CHAPTER 18: Neurodegeneration

• Neurodegenerative diseases, of which Alzheimer disease and Parkinson disease are the most common, are a large and diverse group of neurologic disorders characterized by death of neurons.

• The major processes by which neurons die are necrosis and apoptosis (programmed cell death); both processes involve cascades of proteins including proteolytic enzymes of the caspase family. These mechanisms, once thought entirely distinct, have significant overlap.

• Processes that can initiate cell death include mitochondrial dysfunction, excitotoxicity, oxidative stress, and abnormal protein aggregation.

• Abnormal mitochondrial function plays a key role in apoptosis and other forms of neurodegeneration.

• Excitotoxicity involves excessive activation of neurons via stimulation of glutamate receptors or other mechanisms of depolarization, which trigger biochemical cascades—many of them secondary to excessive Ca²⁺ flux into neurons—that lead to cell death.

• The pathologic hallmarks of Alzheimer disease, the most common cause of severe memory impairment in the elderly, are senile plaques, neurofibrillary tangles, dystrophic neurites, and neuronal loss.

• The development of Alzheimer disease may be due to the improper biochemical processing of amyloid precursor protein (APP) and the subsequent accumulation of β amyloid. A role for the microtubule-associated protein tau has also been implicated.

• Inherited forms of Alzheimer disease have been linked to mutations in APP or in proteins called presenilins, which regulate the processing of APP and several other proteins.

• Currently there is no effective treatment for Alzheimer disease, although several pharmacologic strategies for preventing the buildup of β amyloid, promoting its clearance, or counteracting its deleterious effects appear promising.

• Frontotemporal dementias comprise several other neurodegenerative disorders, roughly half of which involve abnormal aggregation of tau.

• Loss of dopamine input to the basal ganglia causes the symptoms of Parkinson disease; restoration of dopamine function, for example, with the dopamine precursor L-dopa, remains the mainstay of therapy for the disease.

• Rare familial forms of Parkinson disease are caused by mutation in several genes related to protein aggregation, including α-synuclein and parkin.

• Huntington disease is caused by a mutation in the gene for huntingtin; the mutation increases the number of glutamine residues expressed in the protein, which is pathogenic.

• Amyotrophic lateral sclerosis (ALS) is caused by the selective death of upper motor neurons in the cerebral cortex and/or lower motor neurons in the spinal cord.

In general terms, cell death occurs by either of two distinct processes, termed necrosis and apoptosis, which are characterized by very different histologic and biochemical signatures. In necrosis, the stimulus that triggers death is itself often the direct cause of the demise of the cell. In apoptosis, by contrast, the death stimulus activates a cascade of events that orchestrate the destruction of the cell. For this reason, apoptosis is referred to as programmed cell death. Unlike necrosis, which is a pathologic process, apoptosis is part of normal development; however, it also occurs in a variety of diseases.

Because the vast majority of neurons in an adult are postmitotic, the brain has limited capacity to compensate for cells lost during pathologic processes, such as neurodegeneration, stroke, and traumatic injury. As a result, enormous attention has been focused on understanding mechanisms of neuronal cell death in the hope that novel therapeutics can be developed to minimize or prevent cell loss. These efforts have focused on necrotic and apoptotic processes, although recent findings have revealed partly overlapping molecular pathways underlying the two. Identification of multiple forms of nonapoptotic programmed cell death has further blurred the distinction between necrosis and apoptosis. In addition, two forms of cell death appear to be unique to neurons: excitotoxicity and death induced by protein aggregation. This chapter begins with a basic overview of the molecular and cellular mechanisms of these distinct processes of neurodegeneration. The second portion of the chapter focuses on several severe brain disorders, including Alzheimer disease, frontotemporal dementia (FTD), Parkinson disease, Huntington disease, and amyotrophic lateral sclerosis (ALS), each of which is characterized by a unique pattern and mechanism of neurodegeneration. Although disease-altering treatments for these disorders remain lacking, we will discuss new approaches to intervene therapeutically based on our evolving knowledge of the molecular mechanisms of disease pathogenesis.

Necrotic cell death in the central nervous system (CNS) occurs in response to several types of environmental insults, often termed “cellular stress.” Examples of cellular stresses include ischemia (lack of oxygen), UV irradiation, virus infection, free radicals, and various toxins. Necrosis occurs preferentially in areas of the CNS that are most severely affected by abrupt biochemical collapse, which leads to the generation of free radicals and excitotoxins. The histologic features of necrotic cell death are mitochondrial and nuclear swelling, dissolution of organelles, and condensation of chromatin around the nucleus.

Intracellular pathways activated by these cellular stresses are designed to turn off nonessential cellular functions, limit the internal damage, activate repair mechanisms, and allow the cell to survive homeostatic challenges. However, when a neuron receives excessive injury, stress response pathways are overwhelmed, resulting in depletion of energy stores and loss of membrane integrity. The resulting necrotic neurons and the organelles they contain swell until their membranes eventually burst. Necrosis is exactly what might be expected of a cell undergoing failure of active transport, membrane integrity, and osmotic stability, all of which ultimately result in disintegration. Necrosis is accompanied by the leakage of cytoplasmic and organelle contents into the extracellular space; such leakage can be toxic to neighboring cells and can promote inflammatory processes. The molecular mechanisms underlying responses to cellular stress are becoming increasingly well understood 18–1. Given these mechanisms, and the rapidity with which the process occurs, necrotic cell death is extremely difficult to treat or prevent.

Apoptosis, a process of programmed cell death as noted above, is essential for the survival of an organism. In the nervous system, it enables the pruning of excess neurons during development, and thereby contributes to the formation of anatomically precise and correct connections between nerve cells. However, increasing evidence indicates that apoptotic mechanisms of neuronal death also contribute to several neurodegenerative disorders and to decrements in brain function associated with aging. Consequently, considerable attention is focused on understanding the precise molecular basis of apoptosis and on developing medicinal agents that interfere with these processes. A brief overview of the molecular pathways of apoptosis is provided in 18–1.

Apoptotic neurons undergo a series of tightly regulated changes characterized by chromatin aggregation, exocytosis of membrane-bound cytoplasmic fragments (apoptotic bodies), and progressive loss of cell volume. Apoptosis is generally recognized as a series of steps undertaken by a doomed cell in order to prevent necrosis. In contrast to necrosis, during apoptosis potentially dangerous intracellular material is packaged into vesicles that can be safely taken up into surrounding cells by endocytosis and metabolized. Unlike necrosis, which is a passive process, cell death by apoptosis results from the execution of active and well-regulated genetic programs. In many biologic systems, inhibition of gene transcription or protein synthesis can prevent apoptosis, underscoring its active nature.

Apoptosis may result in the death of neurons only moderately affected by injury. For instance, it may affect neurons that undergo a mild or short-lived ischemic event, such as those in a peri-infarct area, which is the area between severely ischemic and normal tissue in the period immediately following a stroke. In cortical cell cultures treated with very high levels of N-methyl-D-aspartate (NMDA) (which activates NMDA glutamate receptors; Chapter 5) or nitric oxide (NO) plus superoxide, neuronal death is generally observed to be necrotic; however, with lower levels of NMDA or NO–superoxide, apoptosis occurs. These findings suggest that severely damaged neurons undergo necrosis before a more controlled process of apoptosis can occur, whereas in moderately damaged neurons apoptotic processes may be initiated as a way to dispose of damaged neurons in a safer manner.

Thus, apoptosis provides a safe means of removing damaged cells, including those that are potentially cancerous. For example, the tumor suppressor p53 is a transcription factor that activates proapoptotic gene expression in response to DNA damage that might lead to cell transformation (see figure in 18–1). For this reason, as the field strives to develop inhibitors of apoptosis to limit neuronal loss in neurodegenerative disorders, caution must be exercised with respect to potentially serious side effects of inhibiting this crucial physiologic process.

Cellular Markers of Apoptosis

One of the hallmark characteristics of apoptosis involves the breakdown of double-stranded DNA into nucleosomal segments (Chapter 4). This breakdown makes it possible for apoptosis to be observed, because the resulting segments are equally spaced and can be visualized as DNA “laddering” on an electrophoretic gel. DNA laddering can also be visualized in tissue sections by use of a technique called terminal transferase-mediated dUTP-biotin nick end-labeling (TUNEL). This technique involves the end-labeling of DNA in tissue sections with a fluorescent marker. Because the fragmented DNA in apoptotic cells has many free ends, TUNEL-labeled cells incorporate more of the fluorescent probe and glow brightly.

Role of Mitochondria

Mitochondria are critically involved in the regulation of apoptosis. The triggering of apoptosis is mediated by altering ratios of proapoptotic and antiapoptotic members of the B-cell lymphoma protein 2 (Bcl-2) family of proteins. Proapoptotic Bcl-2 family members promote cell death by permeabilizing the outer membrane of mitochondria and releasing numerous pro-death proteins, including cytochrome c, which is normally a crucial component of the electron transport chain, into the cytoplasm (see figure in 18–1). These pro-death proteins cannot exert their actions when contained in the intramembrane space between the inner and outer membranes of the mitochrondria due to the actions of antiapoptotic members of the Bcl-2 family. However, once in the cytoplasm, the released mitochondrial proteins promote cell death by systematically dismantling specific targets important to cell functioning. For example, cytochrome c combines with cytosolic binding partners to activate a molecular complex termed the apoptosome. The apoptosome in turn activates a family of proteases, termed caspases, which systematically cleave numerous cellular proteins.

Caspases

Caspases (short for cysteine aspartate proteases) cleave and inactivate prosurvival proteins and activate proapoptotic proteins in an amplifying cascade that leads to the rapid fragmentation of cell structures (see 18–1). Under basal conditions, caspases exist as inactive proenzymes until they are activated by proteolytic cleavage. After it enters the cytoplasm, cytochrome c triggers the cleavage and activation of caspase-9. Activation of caspase-9 then triggers the cleavage and activation of several other caspases including caspases-3, -6, and -7. After they are activated, caspases cleave numerous types of proteins that trigger fragmentation of DNA, cytoskeletal proteins, and other cell structures.

Potential Antiapoptotic Therapies

Researchers are now trying to determine whether interference with apoptotic processes will improve treatment outcomes, such as decreasing the area of infarction caused by stroke. For instance, overexpression of the antiapoptosis Bcl-2 gene, or knockout of the proapoptosis Bax gene, reduces infarction in rodent brains subjected to focal ischemic insults (see Chapter 20 for further discussion of stroke). These findings indicate that inhibition of apoptotic pathways may indeed be neuroprotective. New treatments have focused on combining apoptosis inhibition with additional agents, such as antioxidants, neuronal growth factors, ion channel blockers, or thrombolytic compounds to achieve synergistic benefits.

Caspase inhibitors offer an attractive target for pharmaceutical development in part because of the relative ease in designing a drug to inhibit the active site of a protease. Some of the most effective treatments for human immunodeficiency virus (HIV) are aspartyl protease inhibitors. Currently available caspase inhibitors consist of small peptides that do not readily cross the blood–brain barrier or enter cells. Efforts are under way to develop small molecule inhibitors that might overcome such limitations. However, as stated earlier, such inhibitors must be developed with caution, given the evidence that apoptosis is important in protecting the organism from damaged and potentially cancerous cells. Under some circumstances, interference with apoptosis may worsen stroke-related damage. If neurons were to undergo necrosis instead of apoptosis, increased damage to healthy neighboring neurons might result.

One form of cell death unique to neurons is excitotoxicity—a pathologic process by which nerve cells are damaged and killed by excessive stimulation by excitatory neurotransmitters such as glutamate (Chapter 5). It has morphologic and biochemical features of both apoptosis and necrosis and involves increased mitochondrial membrane permeability as a critical event. Excitotoxicity can occur in response to a variety of neurologic insults, both acute and chronic. In acute injury events such as stroke, damaged neurons within the lesion site spill glutamate into the extracellular space, where glutamate can stimulate presynaptic glutamate receptors to enhance the release of additional glutamate. The resulting excessive excitation of nearby neurons is deleterious by setting in motion a series of biochemical cascades, mediated in part by excessive flux of Ca²⁺ into the cells, which lead to disruption of cellular function and eventually neuronal death. Additionally, in chronic neurodegenerative conditions, the ability of neurons to handle Ca²⁺ flux is compromised, further contributing to excitotoxicity.

When blood flow to brain tissue is interrupted, for example, during a stroke, conduits through which oxygen and nutrients enter cells, and potentially toxic metabolites exit, are severed. Interference with blood flow triggers many biochemical changes in the extracellular environment of the affected area of the brain. These changes lead rapidly to a large number of cellular disruptions including depletion of adenosine triphosphate (ATP) stores, loss of ion gradients, release of glutamate, Ca²⁺ pouring into cells through open channels, and activation of numerous second messenger systems. Many of these biochemical changes are very harmful to neurons and contribute to the neuronal death that results from prolonged ischemia. However, some of the sequelae of ischemia may have little effect on neuronal survival and some may even have neuroprotective effects, representing homeostatic responses to ischemic insult that neurons produce in self-defense. Accordingly, strategies designed to minimize infarct-related neural damage must be based on a clear understanding of how each ischemia-related process affects neuronal survival 18–1.

Depletion of Energy Stores

Under normal conditions, the brain uses oxygen and glucose to produce ATP through the processes of glycolysis and oxidative phosphorylation. Complete interruption of brain blood flow can deplete the brain’s supply of oxygen within 10 seconds. In the absence of oxygen, oxidative phosphorylation ceases, and anaerobic glycolysis becomes the only source of ATP for neurons. Anaerobic glycolysis supplies less than 10% of the ATP garnered from complete oxidation of glucose and leads to the production of lactic acid. Moreover, without incoming blood to supply glucose, neurons must rely on the glycolytic processing of preexisting stores of glucose and glycogen. During ischemia, these reserves are depleted within 2 to 3 minutes. Phosphocreatine stores also are consumed during this time. Consequently, ischemia can lead to the depletion of neuronal ATP within minutes. Because the basal metabolic activity of neurons is so great, such depletion rapidly leads to serious consequences.

Failure of Ion Pumps

One of the most important consequences of the loss of available ATP is the failure of ATP-dependent ion pumps and the subsequent dissipation of the resting membrane potential of neurons. As discussed in Chapter 2, the maintenance of the resting membrane potential is ultimately dependent on Na⁺-K⁺-ATPase, which exports Na⁺ and imports K⁺ against their concentration gradients at the cost of ATP. Although very large neurons can maintain a negative resting potential for some time after the inhibition of Na⁺-K⁺-ATPase, small and highly branched neurons of the mammalian brain are extremely vulnerable to loss of Na⁺-K⁺-ATPase activity because of their very large surface area to volume ratio. Accordingly, many CNS neurons depolarize rapidly after ATP depletion.

Loss of Na⁺-K⁺-ATPase activity has other consequences in addition to depolarization. Many ion pumps, including the Na⁺–Ca²⁺ exchanger, are dependent on the transmembrane Na⁺ gradient for their activity 18–2. As this gradient declines, so does the activity of these exchange systems, which leads to further disruptions of ionic equilibrium. Moreover, many ATP-dependent ion pumps, such as ATP-dependent Ca²⁺ pumps, are disabled when neuronal energy stores are depleted.

Neuronal Depolarization and Firing

As neurons depolarize they fire more action potentials and, as such firing escalates, further depolarization occurs, causing a still further rundown of the many ionic gradients required for normal neuronal function. Increased neuronal firing and increased cellular levels of free Ca²⁺ trigger the uncontrolled release of neurotransmitters including the powerful excitatory neurotransmitter, glutamate (Chapter 5). Released glutamate can then trigger neighboring neurons to depolarize and fire action potentials, further extending the injurious chain of events.

Neural damage caused by excessive release of glutamate is potentiated by the failure of glutamate transporters. As explained in Chapter 5, these transporters, which are expressed mainly by glial but also by neural cells, assist in terminating the actions of synaptically released glutamate by removing glutamate from the synaptic cleft. This reuptake process depends on ATP; thus, when ATP stores in the brain become depleted in response to ischemia, the functioning of these transporters is compromised. Subsequently, higher than normal levels of glutamate accumulate in synapses and cause activation of glutamate receptors for abnormally long periods of time.

The role of glutamate in excitotoxicity has spurred the investigation of drugs that block glutamate action in situations of ischemia such as stroke (Chapter 20). Early studies evaluated the utility of potent NMDA receptor antagonists, such as MK-801. However, all of these drugs can induce cognitive distortions, dissociation, and hallucinations. Indeed, phencyclidine (PCP), also known as angel dust, is abused to produce these symptoms (Chapter 16). Other well-known NMDA receptor antagonists include ketamine and dextromethorphan. Several synthetic opioids have NMDA-antagonist properties including meperidine and tramadol. Unfortunately, it has not been possible to separate the neuroprotective effects of potent NMDA antagonists from their unwanted psychotropic actions (Chapter 17). One potential intervention is memantine, a weaker NMDA antagonist, although the clinical efficacy of memantine is limited, and the mechanism for these modest clinical effects in Alzheimer disease is likely unrelated to NMDA receptor blockade at the dosages utilized clinically as discussed further below.

Increases in Intracellular Calcium

Under normal conditions, concentrations of cytosolic Ca²⁺ are under extremely tight regulation (Chapter 2) with measured increases in Ca2+ influx being crucial for numerous cellular functions (Chapter 4) but excessive Ca2+ influx causing toxicity. In a neuron at rest, Ca2+ concentrations are approximately 100 nM, whereas extracellular Ca²⁺ concentrations are in the millimolar range. With extracellular Ca²⁺ approaching concentrations that are 10,000 times greater than cytosolic concentrations, there is normally a tremendous driving force for Ca²⁺ to enter the neuronal cytoplasm. The great effort that a neuron expends in maintaining a low cytosolic Ca²⁺ concentration is compromised during ischemia (see 18–2). Ca²⁺ that enters the cytosol is removed by a variety of mechanisms, including the ATP-dependent Ca²⁺ pumps and Na⁺–Ca²⁺ exchangers previously mentioned. Both of these pumps are ultimately ATP-dependent because ATP-dependent maintenance of the Na⁺ gradient is required for the effective performance of the Na⁺–Ca²⁺ exchanger; indeed, as the Na⁺ gradient dissipates, it may even reverse and cause Ca²⁺ to be pumped into the cell. Depolarization also can lead to the opening of various voltage-gated Ca²⁺ channels, and glutamate release from neighboring cells can induce the entry of Ca²⁺ through NMDA and certain AMPA glutamate receptors. As a result, Ca²⁺ channel blockers, alone or in combination with other agents, are being studied as potential novel treatments for excitotoxicity, such as that seen during stroke (Chapter 20).

Activation of Intracellular Enzymes by Ca²⁺

Increases in neuronal Ca²⁺ may be the ultimate cause of ischemia-related neuronal death (see 18–1). When Ca²⁺ entry is restricted in vitro, for example, through the removal of Ca²⁺ from the extracellular medium or through the blockage of certain Ca²⁺-conducting ion channels, neurons are protected against excitotoxic damage. Moreover, neurons that express subtypes of AMPA receptors that conduct Ca²⁺ (ie, GluA2 lacking receptors; Chapter 5) exhibit heightened vulnerability to glutamate receptor–induced toxicity. Why does a high cytosolic Ca²⁺ concentration induce neuronal death? This question remains under investigation, but there are several clues to the answer. One clue lies in the fact that Ca²⁺ functions as a signaling molecule (Chapter 4). As described in previous chapters, many neuronal enzymes, and hence many neural processes, respond to this signal. It is possible that excessive activation of these various enzymes and processes, caused by inappropriate Ca²⁺ levels in cells, may contribute to neuronal stress and death. Identification of the key proteins involved would help drug discovery efforts to potentially intervene therapeutically.

Among the many enzymes that have been implicated in Ca²⁺-mediated neuronal damage are Ca²⁺-activated proteases, particularly calpain, which can produce widespread cellular damage. Calpain cleaves cytoskeletal proteins that leads to severe cytoskeletal damage. Phospholipases, such as phospholipase A₂ (PLA₂), also can be activated by Ca²⁺. PLA₂ targets membrane phospholipids and hydrolyzes the bond between the fatty acid portions and the polar head groups. When activated en masse, phospholipases may disrupt membrane integrity by changing the numbers and proportions of phospholipids in the neuronal membrane. Moreover, the resulting free fatty acids, which include arachidonic acid, are metabolized by cyclooxygenases and lipoxygenases to yield numerous biologically active products, such as prostaglandins (Chapter 4). The highly reactive •O₂⁻ radical is formed as part of these reactions.

Nitric oxide synthase (NOS), which is activated on binding to Ca²⁺–calmodulin complexes, produces gaseous NO from arginine. As explained in Chapter 4, NO is an important messenger molecule; among its other actions, it activates guanylyl cyclase, which catalyzes the formation of cGMP. NO also can inhibit many cellular enzymes, particularly those containing iron–sulfur complexes, such as mitochondrial NADH–ubiquinone oxidoreductase and NADH–succinate oxidoreductase that are crucial to metabolism. NO produces some of these effects by nitrosylating specific target proteins. Other reactions of the NO molecule—especially its reaction with oxygen to yield peroxynitrate—also may be important in neuronal demise because they may lead to the formation of free radicals.

Formation of Free Radicals

In vitro studies have demonstrated that excitotoxicity is frequently associated with the production of large numbers of free radicals. Free radicals are molecules with an unpaired electron in an outer shell; this unpaired electron causes the molecule to be unstable and highly reactive. Free radicals can peroxidize membrane fatty acids, and in turn can induce alterations in membrane fluidity and integrity. They also can oxidize protein sulfhydryl groups, the products of which can subsequently be metabolized to yield a variety of products, including •O₂^–. Activation of NOS by Ca²⁺ increases levels of NO, which can react with oxygen or O₂⁻ to yield peroxynitrate (ONOO); ONOO decomposes with the production of •OH, a highly toxic free radical.

Regardless of the source, many experiments have shown that free radical scavengers protect against excitotoxicity in vitro. Such scavengers may effectively reduce ischemic injury only when given soon after the occurrence of an ischemic event and may be good candidates to combine with thrombolytic medications (which dissolve blood clots; Chapter 20). Only a small number of free radical scavengers have been investigated in clinical trials of stroke, and none have yet been shown to be efficacious.

Acidosis

Interruption of blood supply to the brain leads to both increased CO₂ tension—due to decreased CO₂ removal—and increased lactate accumulation caused by forced glycolysis in the absence of aerobic respiration. These two events result in acidosis. Ischemia-related acidosis can be severe; indeed, pH values in an ischemic region often approach 6.0. Acidosis affects cellular function in multiple ways, including the promotion of free radical formation; inhibition of the Na⁺–Ca²⁺ exchanger; decomposition of nicotinamide–adenine dinucleotide (NAD), which is essential for oxidative phosphorylation; and inhibition of neurotransmitter reuptake.

Despite our knowledge of these effects, the significance of acidosis to the ultimate death or survival of ischemic neurons is unclear. In fact, acidosis may have certain neuroprotective functions. Neurons exposed to high levels of glutamate in vitro undergo injury at a neutral pH; however, much of this damage is avoided when such experiments are performed at an acidic pH of 6.6. Although many plausible explanations for the protective effect of acidosis exist, one of the most likely involves the pH sensitivity of the Ca²⁺-conducting NMDA receptor. At acidic pH, NMDA currents are drastically reduced, which may in turn significantly reduce Ca²⁺ accumulation in challenged neurons.

Despite widely divergent etiologies, many neurodegenerative diseases are characterized by the inappropriate accumulation of protein aggregates. The protein aggregates can occur either intracellularly in diseases such as Parkinson disease, Huntington disease, Alzheimer disease (ie, neurofibrillary tangles), ALS, and spinocerebellar ataxia type 1 or extracellularly as in the amyloid-β–containing plaques of Alzheimer disease. Although precise mechanisms have yet to be determined, these protein aggregates have been shown to interfere with a wide variety of intracellular processes that can be deleterious to the functioning of the cell.

Proteasomal Mechanisms

The proteasome endopeptidase complex has been likened to an intracellular “trash can.” When a protein is no longer necessary for cellular functioning, it is targeted for destruction by polyubiquitination, the enzymatic addition of small proteins called ubiquitin by enzymes called ubiquitin ligases 18–3. Once a protein has been tagged with four or more ubiquitins, it is transported to the proteasome, unfolded, and cleaved into short peptides, which can then be further degraded into amino acids by cytosolic peptidases.

As will be seen below, several neurodegenerative disorders (eg, Huntington disease, several spinocerebellar ataxias) are characterized by the genetic expansion of a chain of glutamine residues within a particular protein. These proteins can undergo polyubiquitination, but are unable to be processed by the proteasome due to inefficient cleavage between glutamine residues. Another example for proteasome involvement in neurodegeneration comes from identification of one particular ubiquitin ligase, parkin, as a gene involved in a rare form of Parkinson disease. Loss of function in the parkin gene decreases the ability to add ubiquitin to mitochondrial outer membrane proteins. This in turn appears to disrupt mitochondrial homeostasis. Diminished proteasome activity, by either accumulation of uncleavable aggregates (as with polyglutamine diseases) or genetic loss of function of ubiquitinating enzymes (as with parkin mutations), results in a buildup of unwanted and potentially toxic proteins and organelles within the cell. Intensive research efforts are now aimed to identify the mechanism by which protein aggregates lead to neuronal death and to develop novel methods to intervene to limit such mechanisms of neurodegeneration.

Endoplasmic Reticulum Stress and the Unfolded Protein Response

The endoplasmic reticulum (ER), an intracellular system of membranes (Chapter 2), has unique mechanisms for managing protein aggregates. Unfolded or misfolded proteins are recognized and bound by ER chaperone proteins. However, when all available chaperones are completely bound by misfolded proteins, chaperone-associated signaling proteins are released and activate a process termed the “unfolded protein response.” The unfolded protein response functions to help the cell adapt to the increased load of misfolded protein. Part of this response is to halt protein translation and enhance protein degradation. In neurodegenerative diseases associated with intracellular protein aggregates, it is possible that these normal homeostatic mechanisms for handling misfolded proteins are overwhelmed, enabling the proteins to aggregate and damage the cells in other ways. There is also evidence that chronic activation of the unfolded protein response can induce apoptosis through a mitochondrial pathway dependent on proapoptotic Bcl-2 family members (see 18–1).

Autophagy

Autophagy refers to a cellular method for clearing items too large for proteasome degradation, such as damaged organelles or large protein aggregates. In this process the target is engulfed by an autophagosome that then fuses to a lysosome to allow for degradation of the target by lysosomal enzymes. In addition to eliminating large structures, autophagy is used by the cell to free amino acids for synthesis of critical proteins during periods of protein starvation. While this process is initially adaptive for the cell, excessive activation of autophagy can lead to a form of programmed cell death through inappropriate degradation of proteins and organelles. Abnormal autophagy has been implicated in Parkinson disease (see below) as well as in other neurodegenerative diseases. Importantly, pharmacologic inhibition of apoptotic caspases has been shown in some systems to induce autophagy, indicating that redundant pathways exist to dispose of damaged cells.

Potential Therapies for Protein Aggregation

Several nonspecific strategies to treat abnormal, pathogenic protein aggregation are currently in development. Most approaches involve the use of drugs that prevent protein aggregation in cultured cells or in mouse models of neurodegeneration. Some medications in this class include cystamine, geldanamycin, and minocycline, which are now in clinical trials. Cystamine forms disulfide bonds with and inhibits several enzymes, including certain caspases. Geldanamycin activates heat shock protein-90, an important chaperone, which, by binding to toxic proteins, may protect cells from proapoptotic signals. Although the molecular mechanism of action of minocycline is unknown, the drug has been shown to inhibit microglial activation in vitro and in vivo.

Another potential therapy currently under study in Huntington disease is rapamycin that can induce autophagy and potentially speed the clearance of toxic aggregates. Rapamycin acts by inhibiting mTor (mammalian target of rapamycin), a protein downstream of several neurotrophic factor signaling pathways, including AKT (see figure in 18–1; see also Chapters 4 and 8), which exerts powerful control over protein synthesis and autophagy. Because of their central role in apoptosis, medications that stabilize mitochondrial function, such as coenzyme Q10 and creatine, are of great interest. Both compounds are antioxidants.

Recent work has implicated the generation of toxic peptides from protein aggregates in the pathogenesis of neurodegeneration. For example, mice carrying mutant huntingtin (the cause of Huntington disease; see below) exhibit increased cell death. When the caspase recognition sites in the mutant protein are altered to prevent their cleavage by caspases, neurons become resistant to cell death in these mouse models. These findings suggest that release of toxic peptides by proteolytic cleavage of protein aggregates may be important for the progression of certain neurodegenerative diseases. The findings further substantiate the potential utility of caspase inhibitors in the treatment of these illnesses.

Alzheimer disease has become a major public health problem as human life span has been extended by modern medicine. It is the most common cause of dementia and is characterized by a gradual decline in memory and other cognitive functions over a period of years. The disease processes in the brain begin about 15 years prior to the appearance of symptoms, which are generally noticed later in life. Among individuals older than 65 years of age, 6% to 8% have Alzheimer disease; among persons aged 85 and older, the prevalence of Alzheimer disease is approximately 50%. Rare monogenic forms of the disease can begin in midlife.

Among the earliest symptoms of Alzheimer disease are memory impairment and problems with executive function (problem solving). As Alzheimer disease advances, the ability to learn new information is increasingly compromised. Access to distant memories, which is relatively intact in the initial stages of the disease, begins to diminish. As cognitive impairment progresses, patients may become lost while walking or driving, and become increasingly unable to perform tasks such as preparing meals, taking regular medications, or managing finances. As cognition declines, patients may experience depression and irritability and later delusions and hallucinations. Patients also may become aggressive, even toward their caretakers. The end stage of Alzheimer disease is generally characterized by a complete loss of independence. The disease exacts an enormous toll from not only affected individuals but also the friends and family members who care for them and society at large.

As the number and percentage of elderly persons increase steadily, the prevalence of Alzheimer disease almost certainly will rise. It is estimated that by the year 2040, there will be 11 million people in the United States and 80 million people worldwide with Alzheimer disease. Thus, there is an enormous need to develop effective medications to slow or halt the disease process.

Pathology

Even cursory inspection reveals that the brain of a patient with mid- to late-stage Alzheimer disease is noticeably different from that of a normal person of the same age. The diseased brain appears shrunken compared with the normal brain, and has wider sulci, larger ventricles, and less overall mass. However, the pathologic hallmarks of Alzheimer disease and clues as to its etiology can be observed only at the microscopic level. In addition to loss of neurons and synapses, these hallmarks include the presence of various abnormal protein deposits. Senile plaques are dense, extracellular deposits that are composed primarily of the 38- to 43-amino-acid amyloid-β peptide (Aβ), the biochemistry of which is discussed later in this chapter. Two types of senile plaques—diffuse plaques and neuritic plaques—have been identified. Diffuse plaques are extracellular deposits of Aβ in which the Aβ peptide is aggregated but is not present in a β-sheet conformation. Neuritic plaques also consist of extracellular masses of Aβ, but are distinguished from diffuse plaques by the presence of Aβ in a β-sheet conformation as well as the presence of dystrophic dendrites and activated astrocytes and microglia. Intracellular accumulations of abnormally phosphorylated helical filaments in the form of neurofibrillary tangles and neuropil threads also are hallmarks of Alzheimer disease. The major constituent of the tangles is the microtubule-associated protein tau, a protein present in normal brain tissue that is both abnormally phosphorylated and aggregated in Alzheimer disease.

For reasons that currently are not entirely clear, tangle formation and neuron loss tend to develop preferentially in the hippocampus and medial temporal neocortex, and plaques form first in the frontal, parietal, and other association cortices. The distribution of plaques may be related to the relative amount of neuronal and synaptic activity that occurs over a lifetime. This is supported by data showing that Aβ levels are regulated by synaptic activity and that the highest levels of Aβ and subsequent amyloid plaque development occur in brain regions with the highest levels of neuronal activity such as in a cortical circuit known as the default mode network. Cholinergic nuclei such as the nucleus basalis (Chapter 6) are highly impaired while primary sensory cortices and subcortical brain regions tend to be spared. This pattern correlates with early onset memory impairment (declarative memory is dependent on the medial temporal lobe) and progressive cognitive decline (Chapter 14).

The significance of the defining pathologic features of Alzheimer disease has been the topic of intense research. Among the most important questions that investigators have attempted to answer are those related to causality. Are plaques and tangles toxic to neurons? Or are they coping mechanisms that the cell uses to detoxify other, more toxic forms of Aβ and tau? While answers to these questions are still unclear, there is increasing evidence that small, soluble oligomers of Aβ (ranging from dimers to dodecamers and larger species) are particularly pathogenic and may be a major instigator of Alzheimer disease, whereas tau aggregation and its spread to different regions of the brain may be critical in the progression of cognitive loss.

Genetics

Two primary risk factors for Alzheimer disease have long been recognized: advanced age and genetics. Common forms of Alzheimer disease begin later in life and appear to be genetically complex. There are, however, inherited forms in which symptoms appear in middle age and follow autosomal dominant inheritance patterns. More than 100 different mutations in three known genes (amyloid precursor protein [APP], presenilin-1, and presenilin-2) can each cause early onset, familial Alzheimer disease, with some differences in age of onset and rate of progression.

In late-onset forms, the disease does not occur in a Mendelian inheritance pattern. Nonetheless, family studies have revealed that between 25% and 50% of relatives of patients with Alzheimer disease eventually are afflicted with the disease themselves, compared with approximately 10% among control groups. The apolipoprotein E (APOE) gene is the most important known genetic factor that modifies the risk for Alzheimer disease and the age at which the disease emerges. The APOE4 allele increases risk, whereas the APOE2 allele decreases risk, relative to the APOE3 allele. Numerous other genes identified in genome-wide investigations appear to modify the risk for Alzheimer disease, although none yet have been shown to exert as large an effect as APOE. Environmental factors such as sleep and exercise are probably also important in determining whether or not an individual will develop the disease.

Role of APP and Aβ

The first major genetic advance in the study of Alzheimer disease occurred with the discovery of a linkage in some families with the region of chromosome 21 that codes for APP, whose products include Aβ peptides. Duplication of the APP locus, which increases APP gene dose, also causes early onset Alzheimer disease. Further evidence consistent with a role for APP in Alzheimer disease is that patients with Down syndrome (trisomy 21) and thus an extra copy of the APP gene develop amyloid plaques by their 30s and become demented with a mean age of ~50.

The cellular functions of APP and its metabolites in healthy individuals are unknown, but may include regulating synaptic pruning, gene expression, and neuronal growth. APP is a transmembrane protein that is expressed in many tissues, including the brain 18–4. Several different mutations of the APP gene lead to early onset Alzheimer disease 18–5. The biochemical effects of these mutations on APP processing have been studied extensively. One of these mutations, the Swedish mutation adjacent to the β-secretase site in APP, leads to increased production of all Aβ species. As Aβ aggregation is a concentration-dependent process, this likely drives earlier onset of Aβ aggregation. More common mutations in APP just distal to the N-terminus of the Aβ sequence lead to an increase in relative production of Aβ₄₂ to Aβ₄₀. Aβ₄₂ is longer, more hydrophobic, and more likely to aggregate than Aβ₄₀. Another group of mutations changes the sequence of Aβ and its aggregation properties. The mutations cause Alzheimer disease or cerebral amyloid angiopathy, a condition in which amyloid deposition in blood vessel walls leads to frequent intracerebral hemorrhages and strokes. Together with the known neurotoxic effects of Aβ, these genetic observations have led to the “amyloid hypothesis” that Alzheimer disease is caused by excessive accumulation of Aβ, particularly Aβ₄₂, which ultimately leads to neural damage.

How does Aβ₄₂ harm neurons? Aggregation seems to be a key step. Aβ can aggregate into small oligomers that remain soluble, and also into large, insoluble filaments that form plaques. Aβ oligomers and plaques appear to be in some equilibrium. Both types of Aβ aggregates are toxic to neurons. Amyloid plaques are associated with damage to nearby neurites, potentially interfering with proper signaling. Aβ oligomers appear to be even more toxic, and data from animal models indicate that they can disrupt learning and memory. Aβ oligomers, for example, block long-term potentiation and other forms of synaptic plasticity. Aβ binds to many different cellular proteins, and it is not yet known which is the primary effector of its toxic effects. Other factors may also contribute to Aβ-induced neuronal injury. Microglial activation and inflammatory responses help clear Aβ aggregates, but release of cytokines and other mediators may disrupt normal neuronal function (Chapter 12). Aβ-initiated neurotoxic and inflammatory processes may in addition cause the generation of free radicals, which are also damaging to neurons, as outlined earlier in this chapter.

Role of Presenilins

Shortly after mutations in APP were linked to the inheritance of early onset Alzheimer disease, germline mutations in two other genes, presenilin 1 (PSEN1) and presenilin 2 (PSEN2), were also found to cause Alzheimer disease. Mutations in PSEN1 account for the majority of autosomal dominant cases of early onset Alzheimer disease. PSEN1 and PSEN2 are integral membrane proteins that are widely expressed and have multiple functions.

Presenilin is the catalytically active component of the γ-secretase complex, which is formed along with other proteins including nicastrin, Aph-1, and Pen-2 (see 18–4). γ-Secretase mediates intramembranous proteolytic cleavage of type I membrane proteins with small ectodomains. One substrate is APP, after its ectodomain has been shed by β-secretase; γ-secretase cleavage of this APP carboxy-terminal fragment yields Aβ. Mutations in PSEN1 seem to increase its tendency to generate Aβ₄₂, relative to Aβ₄₀. In this way, the resulting increased production of more aggregation-prone Aβ₄₂ is similar to the effect of APP mutations near the γ-secretase cleavage site. This effect is believed to be a major mechanism through which presenilin mutations cause Alzheimer disease. Thus, inhibiting γ-secretase has until recently been a leading therapeutic strategy for Alzheimer disease. However, the situation is complicated by several factors, including the fact that γ-secretase also cleaves many other proteins besides APP, including notch (see below). As well, some experts question whether increased production of Aβ₄₂ is the predominant mechanism by which presenilin mutations cause early onset Alzheimer disease.

Role of ApoE

Apolipoprotein E (ApoE) is a 34-kDa protein that plays an important role in cholesterol transport, uptake, and redistribution in blood. The three common alleles of APOE (ε2, ε3, and ε4) are inherited in a codominant fashion and, as noted above, contribute importantly to the risk for Alzheimer disease 18–6. The inheritance of one copy of the ε4 allele increases risk of disease by ~4-fold and two copies by ~12-fold relative to the ε3 allele. One copy of the ε2 allele decreases risk by ~0.6-fold relative to the ε3 allele. Thus, APOE genotype is the strongest known risk factor for common forms of Alzheimer disease. How might a lipoprotein have such a profound influence on the risk of developing Alzheimer disease? One strong possibility is through effects on Aβ deposition and plaque formation. Three principal lines of evidence support the hypothesis that ApoE promotes amyloidogenesis. First, unlike wild-type mice, mice that lack the gene for ApoE are protected against amyloid deposition. Second, in vitro experiments have demonstrated that ApoE, especially ApoE4 (the protein product of ε4), can enhance the ability of Aβ to form fibrils. Third, ApoE influences the clearance of Aβ in vivo, with ApoE4 resulting in slower clearance than ApoE2 or ApoE3. Fourth, transgenic mice that overexpress various forms of Aβ develop more plaques when they also express ApoE4 and less when they express ApoE2.

Although its effect on amyloid deposition is probably the most well-studied role of ApoE in Alzheimer disease, there are other mechanisms that may contribute. Possible mechanisms include effects of ApoE on synaptic plasticity, inflammation, and tau aggregation. More work needs to be done in these areas to understand their relevance in mediating the role of ApoE in Alzheimer disease.

Role of Tau

The microtubule-associated protein tau is involved in several neurodegenerative diseases. Some, including certain forms of FTD (discussed later), can be caused by tau gene mutations and are termed primary tauopathies. Alzheimer disease is a secondary tauopathy; tau mutations apparently do not cause the disease, but tau protein nonetheless accumulates and plays an important pathogenic role.

The tau gene on chromosome 17 is very complex and alternatively spliced to generate six different isoforms 18–7. Three of the isoforms contain three microtubule-binding domains (3R tau), while the other three have four such domains (4R tau). Tau is expressed primarily in neurons and at lower levels in glia. It is involved in neurite outgrowth and regulates axonal transport, yet tau knockout mice have no dramatic defects.

Interest in tau has focused on neurofibrillary tangles, which are composed primarily of aberrantly phosphorylated and aggregated tau. Several protein kinases are thought to contribute to tau hyperphosphorylation, including cyclin-dependent kinase 5 (CDK5) and glycogen synthase kinase 3β (GSK3β) (Chapter 4). The amount of tangle pathology is highly correlated with the degree of neuronal loss and the severity of cognitive impairment in Alzheimer disease patients. This suggests that pathologic aggregation of tau may be a final pathway leading to neurodegeneration in Alzheimer disease. There is abundant evidence that Aβ can stimulate modification and aggregation of tau, although mechanisms linking the two are not clear. Aβ activates many of the kinases that phosphorylate tau, and also increases pathologic tau cleavage. Mouse models expressing high levels of both Aβ and human tau develop plaques and tangles, and treatments aimed at Aβ reverse some aspects of the tau pathology. Thus, one model for Alzheimer pathogenesis postulates that Aβ triggers downstream changes in tau, leading to tangle formation and neuronal death, thereby causing cognitive impairment.

While many aspects of this model have been validated, other mechanisms undoubtedly also contribute. For example, animal models indicate that, much like soluble oligomers of Aβ, soluble forms of tau contribute to neuronal dysfunction and cognitive deficits, even apart from any role in tangle formation and cell death. Other important questions about tau’s role in Alzheimer disease remain, such as which phosphorylation sites are most important in the disease, the extent to which various tau posttranslational modifications contribute to pathogenesis, and the extent to which tau aggregates may spread from cell to cell in the brain and propagate in a prion-like manner 18–2. Recent experiments that demonstrate protective effects of antitau antibodies in animal models of tauopathy suggest the potential importance of the latter mechanism.

Treatment

Because Alzheimer disease is so disabling and yet so common, intense research efforts in the pharmaceutical industry and in academic settings have been devoted to developing agents that will prevent or retard the disease process. Currently used agents focus on symptomatic improvement only and include drugs that increase cholinergic neurotransmission or block glutamate receptors. Still in development are several strategies aimed at altering the disease processes themselves, such as regulating Aβ or tau.

Cholinergic agents

Because cholinergic neurons of the nucleus basalis are prominently affected in Alzheimer disease, and because cholinergic activity regulates memory processing, enhancement of cholinergic functioning was one of the earliest treatment goals. The initial attempts to increase cholinergic functioning involved administration of the acetylcholine precursors, choline and lecithin. Although the analogous strategy has proven effective for Parkinson disease—L-dopa, a precursor of dopamine, improves symptoms of the illness (see below)—multiple studies have established that cholinergic precursors are not effective in the treatment of Alzheimer disease.

Subsequent efforts met with partial success by inhibiting the cholinesterase activity that clears acetylcholine from the synaptic cleft (see Chapter 6 for a general discussion of cholinergic systems). Four cholinesterase inhibitors are currently approved for the treatment of Alzheimer disease in the United States: tacrine, donepezil, galantamine, and rivastigmine 18–8. These agents do not reverse the course of the disease, nor are they dramatically effective at restoring cognitive function, but in some patients they can temporarily improve attention and memory, delaying the inevitable symptomatic deterioration.

Other cholinergic treatment strategies include agonists or positive allosteric modulators at particular muscarinic or nicotinic cholinergic receptors (Chapter 14). Still another strategy is targeting the nerve growth factor (NGF) signaling pathway, which provides trophic support for cholinergic neurons. However, as discussed in Chapter 8, it is challenging to deliver growth factors directly into the brain and to design small molecules that activate endogenous growth factor signaling pathways. Nevertheless, there are clinical trials under way in which NGF is being delivered via gene therapy directly into the brain in patients with Alzheimer disease.

NMDA receptor antagonists

In addition to cholinesterase inhibitors, the other approved treatment for Alzheimer disease is memantine. Memantine is a low-affinity antagonist of the NMDA glutamate receptor, which is integrally involved in synaptic plasticity (Chapter 14) but also mediates excitotoxic injury as discussed earlier. Because its effects are voltage dependent, it has been proposed that memantine preferentially inhibits excessive NMDA receptor activation associated with excitotoxicity, without blocking the receptor’s function under most normal conditions. However, this speculation is counter to our knowledge of NMDA receptor function in brain and the use of memantine to treat the cognitive symptoms of Alzheimer disease: one would predict that NMDA agonists might improve cognitive function, while NMDA antagonists might reduce longer-term excitotoxicity. Hence, the use of an NMDA antagonist for symptomatic improvement, not as a disease-altering agent, does not make sense based on the reasoning above. It is, of course, quite possible that memantine’s modest clinical effects are mediated via alternative mechanisms not yet identified.

Anti-inflammatory agents and antioxidants

The buildup of Aβ into plaques causes some local microglial and astrocytic activation, with concomitant release of cytokines and acute-phase proteins (Chapter 12). Thus, inflammatory processes are implicated in the neurodegeneration that is characteristic of Alzheimer disease. Consistent with this suspicion, retrospective epidemiologic studies suggest that patients taking nonsteroidal anti-inflammatory drugs (NSAIDs), for example, ibuprofen, on a regular basis—for example, for arthritis—have a reduced risk of developing Alzheimer disease. Unfortunately, prospective trials for people with symptomatic Alzheimer disease have failed to support the efficacy of several different NSAIDs. Future trials of anti-inflammatory agents as preventive measures may prove more fruitful. It is likely that inflammation can in some ways be beneficial, perhaps by contributing to plaque clearance, and that anti-inflammatory therapies will need to be specifically targeted to the harmful aspects of these processes.

A related treatment strategy involves the use of antioxidants and free radical scavengers. Activated microglia secrete hydrogen peroxide, superoxides, and hydroxyl radicals; these are useful in combating foreign cells or organisms but can damage neurons. It is therefore possible that antioxidants and free radical scavengers may serve a protective function in Alzheimer disease and in other neurodegenerative disorders. Vitamin E (α-tocopherol) has been extensively investigated, with recent studies suggesting that it might slow progression of the illness. There has been considerable interest in several natural products called polyphenols, such as resveratrol and curcumin, which are reported to have anti-inflammatory effects and be neuroprotective in cell culture and in animal models of Alzheimer and Parkinson disease. Resveratrol activates sirtuins, which are categorized as Class III histone deacetylases (HDACs), although they deaceylate numerous non-histone substrate proteins as well. Resveratrol is also reported to inhibit the mTOR and TNF-α signaling pathways (Chapters 4 and 8). Curcumin is an inhibitor of histone acetyltransferases (HATs), among several other actions (Chapter 4). Despite the attention given to these agents, there is no convincing evidence for their therapeutic efficacy in humans, although clinical trials continue for several CNS and peripheral disorders.

Disease-altering strategies: targeting Aβ

Much effort has been devoted to strategies aimed at decreasing the production, enhancing the clearance, or neutralizing the neurotoxic properties of Aβ. Several different compounds, including cyclohexanehexols and tramiprosate, are able to block aggregation of Aβ into oligomers or fibrillar plaques in vitro and in animal models. However, studies in humans with mild-to-moderate Alzheimer disease showed no positive effects.

Immune-based approaches to stimulate Aβ clearance are also under clinical investigation. Vaccination with Aβ plus adjuvant was highly effective in mouse models, profoundly reducing amyloid plaque load and improving cognitive performance. However, a clinical trial in patients was terminated early because a subset of patients developed encephalitis related to the immune reaction. Because the trial still gave hints of benefit in at least some patients, second-generation approaches using different immunologic strategies (namely, passive immunization) are under investigation. In two large phase 3 trials, systemic administration of anti-Aβ monoclonal antibodies bapineuzumab and solanezumab did not meet their primary end points in patients with mild-to-moderate Alzheimer disease. However, solanezumab treatment resulted in significant slowing of cognitive decline in patients with mild impairments. An additional phase 3 trial with solanezumab in such patients is therefore now under way. In addition, two prevention trials in early onset familial Alzheimer disease have started in which anti-Aβ antibodies are being assessed. Also, a large prevention trial in cognitively normal elderly individuals who have amyloid deposition in their brains based on new amyloid imaging agents (eg, Amyvid) is slated to further assess the effects of solanezumab in still earlier phases of the illness. A challenge with immunotherapies is determining their mechanism of action. Several models have been proposed. One hypothesis holds that peripherally administered anti-Aβ antibody acts as a “sink,” altering the equilibrium between Aβ in the blood and that in the cerebrospinal fluid (CSF) and brain parenchyma. An alternative hypothesis is that anti-Aβ antibodies, like all other immunoglobulins (IgGs) in the blood, get into the CNS at concentrations of 0.1% to 0.2% of plasma concentrations and contribute to microglial-mediated phagocytic clearance of amyloid plaque. A third hypothesis is that antibodies that enter the CNS alter Aβ aggregation and neutralization of Aβ toxicity. Since only a small fraction of systemically administered antibody crosses the blood–brain barrier, efforts are under way to develop methods to increase delivery of antibodies directly into the brain.

Considerable effort is being devoted to inhibiting Aβ production by blocking β- or γ-secretase. Challenges in the chemical synthesis of β-secretase inhibitors (termed BACE inhibitors) initially slowed their development. However, there are now several such inhibitors in phase 1 to phase 3 clinical trials. This approach holds a great deal of promise, especially if initiated early in the course of illness. However, there is some concern over potential side effects in that, during development, β-secretase regulates myelin thickness as well as other processes.

It has proven easier to block γ-secretase, but concerns over side effects related to substrate specificity have been the limiting issue. Besides APP, γ-secretase cleaves many other type I transmembrane proteins with small extracellular domains. One of the most important is Notch, a signaling molecule with critical roles in differentiation and development. Notch cleavage by γ-secretase releases the Notch intracellular domain (NICD), which acts in the nucleus to regulate gene expression. Presumably through inhibition of Notch cleavage, γ-secretase inhibitors have demonstrated adverse effects on lymphoid and gastrointestinal tissues, which have large populations of rapidly dividing cells. Attempts to create γ-secretase inhibitors that are more selective for APP have thus far not been successful. A related approach is to develop modulators of γ-secretase rather than direct inhibitors of the enzyme’s catalytic activity. These compounds cause γ-secretase to produce less Aβ₄₂ and more shorter Aβ species, essentially the reverse purported effect of presenilin mutations, and thus may lessen Aβ aggregation and toxicity.

Additional approaches to reducing Aβ levels are being considered. α-Secretase cleavage of APP initiates a nonamyloidogenic pathway by cleaving within the Aβ sequence (see 18–4), so increasing α-secretase activity should reduce Aβ levels. In addition, many endogenous peptidases that serve to degrade Aβ have been identified, including neprilysin and insulin-degrading enzyme. Activating either α-secretase or one of these peptidases may be effective in lowering Aβ levels and treating Alzheimer disease.

Other disease-altering strategies

Pharmaceutical approaches to directly target tau or ApoE are not as advanced as approaches toward Aβ. However, recent work by several laboratories has demonstrated the potential effectiveness of antitau antibodies in animal models of tauopathy. This is a promising approach that is expected to move into human trials over the next 1 or 2 years. Other attempts are focused on identifying specific conformations or posttranslational modifications of tau, or interactions between tau and other proteins, which might be pathogenic and could be targeted to treat the disease 18–9. For example, because tau phosphorylation is such a hallmark of Alzheimer disease, inhibitors of the several protein kinases that are known to phosphorylate tau (eg, CDK5, GSK3β) are being investigated, but the multiple roles of these protein kinases raise significant challenges for developing drugs that are safe. Inhibitors of tau aggregation or agents that stimulate tau clearance are also under investigation. Examples include drugs that would increase tau acetylation or O-linked N-acetylglucosamination (O-GlcNAc), among others. Finally, there are efforts to develop drugs that increase the lipidation state of ApoE, which has been shown to be protective in animal models.

Frontotemporal Dementia

The FTDs are a group of related neurodegenerative disorders that are distinguished in several ways from Alzheimer disease, perhaps foremost because of their heterogeneity, both clinically and pathologically.

Clinical syndromes

Whereas the signature cognitive deficit in Alzheimer disease usually begins with memory impairment, FTD patients classically have preserved memory initially, but severe problems in other cognitive domains. Several different syndromes can be diagnosed, depending on which symptoms occur earliest or are most severe. In the behavioral variant of FTD, patients withdraw from their friends and families and lack normal emotional responses, such as empathy when a loved one is injured. They often develop unusual compulsions, personality changes, altered food preferences, and disinhibited behaviors, and lack insight into their own illnesses. Another variant of FTD is semantic dementia. These individuals show some of the same behavioral abnormalities, but are affected primarily by a loss of semantic knowledge, or information about what things are. This leads to problems in naming objects that are shown to them, and eventually with being able to describe anything about the item. A third variant of FTD is characterized by progressive aphasia. These patients show problems primarily in language expression. Finally, some patients with tauopathies present not with FTD but with parkinsonism and other motor symptoms and are diagnosed with progressive supranuclear palsy (PSP) or corticobasal degeneration (CBD) (see the section “Parkinson disease”). Patients who present with one of these syndromes often go on to develop one or more of the others as their disease worsens.

FTD tends to occur in somewhat younger patients than Alzheimer disease, with an average age of onset in the late 50s. In fact, among patients under 65, FTD is as common as Alzheimer disease.

Pathology

The neuropathology of FTD syndromes is also more heterogeneous than that of Alzheimer disease. Amyloid plaques are not observed, and Aβ is not believed to play a prominent role in causing these diseases. About half of patients show some type of tau-positive inclusions, although usually not the classic neurofibrillary tangles seen in Alzheimer disease. The tau pathology can occur in glial cells or takes the form of spherical neuronal inclusions termed Pick bodies. (FTDs were originally referred to as Pick disease.) The other half of FTD patients do not have tau-positive inclusions, but most have inclusions composed of certain ubiquitinated proteins. For years, the identity of the protein accumulating in FTD was unknown, but it has now been identified as TDP-43 (TAR DNA-binding protein of 43 kD). TDP-43 is a nuclear protein, the exact role of which in neuropathogenesis remains to be determined.

Genetics

Mutations in the tau gene are one cause of FTD. Because these cases often have parkinsonism and the tau gene is on chromosome 17, these cases are often referred to as FTDP-17. Many different tau mutations have been described, most of which are around the microtubule-binding domains of the protein 18–7. Many mutations seem to increase the proportion of 4R tau, while others seem to favor tau aggregation. Current research is aimed at understanding precise molecular mechanisms through which tau mutations cause FTD.

The other major genetic cause of FTD is mutations in the progranulin gene. Patients with progranulin mutations develop TDP-43 pathology. Progranulin is a secreted growth factor, and the many different mutations that have been described lead to a destabilization of its mRNA and resulting haploinsufficiency of progranulin. The leading hypothesis is that a ~50% loss of progranulin function leads to neurodegeneration; however, possible mechanisms remain unknown. Less common genetic forms of FTD are caused by mutations in valosin-containing protein (VCP) or charged multivesicular body protein 2B (CHMP2B), both of which have roles in protein handling.

Treatment

There are no approved drugs for FTD. Cholinesterase inhibitors used for Alzheimer disease are not effective, although memantine may be. Serotonergic systems seem to be strongly affected by FTD, and selective serotonin reuptake inhibitors are commonly utilized as symptomatic therapies. Tau-based therapeutics, discussed above under treatments for Alzheimer disease, have yet to be tested in FTDs.

Other Causes of Dementia

Vascular disease is a major cause of cognitive impairment in the elderly. Both large infarctions and chronic ischemia due to changes in small vessels, as occurs with long-standing hypertension, can lead to dementia. These cases are termed vascular dementia or multi-infarct dementia. They are usually diagnosed based on a combination of stepwise (as opposed to steadily progressive) decline, the presence of cardiovascular risk factors, and neuroimaging evidence of vascular disease. The primary form of treatment is prevention (see Chapter 20).

Dementia with Lewy bodies may clinically resemble Alzheimer disease. Distinguishing features include parkinsonism, frequent fluctuations, and the occurrence of visual hallucinations. Pathologically, Lewy bodies containing α-synuclein, identical to those seen in Parkinson disease, are found throughout the neocortex. Treatment is similar to that for Alzheimer disease.

Parkinson disease is characterized by tremor at rest, bradykinesia, and cogwheel rigidity. The characteristic resting tremor is typified by a pill-rolling motion at a frequency of 3 to 5 Hz. Bradykinesia refers to a slowness of movements—in particular, a difficulty in initiating movements—which interferes with the performance of daily tasks. Rigidity and impaired reflexes are partly responsible for the parkinsonian gait, which is traditionally described as festinating, or characterized by short rapid steps. Patients with Parkinson disease also may experience micrography, or small handwriting; weight loss; and alterations in autonomic function; masked facies, or a blank facial expression caused by less frequent eye blinking, and bradykinesia of the facial muscles also are common among affected individuals. Clinical definitions of abnormal movements are given in 18–1.

Pathophysiology

Parkinson disease is caused by the death of dopamine neurons in the substantia nigra pars compacta, with consequent dopamine depletion in the caudate nucleus and putamen, the region where these neurons send their projections (Chapter 6). Pathologic manifestations include degenerative changes, such as neuronal deterioration and depigmentation in the substantia nigra, and the appearance of intracellular inclusions called Lewy bodies in dopamine neurons. Death of dopaminergic neurons in the VTA and that of other monoaminergic neurons (eg, noradrenergic and serotonergic neurons in the brainstem) also may occur on a much smaller scale.

The functional effects of the loss of dopamine in patients with Parkinson disease can be understood, to some degree, based on our knowledge of striatal circuits (see 18–10 and 18–11; see also Chapter 14). Normally, dopamine exerts an inhibitory influence on GABAergic projection neurons of the striatum. Consequently, the loss of dopamine causes increased activity among these neurons, and in turn causes increased inhibitory output from the basal ganglia to the thalamus (see 18–10). Increased activity among striatal GABAergic neurons decreases the activity of GABAergic neurons in the globus pallidus. These events lead to increased activity among glutamatergic neurons in the subthalamic nucleus, which in turn leads to increased activity among GABAergic neurons of the substantia nigra pars reticulata. Such activity causes increased inhibition of the thalamic ventral tier nuclei. Because these nuclei are responsible for the activation of cortical areas involved in the initiation of movements, the ultimate effect of dopamine deficiency is paucity of movement.

Healthy individuals possess a large reserve of nigrostriatal dopaminergic neurons; consequently, it is estimated that overt parkinsonian symptoms do not occur until greater than 50% of these neurons have been lost. Compensatory responses in remaining neurons, such as the upregulation of tyrosine hydroxylase (Chapter 6), enable these cells to make more dopamine. The functional reserve provided by these adaptations causes most cases of Parkinson disease to be characterized by a slow, progressive loss of function. Rapidly evolving lesions, such as those produced by neurotoxins, can produce parkinsonian symptoms in response to the loss of many fewer dopaminergic neurons. Interestingly, a very slow, progressive loss of dopamine neurons occurs—even in the absence of Parkinson disease—in normal individuals throughout adult life.

Genetic Factors

Increasing evidence suggests that there is a genetic contribution to the etiology of common forms of Parkinson disease, but the exact nature of this contribution remains unclear. In most cases, the disease appears to be sporadic; that is, it occurs in individuals who do not have a family history of the disease. However, this pattern of disease may reflect genetic susceptibility that only gives rise to illness when certain environmental factors, such as exposure to dopaminergic neurotoxins, are also present.

In addition to the sporadic, common forms of Parkinson disease, there are rare familial forms that are characterized by Mendelian transmission. Several genes responsible for such transmission have been identified by genetic linkage analysis, and several more genes are likely to be identified from families known to have Mendelian patterns of inherited Parkinson disease or related syndromes without mutations in the several genes already identified 18–2. The normal cellular functions of the proteins encoded by these genes remain largely unknown, and the pathophysiologic mechanisms by which mutations in these genes cause Parkinson disease remain the subject of intense research. Current leading hypotheses for the causative mechanisms include protein aggregation or disruption of protein degradation, mitochondrial dysfunction, and oxidative stress. Another major unanswered question is why mutations in proteins widely expressed throughout the brain and in other tissues cause selective death of dopamine neurons in the substantia nigra.

Evidence for protein aggregation as a causative mechanism emerged on the identification of point mutations in α-synuclein, which were the first mutations linked to familial Parkinson disease. α-Synuclein is a major component of Lewy bodies, the pathognomonic inclusions of sporadic Parkinson disease. This suggested that α-synuclein aggregation may cause sporadic Parkinson disease and that α-synuclein point mutations linked to familial Parkinson disease increase the propensity of α-synuclein to form intracellular aggregates that may be toxic and may also lead to Lewy body formation. The subsequent identification of α-synuclein gene duplication and triplication mutations linked to familial Parkinson disease reinforced the hypothesis that increased cellular concentrations of even wild-type α-synuclein may cause disease by increasing the rate of formation of α-synuclein protein aggregates. Consistent with this idea, transgenic animals overexpressing wild-type or mutant α-synuclein develop intracellular α-synuclein inclusions and neurodegeneration.

Further evidence supporting protein aggregation or disruption of protein degradation as a pathogenic mechanism comes from the linkage of familial Parkinson disease to loss-of-function mutations in a novel E3 ubiquitin ligase named parkin. As stated in an earlier section of this chapter, ubiquitin ligases are critical for protein degradation via the proteasome (see 18–3). It is thought that one or more proteins accumulate and perhaps aggregate in the absence of normal parkin function, thus leading to cell death. Recently, parkin function has also been linked with mitochondrial clearance. Lysosomes are the other major cellular mechanism for protein degradation. Recessively inherited mutations in a gene encoding a putative lysosomal membrane adenosine triphosphatase (ATPase) of unknown function, ATP13A2, have also been linked to familial Parkinson disease.

The first evidence that mitochondrial dysfunction may cause Parkinson disease came not from genetics but from the discovery of a severe form of parkinsonism that developed in a group of young adults after intravenous use of a preparation of meperidine, a synthetic opiate, which contained the harmful by-product 1- methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Such patients exhibited rapid and extensive loss of dopamine neurons, resulting in severe parkinsonism. Investigators have since learned that MPTP is converted within the brain to its active metabolite MPP⁺, which selectively accumulates in dopaminergic neurons via the dopamine transporter and is directly neurotoxic by inhibiting mitochondrial respiration (Chapter 6). MPTP and another selective dopaminergic toxin, 6-hydroxydopamine, are used to induce Parkinson-like syndromes in animals. Mitochondrial respiration defects also occur in sporadic Parkinson disease and in patients bearing parkin mutations.

Direct genetic evidence of mitochondrial dysfunction as a cause of Parkinson disease comes from the linkage of loss-of-function mutations in the PINK1 (PTEN-induced kinase-1) gene to recessively inherited Parkinson disease. PINK1 is a kinase that localizes to mitochondria, and is induced by PTEN, a dual function lipid and protein phosphatase upstream of the AKT cell survival pathway mentioned earlier (see 18–1; Chapter 4). Although the precise function of PINK1 remains unknown, it may be important for preventing cell death through regulation of mitochondrial function.

Another kinase of unknown function with mutations linked to familial Parkinson disease, leucine-rich repeat kinase 2 (LRRK2; also called dardarin), partially localizes to mitochondria. Although LRRK2 mutations were initially believed to be rare, the G2019S point mutation in LRRK2 has been reported in 30% to 40% of familial Parkinson disease patients of Ashkenazi Jewish or North African Arab origin. LRRK2 polymorphisms may also be a significant risk factor for Parkinson disease in the Asian population.

Oxidative stress has long been hypothesized to be a cause of sporadic Parkinson disease because dopamine itself readily reacts to form chemical species that can cause oxidative damage to cells. Age is the greatest risk factor for Parkinson disease, which implicates cumulative oxidative damage to cells. DJ-1 is a protein, first identified as a product of an oncogene, believed to protect cells from oxidative stress. Loss-of-function mutations in DJ-1 have been linked to parkinsonism. Mice deficient in DJ-1 are hypersensitive to MPTP and oxidative stress.

Recent work has demonstrated that mutations in GBA1, which encodes β-glucocerebrosidase, are a common risk factor for Parkinson disease, with as many as 5% to 10% of all patients with the illness being heterozygous for an abnormality in this gene. Individuals homozygous for GBA1 mutations typically develop Gaucher disease, a lysosomal storage disease characterized by the intracellular accumulation of lipids in brain and several peripheral tissues.

Environmental Factors

Prior to the identification of genes with mutations linked to familial Parkinson disease, it was commonly thought that Parkinson disease was strictly caused by environmental factors. This belief was reinforced by the discovery in the early 1980s that MPTP exposure can cause a severe and rapid-onset form of parkinsonism.

The possibility that environmental neurotoxins may cause sporadic cases of Parkinson disease—albeit with less dramatic effects than MPTP—has been further supported by epidemiologic evidence, not always verified, that the disease may be more common in people who reside in rural areas, drink well water, and have regular exposure to herbicides and pesticides. Potential disease-causing toxins have been identified. The pesticide rotenone can reproduce features of Parkinson disease in animal models. Heavy metal poisoning, particularly from exposure to Mn²⁺, also has been associated with Parkinson disease, particularly in welders. Moreover, a high incidence of the disease has been noted after certain viral epidemics; thus, a viral contribution to the disease is conceivable. However, specific toxins and viruses that cause common forms of Parkinson disease have yet to be identified with certainty.

Epidemiologic studies suggest that inflammation may be involved in sporadic Parkinson disease, because users of NSAIDs are less likely to have Parkinson disease. Smoking and increased caffeine intake are also associated with reduced risk of the illness. The putative protective mechanisms of caffeine and smoking remain speculative, and it cannot be excluded that patients with Parkinson disease are less prone to caffeine or nicotine addiction, especially given the well-established role of dopamine in reward and addiction. Nicotine has been reported to protect against neurodegeneration in the striatum but not in the substantia nigra of animals treated with MPTP. By contrast, caffeine and other antagonists of adenosine A_2A receptors, which are highly enriched in striatum, have been reported to protect against nigral cell loss as well as depletion of dopamine in striatum of animals treated with MPTP. Targeted disruption of the A_2A receptor gene also confers protection from MPTP toxicity in mice.

Drug-Induced Parkinsonism and Dyskinesias

As described in Chapter 6, antipsychotic drugs that potently block D₂ dopamine receptors can cause a reversible parkinsonian-like syndrome. Drug-induced parkinsonism is most common among elderly patients, presumably because they have a smaller reserve of dopamine neurons. Symptoms develop within days to months after use of the offending drug commences 18–3 and disappear over weeks or months after cessation of the drug.

Parkinsonian-like symptoms caused by first-generation antipsychotic medications are among a large number of so-called extrapyramidal side effects produced by these drugs 18–3. The most dramatic such effect produced in rodents is catalepsy, a waxy-like rigidity of the extremities. Such side effects are caused by the D₂ dopamine receptor antagonist properties of these agents. Blockade of these receptors produces functional effects that mimic those of increased activity of GABAergic projection neurons, such as occurs with the loss of dopaminergic neurons 18–11.

The fact that antipsychotic drugs reduce psychosis, presumably through D₂ antagonism in the mesocorticolimbic dopamine system, and cause extrapyramidal side effects, presumably through D₂ antagonism in the nigrostriatal dopamine system, has provided clinicians with a dilemma. Agents that tend to reduce psychosis also tend to increase the severity of motoric side effects, and agents that reduce motoric side effects likewise tend to aggravate psychosis. A similar dilemma is encountered in the treatment of Parkinson disease: although L-dopa is effective in replacing lost dopamine, it can cause dyskinetic side effects and psychotic symptoms after prolonged use. Likewise, agents that are used to reduce the dyskinetic side effects associated with L-dopa tend to increase the severity of parkinsonian symptoms. As discussed in Chapter 17, the introduction of newer antipsychotic medications has helped to reduce some of these complications.

Tardive dyskinesia

Antagonism of D₂ receptors has been implicated in a particularly severe extrapyramidal side effect of traditional antipsychotic medications known as tardive dyskinesia (TD). TD is unlike other extrapyramidal side effects in that it requires prolonged exposure to the offending drug; indeed, the longer the exposure, the greater the likelihood of developing this syndrome. Moreover, unlike other extrapyramidal side effects, it can persist for long periods, even for a lifetime, after the precipitating medication has been discontinued. TD involves excessive, abnormal movements, particularly choreoathetoid movements that are reminiscent of Huntington disease. These abnormal movements, which predominate in the head and neck and typically involve the mouth and tongue, can be debilitating. TD must be distinguished from withdrawal dyskinesias that occur in many individuals after the cessation of antipsychotic treatment and gradually subside over weeks or months.

The Huntington disease–like features of TD symptoms have led to the proposal that TD represents a state of supersensitivity to dopamine in the striatum that develops as an adaptation to persistent D₂ receptor antagonism. Indeed, one treatment for TD symptoms involves raising the dose of the precipitating D₂ antagonist, a practice that is not ultimately beneficial because it leads to a vicious cycle of TD symptoms followed by increasing doses of medication, which in turn cause a worsening of TD symptoms. Despite a great deal of research, the drug-induced changes in the striatum or elsewhere in the brain that cause TD are not yet known. Fortunately, the incidence of TD has decreased dramatically with the introduction of second-generation antipsychotic agents and with the use of lower doses of first-generation antipsychotic drugs (Chapter 17).

Treatment of Parkinson Disease

The treatment of Parkinson disease currently consists of the administration of medications that enhance dopaminergic function in the basal ganglia. As described in Chapter 6, oral administration of L-dopa is the most effective symptomatic treatment available for Parkinson disease 18–12. L-Dopa is transported across the blood–brain barrier and is converted to dopamine within remaining dopaminergic nerve terminals and, perhaps, by other monoaminergic terminals that can transport L-dopa via their plasma membrane transporters.

L-Dopa efficiently restores neurologic function, particularly during the early stages of Parkinson disease. Yet systemically administered L-dopa cannot possibly replicate dopaminergic innervation of the striatum, either spatially or temporally. Moreover, the efficacy of L-dopa decreases after several years and, as previously mentioned, side effects in addition to dyskinesias begin to appear, including visual hallucinations and other psychotic symptoms, sleep disturbances, and confusion. A late complication of L-dopa therapy, referred to as the on–off phenomenon, involves abrupt and transient fluctuations in the severity of parkinsonism at different intervals of the day; such fluctuations are unrelated to drug dosage. To minimize this phenomenon, individuals often restrict their use of L-dopa to times of the day when therapeutic relief is required.

Peripheral side effects caused by the conversion of L-dopa into dopamine by aromatic amino acid decarboxylase expressed in liver and other peripheral tissues may include severe hypotension and nausea. Such problems with L-dopa have been controlled by aromatic amino acid decarboxylase inhibitors such as carbidopa and benserazide 18–12. When administered with L-dopa, such agents, which cannot cross the blood–brain barrier, inhibit peripheral aromatic amino acid decarboxylase and reduce the peripheral conversion of L-dopa into dopamine. Such actions reduce the peripheral side effects of L-dopa and enable the drug to be administered in much smaller doses. L-Dopa and carbidopa are available in a fixed-ratio preparation called Sinemet. Entacapone, an inhibitor of catechol-O-methyltransferase (COMT), is also used in conjunction with L-dopa in the treatment of Parkinson disease. Because L-dopa is metabolized peripherally by COMT (Chapter 6) as well as by aromatic amino acid decarboxylase, entacapone can prolong the efficacy of a dose of L-dopa.

Dopamine agonists, which theoretically should be effective during later stages of Parkinson disease, mimic endogenous dopamine by directly stimulating dopamine receptors and, unlike L-dopa, do not require enzymatic transformation. Many dopamine agonists also are not metabolized by L-dopa oxidative pathways and thus do not produce the potentially toxic metabolites of this agent. Bromocriptine is an ergot derivative that directly stimulates dopamine D₂-like receptors. It is less effective than L-dopa in providing symptom relief, but produces less dyskinesia and less of the on–off phenomenon, possibly because of its direct receptor activation. Like bromocriptine, pergolide is an ergot derivative and dopamine receptor agonist, but it acts on both D₁- and D₂-like receptors, while ropinirole and pramipexole primarily antagonize D₂-like receptors. These medications are used most commonly as an adjunct to L-dopa and, by reducing the amount of L-dopa needed, can decrease the long-term complications of L-dopa therapy. The development of more selective D₁ and D₂ receptor agonists, long a goal of the pharmaceutical industry, has not yet been successful. D₂-like agonists, such as bromocriptine, ropinirole, and pramipexole, are used in the treatment of restless leg syndrome.

Muscarinic cholinergic antagonists are helpful in alleviating tremor and rigidity but are generally less effective than dopaminergic drugs. However, they are the mainstay of treatment for parkinsonism induced by D₂ antagonists because dopaminergic therapy increases the severity of associated psychiatric symptoms. The mechanism by which muscarinic anticholinergic agents reduce parkinsonism can be understood based on the actions represented in 18–11. Normally, cholinergic interneurons provide an excitatory influence on striatal medium spiny neurons; consequently, the loss of dopaminergic inhibition of these medium spiny neurons can be compensated for by a decrease in cholinergic excitation. Muscarinic antagonists commonly used to treat parkinsonism include benztropine 18–12, trihexyphenidyl, and diphenhydramine. There is interest in developing antagonists or negative allosteric modulators that are selective for particular muscarinic receptors, for example, M₄ that is enriched in striatum.

Amantadine, introduced originally as an anti-influenza medication, has more recently proven to be useful in the treatment of Parkinson disease. It may be given alone or in combination with anticholinergic medications and is believed to potentiate the release of dopamine. It appears to exert this effect by acting as a weak antagonist at NMDA glutamate receptors, although precisely how this action leads to enhanced dopamine release is not yet known. Amantadine improves all of the clinical features of parkinsonism, and associated side effects are uncommon. However, the benefits of this drug are relatively modest and short-lived in most patients.

Selegiline (deprenyl) is a selective inhibitor of monoamine oxidase B (MAOB) (Chapter 6). It was introduced into clinical use because MAOB converts MPTP into its active neurotoxic form, MPP⁺. It was hypothesized that the progression of idiopathic Parkinson disease may involve neurotoxins that act like MPTP but are as yet unidentified; accordingly, it was speculated that selegiline might retard the progression of Parkinson disease. Large multicenter studies indicate that although selegiline may offer benefits during early stages of the illness, its efficacy wanes as the disease progresses. In addition, selegiline may not provide a neuroprotective effect but may simply increase dopamine levels by inhibiting monoamine oxidase.

Rasagiline is a newer selective inhibitor of MAOB that shows both symptomatic benefit and potentially neuroprotective properties perhaps by antioxidant or antiapoptotic effects. In initial large-scale clinical trials, new Parkinson patients receiving rasagiline for 12 months showed less functional decline compared with a control group receiving placebo for the first 6 months and rasagiline for the remaining 6 months. Efforts to confirm these promising effects are under way.

The antioxidants coenzyme Q10 and creatine, mentioned earlier in the chapter, are also undergoing clinical trials for Parkinson disease. Coenzyme Q10 is a component of the mitochondrial respiratory chain that is neuroprotective in animal and cell models of dopamine neurotoxicity. A phase 2 trial in Parkinson patients suggested that high doses of coenzyme Q10 may slow the progression of motor symptoms. Further studies are in progress. Like coenzyme Q10, dietary creatine supplementation is thought to promote mitochondrial function that is likely deficient in Parkinson disease. A clinical trial is being conducted with creatine combined with minocycline that is hypothesized to inhibit apoptosis and inflammation.

Drugs that act at various types of adenosine receptors, in particular, A_2A antagonists, are currently under evaluation for use in the treatment of Parkinson disease. Work to date indicates that the beneficial effects of such agents may be limited. These compounds are believed to act by means of interactions with dopamine signaling on striatal neurons, as previously discussed. (See Chapter 8 for further discussion of adenosine systems in brain.)

New approaches to treatment

Because patients treated pharmacologically soon become refractory to L-dopa and related therapies, other treatment strategies have been explored. Excess output from the subthalamic nucleus is postulated to play a critical role in the pathophysiology of Parkinson disease (see 18–10). Surgical lesions of this nucleus, or of the internal segment of the globus pallidus (called a pallidotomy), have reduced major motor disturbances such as akinesia, rigidity, and tremor. However, because such treatment involves an irreversible lesion, it is reserved for the most severe and medication-refractory cases. More recently, deep brain stimulation with a stimulating electrode in the subthalamic nucleus has proven to be a very effective treatment for advanced Parkinson disease. At appropriate stimulation frequencies, subthalamic nucleus function is influenced and symptoms are relieved, at least temporarily. The cellular and circuit mechanisms by which high-frequency stimulation of this brain area alleviates the symptoms of Parkinson disease remain poorly understood.

Cell transplantation, for example, the administration of fetal mesencephalic dopaminergic neurons—or stem cells programmed to differentiate into dopaminergic neurons—into the striatum, remains an area of interest for the treatment of Parkinson disease. This approach is based on the expectation that such cells will proliferate, establish synaptic connections, and synthesize and release dopamine in the striatum. Indeed, Parkinson disease would appear to be particularly well suited, compared with other neurologic or psychiatric disorders, for cell transplantation or stem cell therapies, given the fact that it involves the loss of a highly localized cell population in the adult brain. Yet, clinical trials with fetal dopaminergic neurons have been disappointing. Further, while the use of stem cells, which has not yet been evaluated in humans, offers some promise, it also remains very speculative; a tremendous amount of progress will be needed to (1) deliver programmed stem cells and target them to the striatum, and (2) prompt those cells to form synaptic connections in the adult brain that normally form early in development.

Gene therapy refers to the transfer of a therapeutic gene into a target tissue and the maintenance of its function for a sufficient length of time. Gene therapy for neurologic disorders can be achieved when genes are delivered into the brain either by transplantation of genetically modified cells or by direct injection of genes (eg, by means of viral vectors) into the striatum. Several strategies for viral-mediated gene transfer are under active investigation. Early studies focused on delivering the tyrosine hydroxylase gene to striatal neurons as a way to increase local dopamine synthesis. More recent efforts have focused on the delivery of various types of neurotrophic factors into the substantia nigra or striatum, either by viral gene transfer or by direct delivery of the growth factor into the brain. Brain-derived neurotrophic factor (BDNF), glial cell line–derived neurotrophic factor (GDNF), and neurturin have received the most attention given their ability to promote the survival of dopamine neurons in animal models (Chapter 8). Still other approaches have involved delivering genes that encode proteins that alter the functional activity of neurons, for example, in the subthalamic nucleus, as a way to correct the abnormal circuit function seen in Parkinson disease. So far, trials with GDNF and neurturin have been negative in patients with moderate-to-severe Parkinson disease, although there is a positive phase 2 study involving viral overexpression of glutamatic acid decarboxylase (GAD) in the subthalamic nucleus.

Increasing evidence suggests a prominent role of oxidative stress, inflammation, apoptosis, mitochondrial dysfunction, and proteasomal dysfunction in the pathogenesis of Parkinson disease and other neurodegenerative disorders. Peroxisome proliferator–activated receptors (PPARs), members of the nuclear receptor superfamily (Chapter 4), regulate numerous cellular processes including tissue differentiation, inflammation, mitochondrial function, wound healing, and lipid and glucose metabolism. PPARγ agonists, pioglitazone and rosiglitazone, have been shown to exert neuroprotective activity in preclinical models of Parkinson disease, Alzheimer disease, cerebral ischemia, and ALS. For example, in the acute MPTP model of Parkinson disease, pioglitazone blocked dopaminergic neurodegeneration and diminished astrocytic and microglial activation after MPTP administration. Another animal study showed that rosiglitazone prevented motor deficits, impairment of olfactory function, and loss of dopaminergic neurons in the substantia nigra. Despite these encouraging results, however, it is unlikely that large clinical trials of rosiglitazone or pioglitazone in Parkinson disease will be started until known safety issues with these specific compounds are addressed. Thus, use of rosiglitazone, which is approved for the treatment of type 2 diabetes, is associated with an increased risk of heart attacks. Certain PPARγ-associated proteins, such as PPARγ coactivator 1α, are also being considered as possible drug targets.

Finally, major efforts are being made to take advantage of the increasing knowledge of the genetic and pathophysiologic basis of Parkinson disease to develop treatments that fundamentally slow the progression of the illness. One approach is to develop LRRK2 inhibitors, based on the view that disease-causing mutations in LRRK2 are associated with increased kinase activity. Generation of agents that promote autophagy—to better dispose of accumulated protein aggregates—represents another potential strategy.

Dystonias comprise a movement disorder characterized by patterned, directional, and often sustained muscle contractions that produce abnormal postures or repetitive movements. The term “patterned” refers to the repeated involvement of the same group of muscles, a feature that helps to differentiate dystonias from other hyperkinetic movement disorders. Dystonias present in many different ways and can vary widely in the number of body areas and motor systems affected, as well as in clinical features, age of onset, natural history, and etiology. Common forms of dystonias include writer’s cramp and torticollis (characterized by twisting of the neck). Accumulating evidence suggests that primary dystonia involves circuit dysfunction of the basal ganglia–thalamocortical and cerebellothalamocortical pathways. Furthermore, while dystonias are primarily thought of as motor disorders, there is abundant evidence that there are also derangements of sensory function. Most cases of dystonia are not considered to be of primary genetic etiology. However, study of familial cases of dystonia has to date identified mutations in >20 distinct genes that cause monogenic forms of the disorder. The first identified genetic cause of dystonia was disruption in the gene that encodes torsin-1A (TOR1A; also known as DYT1). Torsin-1A is an ATPase, and it is not known how mutation in the TOR1A gene leads to dystonia. There is no apparent shared functional abnormality across the many known genetic forms of dystonias, creating a challenge for future research.

Pharmacologic treatment of dystonias includes the use of muscarinic cholinergic antagonists, GABAergic agonists (eg, baclofen), and dopaminergic agonists (eg, L-dopa). For focal dystonias the standard of care therapy has become botulinum toxin injections into the affected muscles. Botulinum toxin irreversibly binds to components of the nerve terminal exocytic machinery, preventing the release of acetylcholine into the neuromuscular junction (Chapter 3). The resulting chemical denervation produces local paralysis, thus relieving the sustained muscle contraction marking the disorder. Nonmedication approaches such as physical therapy are useful in many patients.

Huntington disease is an autosomal dominant disorder caused by a mutation of the huntingtin gene on the short arm of chromosome 4. The clinical manifestations of the disease typically begin in midlife, with a mean age of onset of 40 years. Initially, the symptoms are subtle and include minor coordination problems, jerky eye movements, and occasional choreiform movements of the fingers, limbs, or trunk. Such symptoms may be accompanied by depression, psychotic symptoms, irritability, impulsiveness, and cognitive changes. Deterioration leads to progressive dementia, emotional lability and personality changes, and choreiform movements. Progressive cognitive decline causes disruption of memory and difficulty with complicated tasks; in addition, mood swings, depression, and apathy become more pronounced. During late stages of the disorder, choreiform movements regress and dystonia and rigidity are typical symptoms.

In approximately 10% of cases of Huntington disease, symptoms occur before age 20. Individuals with juvenile-onset disease exhibit a more rapidly progressive course and are more likely to experience dystonia with seizures and cerebellar ataxia than chorea. As discussed in the next section, genetic factors are an important determinant of early onset of disease.

Pathophysiology

Huntington disease is associated with degenerative changes that are most apparent in the caudate nucleus and putamen. Such destruction is selective and is restricted to populations of GABAergic medium spiny projection neurons, particularly those that form the indirect pathway and project to the globus pallidus (see 18–10). Neuronal loss also occurs in the thalamus and cerebral cortex. Striatal cholinergic interneurons, as well as midbrain dopaminergic neurons, are largely unaffected. Indeed, choreiform activity is believed to result from excessive activity of preserved nigrostriatal dopaminergic neurons in conjunction with decreased activity of striatal GABAergic neurons (see 18–10 and 18–11). However, the connection between a loss of GABAergic output from the striatum and choreiform movements is not yet understood.

Genetics and Pathogenesis

Huntington disease is a Mendelian dominant genetic disorder; individuals who inherit only one mutated copy of the gene do not differ clinically from individuals who possess two defective copies. The molecular basis of Huntington disease is an expansion of a trinucleotide repeat sequence (CAG)_n within the coding region of the huntingtin gene, which was first identified in a linkage study of a large lineage with the disease. In normal individuals, the repeated CAG triplet encodes an average of 5 to 30 copies of the amino acid glutamine within the huntingtin protein. Huntington disease results from an expansion of these triplet repeats that yields 37 to 86 or more glutamines. Such expansion of nucleotide repeats is common to several inherited neuropsychiatric diseases 18–4. Longer repeats in individuals with Huntington disease are associated with an earlier age of onset and, in some cases, with more severe symptoms. Moreover, repeat lengths are unstable and tend to increase from one generation to the next through transmission from both sexes. This expansion most likely occurs during gametogenesis and is believed to be characterized by average increases of 0.4 and 9 repeat units in female and male transmissions, respectively. Such changes result in the clinical phenomenon of anticipation—an earlier age of onset or an increase in the severity of disease across successive generations. As expected, individuals with juvenile onset of Huntington disease have the largest number of glutamine repeats in huntingtin.

Although the age of onset of Huntington disease is most dependent on repeat length, it also is influenced by many modifying genes and environmental factors. Accordingly, two individuals with the same number of repeats can exhibit very different courses of the disease when they are members of different families. There is great interest in identifying genetic and environmental factors that lead to such modifications because such knowledge might improve management or prevention of the disease and shed light on its pathogenesis. The expanded repeats in huntingtin can be detected in gene carriers by a simple polymerase chain reaction (PCR)–based assay. Consequently, family members can be tested before they are symptomatic to determine whether they carry the defective gene, and the gene also can be detected in fetuses through prenatal testing.

The mechanism underlying Huntington disease represents a classic example of a gain-of-function mutation. The disease is caused by a single copy of a mutant gene that leads to a new biologic effect, rather than a simple increase or decrease in the normal effect of the gene. The new function of mutant huntingtin remains incompletely understood. However, according to an emerging hypothesis, the longer the glutamine repeat is, the more likely the huntingtin protein is to accumulate within vulnerable neurons as intranuclear inclusion bodies, which are seen in the striatum of patients with Huntington disease. Such inclusion bodies or small aggregates of huntingtin lead to disruptions in critical cellular functions, including RNA processing, and eventually cause cell death. It has been hypothesized that the gain of function in mutant huntingtin is the neural toxicity of the expanded polyglutamine domain of the mutant protein. Consistent with this theory are recent findings from mice that overexpress a mutant huntingtin gene; these animals exhibit nuclear inclusion bodies and morphologic changes in vulnerable neurons—early signs of apoptosis that parallel the patterns of cell death associated with Huntington disease. Such mice represent models that can be used to develop new therapies. Interestingly, the huntingtin gene is widely expressed throughout the brain and most other organs; therefore, why mutant huntingtin leads to a relatively selective pathologic process within striatal GABAergic neurons remains a mystery.

The normal function of huntingtin is also something of a mystery. Knockout mice that lack huntingtin die early during embryonic development, exhibiting features of apoptotic cell death. Such findings indicate that huntingtin plays a critical role in many cell types, yet investigators have yet to uncover clues to its function.

Treatment of Huntington Disease

Unfortunately, no specific treatment is available for Huntington disease. The drugs most commonly used for controlling associated dyskinesias are dopamine receptor antagonists, including first-generation antipsychotic agents such as haloperidol and chlorpromazine. These drugs are effective because the blockade of dopamine action relieves dopamine’s inhibitory influence on dying GABAergic neurons, thereby increasing net GABAergic output from the striatum (see 18–11). The use of antipsychotic drugs also has the beneficial effect of reducing psychosis in patients who experience such symptoms. Other pharmacologic treatments of Huntington disease target specific symptoms, such as depression and impulsive behavior, for which antidepressants and propranolol have been used, respectively. However, the therapeutic efficacy of the latter agent, a β-adrenergic antagonist, is limited.

The interactions represented in 18–11 suggest that drugs that increase GABAergic or cholinergic function in the striatum should ameliorate the symptoms of Huntington disease. However, attempts to alleviate chorea and other dyskinesias with drugs that exert such effects have not yielded consistent clinical responses.

Lessons From Huntington Disease

Experience with Huntington disease has raised several points that are essential to remember as we strive to identify the causes of other neuropsychiatric disorders. First, the gene that causes Huntington disease was previously unknown; its discovery illustrates the power of “reverse genetics”—that is, approaches to finding a disease-related gene that can proceed without prior knowledge of the disease’s pathophysiology. Second, the discovery of a disease-related gene provides essential tools for investigating pathogenesis. Indeed, classic approaches to understanding disease pathophysiology did not succeed in explaining Huntington disease. Classic neuropharmacologic approaches would have been based on the assumption that GABAergic and dopaminergic mechanisms were central to the disease. Yet the culprit is a protein that is expressed throughout the body, is not even enriched in the striatum, and is not directly involved in neurotransmission.

Third, the path that leads from the identification of a disease-related gene to definitive treatment can be long and arduous. Close to 20 years after identification of the gene, little is known about the normal product of the gene and we are just beginning to understand how the mutant product generates disease. Definitive treatment of Huntington disease and preventive measures await our ability to discover and exploit such information.

Finally, it should be emphasized that Huntington disease is caused by a single genetic mutation and characterized by a straightforward diagnosis, relatively uncomplicated familial transmission, and definitive pathologic findings. The process of determining the mechanisms that underlie other neuropsychiatric disorders and of developing pathophysiologically based treatments for the disorders, particularly those that represent heterogeneous, genetically complex syndromes, is likely to be far more challenging.

Also known as Lou Gehrig disease in the United States, ALS is a motor neuron disease that generally occurs in individuals between 30 and 60 years of age. It is characterized by the degeneration of upper and lower motor neurons in both the corticospinal and corticobulbar tracts 18–13. The distribution of resulting deficits in movement can be traced to predominant involvement of either the limbs or cranial nerves, and the predominant involvement of either upper or lower motor neurons, and can be used to distinguish the clinical variations of this disorder. In approximately 20% of patients, for example, the disease is limited to weakness of the bulbar muscles; bulbar involvement is characterized by difficulty in swallowing, chewing, coughing, breathing, and speaking. ALS is progressive and typically results in a fatal outcome within 3 to 5 years. Patients with bulbar involvement generally are given a poorer prognosis because of breathing and swallowing difficulties and associated aspiration and pulmonary infections. Motor neuron disease in children, such as Werdnig–Hoffmann disease (infantile spinal muscular atrophy), typically presents early in life and is characterized by impaired swallowing or sucking and muscle wasting of the limbs.

Most cases of ALS are sporadic; only 10% to 20% are characterized by familial transmission. The first identified hereditary cause of ALS, accounting for ~10% of familial cases (and a very small percentage of all cases), is mutations in the gene for the enzyme Cu/Zn superoxide dismutase (SOD), which is involved in removing reactive oxygen species from cells. Accordingly it was believed that mutations in SOD lead to a loss of its function, which in turn increases oxidative stress on cells. However, further investigation indicates a more complicated situation, because knockout mice that lack SOD do not develop signs of ALS, some disease-causing mutations in the protein do not result in reduced enzyme activity, and transgenic mice that overexpress such disease-causing mutant forms of SOD exhibit a loss of motor neurons. The pathogenesis of mutant SOD-related ALS appears to involve aggregation of the SOD protein in affected neurons.

Since the discovery of SOD 20 years ago, the list of genes associated with ALS has grown to well over 20. Recently, ALS has joined the list of other neurodegenerative disorders marked by protein aggregate formation. Certain familial cases of ALS are associated with a hexanucleotide expansion in the chromosome 9 open reading frame 72 gene (c9orf72). As with all of the other known ALS genes, the precise function of c9orf72 and its encoded protein, and how the mutation causes disease, remain areas of intense investigation. In contrast to the other identified ALS genes, the c9orf72 mutation has garnered much attention because of the finding that it is the most common form of hereditary ALS in several large population studies and may explain a significant percentage of sporadic ALS cases as well. Of note, recent studies show that patients with c9orf72 mutations develop, within the CNS, aggregates of TDP-43 as well as aggregates of the peptide repeat encoded by the mutant forms of c9orf72. The role of these aggregates in disease pathogenesis remains to be elucidated.

Until recently, treatment of ALS has been symptomatic. There are no treatments that halt motor neuron loss; however, the one medication currently approved by the US Food and Drug Administration for the treatment of ALS, riluzole, has been shown to prolong survival by approximately 3 months. Riluzole is believed to exert ameliorative effects by decreasing synaptic levels of glutamate; however, the mechanism responsible for this effect is uncertain. Riluzole antagonizes voltage-gated Na⁺ channels as well as SK (small K⁺) channels (Chapter 2). The therapeutic actions of riluzole are consistent with the still unproven hypothesis that ALS may be caused in part by glutamate-mediated excitotoxicity. The recent reports that sporadic cases of ALS might be associated with increased expression of unedited forms of the AMPA glutamate receptor subunit GluA2, which would be expected to increase Ca²⁺ flux into neurons, further support this hypothesis (Chapter 5). Other treatments that have shown some ability to increase survival time in ALS animal models, such as anti-inflammatory agents, several antibiotics, and lithium, have yet to show significant benefit or failed in human clinical trials. There is hope that multidrug combinations will prove to be more effective.