Huntingtin protein is degraded to small fragments by calpain after ischemic injury. The huntingtin fragments generated from calpain cleavage are smaller than those from caspase cleavage and are more toxic.

2.2.2. Cathepsins

Cathepsins, which are two classes of lysosomal proteases including aspartyl (cathepsin D) and cysteine (cathepsin B, H and L) proteases, play key roles in neurodegeneration. Cathepsins have been implicated in both intracellular proteolysis and extracellular matrix remodeling. Dysregulation or absence of cathepsins has important consequences on the maintenance and function of the nervous system. Mice deficient in cathepsin B and L die 2 to 4 weeks after birth and display neuronal loss and brain atrophy. Cathepsin D deficiency induces a lysosomal storage disease in mouse central nervous system neurons and degeneration of neurons in the mouse retina.

In models of neurological disorders, cathepsins B and L have been implicated in delayed neuronal death after global and focal cerebral ischemia. Specific inhibitors of cathepsins B and L effectively reduce ischemia cerebral damage. Cathepsin B release is an early event after occlusion of cerebral arteries, which eventually triggers the activation of proinflammatory caspases, caspase-1 and caspase-11, in focal cerebral ischemia. Cathepsin D is involved in neuronal death induced by aging, transient forebrain ischemia, and excessive stimulation of glutamate receptors during excitotoxicity. Lysosome numbers and the concentration of cathepsin D increase in neurons that are vulnerable to AD before the onset of pathology.

What signals trigger the activation of multiple proteases that lead to demise of neurons in both acute and chronic neurodegenerative diseases? Because an increase in intracellular calcium and abnormal activation of calcium-regulated processes are among the most ubiquitous features in neurodegeneration, calpain activation represents a critical step in both apoptosis and necrosis. Mild calcium elevation favors apoptosis, whereas acute calpain activation precipitates necrosis, probably via catastrophic cleavage of regulatory and structure proteins. Studies in primates also show that calpains localize to lysosomal membranes after the onset of ischemic episodes, with subsequent spillage of cathepsins to the cytoplasm to execute necrosis. Protease activation in neurodegeneration is very complex and is likely to involve cross-talks of caspases, calpains, and cathepsins in a manner depending on the neuronal population and the nature or severity of the insult.

3. CONCLUSIONS

In this chapter, we discussed the roles of the key components of the apoptosis program in neuronal development, Apaf1, Bcl-2 family proteins, and caspases. The role and mechanism by which neurotrophins suppress apoptosis and regulate cell survival signaling are well appreciated. Evidence has been presented that supports the involvement of activation of apoptosis in chronic neurodegenerative diseases such as AD, PD, HD, and ALS. However, it is important to understand that such evidence is by no means conclusive; other cell death mechanisms such as necrosis and other deleterious mechanisms may also contribute to the degenerative process. Furthermore, one must also understand that although neuronal cell death plays a very important role in the progression and in the final stages of these universally lethal diseases, it might not be the primary or sole factor that determines the onset of these chronic neurodegenerative diseases. In many neurodegenerative diseases, axonal dysfunction long precedes neuronal death. The molecular mechanisms underlying axonal damage are distinct from those underlying cell death. Additionally, an early loss of synapses has been observed in various animal models of developmental neurodegeneration. Therefore, targeting cell death alone is unlikely to be sufficient to obtain a beneficial therapeutic effect. Optimal neuroprotection might require administration of a pharmacological cocktail against multiple pathogenic events, including axonal/synaptic dysfunction, inflammation, and cell death.

ACKNOWLEDGMENT

We would like to thank Prof. Ronald W. Oppenheim of Wake Forest University School of Medicine for reading and valuable comments on the draft of this chapter.

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1. INTRODUCTION: PROGRAMMED CELL DEATH, CELL

DEATH SIGNALING, AND NEURODEGENERATIVE DISEASE

Many of the diseases that affect the nervous system feature an abnormality of cell death of one sort or another. For example, developmental and neoplastic disorders of the nervous system feature dysregulation of the intrinsic cellular programs that mediate cell death. Furthermore, there is increasing evidence to suggest that such dysregulation may also occur in neurodegenerative, infectious, traumatic, ischemic, metabolic, and demyelinating disorders. Therefore, targeting the central biochemical controls of cell survival and death may represent a productive therapeutic approach, especially if combined with other therapeutic strategies. Furthermore, recent results from stem cell studies suggest that the fate of neural stem cells may also play an important role in disease outcomes, and therefore, cell death apparently plays a central role in many neurological diseases and potentially in their prevention and treatment.

Early studies of neuronal survival focused on the status of external factors such as glucose availability, pH, and the partial pressure of oxygen. However, although these are clearly critical determinants, research over the past few decades has revealed a more active, and more plastic, role for the cell in its own decision to survive or die than was previously appreciated. Complementing this concept, studies of the internal suicide programs of neural cells have offered new potential targets for therapeutic development.

In neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, frontotemporal dementia, Huntington’s disease, and amyotrophic lateral sclerosis (Lou Gehrig’s disease), neurons in various brain or spinal cord nuclei are lost in disease-specific distributions. However, the neuronal loss is a relatively late

event, typically following synaptic dysfunction, synaptic loss, neurite retraction, and the appearance of other abnormalities such as axonal transport defects. This progression argues that cell death programs may play at best only a secondary role in the neurodegenerative process. However, emerging evidence from numerous laboratories has suggested an alternative possibility: that although cell death itself occurs late in the degenerative process, the pathways involved in cell death signaling do indeed play critical roles in neurodegeneration, both in sub-apoptotic events such as synapse loss and in the ultimate neuronal loss itself.

Although initial comparisons of the intrinsic suicide program in genetically tractable organisms such as the nematode Caenorhabditis elegans failed to disclose obvious relationships to genes associated with human neurodegenerative diseases (e.g., presenilin-1 does not bear an obvious relationship to the major cell death genes ced-3, ced-4, or ced-9 in C. elegans), more recent studies support such a relationship. For example, the mammalian homologs of ced-3 comprise a family of cell death proteases, the caspases, and mutation of a single caspase cleavage site in huntingtin blocks the development of the Huntington’s phenotype in transgenic mice. A detailed understanding of the interrelationship between fundamental cell death programs and neurodegenerative processes is still evolving, and it promises to offer novel approaches to the treatment of these diseases.

2. MECHANISTIC TAXONOMY OF CELL DEATH: HOW

MANY TYPES OF PROGRAMMED CELL DEATH CAN

BE DISTINGUISHED?

Classical developmental studies support the view that at least three different programmed cell death (PCD) forms are distinguishable: type I, also called nuclear

or apoptotic; type II, also called autophagic; and type III, also called cytoplasmic. These occur reproducibly within specific neuroanatomical nuclei and with specific frequencies, at specific times of nervous system development. However, these developmental or physiologic cell death pathways may also be activated by various insults, such as ischemia, DNA damage, or the accumulation of misfolded proteins. Mechanistic requirements for type I cell death center on caspase-dependent pathways (extrinsic and intrinsic), though some have argued that cellular morphologies resembling apoptosis can occur independent of these proteases. Types II and III do not require caspase activation, but the possibility that they may in some cases be accompanied by caspase activation has not been excluded.

Beyond the three types of developmental cell death, other forms have been described that do not fit the criteria for any of them. For example, a nonapoptotic, caspase-independent form of cell death that does not resemble type II or type III developmental PCD has been described by Driscoll and colleagues in C. elegans that express mutant channel proteins, such as mec-4(d), that mediate neurodegeneration. A uniform, necrosislike cell death ensues, characterized morphologically by membranous whorls lacking in other cell death types, triggered by calcium entry, mediated by specific calpains and cathepsins, and inhibited by calreticulin. Although it is possible that this alternative form of PCD will ultimately turn out to proceed via one of the previously described pathways (e.g., type II or type III), the morphological characteristics suggest that it may be a distinct form of nonapoptotic PCD.

A fifth apparent form of PCD has been described by Yu, the Dawsons, and their colleagues, who showed that a nonapoptotic form of cell death depends on the activation of poly-(ADP-ribose) polymerase (PARP) and the consequent translocation of apoptosis-inducing factor (AIF) from mitochondria to nucleus. AIF is a flavoprotein, discovered by Kroemer and his colleagues, that is involved with DNA fragmentation, along with endonuclease G and DNA fragmentation factor. This form of PCD was shown to be activated by agents that induce DNA damage, such as hydrogen peroxide, N-methyl-D- aspartate, and N-methyl-N -nitro-N-nitrosoguanidine. Just as in the case of the calcium-activated PCD referred to previously, PARP-dependent PCD displays a morphology and biochemistry that appear to be distinct from types I, II, and III PCD.

As additional data are gathered from other cell death paradigms, novel biochemical pathways of PCD may be characterized. For example, an extensive literature on the morphological criteria for another potential form of

PCD – oncosis – exists, but the biochemical mediators of oncosis have not yet been described. Oncosis refers to a specific morphology of cell death – cellular swelling – typically induced by ischemia and thought to be mediated by the failure of plasma membrane ionic pumps. One potential mediator of oncosis is a calpain-family protease (possibly a mitochondrial calpain). This finding suggests that oncosis may prove to be similar to the calcium-activated necrosis-like cell death described by Driscoll et al. Another potential PCD pathway has been referred to as autoschizis, a form of cell death shown to be activated in certain tumor cells after treatment with ascorbate and menadione. Altogether, these observations illustrate the difficulty of relying on cell morphology to define cell death mechanisms and suggest a wide diversity of possibilities.

3. PROGRAMMED CELL DEATH SIGNALING IN

NEURODEGENERATION

Evidence for caspase activation in neurodegeneration has been derived both from the use of antibodies directed against newly exposed proteolysis-dependent epitopes (neo-epitopes) generated by caspase cleavage and from the inhibition of neurodegeneration by caspase inhibitors. However, some neurodegenerative models and diseases clearly demonstrate nonapoptotic forms of PCD as well. Determining which PCD pathways are triggered in each neurodegenerative disease, which pathway accounts for each fraction of cell death, the mechanism(s) by which each pathway is triggered, and the interactions of the various pathways should shed new light on the degenerative process and its potential treatment or prevention.

One of the critical goals for dissecting the relationship between PCD and neurodegeneration is to determine the specificity of the trigger: in other words, is PCD activated in neurodegeneration as the result of a relatively nonspecific toxic effect of a peptide or protein aggregate? If so, then secondary neurodegeneration may occur due to loss of trophic support, excitotoxicity, or any number of other secondary effects. Alternatively, by analogy to neoplasia, are specific, physiologically relevant transduction events that underlie neurite retraction and synapse loss triggered directly by neurodegeneration-associated transcriptional and posttranscriptional events? In other words, if neoplasia is the result of an imbalance in physiological signaling events involving oncogenes and tumor suppressor genes, is neurodegeneration an analogous process that is the manifestation of an imbalance in physiologic signals that mediate synaptic maintenance and synaptic

re-organization? Evidence on both sides exists: for example, numerous toxic properties have been attributed to the Aβ peptide implicated in Alzheimer’s disease, such as reactive oxygen species generation and metal binding, among others. However, signal transduction effects have also been attributed to Aβ peptide, such as binding and multimerization of amyloid precursor protein, with resultant complex formation and direct caspase activation.

Because the neurodegenerative process may be induced by widely varying insults – from misfolded proteins to reactive oxygen species to caspase recruitment complexes, as well as other mechanisms – and yet produce a relatively small number of syndromes, the existence of a death network is suggested. Such a network may be entered from many different sites, but once triggered, would follow similar interdependent biochemical pathways, with little dependence on the point of entry. This notion is compatible with the findings that therapeutics aimed at different pathways (caspase activation, mitochondrial release of cytochrome c, metal binding, reactive oxygen species scavenging, etc.) all have partially salutary effects. However, it also suggests that a complete halt of the neurodegenerative process may require therapeutics that address all of the network’s interacting pathways.

4. APOPTOSIS INDUCED BY MISFOLDED, UNFOLDED, OR

ALTERNATIVELY FOLDED PROTEINS

One of the features common to all of the major neurodegenerative diseases is the accumulation of misfolded, unfolded, or alternatively folded proteins (Rao and Bredesen, 2004). These proteins may be the result of many different processes, such as impaired ubiquitinmediated protein degradation, impaired chaperonemediated autophagy or other autophagic degradative pathways, or expression of mutant proteins that may aggregate and resist proteolytic degradation. Misfolded proteins and other activators of endoplasmic reticulum (ER) stress trigger an alternative intrinsic pathway of apoptosis (Figures 12-1 and 12-2) that leads to caspase- 9 activation and displays both cytochrome c/Apaf-1– independent and cytochrome c/Apaf-1–dependent activation of PCD. Cell death pathways triggered by protein misfolding, unfolding, or alternative folding and associated ER stress are of special interest in neurodegenerative disease studies, for the reason noted previously. Neurodegenerative disorders such as Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, and prion protein diseases all share the common feature of ER stress. The presence of

misfolded proteins elicits cellular stress responses that include an ER stress response that serves to protect cells against the toxic accumulation of misfolded proteins and is activated by the exposure of hydrophobic protein regions that bind GRP78/BiP (glucose-regulated protein of 78 kilodaltons/binding protein), relieving its otherwise ongoing inhibition of unfolded protein response–activating proteins PERK (protein kinase R- like endoplasmic reticulum kinase), ATF6 (activating transcription factor 6), and Ire1 (inositol-requiring 1) (Figures 12-1 and 12-2). Accumulation of misfolded proteins in excessive amounts, however, overwhelms the cellular protective response that induces folding, translational, degradative, and aggresomal protection, ultimately triggering cellular suicide pathways.

Because the degradation of cellular proteins is coupled, via the ubiquitin-mediated proteasomal degradation pathway, to ER dislocation (translocating the protein targeted for degradation back out of the ER into the cytosol), any conditions that block the ER retrotranslocation of proteins or proteasome function may also result in the accumulation of misfolded protein substrates within the ER. Therefore, misfolded proteins both within and outside the ER may trigger the ER stress response. Misfolded proteins typically aggregate, initially as oligomers but ultimately as polymers that are deposited as microscopically visible inclusion bodies or plaques within cells or in extracellular spaces. These aggregates may interact with cellular targets, with several potential effects: (1) inhibition of synaptic function,

(2) loss of synapses, (3) sequestration of cellular chaperones and transcription factors, (4) interference with signal transduction pathways, (5) disruption of calcium homeostasis, (6) release of free radicals, (7) dysfunction of the protein degradation pathways, and (8) induction of cell-death proteases. Despite these mechanisms, it is becoming increasingly clear that toxicity may be exerted before the appearance of aggregates, for example, with the production of small oligomers.

Mediators of PCD induced by misfolded or unfolded proteins have been identified (Figures 12-1 and 12-2). As in the classical intrinsic pathway, the Bcl-2 family proteins play a critical role in the cellular suicide decision process, communicating between the ER and the mitochondria. Bax/Bak double knockout cells fail to activate caspases after ER stress, arguing that these are required mediators. Bik may function to activate Bax and Bak in this pathway, whereas BI-1 binds to Ire1, suppressing Bax activation and translocation to the ER. Other Bcl-2 family proteins are also involved: for example, the BH3 protein Puma interacts with an hsp90 (heat shock protein of 90 kilodaltons)–independent fraction of p23, which,

when cleaved by caspases, releases Puma, leading to Bax interaction, oligomerization, and PCD. Noxa and p53 have also been implicated in this pathway.

Given the mitochondrial-ER interplay, it is not surprising that part of the resulting apoptotic pathway is Apaf-1–dependent. However, part is also Apaf-1– independent yet caspase-9–dependent. Caspase-7 is recruited to the ER by an unknown mechanism, where it interacts with caspase-12 (in the murine system; most humans do not express caspase-12, and in humans, caspase-4 may play this role); caspase-12 is cleaved and released, leading to interaction with caspase-9. Of note, murine caspase-12 lacks protease activity and plays a

predominant role as a dominant-negative inhibitor of caspase-1. GRP78/BiP interacts with caspase-7 (requiring the adenosine triphosphate [ATP]–binding domain) and -12, preventing activation, but this inhibition is relieved by (d)ATP. Although the upstream activation of this pathway is not certain, one candidate is the triggering of c-Jun N-terminal kinase activation by IRE1, via TRAF2 and ASK1.

There is also a caspase-8–dependent pathway that is activated in response to misfolded proteins: Bap31, an ER membrane protein, binds Bcl-2 (or Bcl-xL) and a proapoptotic complex that includes caspase-8. After Bap31 cleavage, a proapoptotic p20 fragment is derived,

Figure 12-2. Proteins implicated in ER stress-pcd pathways. Reproduced from Bredesen DE, Rao RV, Mehlen P. Cell death in the nervous system. Nature. Oct 19 2006;443(7113):796–802, with permission. See Color Plate 13.

which, among other effects, induces mitochondrial fission, enhancing cytochrome c release. Conversely, BAR (bifunctional apoptosis regulator), which is expressed primarily in neurons of the CNS, also bridges Bcl-2 and caspase-8 but functions as an antiapoptotic protein.

Other mediators of ER stress-induced PCD have been identified, and these are depicted in (Figures 12-1 and 12-2). One of interest in neurodegeneration is valosincontaining protein (VCP), which functions as both a sensor of abnormally folded proteins and a cell death effector in polyglutamine-induced cell death, as well as being a mediator of ER stress-induced PCD. Mutations in VCP are associated with a syndrome of inclusion body myositis, frontotemporal dementia, and Paget’s disease of bone. Another ER stress mediator is Alix/AIP-1, which is an ALG-2–interacting protein that links developmental motor neuron cell death, as well as neuronal death in a Huntington’s model, to the endolysosomal system.

In addition to protein misfolding, results from Lingappa and colleagues suggest that at least some proteins may trigger PCD via a subset of multiple physiologically relevant conformations. For example, the prion protein exists in three different topologies: a secreted form, a transmembrane form with N-terminus extracellular, and a transmembrane form with C-terminus extracellular (Ctm). The Ctm form is proapoptotic and associated with neurodegeneration in vivo, whereas the secreted form is antiapoptotic. It is not yet clear how many proteins will display this feature, but it is possible that not only prion protein, but also other neurodegenerationassociated proteins will turn out to exist in multiple physiologically relevant conformations (and perhaps even topologies), with the degenerative effect being

attributed to a subset of these conformations rather than true protein misfolding.

5. TROPHIC FACTORS AND CELLULAR DEPENDENCE

IN NEURODEGENERATIVE DISEASE

Neurons, as well as other cells, depend for their survival on stimulation that is mediated by various receptors and sensors, and PCD may be induced in response to the withdrawal of trophic factors, hormonal support, electrical activity, extracellular matrix support, or other trophic stimuli. For years it was generally assumed that cells dying as a result of the withdrawal of required stimuli did so because of the loss of a positive survival signal, for example, mediated by receptor tyrosine kinases. Although such positive survival signals are clearly extremely important, data obtained over the past 15 years argue for a distinct and complementary effect that is proapoptotic, activated or propagated by trophic stimulus withdrawal, and mediated by specific receptors dubbed dependence receptors (Rabizadeh et al., 1993; Bredesen et al., 2004). More than a dozen such receptors have now been identified, and examples include DCC (deleted in colorectal cancer), Unc5H2 (uncoordinated gene 5 homolog 2), Neogenin, RET (rearranged during transfection), Ptc (Patched), and β-amyloid precursor protein (APP). These receptors interact in their intracytoplasmic domains with caspases, including apical caspases such as caspase-9, and may therefore serve as sites of induced proximity and activation of these caspases. Caspase activation leads in turn to receptor cleavage, producing proapoptotic fragments; conversely, mutation of the caspase cleavage sites of dependence receptors suppresses PCD mediated by the receptors. A striking example of this effect was obtained in studies of neural tube development: withdrawal of Sonic hedgehog from the developing chick spinal cord led to apoptosis mediated by its receptor, Patched, preventing spinal cord development; however, transfection of a caspaseuncleavable mutant of Patched blocked apoptosis and restored significant development, even in the absence of Sonic hedgehog. Recently, a caspase activity complex has been reported that associates with Patched, components of which include adapter proteins DRAL and CARD9 (TUCAN/Cardinal) and pro-caspase-9.

Thus cellular dependence on specific signals for survival is mediated, at least in part, by specific dependence receptors that induce apoptosis in the absence of the required stimulus (when unoccupied by a trophic ligand, or when bound by a competing, anti-trophic ligand), but block apoptosis after binding to their respective ligands. Expression of these dependence receptors thus

creates cellular states of dependence on the associated trophic ligands. The states of dependence are not absolute, because they can be blocked downstream in some cases by the expression of antiapoptotic genes such as Bcl-2 or p35; however, they result in a shift of the apostat toward an increased likelihood of triggering apoptosis. In the aggregate, these receptors may serve as a molecular integration system for trophic signals, analogous to the electrical integration system afforded by the dendritic arbors within the nervous system.

Cellular dependence on trophic signals was originally described in the developing nervous system, but does this phenomenon have anything to do with neurodegenerative disease? APP exhibits several features characteristic of dependence receptors: an intracytoplasmic caspase cleavage site (Asp664), co-immunoprecipitation with an apical caspase (caspase-8), caspase activation, derivative proapoptotic peptides (see below, this section), and suppression of apoptosis induction by mutation of the caspase cleavage site.

These findings raise several questions: first, does the caspase cleavage of APP occur in the human brain, and, if so, is this increased in patients with Alzheimer’s disease? Second, if this cleavage is prevented, is the Alzheimer’s phenotype affected? Third, is there a physiologic role for this cleavage event?

Neo-epitope antibodies directed against residues 657–664 of human APP disclosed the presence of caspase-cleaved APP fragments in human brain, especially in the hippocampal region. There was an approximately four-fold increase in Alzheimer’s patients over age-matched controls. However, in brains without Alzheimer’s pathology, there was an inverse relationship between age and immunohistochemical detection of APPneo, and the distribution was different from that of Alzheimer’s disease brains. Whereas in the Alzheimer’s brains the distribution was primarily in somata, in the non-Alzheimer’s brains, the distribution was primarily in the processes. These findings suggest that the caspase cleavage of APP occurs physiologically and is reduced with age but is somehow increased in association with Alzheimer’s disease.

The effect of preventing the caspase cleavage of APP on the Alzheimer’s phenotype was evaluated in Alzheimer’s disease model transgenic mice that express APP with Swedish and Indiana mutations that are associated with familial Alzheimer’s disease. Although the caspase mutation (D664A) had no effect on plaque formation or on the production of Aβ peptides 1– 40 or 1–42, the mutation prevented the synapse loss, dentate gyral atrophy, electrophysiological abnormalities (reduction in excitatory postsynaptic potentials

and long-term potentiation), neophobia, and memory deficits that characterize Alzheimer’s model mice. These findings indicate that key features of the Alzheimer’s phenotype, at least in a standard transgenic mouse model, depend on the presence of the caspase cleavage site within APP. Yet extensive previous work has shown that the phenotype is also dependent on Aβ itself, suggesting that the APP caspase site may lie downstream from the Aβ accumulation. This possibility has received support from studies showing that Aβ interacts directly with APP in the Aβ region itself, leading to multimerization, caspase cleavage, and cell death signaling.

If APP does indeed function as a dependence receptor and Alzheimer’s disease is a “state of altered dependence,” then what is/are the trophic ligand(s) for APP? Several candidate APP interactors have been described, such as collagen (types I and IV), heparan sulfate proteoglycan, laminin, glypican, and F-spondin. In the case of F-spondin, β-secretase activity is reduced. Lourenco et al. (2009) have recently shown that netrin-1, a multifunctional axon guidance and trophic factor, also binds APP. Furthermore, netrin-1 also interacts with Aβ itself, and thus Aβ may interfere with netrin-1 binding to APP. The binding of netrin-1 to APP results in enhanced interaction of APP with Fe65 and Dab, upregulation of KAI1, and a marked reduction of net Aβ production (Lourenco et al., 2009).

These findings suggest a model in which the Aβ peptide functions as an anti-trophin, blocking netrin’s guidance and trophic effects, binding and oligomerizing APP, recruiting and activating caspase-8, engendering the processing of APP at Asp664, and inducing neurite retraction, and, ultimately, neuronal cell death. An alternative possibility, however, is that the D664A mutation altered a protein–protein interaction critical to AD pathogenesis in the mouse model and that caspase cleavage is not critical. In either case, however, the results suggest that APP signal transduction may be important in mediating Alzheimer’s disease, at least in the transgenic mouse model, possibly downstream from Aβ oligomerization and binding of APP.

The results obtained in the transgenic mouse model of AD also suggest an alternative to the classic models of AD. Chemical and physical properties of Aβ have been cited as the proximate cause of AD pathophysiology: reactive oxygen species generation involving Aβ itself, metal binding by Aβ, and direct membrane damage, among others (Butterfield and Bush, 2004). These theories do not explain why Aβ is produced ubiquitously and constitutively, nor do they offer a physiologic function for the Aβ peptide. They also fail to account for