alzheimer 's disease ( ad ) is the most prevalent form of dementia , which affects more than 37 million people worldwide , with an estimated cost of $ 422 billion in 2009 [ 1 , 2 ] .
moreover , the incidence of the illness and the prospect of an aging population will result in rising social and economic demands
.   ad is characterised by the presence of extracellular amyloid plaques and intracellular neurofibrillary tangles within the afflicted brain , which cause neuronal loss in the neocortex , hippocampus , and basal forebrain , leading to progressive cognitive and behavioural decline .  
zinc , in addition to copper and iron , has been shown to be involved in ad . here
, we review the current literature relating to neuronal zinc metabolism and the way in which zinc can modulate normal brain activity . we discuss also the contribution of zinc to the formation , aggregation , and degradation of the amyloid- ( a ) peptide and the contribution of zinc to the pathogenesis of ad .
as the most abundant trace metal in the brain , zinc is found tightly associated with numerous proteins conferring either structural or catalytic properties upon them . however , a significant amount of loosely bound , chelatable zinc can be found sequestered in presynaptic vesicles forming a subpopulation of  zinc enriched 
( zen ) neurones [ 5 , 6 ] , which co - release zinc with the neurotransmitter glutamate upon excitation .
neurones   have their cell bodies located in either the cerebral cortex or the limbic structures of the forebrain , and so an extensive network uniting limbic and cerebrocortical functions is created .
this connection between zinc and glutamatergic neurotransmission allows the ion to modulate the overall excitability of the brain and also influence synaptic plasticity .
the identification of synaptic zinc was first made by mclardy over fifty years ago who identified that a band of zinc dithizonate staining correlated with hippocampal mossy fibre axons . since then
, many more cerebrocortical pathways have been identified which contain zinc - rich synaptic vesicles ; indeed nearly 50% of the glutamatergic synapses are actually  gluzincergic  in some parts of the cerebral cortex .
significantly , only small amounts of chelatable zinc can be determined in glutamatergic pathways which originate outside the cerebral cortex or limbic systems . despite this extensive network of zinc - containing neurons ,
there are two families of zinc transporters : the znt family , which act to decrease intracellular zinc concentrations by exporting zinc from the cytoplasm to the lumen of organelles or the extracellular space , and the zip family , which import the metal from the extracellular space or organellar lumen into the cytoplasm .
whilst many of the transporters have particular distribution patterns , only znt3 expression is restricted to the brain and the testis .
it is located in the vesicular membrane   and is necessary to transport zinc from the cytoplasm into the synaptic vesicle of the neuron .
targeted disruption of znt3 in a mouse model resulted in a complete lack of chelatable zinc .
a number of approaches have been taken to confirm that zinc is coreleased with glutamate from the presynaptic bouton during neuronal excitation : imaging of zinc in boutons before and after stimulation , analytical detection of zinc released into perfusates , and direct imaging of released zinc using fluorescent extracellular probes [ 18 , 19 ] .
stimulation of organotypic cultures from rat hippocampus produced a cloud of green fluorescence as the released zinc bound an apometalloenzyme confirming the release of zinc from the culture .  
a later study by quinta - ferreira and colleagues   demonstrated a release of zinc with each pulse of an action potential . whilst there is now no doubt that zinc is released during synaptic activity
, there is little consensus on the amount or duration of its existence in the synaptic cleft . following an intense burst of neuronal activity ,
the release of glutamate and postsynaptic membrane depolarisation open a variety of zinc - permeable ion channels which contribute to removing the ion from the extracellular fluid .
these include n - methyl - d - aspartate ( nmda ) channels and calcium permeable -amino-3-hydroxy-5-methyl-4-isoxazole propionic acid ( ampa)/kainate channels .
the consequences of zinc acting on these receptors are diverse and demonstrate the significance zinc has in modulating fast excitatory glutamatergic transmission .
zinc can act to either enhance or depress synaptic activity with varying degrees of potency .
initially , zinc was thought to selectively inhibit nmdar - mediated neuronal activity by inducing a voltage - independent noncompetitive inhibition that decreased the probability of the channel being open [ 2224 ] .
a voltage - dependent inhibition of nmdar could be observed at higher concentrations of zinc ( > 20  m ) and was believed to be due to binding of the cation within the channel pore . with the cloning of nmdar subunits
, it was confirmed that zinc could cause both voltage - independent and voltage - dependent inhibition .
the exceptional sensitivity of the glun2a subunit towards zinc suggests that even contaminating ions found in routine laboratory solutions are sufficient to cause inhibition .
significantly , despite being responsible for inhibitory effects at nmdar , it has also been shown that nmdar activation may provide a route of influx for zinc contributing to toxic effects of exposure   ( figure 1 ) .
whilst it is thought that zinc exposure generally attenuates nmdar - mediated neurotoxicity , zinc has been shown to potentiate ampar - mediated toxicity at suggested physiological concentrations ( 50  m ) .
originally , it was proposed that the toxic effect was due to zinc influx via voltage - gated calcium channels , with lin et al .   
subsequently , it has been demonstrated that zinc can carry currents directly via ampar , mostly via the calcium - permeable subtype   ( figure 1 ) . at high supraphysiological doses ( 1  mm )
a few studies have also looked directly at zinc - mediated inhibition of voltage - gated calcium channels , as their proximity to vesicular release sites on the presynaptic membrane suggests they could interact [ 33 , 34 ] .
most recently , neurotoxicity has been attributed to transient receptor potential metastatin 7 ( trpm7 ) channel activation resulting in increased intracellular zinc   ( figure 1 ) .
the significance of synaptically released zinc centres on the amount that is released into the synaptic cleft upon excitation .
some authors argue that the amount of zinc ( 10100  m ) released following an action potential arriving at the glutamatergic bouton is high enough to bring about the voltage - dependent inhibition of nmdar and that there would be no spillover onto neighbouring cells [ 36 , 37 ] .
other authors disagree , suggesting that the zinc concentration would be sufficient to affect nearby cells .
it has also been demonstrated that the zinc level ( low nm ) is only sufficient to block the voltage - independent component of nmdar activity . alternatively
, it could be that there is little or no diffusible zinc released , supporting the notion that zinc behaves in a  tonic  mode .
kay and tth   proposed that zinc is exocytosed from the presynaptic membrane and that instead of diffusing into the extracellular space it remains tightly bound to an as - yet unidentified presynaptic component .
this would create a  veneer    of zinc ions which would build up with synaptic activity or erode with quiescence to modulate plasticity .
first ,  synaptic zinc  acts as a conventional neurotransmitter , is contained within presynaptic vesicles , and is released upon excitation and binds to a variety of receptors on the postsynaptic membrane .
the downstream consequence of receptor binding is one of tonic modulation of glutamatergic excitatory synapses .
the second type is similar to calcium signalling and occurs in conjunction with synaptic zinc signalling . a transmembrane flux of  synaptic zinc  from the extracellular space passes through post - synaptic zinc - permeable ion channels .
this class is difficult to define and as yet has not been identified in neurons but has been demonstrated in mast cells .
it is found in synaptic vesicles , released upon excitation , and has multiple synaptic targets .
the significant inhibitory effect of zinc on nmdar , alongside the crucial function of nmdar in both neurophysiology and pathophysiology , advocates a vital role for zinc in both healthy and diseased brains .
the most prominent lesions in the brain of ad sufferers are the amyloid or  senile  plaques , which predominantly consist of a peptides derived from the proteolytic processing of the amyloid precursor protein ( app ) .
app is an ubiquitously expressed glycosylated transmembrane protein with a large n - terminal extracellular domain , a single hydrophobic transmembrane domain , and a small c - terminal cytoplasmic domain .  
a specific and saturable binding site for zinc ( kd = 750  nm ) has been reported in the cysteine - rich region on the ectodomain of app [ 44 , 45 ] .
it is hypothesised that zinc could have a role in sustaining the adhesiveness of app during cell - cell and cell - matrix interactions [ 46 , 47 ] .
app can be processed by one of two pathways : the amyloidogenic pathway , leading to the production of a , and the non - amyloidogenic pathway ( figure 2(a ) ) , reviewed in . in the amyloidogenic pathway
, app is sequentially cleaved by the aspartyl protease , -site app - cleaving enzyme 1 ( bace1 ) forming the secreted app ( sapp ) fragment and a membrane bound c - terminal fragment of 99 amino acids ( c99 ) .
the c99 fragment is then further processed by the -secretase complex into app intracellular domain ( aicd ) and a peptides , predominantly 40 and 42 amino acids in length .  
it is these aggregation - prone a peptides which form oligomeric and fibrillar structures which deposit in the brain and over time cause ad .  
the -secretase complex comprises four components : presenilin ( ps ) 1 or 2 , nicastrin ( nct ) , presenilin enhancer 2 ( pen-2 ) , and anterior pharynx defective-1 ( aph-1 ) .
sapp is involved in the pruning of synapses during the development of central and peripheral neurons   and aicd is known to be a transcription factor for several genes , including the upregulation of the a-degrading enzyme , neprilysin ( nep ) .
the predominant app - processing pathway in healthy brain is the nonamyloidogenic pathway where app is cleaved by the -secretase within the a region forming the secreted app ( sapp ) fragment and the membrane - bound c - terminal fragment of 83 amino acids ( c83 ) ( figure 2(a ) ) .
c83 is subsequently cleaved by the -secretase complex generating aicd and p3 ( figure 2(a ) ) .
-secretase activity is attributed to the disintegrin and metalloprotease ( adam ) family of zinc metalloproteases ( reviewed in [ 48 , 52 ] ) .
the adams , along with the matrix metalloproteases ( mmps ) , are members of the metzincin clan of metalloproteases as they have a long zinc - binding consensus sequence hebxhxbgbxh   ( h , histidine , zinc ligand ; e , catalytic glutamate ; g , glycine ; b , bulky apolar residue ; x , any amino acid ) which contains three zinc ligands . structurally , the catalytic domain is globular and divided into two subdomains , with the active site cleft running between the two .
the defining feature of the metzincins is the conserved methionine residue which creates a 1,4--turn ( met - turn ) , creating the catalytic cleft .  
the catalytic zinc atom sits at the bottom of the groove between the subdomains , with the subsites in the groove determining specificity for particular amino acid sequences in the substrate ( reviewed in ) .
a number of enzymes in this family , namely , adam9 , 10 , 17 ( also known as tnf- converting enzyme , tace ) and 19 , are known to exert -secretase activity , but it is unclear which enzyme or enzymes are responsible for the -cleavage of app in vivo [ 5558 ] .
adam10 appears to be the predominant enzyme as overexpression of functional adam10 in ad transgenic mice led to an increase in sapp and reduced a production , plaque deposition , and cognitive deficits .
although fibroblasts from adam10 mice showed no deficiency in -secretase activity , probably due to compensation by adam17 , a recent detailed study has provided strong evidence that adam10 is the physiologically relevant constitutive -secretase in primary neurons .
in a healthy brain , the relatively small amount of a-constitutively being produced is rendered safe by a degrading enzymes .
the steady state levels of a synthesis and clearance in cerebrospinal fluid ( csf ) are 7.6% and 8.3% per hour , respectively .   
a large number of candidate a-degrading enzymes have been identified , with the majority being zinc metalloproteases .
these include nep , insulin - degrading enzyme ( ide ) , endothelin - converting enzyme ( ece ) 1 and 2 , mmp2 , 3 , and 9 , and prep ( reviewed in ) .
nep , ece1 and 2 , and mmps have the conserved zinc - binding motif hexxh ( h , histidine , zinc ligand ; e , catalytic glutamate ; x , any amino acid ) , and ide and its homolog prep are both inverzincins , as they contain the inverted zinc - binding motif hxxeh .
nep appears to be the dominant a protease [ 6366 ] , and is capable of degrading monomeric and oligomeric a [ 63 , 67 ] ( figure 2(b ) ) .
nep knockout mice have significantly elevated a levels [ 68 , 69 ] and more importantly , human postmortem tissue analysis suggests that nep expression is inversely related to the extent of ad pathology [ 7072 ] .
ide is predominantly a cytosolic enzyme but has been found extracellularly   and can only degrade monomeric a and not oligomers ( figure 2(b ) ) , although it did lower the net production of oligomers by cultured cells .
even though there are conflicting data regarding ide and ad , ide knockout mice have elevated levels of endogenous a [ 76 , 77 ] .
there are two isoforms of ece , 1 and 2 , both of which have similar catalytic activity , with ece 1 being the predominant isoform within the brain [ 65 , 78 ] . however , both eces are primarily localised to endothelial cells but have been detected in neurons and glial .
the mmps , specifically mmp2 and mmp9 , degrade fibrillar a   , whereas prep degrades monomeric a. however , prep is confined to mitochondria , so it would therefore have limited action on the majority of a which is extracellular .
high concentrations of zinc ( up to 1  mm ) have also been found within amyloid plaques , which is thought to have been released from glutamatergic synapses .
aggregation of the a peptides can be rapidly induced in the presence of zinc ions under physiological conditions in vitro [ 83 , 84 ] and studies on ad tissues show that zinc colocalised with a deposits .
the metal binding site for zinc on a is the same as the copper binding site and lies within the n - terminal hydrophilic region , the first 16 residues   ( figure 3 ) .
the majority of studies on zinc binding to a have utilised truncated peptides of 16 ( a116 ) and 28 ( a128 ) amino acids , as the binding region is situated within these sequences .  
a116 does not aggregate under moderate conditions [ 8790 ] , making it an ideal model system , and a128 can undergo aggregation and fibril formation , but at significantly reduced rates , to the full - length peptides .  
zinc binds to a in a 1  :  1 stoichiometry with a mononuclear binding site [ 84 , 8690 , 9294 ] .
nmr studies have shown that all three histidine residues ( residues 6 , 13 and 14 ; see figure 3 ) are involved in zinc binding [ 87 , 88 , 90 , 93 , 95 ] , with confirmation from mutational studies [ 96 , 97 ] .  
other possible ligands have also been reported as zinc coordination classically involves four to six ligands .
the candidate ligands are asp1 [ 91 , 93 ] , arg5 , ser 8 , glu11 , and tyr10   ( figure 3 ) however , tyr10 has been ruled out   and arg5 has been deemed highly unlikely .
the carboxyl side chain of glu11 is a zinc ligand [ 88 , 90 ] however , asp1 is considered the most attractive zinc ligand , either through its n - terminal amino group and/or its side - chain carboxylate group [ 87 , 88 , 91 , 93 ] .  
raman spectroscopy has shown zinc binding to the n site of histidine side chains in senile plaques taken from ad brains , but it is unclear if zinc only binds to free a peptides that subsequently aggregate or whether zinc binds to a in preformed plaques .
the reported apparent binding constants ( kd ) of zinc to a peptides vary from 1 to 300  m ( reviewed in ) .
the published kd values vary greatly due to the in vitro conditions ( e.g. , buffer composition , ph ) , a fragment , and experimental method .  
the high kd ( 20300  m ) values come from tyrosine fluorescence experiments however , even these are contentious and hard to reproduce [ 99101 ] , and any change in tyrosine fluorescence could be due to a aggregation rather than zinc binding . discounting the tyrosine fluorescence measurements ,
the most likely apparent kd value for zinc binding to a peptides is a range of 120  m .  
the binding affinity of zinc for preformed a fibrils is approximately the same as of the peptides with a kd of 120  m [ 89 , 101 ] .
it has been shown that there is a significant increase in serum   and hippocampal   zinc in ad patients compared to age - matched controls .
.   demonstrated a significant decrease in csf zinc but could find no difference in serum zinc levels between ad and age - matched controls . a decrease in serum zinc
has also been reported , though it is possible that some of the ad patients included in one study were malnourished [ 104 , 105 ] .
overall , there is currently no consensus on what happens to zinc concentrations in ad subjects though much of the discrepancy could be put down to differences in patient allocation , sample type , postmortem interval , or type of analysis used .  
lovell and coworkers [ 106 , 107 ] have mapped the expression levels of a number of the znt zinc transporters in ad .
znt-1 , 4 , and 6 were all found to show increased expression in early stages of disease , though znt-1 expression was decreased during mild cognitive impairment [ 106 , 107 ] .  
although it is unknown whether increased expression necessarily correlates with increased activity , these changes in transporter level could result in modified subcellular zinc concentrations .
an increase in znt6 would lead to an increase in zinc in the tgn   which could reduce -secretase activity .
it is well established that amyloid plaques contain increased concentrations of copper , iron , and zinc [ 98 , 109 ] . whilst copper and iron appear to be primarily responsible for the toxicity of a via oxidative - stress - type mechanisms [ 109 , 110 ] , zinc has a crucial role in a aggregation which is the most well - established contribution that zinc may have in ad pathogenesis .
whilst the concentration of zinc required for fibrillisation to occur is contentious , with concentrations differing by 100-fold being suggested , zinc is an unequivocal partner in the process [ 83 , 111 , 112 ] . in 2006 ,
dong and co - workers were able to show that zinc could control the rate of self - assembly of the a peptides and go on to regulate the amyloid morphology via specific coordination sites .
furthermore , it has been demonstrated that zinc can spontaneously coordinate both intra - and inter - molecular bridging between two peptides to promote a aggregation   and that synaptic zinc promotes a oligomer formation and their accumulation at excitatory synapses .
studies with synthetic a showed that chelation chemistry could help solubilise amyloid plaques ,   with depletion of zinc having a more marked effect on extracting a than depletion of copper . oral treatment with 5-chloro-7-iodo-8-hydroxyquinoline ( clioquinol cq ) in tg2576 mice resulted in a 49% reduction in cortical amyloid deposition .
although cq has a fairly low affinity ( nm ) for both copper and zinc , it was still able to release the ions from the a binding site . a pilot phase ii trial in humans showed a decrease in cognitive decline and a reduction in plasma a1 - 42 in moderately severe ad compared with placebo control .
it has been suggested that although cq may chelate copper and zinc from metallated a and promote disaggregation , it may not completely holt the aggregation process . a second generation chelator ( pbt-2 ) , with improved blood brain barrier penetration ,
has just completed a phase ii clinical trial in early ad with promising results showing good tolerance , a reduction in csf a and   neuropsychological testing .
recently , it has been shown that zinc can also accelerate the aggregation of a tau peptide under reducing conditions .
zinc inhibited the formation of intramolecular disulphide bonds but promoted intermolecular bonds between key cysteine residues .
furthermore , zinc exposure has been shown to increase the phosphorylation of pi3k and mapk - dependent pathways which are key players in tau modifications .
the essential requirement for znt3 in loading zinc into synaptic vesicles would suggest that this transporter could have a major impact on zinc signalling in the neuron , even regulating cognitive function .
whilst a lack of zinc signalling in brain slices from znt3 mice confirmed the vesicular origin of the released zinc , the mice failed to express a cognitive phenotype .
initial studies detailed a 20% reduction in total zinc level and a loss of histochemically reactive zinc in the synaptic vesicle ; however , there was no impairment of spatial learning , memory , or sensorimotor function .
the implication being that the vesicular zinc is not required for cognitive function or that compensatory mechanisms made up for the deficits .
however , a follow - up study demonstrated marked differences in learning and memory when an older ( 6 month ) cohort of mice was used , suggesting that the lack of effect in the previous study was due to the young age ( 610 weeks ) of the mice and highlights the importance of aging ( the most significant risk factor ) when modelling ad pathology .  
the results obtained from the older znt3 cohort established a requirement for zinc in memory function and the maintenance of synaptic health upon aging .
adlard and colleagues proposed that -amyloid pathology could cause cognitive impairment by trapping zinc within plaques rather than via a directly toxic mechanism .
the zinc immobilisation by amyloid would then have similar consequences as a loss of znt3 activity with a loss of zinc - dependent synaptic modulation promoting cognitive decline .  
an alternative approach to minimising the consequences of released zinc could be to promote mechanisms which enhance reuptake .
recently , it has been shown that the cellular form of the prion protein ( prp ) is an evolutionary descendent of the zip family of divalent metal transporters .
in particular , zips 5 , 6 , and 10 were found to have a 
prion - like  domain with significant structural similarity . as both prp and the zips bind divalent metal ions via histidine - rich motifs
contained with n - terminal repeating sequences , this could suggest that prp is involved in zinc sensing , scavenging , or transport . in agreement with that possibility
, we have shown that prp promotes zinc uptake ( watt et al . , unpublished ) .  
ensuring efficient clearance of extracellular zinc from the synaptic cleft via prp would exploit an existing physiological process .
furthermore , enhancing zinc uptake would help prevent its ability to contribute to the synaptic targeting of a oligomers , thus preserving synaptic function   and maintaining the proposed ferroxidase activity of app .
as prp levels decrease with age and in sporadic ad , it is possible that zinc is cleared less efficiently from the synaptic cleft enhancing aggregation of a and inhibiting app ferroxidase activity to promote a pro - oxidative environment .
this would suggest that preserving prp function during ad could provide multifactorial benefits , an inhibition of bace1 which would reduce a formation   and ensure efficient clearance of zinc from the synaptic cleft to prevent aggregation of a peptides , as well as provide protection against oxidative stress [ 130 , 131 ] .
it is clear that zinc not only plays critical roles in the structural and functional integrity of many proteins , but that it also modulates the activity of glutamatergic synapses and indeed may act as a neurotransmitter in its own right .
several of the enzymes involved in processing app and a are zinc metalloproteases , with an essential requirement for zinc in their catalytic activity .
although zinc dyshomeostasis may contribute to the development of ad , further work is required to clarify the molecular and cellular mechanisms affected by zinc under both normal and disease situations .
 zinc , the most abundant trace metal in the brain , has numerous functions , both in health and in disease . 
 zinc is released into the synaptic cleft of glutamatergic neurons alongside glutamate from where it interacts and modulates nmda and ampa receptors . 
 in addition , zinc has multifactorial functions in alzheimer 's disease ( ad ) . 
 zinc is critical in the enzymatic nonamyloidogenic processing of the amyloid precursor protein ( app ) and in the enzymatic degradation of the amyloid- ( a ) peptide . 
 zinc binds to a promoting its aggregation into neurotoxic species , and disruption of zinc homeostasis in the brain results in synaptic and memory deficits . 
 thus , zinc dyshomeostasis may have a critical role to play in the pathogenesis of ad , and the chelation of zinc is a potential therapeutic approach . 
1. Introduction
2. Physiological Role of Zinc in the Brain
3. Role of Zinc in APP Processing
4. Role of Zinc in A
5. Zinc Binding to A
6. Role of Zinc in Alzheimer's Disease
7. Conclusions