portada-contrapo 59 (2) (Page 5)

Correspondence to: J. Jordán (Tel.: +34 967 599 200;e-mail: [email protected]).

Mitochondrial control of neuron death andits role in neurodegenerative disorders

J. Jordán, V. Ceña and J. H. M. Prehn1

Centro Regional de Investigaciones Biomédicas and Dpto. de Ciencias Médicas,Facultad de Medicina, Universidad de Castilla-La Mancha, Albacete, Spain.

1Department of Physiology, Royal College of Surgeons, 123,St Stephens Green, Dublin 2, Ireland

(Received on February 18, 2003)

J. JORDÁN, V. CEÑA and J. H. M. PREHN. Mitochondrial control of neurondeath and its role in neurodegenerative disorders (minireview). J. Physiol. Biochem.,59 (2), 129-142, 2003

Genetic or functional mitochondrial alterations can result in the initiation of celldeath programs that are believed to contribute to cell death in diabetes, ageing andneurodegenerative disorders. Mitochondria are being considered the main linkbetween cellular stress signals activated during acute and chronic nerve cell injury,and the execution of nerve cell death. This second function of mitochondria is regu-lated by several families of proteins that can trigger an increase in permeability of theouter and/or inner mitochondrial membrane. One example of this is the formation ofthe mitochondrial permeability transition pore (MPTP). This process can trigger therelease of cell death-inducing factors from mitochondria, as well as a dissipation ofthe mitochondrial transmembrane potential, depletion of ATP, and increased freeradical formation. Among the factors released from mitochondria are cytochrome c,the apoptosis inductor factor (AIF), and caspases. We review the role of the MPTPin diverse physiological and pathological processes, including neurodegenerative dis-orders such as Alzheimer’s disease, Parkinson’s disease and amyotrophic lateral scle-rosis (ALS). The design of drugs that could interfere with the functions of the MPTPcould allow novel therapeutic approaches for the treatment of acute and chronicnerve cell injury.

Key words: Apoptosis, Necrosis, Apoptosome, Caspase, Permeability transition pore,Mitochondria.

J. Physiol. Biochem., 59 (2), 129-142, 2003

Mitochondria are cytoplasmic organ-elles in eukaryotic cells that make up as

much as 10% of the cell volume. They arepleiomorphic organelles with structuralvariations depending on cell type, cell-cycle stage and the intracellular metabolicstate. The key function of mitochondria is

energy production through oxidativephosphorylation (OxPhos) and lipid oxi-dation (23, 64). Several other metabolicfunctions are performed by mitochondria,including urea production, heme, non-heme iron and steroid biogenesis, as wellas intracellular Ca2+ homeostasis regula-tion. In addition, mitochondria also play apivotal role in apoptosis and necrosis.Our understanding of the componentsinvolved in these mitochondrial functions,especially mechanisms of regulation, isstill incomplete. In the last years, thisorganelle is being considered the placewhere different apoptosis signals path-ways converge (26, 39). Due to the abilityto increase its permeability, mitochondriacontrol the release of molecules into thecytoplasm that are able to activate path-ways that result in cell death. The objec-tive of this work is to review the role ofmitochondria and the regulation of thepermeability of its membranes by meansof the formation of multiprotein complex-es, in particular the mitochondrial perme-ability transition pore (MPTP) duringnerve cell death. This review is dividedinto three sections. First, we demonstrategeneral concepts of mitochondria; in thesecond part we discuss aspects of struc-ture and regulation of MPTP formation;and finally in the third section we consid-er mitochondria and the MPTP as centralregulators of apoptosis and necrosis sig-naling pathways.

Mitochondrial structure

Mitochondria present a structure withtwo well-defined compartments, themitochondrial matrix and the intermem-brane space. These structures are delineat-ed by two membranes, the outer and innermitochondrial membrane. Each mem-brane has distinct biochemical and func-

tional characteristics. In the mitochondri-al matrix several copies of the circularmitochondrial genome, ribosomes andnumerous enzymatic complexes necessaryfor energetic functions, for gene expres-sion and replication are located (25). Theintermembrane space contains enzymesthat control the traffic of substancesamong the mitochondrial matrix and thecytosol, complexes with kinase activity,and soluble proteins required for electrontransfer, in particular cytochrome c. Theouter membrane is of limited surface areaand contains cholesterol, albeit at a lowerconcentration than the plasma membrane.At various points, termed contact sites, itis tightly juxtaposed to the inner mem-brane; a variety of metabolic processes areclustered at these contact sites, includingtransport of proteins, fatty acids, andsmall molecules (6). The inner mitochon-drial membrane, which contains a largeamount of cardiolipin and virtually nocholesterol, comprises two functionallydistinct domains. The inner boundarymembrane, which parallels the outermembrane, and the crista membranes,which are long tubular and lamellar struc-tures that represent a vast surface area forthe electron transfer complexes (55) andproteins that regulate the metabolitesuptake, such as the adenine nucleotidetranslocator (ANT). The inner membraneis especially non permeable to ions owingto its high content in the phospholipidcardiolipin, an essential characteristic thatpermits it to bear a electrochemical gradi-ent necessary for the execution of theenergetic function. However, the outermembrane lacks mitochondrial cristamembranes and, in physiological condi-tions, is permeable to ions and metaboliteswith molecular weights lower than 6000Da due to the presence of the proteinporin (18), also referred to as the voltage

J. JORDAN, V. CEÑA AND J. H. M. PREHN130

J. Physiol. Biochem., 59 (2), 2003

dependent anionic channel (VDAC).VDAC forms an aqueous channelthrough which proteins up to 10,000 Dacan pass and go into the intermembranespace.

In spite of the existence of these com-partments, that to a first view seems rigid,the mitochondrial structure permits theregular flux of proteins by means ofprocesses controlled by transmembranecomplexes. These are the translocase ofthe inner membrane (Tim) and translocaseof the outer membrane (Tom). Thesecomplexes are able to discriminate amongthose proteins that should be directed tothe mitochondrial matrix and the onesthat will reside in the intermembranespace (51). Proteins that cross to thematrix have an NH2 cleavable signalsequence. Most proteins must be uncoiledor stretched out to go through the translo-cators. This involves ATP binding and ismonitored and stabilized by chaperoneproteins, including hsp70. Thus, beforethe protein can go through the Tom com-plex, it must become “translocation com-petent”. Not surprisingly, the Tom com-plex will include import receptors that ini-tially recognize the signal peptide or a sig-nal sequence. These receptors includeTom20, Tom22, and Tom70, each recog-nizing a different set of proteins. With theaid of these receptors, the proteins aresubsequently brought into the region con-taining the translocator proteins. Thiscomplex, called the General Import Pore(GIP), facilitates the translocation of thepresequence of the protein across theouter membrane. The GIP is comprised ofTom40, Tom5, Tom 6, and Tom7. Tom40appears to be the core element of the pore.It also interacts with polypeptide chainspassing through the pore. All of the otherTom components in GIP are anchored to

the outer membrane by helical transmem-brane segments.

Mitochondrial proteins destined for thematrix often have a cleavable signal pep-tide on the protein which must be recog-nized before it will be admitted throughthe mitochondrial translocator. Theseproteins with “amino terminal signals”,“preproteins”, or “presequences” usuallyinteract with Tom20 first before beingtranslocated through the outer membrane.To do so, they are transferred to the GIPcomplex by interaction with Tom22 andTom5 which ushers them to the poreformed by Tom40. The proteins subse-quently enter the matrix using the porecomplex. The entry of proteins dependson the mitochondrial transmembranepotential which is generated by the elec-tron transport complexes. This membranepotential actually helps to pull the proteininto the GIP. The protein then enters thematrix where the cleavable preprotein isclipped off by a protease and a chaperoneprotein (mt-hsp70) in the matrix thatcooperates with Tim44 to complete thefull transfer into the matrix. Mt-hsp70 andTim 44 “pull” the protein into the matrixby a process that requires ATP. It alsorequires the membrane potential set up bythe electron transport chain. Negativecharges in the matrix, set up by the pump-ing of hydrogen ions to the intercristalspace, attract the positively charged pro-tein that enters the GIP.

Some mitochondrial proteins destinedfor the inner membrane have a cleavablesignal peptide followed by one or moremembrane-spanning segments that serveto insert the polypeptide into the innermembrane after it gets into the matrix.Proteins that are targeted through thismechanisms include the proteins of theelectron transport chain.

MPTP IN NEURODEGENERATIVE DISORDERS 131


At the contact sites between outer andinner mitochondrial membrane, cytoplas-mic and mitochondria matrix proteins areable to interact giving rise to the forma-tion of a channel called the mitochondrialpermeability transition pore, MPTP (32).

Energetic functions of mitochondria

The mitochondria are the cellular com-partment in which oxidative phosphoryla-tion takes place. During OxPhos reductorcapacity is generated, in form of NADHand FADH2, that is subsequently utilizedin the respiratory chain. During electrontransport, three protons from the mito-chondrial matrix exit into the intermem-brane space as a consequence of differ-ences in redox levels and the energy liber-ated. This results in an electrochemicalgradient between the two compartments(28) that is translated into a ∆pH of oneunit, and into an electric difference with apotential of 140 mV. This energy, approx-imately ∆G of –5Kcal, is utilized by theATP synthase (complex V), located in theinner membrane, for phosphorylatingADP to give rise to ATP, and at the sametime protons return to the mitochondrialmatrix (62).

The correct function of these enzymat-ic complexes is essential for the life of thecell. Any alteration in enzyme functionthat influences its function can result indegenerative processes. In fact, there aremany inhibitors of the different compo-nents of the mitochondrial electron trans-port chain (ETC) that are utilized in thestudy of neurodegenerative processes.

Mitochondria participate in the regulation of second messengers

The mitochondria, due to its capacityto precipitate Ca2+ in its interior in form

of phosphates, participate, together withendoplasmic reticulum and ion pumpsand channels located in the cellular mem-brane, in the regulation of the cytoplas-matic free calcium levels [Ca2+]c (24, 71).

At the moment, three transport sys-tems that control the entrance and exit ofthis cation into the mitochondria havebeen described: the Ca2+ uniporter, the2Na+/Ca2+ antiporter and the Ca2+/2H+

antiporter (Fig. 1). The Ca2+ uniporter isan electrogenic transporter that shows alow affinity for Ca2+ and can be inhibitedby Ruthenium Red and Mg2+. On theother hand, the Na+/Ca2+ antiporter,which can be inhibited by L-type Ca2+

channel blockers such as diltiazen, and theenergy dependent Ca2+/2H+ antiportersystem, located in the mitochondria ofsome cells, carry out Ca2+ release inresponse to specific stimuli (5). In somepathological situations, an alternativemechanism can trigger Ca2+ extrusionfrom mitochondria, the MPTP formation.This pathway is outlined below, and isbelieved to be a crucial process in cellulardeath programs. Its formation results inelectric potential dissipation and release ofsubstances from the mitochondrial matrixand the intermembrane space.

Taken together, these systems allow themitochondria to respond to [Ca2+]c fluc-tuations with a variation in [Ca2+]m levelsin a slow manner. Due to the kinetic andthermodynamic properties of the trans-port systems and to the [Ca2+]c valuesduring resting conditions (0,1-0,2 mM) orstimulated (0,5-3 mM) conditions, theprediction is that, under physiologicalconditions, Ca2+ accumulation in mito-chondria turns out to be negligible. How-ever, experiments in which [Ca2+]m wasdetermined using specifically targetedrecombinant aequorin (60), have shownthat quick changes in the [Ca2+]c pro-



duced by the stimulation of nerve cells,have its representation in mitochondria(81), exhibit a high amplitude (> 10 mM),a very short period of time ( <5 s) (58, 59)and is balanced by Ca2+ extrusion. Theresponse to the [Ca2+]m increment wouldhave as a result the metabolic route activa-tion, generating more quantity of energyinto the cell, without activating death pro-grams. Nevertheless, in pathological con-ditions, where the [Ca2+]c is found high ina maintained manner, Ca2+ enters mito-chondria excessively. This results in sever-al different effects, such as activation ofthe mitochondrial Ca2+ uniporter system,saturation of the calcium efflux systems,and eventually drastic changes in themitochondrial interior. Mitochondriahave been proposed to be the most impor-

tant source of ROS in cells of the nervoussystem (4, 43). In the mitochondrial ETC,a non-enzymatic reduction of O2 generat-ing superoxide anion may occur, parallelto the enzymatic reduction of O2 to H2Oby cytochrome oxidase. In some cases, upto a 2% of the total O2 consumed by ETCcan be transformed to O2

- by the coen-zyme Q. This reaction occurs due to thefact that electron transport is generally avery reactive process and involves compo-nents with a negative redox potential, suchas flavine (complex I) and ubiquinone(complex III) (48). Moreover, in condi-tions where ADP levels have diminished,such as during treatment with differentETC complex inhibitors, an increasedsuperoxide anions production has beenobserved.



Fig. 1. Mitochondrial role in regulation of cytosolic free Ca2+. A. Mitochondrial Ca2+ entry and extrusion mech-anisms. The outer membrane is permeable to Ca2+ due to the presence of a voltage dependent anionic channel,VDAC, while the inner membrane requires the activity of cation transporters. In the inner membrane, Ca2+

entry is regulated by the mitochondrial Ca2+ uniporter system (1) and Ca2+ release is mediated by Ca2+/Na+ (2)or Ca2+/H+ (3) antiporter systems. Electron transport chain, ETC (4) and Na+/H+ antiporter system (5) arerequired for the correct function of the other systems. B. The formation of the channel mitochondrial perme-ability transition pore, MPTP, results in intramitochondrial content release. Drugs that inhibit MPTP are

indicated.

Mitochondria in apoptosis pathways

The first indication that mitochondriaplay an important role in the induction ofapoptosis processes came from the obser-vation that a mitochondrial fraction wasrequired to induce apoptosis in an in vitrosystem (52). Although apoptotic stimulican be from different origin, most of themconverge at the mitochondria (Fig. 2).

When the nervous system is submittedto overstimulation by glutamate, as takesplace in ischemic processes, a massive andprolonged Ca2 + influx to the cytoplasm(82) has been observed. This entrance is“detected” by the mitochondria, as dis-cussed above. During ischemia, deathreceptors including TNF-R1 and Fas mayalso be activated. These receptors triggercaspase-8 activation and proteolytic cleav-

age of Bid, which subsequently translo-cates to mitochondria and increases theirpermeability (27, 30).

On the other hand, trophic factors andcytokines are able to activate the phos-phoinositide 3-kinase pathway. Thisresults in the generation of two lipidproducts (PI-3,4-P2 and PI-3,4,5-P3) thatact as second messengers and activate theserine threonine kinases Akt and PDK1(19). When a cell is deprived of trophicfactors and cytokines, these pathways arenot activated and Bcl-xL, an inhibitor ofmitochondrial permeability increases, isneutralized by the protein Bad. Finally,damage in the cellular genome can bedetected by the transcription factor p53,which is able to transcriptionally activatethe pore-forming protein Bax and otherpro-apoptotic Bcl-2-family members.



Fig. 2. Mitochondrion, headquarter in apoptotic pathways.

Mitochondrial permeabilitytransition pore

The MPTP is a multiprotein complexformed in the contact sites among theinner and outer mitochondria membranes.Cytoplasmic (hexokinase), outer mem-brane (VDAC), inner membrane (ANT)and mitochondrial matrix (cyclophilin D)proteins participate in its structure (32).

Under physiological conditions, thedifferent components of the MPTP arefound disaggregated (16). VDAC con-tributes to outer membrane permeabiliza-tion, ANT controls, in a specific way, theinflux through the inner membrane ofphosphorylated and non-phosphorylatedadenine nucleotides, and cyclophilin Dexhibit a peptidyl propyl isomerase activ-ity, which is crucial for protein folding(1). Some of these components can befound bound to other proteins. VDACassociates with the mitochondrial benzo-diazepine receptor and, in this manner,regulates extramitochondrial cholesteroltransfer to the intermembrane space (53).Moreover, at the contact sites, creatinephophokinase association facilitates theenergy transport through the creatine/phosphocreatine system (83).

When an apoptotical stimulus reachesthe mitochondria, the different MPTPprotein components can join to form apore of aprrox. 1.0 to 1.3 nm ratio. Thispore triggers the flux of molecules smallerthan 1500 Da in a non-selective manner.Its opening produces an inner mitochon-drial membrane permeabilization, result-ing in i) release of proteins and othersolutes from the mitochondrial matrixinto the cytosol; ii) a decrease in the elec-tric transmembrane potential, with theconsequence of ATP depletion; and iii)mitochondrial swelling, due to entrance ofwater, that eventually will break the outer

membrane, releasing several intermem-brane space components into the cyto-plasm.

There have been characterized at least79 peptide nature components that arereleased during MPTP opening (54).These include catabolic enzymes,endozepine and a multitude of ions andmolecules with known apoptotic activity,such as cytochrome c (Cyt c), Smac/Dia-blo (78), apoptosis inductor factor (AIF)(74) and some members of the caspasefamily, such as caspase-2, capase-3 andcaspase-9 (38, 73).

Once they have been released into thecytoplasm, these agents could activate dif-ferent apoptotic pathways. Cyt c is acofactor of a caspase-activating, cytosolicmultiprotein complex, the apoptosome.AIF release yields in chromatin condensa-tion and DNA fragmentation, and canfunction independently of caspases toinduce apoptosis (74).

MPTP regulation.– There are manyfactors that regulate the formation andopening of the MPTP. Calcium, ROS andmembers of the Bcl-2 family proteinsrelated to neuronal death processes, mod-ulate MPTP formation. In conditionswhere [Ca2+]c are found high in a main-tained manner, as it takes place in neu-rodegenerative processes, ischemia andexcitotoxicity, uptake uniporter systemsare activated, allowing Ca2+ entry intomitochondria, while the responsible sys-tems for the mitochondrial Ca2 + extru-sion are saturated. In these situations,mitochondria are not able to increase[Ca2+]m indefinitely, and eventuallyrelease their Ca2+. The mechanismresponsible for this liberation implies theincrease of the inner mitochondria mem-brane permeability though the MPTP for-mation. Conditions of oxidative stress andsubstrates such as phosphate, acetoacetate



and oxalacetate, are factors that stimulateCa2+ release from mitochondria throughMPTP formation. ROS are able to induceMPTP opening (3, 33), a process that isprevented by antioxidant agents such asvitamin E or glutathione.

Increases in [Ca2+]m or ROS levels areable to eliminate ANT specificity andpositively modulate its association withcyclophilin D (Cyp D). This association ismodulated by other factors such as ROSthat enhance the sensibility of ANT toCa2+. On the other hand, adeninenucleotides of the mitochondrial matrixand decreases in intracellular pH con-tribute to its insensitivity. Some Bcl-2protein family members have been relatedto MPTP. Bcl-2 inhibits MPTP forma-tion, producing mitochondrial membranestabilization, increases the capacity tobuffer Ca2+ (47, 49) and protects against∆Ψm depolarization (85). There are twohypotheses that help to explain how bcl-2inhibits the MPTP: its direct interactionwith a thiol residue of ANT sequence (15)or the capacity to close VDAC through itsBH4 region (69). Bcl-2 has been shown toefficiently block several apoptotic mecha-nisms, including ischemic injury, expo-sure to NMDA and neurodegenerationassociated with amyotrophic lateral scle-rosis, among many others (35, 36, 57).Bcl-xL, depending on its phosphorylationstate, regulates VDAC opening. Thus,Bcl-xL prevents the release of Cyt c fromthe mitochondria, induced by chemicalagents (70). Its BH1 domain is required toestablish the binding to the VDAC, whilethe BH4 is necessary and sufficient toocclude it (69).

The pro-apoptotic BCL-2 family mem-bers Bax, Bak, Bid and Bad (79), are usu-ally localized in the cytoplasm or areloosely associated with mitochondria.After post-translational modification(cleavage, phosphorylation processes)

they are able to translocate to mitochon-dria and to induce pore opening andrelease of Cyt c (34), processes inhibitedby Bcl-2 (7) and Bcl- xL (13, 22). It seemsthat this release induced by these factorsconcerns only the outer membrane, anddoes not imply changes in the potential ofthe mitochondrial membrane or mito-chondrial volume prior to the release ofCyt c. The Bax pathway seems to be relat-ed to its capacity to form channels in lipidmembranes. Its structure displays similar-ity with the pore-forming domains ofdiphtheria toxin and the bacterial colicins(2). Bax can be present in monomeric oroligomeric forms, and only the last one isable to form channels in lipid membranes(2). Bid, similar to Bax, displays a cyto-plasmic localization, but to activate apop-tosis, it requires partially hydrolysationby proteases, such as caspase-8 orgranzyme B (29), and to undergo subse-quent NH2-terminal myristilation inorder to translocate to mitochondria. Itsaction can be mediated by different mech-anisms, such as interaction with ANT(84), enlarging its activity (68) or induc-tion of the mitochondrial insertion of Bax(20). These effects are sensitive to thepresence of MPTP inhibitors (67).Although Bax, Bak and Bid induce Cyt crelease, the different proteins may carryout this process via different mechanisms.Under certain circumstances, Bax mayrequire the presence of ANT (46), induc-ing a drop in ∆Ψm, and its effects can beinhibited by inhibitors of the ETC such asantimycin, protonophores, KCN andoligomycin, while Bid does not inducechanges in ∆Ψm and can act independentof these drugs (67). Mitochondria are alsotargets of other protein kinases (proteinkinase C (44)), of transcription factors(p53 and Nur 77) and of viral proteins(protein R, Vpr (21)). The protein p53, in



conditions where it induces apoptosis,undergoes a quick and selective transloca-tion to the mitochondria, before changesin ∆Ψm, Cyt c release and caspase-3 acti-vation are observed (45). Other examplesare, Nur 77, also refered as TR3, thatundergoes translocation from the nucleusto the mitochondria, where it induces Cytc release (42); and the protein Vpr that isable of directly interact with ANT, pro-ducing a quick ∆Ψm dissipation (31).Currently there are drugs that modulateMPTP formation (75). Among them arecyclosporin A and bongkrekic acid whichexhibit significant pharmacological differ-ences. While the cyclosporin A binds withCyp D when associated with ANT (17),bongkrekic acid directly inhibits the ade-nine nucleotide translocator (9).

Mitochondria in neurodegenerativeproceses

Due to the multiple functions andnumerous proteins present in the mito-chondria, it is not surprising that geneti-cally inherited defects of mitochondrialfunction are a major cause of morbidityand mortality in humans. In particular,there are several human diseases that haveknown defects in the proteins responsiblefor oxidative phosphorylation (OxPhos)in cells. Typically, such defects producelactic acidemia, exercise intolerance orneurological disorders. As a result,mtDNA in somatic cells builds up muta-tions over time due to errors in replicationthat are not repaired. Such accumulatedmutations are implicated in a number ofneurodegenerative diseases (notablyParkinson’s and Alzheimer’s diseases)where the mutation load triggers prema-ture apoptotic or necrotic cell death. Forexample, a strong link has been estab-lished between exposure to the pesticide

rotenone, a well-defined and specificinhibitor of OxPhos, and Parkinson’s dis-ease. mtDNA mutations function byreducing energy production within thecell and are thought to contribute to can-cer and to ageing. Likewise, mutations inthe nuclear-encoded subunits of OxPhoshave been found to regulate the life spanin flies and worms.

For example, in the ataxia of Friedreich,the protein frataxin is mutated. Frataxinlocalizes to mitochondria and is requiredfor the maintenance of iron homeostasisand DNA content (37). Another exampleis the hereditary spastic paraplegia, wheremutations of paraplegin occur, a mito-chondrial metalloprotease, that give riseto defects in the oxidative phosphoryla-tion (11). In postmitotic systems, such asthe nervous system, mitochondrial somat-ic mutations accumulate and, added to thedescent in mitochondrial function, can because of ageing and senescence (80). TheETC activity is reduced or compromiseddue to an inadequate oxygen level in thecells as takes place in arteriosclerosisprocesses, anaemia, as well as in alco-holism situations (77). Patients withhereditary optic neuropathy of Leber pre-sent mutations in the ETC complex I (8),and a reduction of its activity has beenobserved in subjects with Parkinson’sdisease (66). In this sense, some neurotox-ins or their metabolites may cause orcontribute to Parkinson’s disease. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyri-dine (MPTP), and its metabolite MMP+

are inhibitors of complex I and are drugsutilized to induce symptoms of Parkin-son’s disease. In neuronal cultures, MPP+

is able to induce Cyt c release and adecrease in cellular viability (40). A loss of∆Ψm, which may be a consequence of for-mation and opening of MPTP, has beendescribed in cells originating from indi-



viduals that suffer from Chorea Hunting-ton or Alzheimer’s disease (12, 63), as wellas in experimental models for these disor-ders such as 3-nitropropionic (14) or β-amyloid peptide toxicity (57). The MPTPalso performs an important role in theprocesses of cellular death induced byactivation of receptors located in the cel-lular membrane, such as Fas (FasR). Theactivation of these receptors implies theactivation of the caspase pathway, andmitochondria function here in amplifyingthe death signal (76). The activation ofFasR is triggered by the trimerization ofthe receptor, recruitment of Fas associateddeath domain (FADD), and formation ofa caspase-8-activating complex, the deathinducing signal complex (DISC). Cas-pase-8 is able to cleave Bid into a truncat-ed form that induces Cyt c release frommitochondria. The blockade of MPTPformation protects cell cultures fromapoptotic stimuli. As we have commentedpreviously, in the ischemic processes anexcessive glutamate receptor activationoccurs that results in an increase in [Ca2+]cinducing its accumulation in the mito-chondria and subsequent MPTP opening.In cell cultures where accumulation is pre-vented by means of drugs able to dissipatethe electric potential, such as theprotonophore FCCP, cellular deathprocesses are prevented. Hence, MPTPformation may be crucial for the activa-tion of signaling cascades that result in celldeath, while excessive cytoplasmic calci-um levels may be tolerated (72). The inhi-bition of ROS production, by means ofdrugs with catalase or superoxide dismu-tase activities, has been shown to efficient-ly prevent MPTP formation, both in iso-lated mitochondria, and in cell culture.EU-134, a drug with superoxide dismu-tase activity, blocks neuronal deathprocesses in different models, such as

NMDA exposure or by treatment withthe protein kinase C inhibitor, stau-rosporine, that has extensively been usedas an apoptosis inductor in several cellularmodels (56, 61). Finally, MPTP blockers,such as cyclosporin A (CsA) andbongkrekic acid, also exhibit neuropro-tective effects. Although CsA has manyother actions, it is likely that its capacityto bind to Cyp D is responsible for itscytoprotective effects. Other drugs that,like CsA, are able to inhibit calcineurin(such as the inmunophilin FK506), do notshow comparable protective effects. CsAprevents decreases in cellular viability inneurotoxicity models of Huntington dis-ease (10, 12, 41 63, 65, 82) or in hip-pocampal areas affected by ischemicprocesses (50). Bongkrekic acid, withcapacity to bind to ANT, blocks the inter-action of the translocator with Bax andhas been shown to inhibit caspase-9release from brain mitochondria. Thisdrug offers neuroprotection againstischemic injury in the brain, and protectsneurons against NMDA-induced excito-toxic injury. In the latter setting, it alsoprotects against the drop in ∆Ψm andATP levels (9).

We have summarized the currentknowledge supporting the key role thatmitochondria play in multiple cellulardeath processes and that many differentmitochondrial cell death pathways involvethe formation of the MPTP. The fact thatthe formation and opening of MPTP is aprocess controlled by a great variety ofstimuli, may make it an interesting phar-macological target that would allow inter-ference with neurodegenerative processesand acute nerve cell injury.

Acknowledgements

This work has been supported, in part, by grantsSAF2002-04721 from CICYT to J.J. SAF99-0060



from CICYT, BFI2001-1565 from Ministerio deCiencia y Tecnología; G03/167 from Ministerio deSanidad, GC-02-019 and PAI-02-031 from Conse-jería de Ciencia y Tecnología, JCCM and from Fun-dación Campollano-Banco Santander Central His-pano to V.C. and JHMP is supported by theDeutsche Forschungsgemeinschaft. We are gratefulto Juana Rozalén and Daniel Tornero for invaluablediscussions.

J. JORDÁN, V. CEÑA y J. H. M.PREHN. Control mitocondrial de la muerteneuronal y su papel en las enfermedades neu-rodegenerativas. J. Physiol. Biochem., 59 (2),129-142, 2003.

Las alteraciones mitocondriales, genéticas ofuncionales, pueden dar lugar a iniciación deprogramas de muerte celular que han sido rela-cionados con patologías como diabetes, enve-jecimiento y alteraciones neurodegenerativas.La mitocondria está siendo considerada comoel principal lugar de unión de las señales deestrés activadas en el daño neuronal, tantoagudo como crónico y en los procesos de eje-cución de la muerte celular. Esta segunda fun-ción mitocondrial está regulada por variasfamilias de proteínas que conllevan a un incre-mento en la permeabilidad de las membranasmitocondriales externas y/o internas. Un ejem-plo de estos procesos es la formación del porode permeabilidad transitoria mitocondrial(MPTP). Este proceso puede deteminar la libe-ración de factores inductores de muerte celulardesde la mitocondria, así como la disipacióndel potencial de transmembranal mitocondrial,disminución de los niveles de ATP e incremen-to en la producción de radicales libres. Dentrode los factores liberados desde la mitocondriase encuentra el citocromo c, el factor inductorde apoptosis (AIF) y las caspasas. Se revisa eneste trabajo el papel del MPTP en diversosprocesos fisiológicos y patológicos, que inclu-yen enfermedades neurodegenerativas como laenfermedad de Alzheimer, de Parkinson y laesclerosis lateral amiotrófica (ALS). El diseñode fármacos que puedan interferir con las fun-ciones del MPTP podrían permitir nuevasaproximaciones terapéuticas en el tratamientodel daño neuronal.

Palabras clave: Apoptosis, Necrosis, Apoptosoma,Caspasa, Poro de permeabilidad transitoria,

Mitocondria.

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