulation of insulin secretion is accomplished by modulation of the magnitude of insulin secretory bursts with amplification following meal ingestion, glucose infusion, or GLP-1 infusion and inhibition with somatostatin, a hormone that inhibits insulin secretion.

In numerous studies of insulin secretion in vivo, the technique of an acute bolus injection of glucose, arginine, or glucagon (all insulin secretagogues) has been used. Following this approach, insulin is secreted in a biphasic manner with a first phase of insulin secretion over ∼15 min generally ascribed to release of insulin from the readily releasable insulin vesicles already docked to the beta-cell membrane. The first-phase insulin release is decreased in the setting of a partial loss of beta-cell mass, as is the magnitude of insulin pulses in response to an increment in glucose. Taken together, these data indicate that the insulin secretory bursts and first-phase insulin release are derived from a physiologically related pool of insulin vesicles.

The second phase of insulin secretion is separated from the first phase by a nadir. It is characterized by a lower amplitude and longer duration and lasts as long as the secretory stimulus is active on the beta cells. The underlying mechanism that leads to the more sustained nature of the second phase is under active investigation. There is evidence from studies at the single-cell level to suggest that vesicle maturation and mobilization from a reserve pool into the membrane-bound readily releasable pool may be the rate-limiting steps determining Vmax of the second-phase secretion process. The characteristic physiology of insulin secretion with a biphasic response to a step increase in the ambient glucose concentration and a typical pulsatile pattern is preserved in isolated human islets, which are independent of innervation and vascularization. In vivo, insulin secretion follows a circadian rhythm (∼24 h period), an ultradian rhythm (∼40 min period), and a high-frequency rhythm (∼4 min period). Insulin secretion is regulated by modulation of the mass of the high-frequency pulses and pulse frequency remains remarkably stable. Correlative studies at the level of the single beta cell should help establish the basis of regulation of the insulin secretion with each pulse.

Insulin Secretion

Regulated insulin secretion from pancreatic beta cells is critical to health. Both insufficient insulin secretion (resulting in diabetes mellitus) and excess insulin secretion (leading to hypoglycemia) are life threatening. The complexity of regulated insulin secretion in health becomes apparent with the difficulty of reproducing it in patients with insulin deficiency. Appropriate regulated insulin secretion depends on several components. First, development and maintenance of an appropriate number of functional insulin-secreting beta cells is necessary, often collectively referred to as the beta cell mass.1 Second, beta cells need to sense the key regulators of insulin secretion, most importantly the prevailing blood glucose concentration.2 Third, proinsulin synthesis and processing (see Chapter 31) must proceed at a rate to provide sufficient insulin for secretion, the insulin being targeted to insulin vesicles that are available for secretion (secretion competent).3 As the majority of insulin secretory granules are not secretion competent (presumably because of aging or other factors),4-6 the focus for regulation of insulin secretion is the pool of insulin secretory vesicles that are primed, docked, and available for secretion.5 Finally, minute-by-minute changes in insulin release from these primed and docked vesicles need to be tightly linked to the regulating signals that impact the beta cell. Predominant among these is the circulating glucose concentration.7 In addition, other circulating fuels (free fatty acids, amino acids),8-11 other circulating hormones including glucagon like peptide-1 (GLP-1),12-14 glucose-dependent insulinotropic polypeptide (GIP),15 epinephrine,16 innervation by adrenergic and cholinergic fibers,17-19 and paracrine effects including islet amyloid polypeptide (IAPP), somatostatin, and insulin itself, 20-23 are all regulators of insulin secretion.

Our understanding of these complex processes that underlie successful regulated insulin secretion is hampered by the complexity of the anatomy of the endocrine organ that subserves regulated insulin secretion (Fig. 32-1). The islet of Langerhans was named after Paul Langerhans (1847 to 1888), a German pathologist (Fig. 32-2) who first described the appearance of these islets scattered in the pancreas.24

Insulin secretion

Insulin secretion is governed by the interaction of nutrients, hormones, and the autonomic nervous system. Glucose, as well as certain other sugars metabolized by islets, stimulates insulin release. Basal and peak insulin levels are closely related to the glucose concentration, and prolonged fasting will further reduce glucose and insulin levels—which, however, remain in the measurable range at 2 to 5 mU/mL. There is evidence that a product or products of glucose metabolism may be involved in maintaining insulin secretion and that sugars not metabolized by islet cells do not promote insulin release.100-102

The initial steps of glucose-stimulated insulin release are depicted in Figure 19-7 and discussed in detail in Chapter 6 in connection with mutations in the sulfonylurea receptor (SUR)-Kir6 (inward-rectifying potassium channel) complex of the adenosine triphosphate-regulated potassium channel KATP, along with the subsequent steps that may cause activation of glucose or amino acid-stimulated insulin secretion.103 This schema involves glucose transport into the beta cell through the GLUT2 glucose transporter and phosphorylation of glucose by means of glucokinase. Defects in the former are associated with type 2 diabetes, whereas heterozygous mutations in the latter are associated with MODY. Homozygous mutations in glucokinase result in permanent neonatal diabetes mellitus (as described in detail in Chapter 9). Glucokinase defects are generally associated with normal insulin release at higher glucose concentrations, and therefore with a milder type of diabetes.12,13After intravenous glucose infusion in normal persons, insulin secretion is biphasic—with an initial spike followed by a sustained plateau. It is proposed that the initial spike represents preformed insulin, whereas the sustained plateau represents newly synthesized insulin.

Cyclic adenosine monophosphate (AMP) is involved in stimulating insulin release. Therefore, agents that inhibit phosphodiesterase and reduce cyclic AMP destruction (such as theophylline) augment insulin release. Translocation of calcium ions into the cytoplasm from the exterior, as well as from the intracellular organelles (see Figure 19-7) plays a key role in the contractile forces that propel insulin to the cell surface.103 There, the membrane of the insulin vesicle fuses with the cell membrane—allowing extrusion of insulin granules into the surrounding vascular space, a process known as emiocytosis. Other ions, including potassium and magnesium, are involved in the insulin secretion.103-106 The sulfonylurea receptor is closely linked to potassium channels in the beta cell.103-106 Amino acids also stimulate insulin release, although the potency of individual amino acids varies.107 A group of amino acids is more potent than any single one, and the insulin-secretory response is potentiated in the presence of glucose.107 Free fatty acids and ketone bodies may also stimulate insulin release.107 Insulin responses to oral glucose administration are always greater than responses to intravenous administration of glucose that result in the same blood glucose profile a finding that led to the concept that gut factors (incretins) modulate and increment insulin secretion.108 Although a variety of gut hormones participate in promoting insulin release,108 gastrointestinal polypeptide (GIP) pancreatic glucagon and the glucagon-like peptides (GLP) play a major role in stimulating insulin release.108 These properties have found application as agents, collectively named incretins, in augmenting insulin secretion in persons with T2DM and in some persons with T1DM. Somatotropin release-inhibiting factor (somatostatin), produced in the delta cells of islets, inhibits insulin and glucagon release and reduces splanchnic blood flow. These properties have found application to reduce insulin secretion in neonates with hyperinsulinemic hypoglycemia of infancy (see Chapter 6). Together, these factors may finely regulate nutrient intake and its disposition and form an enteroinsular axis for metabolic homeostasis.108 In addition to these gut hormones, several other hormones modulate insulin secretion. Growth hormone is involved in insulin synthesis and storage. Persons with congenital growth hormone deficiency have subnormal basal and stimulated insulin responses, whereas in acromegaly basal and stimulated insulin levels are increased. Human chorionic somatomammotropin (also known as human placental lactogen), structurally related to growth hormone, likewise affects insulin release. The stimulatory effect of each hormone on insulin secretion is antagonized by the anti-insulin effect at the peripheral level, however. Similarly, glucocorticoids and estrogens evoke greater insulin secretion while inducing peripheral insulin resistance—in part by decreasing insulin receptors on target cells.

Insulin secretion is constantly modulated by the autonomic nervous system.102,109 The parasympathetic arm, through the vagus, directly stimulates insulin release. Modulation of insulin secretion by the sympathetic arm depends on whether α- or β-adrenergic receptors are activated. Activation of β2 receptors by agents such as isoproterenol stimulates insulin secretion by a process that involves cyclic AMP generation. Blockade of β-adrenergic receptors by propranolol blunts basal and stimulated insulin release. Conversely, activation of α-adrenergic receptors blunts insulin secretion, and blockade of these receptors by agents such as phentolamine augments basal and glucose-stimulated insulin release. Epinephrine and norepinephrine stimulate predominantly α-adrenergic receptors in islets, resulting in impaired insulin secretion—as observed during stress or in patients with pheochromocytoma.102

In summary, in normal humans insulin secretion is constantly modulated by the quantity, quality, and frequency of nutrient intake; by the hormonal milieu; and by autonomic impulses. The ingestion of nutrients, principally carbohydrate and protein, produces intestinal hormonal signals that prime and initiate insulin release. The entry of glucose into the beta cell, the phosphorylation of glucose, and the generation of adenosine triphosphate (ATP) by this or other nutrients result in insulin release. This sequence involves cyclic AMP, β-adrenergic receptors, and ions—principally calcium and potassium. Glucose metabolism within the beta cell provides energy for further synthesis and release of insulin.103

Why the Liver?

Insulin secretion from beta cells within native islets is relatively complex. Although it is clear that insulin is secreted in response to increases in blood glucose levels, other secretagogues (e.g., arginine) also stimulate insulin secretion. The arrangement of alpha, beta, and delta cells within islets is not random [88]. Somatostatin secreted from delta cells has paracrine actions modulating insulin secretion [50]. The autonomic nervous system represents a further influence on insulin secretion and may influence pancreas regeneration [62]. Variations in pulse frequency and amplitude are seen to occur as insulin secretion changes in different physiologic states [61].

In the native pancreas, islets receive their blood supply from the systemic circulation while insulin secreted by islets drains into the portal vein and thence to the liver. The liver extracts much of the insulin during the first pass. Consequently, in the postprandial state the concentration of insulin in the systemic circulation is significantly lower than in the portal vein. Circulating insulin levels are significantly lower in whole-pancreas transplant recipients where the venous drainage of the graft is into the portal rather than the systemic circulation [95]. First-pass metabolism in the liver affects other substances relevant to intraportal islet transplantation, including drugs and nutrients.

Intraportal infusion of islets may permit delivery of pancreatic hormones, particularly insulin, directly to the liver. The ability to deliver insulin and glucagon directly to the liver is attractive from a physiological perspective. The liver is a key site of insulin action. Insulin promotes glycogen synthesis and turns off both glycogenolysis and gluconeogenesis. Glucagon has the opposite effects to insulin on glycogenolysis and gluconeogenesis, raising blood glucose levels. Glucagon plays an important role in preventing hypoglycemia. (In addition to the loss of insulin secretion, normal glucagon secretion is lost in type 1 diabetes and contributes to the susceptibility to hypoglycemia seen in some patients [46].

The secretion of insulin from intrahepatic islets may not be exactly equivalent to portal delivery of insulin from the native pancreas. Normally, insulin secretion will result in uniform concentration of insulin being delivered throughout the liver. With intrahepatic islet transplants, there may be areas of the liver surrounding engrafted islets that have relative hyperinsulinemia. This localized hyperinsulinemia may explain the patchy focal steatosis observed in some patients after clinical islet transplantation [15].

The intrahepatic location of transplanted islets has a number of potential advantages and disadvantages. The concentration of orally administered immunosuppressant drugs in the portal circulation is significantly higher than in the systemic circulation because of hepatic first-pass metabolism [35]. Potentially this could be advantageous, in that islets and the liver could be exposed to effective concentrations of drugs using lower doses with reduced potential for systemic toxicity. In fact, monitoring venous trough levels assesses dosing of most immunosuppressive drugs. The current recommended levels are relatively high in comparison to solid organ transplants. Peak drug levels in the portal system are likely very high and potentially may be toxic to islets in the longer term [11].

Another potential disadvantage is the exposure of islets to high concentrations of nutrients in the portal vein, not yet subject to the first-pass metabolism in the liver. Potentially, this may be associated with supramaximal stimulation of islets in the postabsorptive state, which may have deleterious effects on long-term islet function.

Insulin secretory defect

Insulin secretion and β-cell mass increase to compensate for states of insulin resistance such as obesity, pregnancy, or cortisol excess, so that fasting and meal-stimulated insulin levels are elevated even as glucose levels remain normal. When insulin secretion fails to fully compensate for the degree of insulin resistance, glucose levels rise, first to those of mild glucose intolerance (glucose levels above normal but below criteria for a diagnosis of diabetes), and then to those of diabetes. Insulin levels early in this progression are still elevated compared to individuals with normal insulin sensitivity, but are not elevated sufficient to maintain normal glucose control. With time, in most patients, the insulin secretory defect continues to worsen, including loss of β-cells, ultimately requiring exogenous insulin treatment to maintain control of blood glucose levels.

The development of impaired insulin secretion is likely due to both genetic and environmental factors. Indeed, variants at more than 20 genetic loci have recently been identified as playing a role in type 2 diabetes risk; of these, most appear to affect insulin secretion, but detailed mechanisms of action remain to be elucidated. Gene variants may also affect an individual's response to nongenetic factors. One factor contributing to the loss of β-cell function may be a persistent stimulus to oversecrete insulin caused by insulin resistance (Figure 1). This may be due in part to the effect of excessive secretion of islet amyloid polypeptide (IAPP), which is cosecreted with insulin; in the islets in the majority of patients with type 2 diabetes there is the accumulation of amyloid, containing IAPP. Another factor is the elevated free fatty acid level present in insulin resistance, which can contribute to the defective insulin secretion, just as it can contribute to insulin resistance. While short-term exposure to lipids increases insulin secretion, long-term exposure impairs β-cell function, and may be responsible for β-cell death by apoptosis. Similarly, prolonged exposure to increased glucose levels imposes a glucotoxicity on the β-cell; if glucose levels are normalized in a patient with diabetes, endogenous insulin secretion will improve. Over time, however, this glucotoxicity may lead to irreversible β-cell damage, perhaps through oxidative injury. Inflammatory cytokines such as IL-1β also appear to induce β-cell apoptosis, which may be important in proinflammatory states such as obesity. Finally, insulin resistance of the β-cell may itself lead to impaired β-cell function, as demonstrated in mice carrying genetic defects of insulin signaling in β-cells.

Recent investigations have implicated endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) in the pathogenesis of type 2 diabetes. The strongest evidence supports a role for the UPR in contributing to β-cell loss, although there are also data indicating that it can contribute to insulin resistance. The UPR is activated in cells when protein synthesis exceeds the capacity of the ER to properly fold them. The response includes a global decrease in protein translation, an increase in the degradation of misfolded proteins, and a specific increase in production of proteins that assist in the folding of proteins within the ER, such as chaperones. When these steps fail to prevent the accumulation of misfolded proteins, apoptosis is triggered. The rate of proinsulin production can approach 1 million molecules per minute per cell, making it clear why β-cells are at risk of activation of the UPR. A clear demonstration that the UPR can contribute to β-cell loss is the monogenic disease Wolcott–Rallison syndrome, a rare disorder that includes insulin-deficient diabetes with an onset in infancy. It is caused by a mutation in the EIF2AK3 gene, which encodes a protein (eukaryotic initiation factor 2α (eIF2α) kinase 3, equivalent to rodent protein kinase RNA-like endoplasmic reticulum kinase (PERK)) that is central to the UPR.

1 Regulation of Release

Insulin secretion by the β cells of the islets of Langerhans is primarily regulated by the d-glucose level in the extracellular fluid bathing the β cells. Glucagon increases and somatostatin decreases insulin release via paracrine actions. Insulin release is stimulated by GH, cortisol, PRL, and the gonadal steroids. It is decreased by PTH. The effects of thyroid hormones are more variable. Epinephrine inhibits insulin release. Sympathetic nerve stimulation inhibits insulin release. Cholinergic stimulation promotes insulin release. Pro-inflammatory cytokines diminish the effect of insulin and have been implicated in the phenomenon of insulin resistance.

Insulin

Insulin secretion increases rapidly with eating as a result of cephalic and intestinal phase reflexes (the latter arise from excitation of intestinal receptors by preabsorptive food stimuli) and of direct substrate actions on the pancreatic beta cells. Acute insulin administration has been demonstrated to decrease meal size under some test conditions in rats, but it has been difficult to establish a reliable dose–response relationship and no satiating effect has been found in humans. Nevertheless, prandial antagonism of endogenous insulin by infusion of specific antibodies increases meal size in rats, suggesting that insulin has at least a permissive effect in normal satiation.

Diabetic pathogenesis

Dietary carbohydrate is the major source of exogenous glucose in the body. Due to the importance of glucose to meet the energy requirements of the cells, mammals have developed an advance mechanism to maintain the glucose level within a certain threshold in the blood during the fast and fed states. This mechanism involves the hormonal modulation of glucose production by the liver (during the fast state) or glucose uptake and utilization by the muscle and the peripheral tissues (during the fed state).

Insulin is the main source of hormonal regulation of the energy metabolism. It is produced by the β-cells of the pancreas in response to elevated blood glucose. In vitro and in vivo studies have shown that the release of insulin is stimulated by the exposure of the pancreatic islet to elevated glucose concentrations.34 The primary effect of insulin is to facilitate the uptake of glucose by the glucose transporter (GLUT4), which is predominant in the skeletal muscle and adipose tissues.

An impairment in maintaining the blood glucose threshold leads to the multifactorial physiological state called hyperglycemia. Chronic exposure of β-cells to fatty acid and glucose beyond the physiological concentrations (hyperglycemia and hyperlipidemia) causes glucolipotoxicity, which leads to a cascade of events resulting in β-cell dysfunction and insulin resistance.35 Both β-cell dysfunction and insulin resistance are the underlying pathology of type 2 diabetes, which accounts for 90%–95% of diabetic cases.36

VA1 Secretion of Insulin

Insulin secretion by the pancreatic β cells provides an excellent example of a cellular activity that requires direction. Insulin is packaged in secretory vesicles which have to migrate to the plasma membrane and fuse with it to release the entrapped insulin. Both microscopic and biochemical studies have shown that secretory granules are linked to microtubules which direct attached vesicles to the cell surface. However, a cortical band of fine microfilaments is consistently observed in β cells. Alteration of this cell web by cytochalasin B is associated with an enhancement of glucose-induced secretion of insulin by isolated islets. This microfilamentous web plays an important role in the exocytosis of insulin secretory granules by controlling access to the cell membrane via a mechanism probably similar to that previously described for chromaffin cells. Ca2+ appears to initiate the cascade of events by which microtubules facilitate the displacement of granules toward the cell membrane. Glucose metabolism increases intracellular concentration of ATP, which closes the ATP-sensitive K+ channels, consequently inducing cell depolarization and Ca2+ influx, while cAMP modifies the intracellular distribution of Ca2+ by increasing the cytosolic pool at the expense of Ca2+ bound to intracellular organelles. Protein kinase C also appears to be involved in the secretion of insulin.

Insulin Secretion, Diabetes and Ketone Body Metabolism

Insulin secretion occurs through two mechanisms. The first is a process that involves closing of the cell-surface ATP-sensitive potassium channels in response to increases in the circulating glucose concentrations, which stimulates exocytosis of insulin-containing vesicles from the β-islet cells through an increase in the cytosolic calcium concentration. The second mechanism is dependent on pyruvate carboxylase, which is highly expressed in β-islet cells; it has been estimated that 35–45% of pyruvate enters the citric acid cycle through this anaplerotic pathway in β-islet cells. Inhibition of pyruvate carboxylase with phenylacetic acid decreases glucose-stimulated insulin release from β-islet cells. Furthermore, there is evidence that pyruvate carboxylase plays an important role in the early stages of type 2 diabetes. Specifically, in Zucker fatty rats with insulin resistance, the hyperfunctioning β-islet cells increase insulin production in part through increases in pyruvate carboxylase activity.

Citric acid cycle pool size increases in hearts of rats with experimentally induced diabetes, suggesting enrichment by anaplerotic pathways. We have suggested that an increase in anaplerotic flux, which primarily occurs through pyruvate carboxylation (via malic enzyme), plays an important role in maintaining flux through the second span of the citric acid cycle. Acutely, the metabolic derangement of ketoacidosis that occurs with diabetes inhibits flux through α-ketoglutarate dehydrogenase by sequestration of coenzyme A (CoASH). This phenomenon is associated with contractile dysfunction of the heart that can be readily reversed by the addition of glucose, lactate, or pyruvate (all of which are anaplerotic substrates). The effects of pyruvate are mediated by enrichment of malate in the citric acid cycle pool, which occurs by carboxylation of pyruvate to form malate and oxaloacetate through the actions of malic enzyme and pyruvate carboxylase, respectively (Figure 5). The citric acid cycle is thereby able to operate once again in a span that can generate reducing equivalents to support oxidative phosphorylation of adenosine diphosphate to form the ATP necessary to drive the contractile machinery of the heart.