caffeine and coffee as therapeutics against alzheimer’s ...€¦ · g.w. arendash and c. cao /...

Journal of Alzheimer’s Disease 20 (2010) S117–S126 S117DOI 10.3233/JAD-2010-091249IOS Press

Caffeine and Coffee as Therapeutics AgainstAlzheimer’s Disease

Gary W. Arendasha,b,∗ and Chuanhai Caob,c

aThe Florida Alzheimer’s Disease Research Center, Tampa, FL, USAbDepartment of Cell Biology, Microbiology, and Molecular Biology, University of South Florida, Tampa, FL, USAcThe Byrd Alzheimer’s Center and Research Institute, Tampa, FL, USA

Abstract. Epidemiologic studies have increasingly suggested that caffeine/coffee could be an effective therapeutic againstAlzheimer’s disease (AD). We have utilized a transgenic mouse model for AD in well-controlled studies to determine if caffeineand/or coffee have beneficial actions to protect against or reverse AD-like cognitive impairment and AD pathology. AD micegiven caffeine in their drinking water from young adulthood into older age showed protection against memory impairmentand lower brain levels of the abnormal protein (amyloid-β; Aβ) thought to be central to AD pathogenesis. Moreover, “aged”cognitively-impaired AD mice exhibited memory restoration and lower brain Aβ levels following only 1–2 months of caffeinetreatment. We believe that the cognitive benefits of chronic caffeine administration in AD mice are due to caffeine itself, and notmetabolites of caffeine; this, because our long-term administration of theophylline to AD mice provided no cognitive benefits. Inacute studies involving AD mice, one oral caffeine treatment quickly reduced both brain and plasma Aβ levels – similarly rapidalterations in plasma Aβ levels were seen in humans following acute caffeine administration. “Caffeinated” coffee provided toAD mice also quickly decreased plasma Aβ levels, but not “decaffeinated” coffee, suggesting that caffeine is critical to decreasingblood Aβ levels. Caffeine appears to provide its disease-modifying effects through multiple mechanisms, including a directreduction of Aβ production through suppression of both β- and γ-secretase levels. These results indicate a surprising ability ofmoderate caffeine intake (the human equivalent of 500 mg caffeine or 5 cups of coffee per day) to protect against or treat AD ina mouse model for the disease and a therapeutic potential for caffeine against AD in humans.

Keywords: Alzheimer’s disease, amyloid-β, caffeine, coffee, memory

INTRODUCTION

The premise that caffeine/coffee may provide a safe,effective, readily available, and inexpensive therapeu-tic against Alzheimer’s disease (AD) is underscored bya growing body of epidemiological literature. Severalcross-sectional human studies have reported that caf-feine/coffee consumption in both young and aged nor-mal adults is associated with better cognitive perfor-mance [1,2]. Caffeine/coffee has been shown to im-prove information processing in the elderly, as well as

∗Correspondence to: Gary W. Arendash, Ph.D., Department ofCell Biology, Microbiology, and Molecular Biology, University ofSouth Florida, Tampa, FL 33620, USA. Tel.: +1 813 974 1584; Fax:+1 813 974 1614; E-mail: [email protected].

in young adults [3]. In a 10-year prospective study in-volving aging men, Van Gelder et al. [4] found that cof-fee consumption was linked to slower cognitive decline,particularly with modest coffee consumption of 3 cupsper day (approximately 300 mg caffeine). More recent-ly, women with higher coffee consumption over a fouryear period showed less cognitive decline than thoseconsuming little or no coffee [5] and mid-life coffeeconsumption has been linked to a 65% decreased riskof AD [6]. Most convincing of the epidemiologic stud-ies in establishing an association between caffeine andAD has been Maia and de Mendonca [7], which report-ed that AD patients consumed markedly less caffeineduring the 20 years preceding diagnosis of AD, com-pared with age-matched individuals without AD. Al-though these epidemiologic studies collectively suggestthat chronic caffeine/coffee intake delays or reduces the

ISSN 1387-2877/10/$27.50 2010 – IOS Press and the authors. All rights reserved

S118 G.W. Arendash and C. Cao / Caffeine and Coffee as Therapeutics Against AD

risk of AD, the extent to which caffeine/coffee protectsagainst AD is problematic to assess in humans because:1) retrospective studies are based on recall and cannotunequivocally isolate caffeine/coffee intake from otherfactors affecting cognition over years, and 2) controlledlongitudinal studies involving caffeine/coffee admin-istration over decades are impractical. Moreover, thepotential for caffeine/coffee to treat established casesof AD has not been reported to date in the scientificliterature.

The aforementioned issues concerning epidemiolog-ic, longitudinal, and controlled studies in humans toevaluate the therapeutic potential of caffeine/coffeeagainst AD are at least partially addressed by humanamyloid-β protein precursor (AβPP) transgenic mousemodels for AD. These AD mice bear one or more hu-man genetic mutations that cause the familial form ofAD. One such mutation is the “Swedish” mutation, re-sulting in a double amino acid substitution at the β-secretase cleavage site of AβPP, making such cleavagemuch more likely and thus greatly increasing produc-tion of human Aβ in brains of these transgenic mice.The human Aβ that is produced then aggregates intothe Aβ cores of neuritic plaques, as in human brains.During the process of Aβ aggregation, these “AβPPsw”mice become cognitively impaired in a variety of cog-nitive domains [8–10], similar to human AD patients.Although AβPPsw mice are only a model for the Aβdeposition of AD (and do not mimic the full patholog-ic complexity of the human disease), they do modelwhat is thought to be the primary disease mechanism –Aβ production and aggregation in the brain. As such,highly controlled studies can be done by utilizing thesemice to develop insight into what therapeutics couldprotect against or treat AD in humans. This paper pro-vides a brief overview of the studies we have performedwith AD transgenic mice to evaluate the potential ofcaffeine/coffee for protection and treatment against ADin humans. As well, we mention initial results fromour on-going clinical studies involving caffeine admin-istration.

MATERIALS AND METHODS

The Materials and Methods for the studies/data in-volving caffeine administration have been described indetail in our previous work [11–13], which is referredto specifically when a particular study is presented anddiscussed. For our new behavioral study involvinglong-term theophylline administration to AD mice, ap-propriate methods are included in the section detailingresults of that study.

RESULTS

Caffeine to protect against Alzheimer’s disease:transgenic mouse studies

Our initial studies were designed to determinewhether caffeine could protect against the developmentof Aβ neuropathology and cognitive impairment inAβPPsw transgenic mice. In those studies, which aredetailed in Arendash et al. [11], we put caffeine intothe drinking water of AβPPsw transgenic (Tg) micebetween 4 and 9 months of age (300 mg/L). The re-sultant amount of caffeine intake for each mouse (ap-proximately 1.5 mg/day) was the human equivalent of500 mg or 5 cups of coffee per day. Cognitive testingin a comprehensive battery during the final 6 weeksof treatment revealed surprisingly better performanceof Tg mice being given caffeine compared to con-trol Tg mice given non-caffeinated water. In caffeine-treated Tg mice, cognitive domains of spatial learn-ing/reference memory, working memory, and recog-nition/identification were in fact at or near the per-formance level of non-transgenic control mice, whichwere being given water. Even when “overall” cogni-tive performance was assessed by evaluating many cog-nitive measures collectively in discriminant functionanalysis (DFA), the superior overall performance ofcaffeine-treated Tg mice could easily be distinguishedfrom the overall performance of Tg controls. Thus,long-term caffeine treatment protected AD mice fromcognitive impairment overall and in multiple cognitivedomains.

There are several reasons why caffeine-induced cog-nitive protection in AD Tg mice is unlikely to involvegeneralized mechanisms (e.g., increased glucose uti-lization) and/or the well-known stimulant effects of caf-feine to increase alertness/focus. First, long-term caf-feine administration had no effect on sensorimotor oranxiety function in such Tg mice [11]. Second, wehave found that life-long administration of caffeine indrinking water to normal (non-transgenic) mice doesnot improve their cognitive performance when testedin older age [12].

Accompanying the cognitive protection providedby long-term caffeine treatment to Tg mice, therewere significant 32–37% reductions in their hippocam-pal (brain) levels of Aβ compared to untreated Tgmice [11]. Importantly, the mechanism for this caffeine-induced reduction in brain Aβ involves suppression ofAβ production because both β- and γ-secretase levelswere reduced in the hippocampus of caffeine-treated

G.W. Arendash and C. Cao / Caffeine and Coffee as Therapeutics Against AD S119

Fig. 1. Protective/treatment effects of caffeine intake on Aβ production/aggregation and memory in AD transgenic (Tg) mice when caffeineconsumption is started during young adulthood (life-long caffeine) or after development of AD pathology/memory impairment.

Tg mice [11]. Exactly how caffeine suppresses both ofthese secretases was explored in our follow-up studies(see next section). Our initial study [11] provided thefirst evidence that caffeine can directly affect AD patho-genesis by suppressing levels of both enzymes need-ed for Aβ production. Moreover, we have shown thatthis suppression of Aβ production by caffeine occurs atphysiologic concentrations (1–10 µM) – caffeine con-centrations that are present in the plasma and brain fol-lowing only 1–2 cups of coffee. Parenthetically, we arenot aware of any AD drug under development by thepharmaceutical industry that has caffeine’s profoundability to reduce both Aβ − producing enzymes. Asdepicted in Fig. 1, “unmodified” brain Aβ productionand aggregation during aging in Tg mice inevitably re-sults in memory impairment. By contrast, long-term(“life-long”) caffeine intake to Tg mice reduces brainAβ production/aggregation, thus protecting against de-velopment of Aβ-induced memory impairment (Fig. 1).

Caffeine to treat Alzheimer’s disease: transgenicmouse studies

To determine if caffeine treatment has the abilityto reduce pre-existing memory impairment and Aβneuropathology, caffeine treatment in drinking water(300 mg/L) was administered for 2 months to “aged”Tg mice already exhibiting both of these AD character-istics [12]. This concentration of caffeine in drinkingwater, which is identical to that utilized in our prior

protection-based study, results in mice receiving a dai-ly caffeine intake of approximately 1.5 mg/day (e.g.,the human equivalent of 500 mg caffeine or 5 cups ofcoffee per day). Prior to caffeine treatment, aged Tgmice were substantially impaired in a working mem-ory task, the radial arm water maze. Following caf-feine treatment, however, these same mice exhibited arestoration of working memory to the level of normal,aged mice [12]. In these same aged Tg mice, whichhad substantial brain Aβ deposition prior to caffeinetreatment, reductions of 40% and above for both sol-uble and deposited (insoluble) brain Aβ were evidentfollowing caffeine treatment [12].

The profound cognitive restoration by caffeine inaged AD mice is likely due to caffeine’s aforemen-tioned ability to reduce Aβproduction by reducing bothβ- and γ-secretase protein expression [11]. We havefurther extended these mechanistic findings by showingthat 1) caffeine administration to Tg mice was able toreduce the Raf-1/NFκB inflammatory pathway, whichstimulates brain β-secretase, and 2) caffeine suppress-es both α and β isoforms of GSK-3 in neuronal cellcultures, which are linked to PS1/γ-secretase activityand tau hyperphosphorylation, respectively. Regard-ing suppression of the Raf-1 inflammatory pathway, wehave determined that caffeine stimulates PKA activityin the hippocampus of treated AD Tg mice [12]. Thisenhanced PKA activity then inhibits (inactivate) cRaf-1 by phosphorylation at serine259, decreasing NFκBactivity and the expression of NFκB-controlled genessuch as BACE1, thus resulting in decreased Aβ pro-


duction. Regarding the suppression of GSK-3, caffeineaffects both active and holoprotein (total) GSK-3 lev-els [12], which suggests that caffeine does not dimin-ish GSK-3 activity through direct substrate-level phos-phorylation, but rather through alternations in proteinproduction or degradation The extent to which either orboth of these caffeine mechanisms involve adenosinereceptor antagonism is unknown and currently beinginvestigated.

Our measurement of plasma caffeine levels ofaged, behaviorally-evaluated AβPPsw mice given twomonths of caffeine treatment underscores the humanrelevance and physiologic significance of the caffeinetreatment in this long-term study. At the caffeine con-centration in drinking water that was employed, plasmacaffeine levels averaged 26 µM [13], equivalent to theplasma caffeine concentration expected in humans fol-lowing intake of several cups of coffee. This caffeineconcentration is very close to the 20 µM concentrationthat we found to be optimal in our cell culture studiesfor suppression of Aβ1−40 and Aβ1−42 production, al-though caffeine concentrations in the much lower nMrange (250–500 nM caffeine) were also effective [11].This very low nM range of caffeine effectiveness toreduce Aβ production in vitro is below the level of anyknown molecular target, including adenosine receptorantagonism. As such, caffeine’s reduction of Aβ pro-duction appears to be novel in being independent ofadenosine receptors actions. It is important to note thattheophylline, one of several active metabolites of caf-feine, may be responsible for at least some of the cog-nitive, neuropathologic, and neurochemical benefits ofcaffeine. This premise is underscored by the fact thatcaffeine metabolites such as theophylline have a con-siderably longer half-life than caffeine. We have ex-plored the contribution of theophylline to the benefitscurrently being attributed to caffeine in our AD mice,and report our findings in the next section.

Also in this long-term caffeine treatment study inaged AD mice, we found a clear reduction in hip-pocampal inflammatory cytokines in caffeine-treatedmice with higher plasma caffeine levels compared tothose with lower caffeine levels. The ability of caf-feine to provide beneficial anti-inflammatory effects inthe brains of AD mice is consistent with prior studiessupportive of its anti-inflammatory capacities [14] and,as such, could represent a potent “non-amyloidogenic”mechanism of caffeine action that may contribute toits ability to protect against/reverse cognitive impair-ment. Other beneficial mechanisms of caffeine ac-tion in AD mice (and perhaps in human AD) include

the antioxidant actions of caffeine [15], its ability toblock disruptions of the blood-brainbarrier [16], and itswell-established antagonism of brain A1 and/or A2Aadenosine receptors [1,2,17,18]. Regarding the latter,adenosine receptor antagonism may be central to mostor all of the beneficial actions of caffeine against AD.Any combination of such mechanisms of caffeine ac-tion may collectively provide the cognitive benefits wehave documented in AD mice.

The effects of caffeine on established AD charac-teristics are summarized in Fig. 1 (Caffeine after AD),which shows caffeine-induced suppression of Aβ pro-duction leading to removal of Aβ from the brain,resulting in reversal of cognitive impairment. Thisstudy [12], performed in aged AD mice, provides thefirst evidence that caffeine treatment can reverse cog-nitive impairment and AD neuropathology in a modelfor the disease.

Theophylline and cognitive function in AD transgenicmice

As a major active metabolite of caffeine, theo-phylline may be responsible for a significant part ofthe cognitive benefits we found through long-term caf-feine administration. In the aged Tg mouse study justpresented [12,13], we found plasma theophylline levelsto be strongly correlated with plasma caffeine levels.Moreover, theophylline has a longer half-life than caf-feine, with brain concentrations of theophylline usual-ly being higher than those for caffeine following long-term caffeine treatment [19]. Thus, it is important todetermine the extent to which theophylline, contributesto the cognitive benefit we are attributing to caffeineadministration in our AD mice. We therefore admin-istered theophylline in drinking water (200 mg/L) toAβPPsw+PS1 double transgenic mice (Jackson Lab-oratories, Bar Harbor, Maine) or non-transgenic lit-termates beginning at 5.5 months of age. The re-sulting dose of theophylline (approximately 1 mg/day)is just slightly less than the 1.5 mg/day dose of caf-feine we routinely administer long-term to AD mice.This theophylline dosage presumably resulted in plas-ma theophylline levels at least as high as those result-ing from chronic caffeine administration because ADmice chronically treated with caffeine have a plasmacaffeine-to-theophylline ratio of approximately 8-to-1 [13].

At 6 months into theophylline treatment, mice wereevaluated in our standard 6-week behavioral batteryof sensorimotor, anxiety, and cognitive-based tasks, as


Fig. 2. Inability of long-term treatment with the active caffeine metabolite theophylline to provide cognitive benefit to AD transgenic mice ornon-transgenic littermates, as demonstrated by the radial arm water maze task of working memory. The data presented are for Trial 1 (naıvefirst trial), and Trials 4 and 5 (working memory trials) overall across 12 days of testing at 6 months into theophylline treatment. The number ofanimals in each group were: NT controls (n = 7), NT+theophylline (n = 8), Tg controls (n = 7), and Tg+theophylline (n = 9).

done in our initial caffeine study [11]. Across multiplecognitive-based tasks [Y-maze spontaneous alternation,Morris maze acquisition/retention, circular platform,platform recognition, and radial arm water maze], therewere no effects of theophylline treatment in either Tgor non-transgenic mice compared to controls. An ex-ample of this surprising lack of cognitive benefit withlong-term theophylline treatment is shown in Fig. 2 forthe radial arm water maze task of working memory.For both errors and latency during working memorytrials T4 and T5, theophylline-treated mice performedno different from controls across 12 days of testing. Inaddition, there were no effects of theophylline treat-ment on sensorimotor tasks (open field activity, balancebeam, string agility) or on the elevated plus-maze taskof anxiety (data not shown). Thus, we conclude that

the cognitive benefits of caffeine administration in ourprior protection-based [11] and treatment-based [12]studies were likely due to caffeine itself, and not theo-phylline. Whether or not the other major caffeinemetabolite (paraxanthine) may be responsible for someof caffeine’s cognitive benefits in Tg mice remains tobe determined.

It is important to put our long-term theophyllinetreatment study (> 7 months of treatment) in propercontext of prior work investigating theophylline’s ef-fects on cognitive function. Our search of the scien-tific literature has, firstly, revealed a wealth of stud-ies showing no effect of theophylline administration onthe cognitive performance of humans (both adults andchildren). This is consistent with our present finding oflong-term theophylline treatment having no cognitive


Fig. 3. Brain Aβ production/clearance, the suppressive actions of caffeine on Aβ production, and resultant effects on brain and plasma Aβlevels. (A) Unmodulated: Aβ is primarily produced in neurons, secreted into the brain extracellular space in soluble form, then enters a dynamicequilibrium between soluble and deposited (insoluble) Aβ. Continual transport of soluble Aβ occurs into plasma. (B) Caffeine: Short-Term:Caffeine suppression of both β- and γ-secretase activities reduces Aβ production, resulting in lower soluble Aβ in brain and plasma. Theequilibrium between soluble and deposited Aβ is not impacted by this short-term reduction in brain soluble Aβ levels. (C) Caffeine: Long-Term:Continuous caffeine suppression of Aβ production and resultant chronically lower levels of brain soluble Aβ induce a flux of deposited (insoluble)Aβ to the soluble form, which is cleared from brain into plasma via soluble Aβ transport. Plasma Aβ levels may be reduced or not changed,depending on degree of caffeine-induced suppression of Aβ production. Reproduced with permission from IOS Pres.

effects in normal mice across multiple tasks and cogni-tive domains. Secondly, no studies had investigated theeffects of “long-term” theophylline on cognitive func-tion prior to our present work. Earlier studies only in-volved “acute/single treatment” theophylline adminis-tration to normal mice or as a pre-treatment to mice ex-posed to a noxious stimulus (e.g., ischemia, microwaveirradiation). Particularly since acute administration ofcaffeine (and presumably theophylline) may have cog-nitive effects that are different from long-term adminis-tration, the behaviorally-based theophylline study thatwe present in the current manuscript appears to be thefirst long-term theophylline treatment study investigat-ing cognitive performance in any rodent, either normal,genetically-manipulated, or CNS-injured.

Acute effects of caffeine/coffee on blood Aβ levels inAD mice and humans

Following formation of monomeric Aβ in the brain,newly produced Aβ enters into a dynamic equilibriumbetween soluble and deposited Aβ in the brain, withcontinual transport of soluble Aβ out of the brain andinto plasma down a concentration gradient (Fig. 3A).Therefore, blood levels of Aβ may be indicative of caf-feine’s therapeutic actions on brain levels of Aβ. Inview of our findings that caffeine decreases Aβ pro-duction in AD transgenic mice through suppression of

both β- and γ-secretase [11,12], we have hypothesizedthat ensuing lower brain levels of soluble Aβ will resultin lower plasma Aβ levels acutely (Fig. 3B). This hy-pothesis is supported by the finding that Aβ is rapidlyproduced and cleared from the brain [20]. Moreover,following almost 2 months of caffeine administrationto “aged” Tg transgenic mice (see preceding section),we have found that higher plasma caffeine levels werestrongly associated with lower plasma Aβ levels in in-dividual Tg mice (r = −0.761; p = 0.011).

Considering the above findings, we determined theeffects of acute caffeine [13] or coffee administra-tion on plasma and brain Aβ levels in AD transgenicmice. Acute (single treatment) administration of caf-feine (1.5 mg/0.2 cc) or saline vehicle was given byi.p. injection or gavage to AβPPsw transgenic (Tg)mice in young adulthood (3 months old) or in older age(14 months old). At 3 hours following treatment, plas-ma Aβ1−40 levels in 3-month old Tg mice were sub-stantially reduced by 41%. Even the older 14-monthold Tg mice showed significant reductions in plasmaAβ following acute caffeine treatment. We have sincerepeated these studies in additional Tg mice, acutelyadministering caffeinated or de-caffeinated coffee (i.p.)instead of caffeine (Caffeinated coffee contained thesame 1.5 mg caffeine/0.2 cc as in pure caffeine admin-istration). At 3 hours following an initial treatment and3 hours after another treatment 3 days later, caffeinated


Fig. 4. Ability of (caffeinated) coffee to acutely suppress plasma Aβlevels, but not de-caffeinated coffee or saline control. Treatments(i.p.) were administered on Days 1 and 4, 3 hours prior to takinga blood sample on both days. Percent change from pre-treatmentAβ levels are graphed. Each group consisted of 6–7 AβPPsw mice.∗p < 0.02 for Pre vs. Day 1, coffee group only; ∗∗p < 0.02 forcoffee group vs. other two groups on that day. Absolute values (inpg/ml) for pre – versus post-treatment (Day 1) plasma Aβ1−40 levelsand associated paired t-test statistics were: Saline treatment (6055 ±1180 vs. 5400 ± 381, N.S.), Decaffeinated coffee (6036 ± 1029 vs.5662 ± 809, N.S.), Caffeinated coffee (6851 ± 1446 vs. 5090 ±637, p < 0.02).

coffee provided the same beneficial reduction in plas-ma Aβ-40 levels as we had previously reported for purecaffeine (Fig. 4). By contrast, decaffeinated coffee andsaline treatments had no effect on plasma Aβ1−40 lev-els. Thus, caffeine is the likely coffee constituent thatis critical for decreasing brain and plasma Aβ levels.Although this acute study did not involve measurementof “brain” Aβ levels as in our prior acute and long-termstudies [11–13], the fact that acute caffeine adminis-tration in our prior work resulted in reductions in bothplasma and brain Aβ levels within the same 2–3 hourtime course [13] strongly suggests that a similar reduc-tion in brain Aβ occurred with coffee administration,but not with decaffeinated coffee administration.

In a separate group of 3-month old Tg mice, mea-surement of brain interstitial fluid (ISF) levels of Aβthrough in vivo microdialysis was performed bothbefore and following caffeine administration (1 mg,i.p.). This single caffeine treatment significantly low-ered brain ISF Aβ levels by 32% at 2–3 hours post-treatment, compared to the basal ISF Aβ levels in each

mouse [13]. Follow-up studies showed that this rapidsuppression of brain Aβ levels by caffeine was not dueto increased Aβ elimination from the brain, but ratherwas due to decreased Aβ production. These acute stud-ies collectively indicate that plasma Aβ can be a viablebiomarker for the efficacy of anti-Aβ therapeutics suchas caffeine. It is nonetheless important to underscorethat our acute caffeine administration studies involvedAD transgenic mice “naıve” to caffeine, whereas anyclinical trials in humans would involve many occasion-al or habitual caffeine users.

As a clinical extension of our studies in AD miceand in view of the aforementioned clinical caveat, wehave embarked on acute caffeine and coffee adminis-tration studies in humans. Similar to our AD mousestudies, we find that acute oral caffeine administration(400 mg) to aged humans induces an immediate de-crease in blood Aβ levels in approximately half of sub-jects (data not shown). In both young adult and agedhumans, we find that blood Aβ levels continue to bemodified many hours following caffeine or coffee ad-ministration. These initial clinical studies are consis-tent with caffeine/coffee having an immediate and di-rect effect on what could be the primary pathogenicmechanism of AD. Inasmuch as the protocols and re-sults of these acute caffeine administration studies willbe presented in detail within upcoming manuscripts,they should currently be considered as preliminary ob-servations that suggest a similar response of human andmouse subjects to acute caffeine administration. Addi-tional clinical studies are underway to determine morelong-term effects of caffeine and coffee administrationto aged humans and AD patients.

DISCUSSION

The results of our studies indicate a surprising abilityof moderate caffeine intake to protect against or reverseAD-like cognitive impairment and Aβ neuropathologyin established mouse models for AD. The equivalentamount of daily caffeine intake that our studies sug-gest is required for human therapeutic benefit againstAD is approximately 500 mg. The average US intakeof caffeine is only around 150 mg/day and decreasesduring aging – thus, average caffeine intake appearsto be significantly below levels necessary for benefitsagainst AD. An 8 oz. cup of coffee contains 100 mgcaffeine, while a similar serving of tea or soft drinkscontain only 35 and 25 mg caffeine, respectively. Assuch, only five cups of coffee per day (a moderate con-


sumption level) are needed to reach the 500 mg level,while many more servings of tea or soft drinks wouldbe required. Moreover, given coffee’s significant con-tent of antioxidants and other beneficial phytochemi-cals, this source of caffeine is clearly preferable overessentially any other source.

Caffeine’s beneficial effects against cognitive im-pairment and AD neuropathologyprobably involve firstand foremost the unique ability of caffeine to suppressboth enzymes needed for brain Aβ generation, name-ly β-secretase and γ-secretase. Nonetheless, multipleother mechanisms are probably contributing to the ther-apeutic actions of caffeine in AD animal models, in-cluding caffeine’s ability to act as: 1) a strong anti-inflammatory agent, 2) an anti-oxidant, 3) a mitochon-drial activator, and 4) a stimulator of both neuronal ac-tivity and glucose utilization. Although adenosine re-ceptor antagonism should certainly be included in thislist, caffeine’s actions almost certainly do not involvesimply the increase in alertness provided by such recep-tor blockade. Indeed, if this were a primary mechanismof caffeine’s cognitive benefit,normal mice should havebenefited cognitively from a lifetime of caffeine ad-ministration, but this was not the case [12]. Thus, wepropose that caffeine does not act as a general nootrop-ic agent to enhance “normal” cognitive performance.Rather, we believe that caffeine protects against andminimizes brain cognitive impairment, such as that dueto brain Aβ accumulation/aggregation in AD.

Studies in this paper present evidence that caffeineis providing cognitive benefits to AD transgenic micedirectly, since a major active metabolite of caffeine(theophylline) was found to be cognitively ineffective.It is important to note, however, that our theophyllineadministration studies utilized mice of a different trans-genic line (AβPPsw + PS1), bearing a different strainbackground (C57/BL6), and obtained from a commer-cial breeder (Jackson Laboratories). By contrast, allof our other behavioral studies with caffeine adminis-tration [11–13] involved inbred mice of our own trans-genic colonies. We have previously determined thatAβPPsw or AβPPsw + PS1 mice on a C57/BL6 back-ground (e.g., those from commercial breeders) requireconsiderably longer aging (compared to our inbredmice on a mixed C57/BL6/SW/SJL background) be-fore cognitive impairment becomes apparent. Thus,it is possible that results of this study could havebeen different had we utilized mice of our own inbredcolonies and/or began the study with aged, already-impaired transgenic mice for examining the effects oftheophylline administration on cognitive function.

If the inability of long-term theophylline adminis-tration to mimic the therapeutic effects of caffeine isindependent of the AD mice and treatment paradigmutilized (as we suspect is the case), this would argue forsuch benefits of caffeine to be independentof adenosinereceptors. Supportive of this premise, theophylline is 3to 5 times more potent than caffeine as an antagonist ofboth adenosine A1 and A2a receptors. Alternatively,it is possible that brain theophylline levels (which wedid not measure in this study) in theophylline-treatedTg mice were insufficient for providing behavioral andneuropathologic benefit. Either scenario would be con-sistent with a mechanism that does not primarily in-volve adenosine receptor antagonism.

In our ‘acute” studies, AD mice given a singletreatment with caffeine exhibited quick reductions inboth brain interstitial fluid and plasma Aβ levels (i.e.,within 2–3 hours). Since caffeine did not affectthe half-life of ISF Aβ, caffeine had directly affect-ed brain Aβ production rather than its elimination.Underscoring this premise are our findings that caf-feine affects Aβ production through suppression ofboth β-secretase (BACE1) and γ-secretase/PS1 expres-sion [11,12]. Acutely, this caffeine-induced decreasein Aβ production would result in lower soluble Aβ inbrain ISF and consequently lower plasma Aβ levels(Fig. 2, Caffeine – Short-Term). This is exactly whatwe observed in AD mice that were young adults (withno brain Aβ deposition), as well as aged AD mice(bearing robust Aβ deposition). Whether or not thiscaffeine-induced suppression of Aβ production is di-rect or involves adenosine receptor blockade/mediationis currently unknown. Evidence against involvementof adenosine receptors is provided by studies showingthat acute and chronic caffeine administration to micegiven noxious brain conditions (e.g., ischemia, exci-totoxin exposure) results in opposite effects (worse orlesser damage) due to antagonism of A1 and A2a re-ceptors, respectively [21]. In sharp contrast, both acuteand chronic caffeine administration decrease plasmaand brain Aβ levels.

In our long-term “treatment” study in aged/cognitive-ly-impaired AD mice, caffeine reversed their workingmemory impairment. This 2-month caffeine adminis-tration regime not only reduced brain soluble Aβ levels,but also reversed brain Aβ neuropathology. Althoughthe mechanism(s) for caffeine-induced reversal of Aβdeposition requires further investigation, we hypothe-size that “chronic” caffeine suppresses Aβ productionlong-term, resulting in consistently lower brain levelsof soluble ISF Aβ (Fig. 2, Caffeine – Long-Term). This


would then induce a flux of insoluble Aβ out of thedeposited form and into the soluble form due to dy-namic equilibrium. Newly-solubilized Aβ would thenbe cleared into the plasma via both blood-brain-barriertransport and bulk fluid flow. The dynamic equilibri-um between soluble (ISF) and insoluble/deposited Aβin the brain is underscored by previous work showingthat young adult and aged AβPPsw mice had similarsteady-state levels of ISF Aβ, but rapid inhibition of Aβproduction resulted in a two-fold longer Aβ half-lifein the aged, Aβ deposit-bearing mice [22]. We believethis longer Aβ half-life is caused by solubilization ofa portion of the deposited Aβ pool due to the dynamicISF Aβ ↔ deposited Aβ equilibrium.

The effects of “chronic” caffeine intake on healthhave been reported in numerous retrospective andprospective studies. A comprehensive review of theliterature [23] found that for healthy adults, moderatedaily caffeine intake poses no adverse effects on thecardiovascular system, bone status and calcium bal-ance, or incidence of cancer. Prospective studies havenot found significant associations between coffee con-sumption and the risk of coronary heart disease [24,25]. Contrary to public belief, caffeine intake of 500–600 mg daily does not increase the risk, frequency, orseverity of cardiac arrhythmias [26,27]. Indeed, thereis a growing list of age-related diseases wherein habit-ual caffeine intake of 400–600 mg (4–6 cups of coffee)daily reduces the risk. For example, strong data from anumber of prospective studies show that both caffeineand coffee reduce the risk of Type 2 Diabetes Melli-tus by 35–79% [28]. Both retrospective and prospec-tive studies report a reduced risk of Parkinson’s dis-ease with habitual caffeine/coffee intake [29,30]. Riskof liver cirrhosis is significantly reduced by both caf-feine and coffee [31]. Thus, far from posing a threatto human health during aging, habitual caffeine/coffeeintake provides a number of health benefits.

Finally, it should be underscored that “acute” effectsof caffeine or coffee administration on plasma Aβ lev-els clearly occur in both AD mouse models and hu-mans. This suggests not only that these mouse mod-els are providing valuable insight into development oftherapeutics against AD, but also that blood levels ofAβ could be a monitor for anti-Aβ therapeutic effectsoccurring in the brain, such as with caffeine. An excit-ing period of clinical research with caffeine and coffeeagainst AD is on the horizon. This research may finallyshow that a safe and readily available natural agent –caffeine – provides the first effective therapeutic againstAD.

In summary, our studies utilized AD transgenic miceto demonstrate that: 1) Caffeine protects AD miceagainst memory loss through mechanisms that directlyaffect the disease process, 2) Caffeine reverses mem-ory loss and AD pathology in “aged” AD mice, and3) Caffeine and/or caffeinated coffee both affect plas-ma levels of Aβ in AD mice and humans. Our resultswarrant the clinical trials in progress with caffeine andcaffeinated coffee as safe, inexpensive, and effectivetherapeutics against AD.

ACKNOWLEDGMENTS

The following collaborators in these studies aregratefully acknowledged: Valentina Echeverria, JohnCirrito, David Holtzman, Huntington Potter, TakashiMori, Maggie Mamcarz, Melissa Runsfeldt, Li Wang,William Schleif, Edwin Jackson, Jennifer Cracchiolo,Daniel Shippy, Alexander Dickson, Ashok Raj, KavonRezia-Zadeh, Jun Tan, Bruce Citron, Xiaoyang Lin,and Chi Zhang. This work was supported by NIH grantAG025711 and funds from the Byrd Alzheimer’s Cen-ter and Research Institute.

Authors’ disclosures available online (http://www.j-alz.com/disclosures/view.php?id=242).

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caffeine and coffee as therapeutics against alzheimer’s ...€¦ · g.w. arendash and c. cao /...

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