Amyloid-senile-plaques and neurofibrillary-tangles in elderly Schizophrenics non-treated and treated by neuroleptics

THIRD

S98

The neuroeclodermderived~trwtws of the eye are in intimatecontact with the ocular fluids (aqueous and vitreous humor). The trace element composition of the fluids I” the human ocular fluid compartment in health and in association with neurodegenerative diseases, has not been investgated in detail to date. Elemental analysis of ocular fluids collected at post-mortem exammation has been undertaken to evaluate 11s uselulness in the assessment of trace element accumulation m a companment relatively protected from the effects of post-mortem contamination by diffusion of serum electrolyte and trace elements the work described. the elements Al, Ca. Cd, Co. Cr. Cu. Fe. Hp. K. Mg. Mn. MO, Na. NI, Pb. Se, SI. Sn and Zn were determined on a commercial ICP-MS system. The technique uees an atmosphervz pressure plasma ion source coupled :o a qwzdrapole mass spectrometer end is cherecterised by a working concentration range of six orders of magnitude. rapId multi-element analysis, simple mess spectra, comparative freedom from spectral

In

interference and sub ng.ml-1 limits of detection for most elements. A mayor problem encountered in biological mineral analysis IS the risk of sample contammetion durmg collection and analysis. The methods developed to overcome this problem are described. In addmon a wet-oxidation method which proved suitable to prepare paired samples of ocular fluld and serum for direct introduction into the ICP.MS instrument will be described together wth system operation. The results of thts study indicate that elemental concentrations I” human ocular fluid are generally of the same order of magnitude es in serum and offer some guidellne values lor Ihe elemental compositmn of a hereto little investigated human body fluid. as well as showing the applicability of ICP-MS to the analysis of biological materials.

INTERNATIONAL

CONFERENCE

ON ALZHEIMER’S

DISEASE

D,L-a-dipalmltoylphosphatldylcholine (DPC) leads to the appearance of two 31~ signals due to the free and complexed ligand. Owing to reproducibility problems observed in the water-based system, we moved to CDC13, in which medium 31P, IH, 27Al and, in principle, l3C NMR is likely to offer detailed and corn lementary information on the AI(III) - biomolecule interaction. 31 P ( pH) spectra of a 5x10-3 M Al(III)-PCD solution exhibit two evident signals, with comparable shape and intensity. Important changes in the 27Al and 1H spectra are also observed. Thus, the 1H signals due to the hydrogen atoms adjacent to the phosphatidic group undergo a marked broadening effect, while the signals due to the remote hydrogen atoms result to be quite unaffected by Al@) - PCD interaction. References 1. R. Vierstra and A. Hang (1978). Biochem. Eiophys Res. Commun., 84, 138. 2. B. Corain et al. (1991) :In “Alzheimer’s Disease: Basic Mechanisms, Diagnosis and Therapeufic Sfrafcgies”, K.Iqbal. D.R.McLachlan, B.Winblad and H.Wisniewski (Ed.s); John Wiley & Sons Ltd. New York, 394 (Chichester, GB). 3. M. Deleers et al.(l986)$iochim. Biophys. Am, 855, 271. 4. B. Corain et al. (1992) Coord. Chem. Rev., 112, 19.

387 The Solution State of Atuminum(III) uAIgdroxyaw%oxylates and its Relevance to Aluminum Toxicology

Prof.Dr.med.E. Y. Funfgeld, ““I”.

Aluminum s0lubility in neutral aqueous solutions in the absence of ligands different fromH~0andOH-isLmowntobeca10~M[1l.lnthepresenceofgoodby~~~c ligands the solubility csntemucbhigher, but the actual molecular identityof tlx species present in solution may not agnx with the thermodynamically predicted metal speciaof kineticeffea [Z]. tiOSaSthec0~ Severalaluminumcarboxylates.amongwhich lactate.,&ate., tar&ateandgluconate, are kii used since long for preparing0.01-0.1 M solutions appsnmdy stable at pH ca 7, to be employed inexperimental pathology investigation. As tbe observed stability is in conb-adictionwith thermodynamic expectations in tbe cases of lactate. tmlrale snd gluconate, we have carded out a thorough study of k solution state of the systems Al(Ill)/H~O/oH~/csrboxylste..Al@c1ate.)3 (1) (commercislly available) was cbamcterized in the solid state by X-my single crystal analysis and lR speclmmetry; the novel complexes &(cibate)@20)6 (2), Al&whate)3(H20)z (3) and A@x0natWX92 (4) have beenprepared and characteti in Ibe solid state by IR spech-ometry. Comb&d lR and NMR (‘H and 13C)analyses in DzO have shown mat 1 and 3 exist mainly as highly metastable aqua-hydmxo species at pH 7.5; on the contrary, 2 and 3 appear to maintain in neutral solutions molecular sm~ctures(related to those observed ln the solid state) in which the carboxylate ligamis persist inside lbe metal coordination sphere [31.

References l.B.Corainetal.(l~)Coord.Chem.Rev., 112.19 2. B.N.k&tin (1991)in “Aluminumin Chemistry,Biologyand Medicine”,vol. 1, M. Nicolini.p. Zaas. B. Cckn (eds).CortinaInternational- RavenPress,Vemns,p. 3 3. B. Cominezal. (1992)J. Chem.SOL,D&on Trans., 169

Ilerburp,

Zinc is one of the most ?? rsentiel co2 trsnrport;

further

Modelling the Interaction of Membrane Phospholipid Components with AI(III) in Water and Aprotic Solvents NMR Approach

Stuttge~t Llteraturc.

Al(III) is known to be an effective aggressor on biological [ 1,2] and artificial phospholipid-based [3] membranes. We have recently proposed[3] that the chemical interaction of Al(III) with the phosphate head groups is likely to be the cause of the observed damage to the delicate membrane supramolecular order.We are utilizing 1~. 31P and 27Al nuclear magnetic resonance spectroscopy (NMR) to study the interaction of membrane phospholipids with the neutral hydrolytically stable Al(II1) complexes (Al(acac)3 (Hacac= 2,4-pentancdione) and Al(malt)3 (Hmalt= 3-hydroxy-2-methyl-4H-pyran-4one) [4] both in aqueous (Tris buffer + Triton X-100) [5] and organic media. In both solvents the interaction of equimolar amounts of Al(III) and

trece

?? lemmts,

ir a nccerrary

WI).

HU~~OUI

c.g.for

co-t

of cell

the cell

respxration

and the

nembrenes.

cMny ~t~“ct”~aL and IeteboLiC procerrea such a(i and even the ryntherir and degradation of nucle~nic

mryra

Zinc is mainly stored in certein ?? rytrocyte*. Given e* rdicetion

ere influnced

acids

we found en inconsistency

and his whoLe blood.

Ye folloued

therapeutic

agent

in LLzheirr

by Zinc - 2L to 200 according

to the

pleccr of the brain, in the Lwer, kidney and Zinc move* quickly tr~n the blocd to tlwre plscer.

In our ~nvertigdtion

e suggestion patients.

of Zinc levelr

I” a patrent’s

by J. Co”rtant?nidls

Our patient

bleed

who prqrored

group (50 patients,

~OPUOI

ZI”C 8% e

age range 36-81

y.1 included ceses of Perkinronin, multiple rclcros~r, early chiLdhood bra?” dirturbancer,ar!d neurologicel systemic dqnerative ~LLnerses. Pat?~“ts seLected project shwed Lw Zinc veLu+t end *Loued brein U~YCI as recorded by ccwcnt~onal well a, by oynanrc mein l!aP@lg systm
for rhlr EEG as

In rare fha” Box of the peticntr, the i”trwewu~ explication of 24 to 30 nq l~nc srpartate rerultd in e topqrephic change of percentages and dirtributlon of the frequmcy

This

band* within

M

to 60 min.

Leeds to the hypothesis:

patmtial

ma, P. Zambenedetdb. B. Cortinb and P.F. Zattac .a Istituto C.N.R.I.C.T.R., b Dipartimento di Biologia, c Dipartimento di Chimica Inorganica Metallorganica Analitica , d Centro C.N.R. Emocianine, 35131-Padova OtalY)

Feculfy

?? specrslly irnuwlopy, Deteits with fevoreble therapeutic findlnps r” internal ledrsinc, obstetric* *nd dervtotogy ~c~c prbli*hcd by H.Y. Wtneier and Y. Kruse-Jerres (“Zink”,

e “cy class

:A

it

zinc a,*0 control* mid rqutete* carbahydretes, Lipids, proteins and hormmcs.

Zinc effects

388

“ed. ~rbllrgleenny

w-,550

ere ultilocetcd. of therapeutic

effects

etso on

Thus Zinc belongs eg~lrtt brein

This should Lead to e “CY therapeutic

with

to

significant

disturbances. concept.

390 AMYLOID-SENILE-PLAQUES AND NEUROFIBRILLARY-TANGLES tN ELDERLY SCHIZOPHRENICS NON-TREATED AND TREATED BY NEUROLEPTICS J. COnSlaOtlnldla Geneva University, Psychiatric InstMiins After the age of 70 years, 60% 01the brains 01 patients dead in the general hospital ot Geneva presented amyloid (AM)-senile plaques (SP). Many authors agree that the PrfmsrY, AH formation in the brain, Induces disturbances, producing secondarily

the neurofibrillarytangles (NFT) [Constantinidis 8 Tissot 1979; Constantinidis & Richard 1985: Mann 1988; Wisniewski 1989; Selkoe 19911. We emitted the

THIRD

INTERNATIONAL

hypothesis

CONFERENCE

that, the AM-induced

by the chronic neuroteptics.

edmlnlstratlon chronically

ON ALZHEIMER’S

be InhIbited

production

of NFT may

of

we tried to verify this hypothesis

drug,,

administered

investigated for the presence of AMSP

to schizophrenics

since

DISEASE

or rnhancrd.

1953,

for the and

and NFT. all the brains of schizophrenics

we who

died at more than 70 years, and came to autopsy in the psychiatric hosptial of Geneva: 41 of them

non treated

treated by neuroleptics

by neuroleptics

(dead

from

1932

through

1952)

and 61

The mean age at death of the

(dead from 1953 through 1990).

lwo groups being identical, about 79 years, as was also the incidence of AMSP: 60% in both groups, identical to that observed in the general population of same age in Geneva, the incidence of NFT smong the brains presenting AM.SP, has been found to be hvlce treated

hlghor

In the group

brains of elderly schizophrenics and without interference enhance

the

by neuroleptics

Potential

for

to the cllnlcel

with the same frequency

with the neuroleptic

AM-Induced

disturbances

therapeutic

drugs (to be discovered)

1979:

treated

then

In the group

non

(74% versus 36% p
Intervention:

may lnhlblt

treatment, leading

to the

the chronic

the AM-induced

Symptoms of dementlb.

as in the general population and that nouroloptlcs

and therefore,

formation

administration

production prevent,

Schweiz. Arch. Neural. Psychiat. 124: 317-333: 1985:

of the

may NFT.

of other

01 the NFT, related

stopordelaythem.

Alzheimer’s disease. in Handb.

Clin. New-al. 46: 247.282; 1988: Alz. Dis. Ass. Disord. 2: 224; I9 8 9: J. Neural. Transm. (P-O) 1: 7; 19 9 1: Nature 353: 432433.

The voltage-dependent anion channel (VDAC) is the most abundant protein in the outer membrane of mitochondria (OMM) where it selves as the major permeability pathway. When studied in planar bilayers(in symmetrical 1 M KCI). this channel displays a high conductance(of approx. 4.0 nS) at low values of the membrane potential, whereas, upon increasing of voltage, it closes to several subconducting states. Given the possible neurotoxicological action of aluminum(Al), we have studied the effect of two different AI(III) compounds, AICl3 and aluminum lactate (AlLaoQ), on the detergent-free OMM isolated from rat brain and inserted in planar bilayer (in symmetrical 1 M KCI. pH 7.2). The experimental conditions used were such that one channel was present in the bilayer, this permitting to follow the behaviour of a single VDAC molecule. The resutts obtained showed that there was a dramatic effect on the channel electrical characteristics when the imposed voltage was positive, relative to the side where AI(III) was added. In fact, at either 1OOpM AICl3 or 10 uM AILact3, imposition of 20-30 mV reduced the conductance of the channel to approximately 50%. Moreover, there was also an increase in the rapkf open-close transitions. At negative voltages, but only prior to imposition of positive potentials, the channel had the tendency to remaining in the open state. Though our experimental conditions have allowed a better definition of the aluminum effect, the results resemble closely those In our work the found by Dill et al.[l] in multiVDAC-containing bilayer. AI(III) species responsible for the action on VDAC seems to be AI(OH)4-. As it appears, at pH 7.2 the concentration of AI(OH)4- is vanishing small so to render this species extremely effective on biological targets. Remarkably, the change in VDAC behavlour provoked by AI(III) remained despite the washing out operation and thus pointing to an apparently irreversible binding with, and alteration of, VDAC protein. Additionally, we found that AI(III) increased the stability of pure phospholiptd bilayer, and hence that it stably interacted also with the polar phospholipid headgroups[2]. 1) Dill et al. (1987) JMembr. Biol. 99: 187-196 2)Corain B, Nicolini M. Zatta P. (1992)Coord. Chem. Rev. lj_2: 33-54.

549 SERUM ALUMINUM-BINDING PROTEINS: CRARACTRRIZATION ALBINDIN, A NEW CARRIER PROTEIN, C. Mizzen,

OF

M. B. Krishnan, T. Kruck and D. Favarato*, E. Jaikaran, McLachlan. Centre for Research in Neurodegenerative Diseases; University of Toronto, Toronto, Ontario, Canada M5S JAB and *Department of Biology; University of Padova, 35131 Padova, Italy. Elevated levels of aluminum in post-mortem brain is a common feature of Alzheimer's disease (AD) and suggests that aluminum may participate in the the role of etiopathogenesis of AD. To investigate serum aluminum in AD pathogenesis, we have developed chromatography (SEC) method for a size exclusion monitoring aluminum speciation in serum. This method reveals that two principle carriers of serum aluminum exist in most individuals, transferrin and a fraction we have named albindin. Albindin is proteinaceous and does not appear to be related to transferrin. The apparent molecular mass of aluminum-albindin complexes is approximately 5,300 daltons. Aluminum binding by albindin in vitro does not require exogenous bicarbonate and aluminum-albindin complexes resist chelation in vitro by EDTA and desferrioxamine. Typically, aluminum-albindin complexes are not detected in individuals whose serum aluminum levels are normal (t200 nmol/l). However, sera of untreated hyperaluminemic patients and sera from patients with either normal or elevated initial serum aluminum levels during therapy with desferrioxamine all contain aluminum-albindin complexes. These observations indicate that further investigation of the role of albindin in aluminum transport, in particular the fate of aluminum-albindin complexes, is necessary to understand mechanisms of aluminum toxicity and to develop appropriate therapies. Acknowledgements: Financial support from the Alsheimer Society of Peterborough and Victoria, Parke-Davis Inc. the Scottish Rite (Italy), Charitable Foundation of Canada, National Health and Welfare Canada and the Hunter Foundation is gratefully acknowledged.

550 EFFECT

OF DIFFERENT

ALUMINUM

COMPOUNDS

ON THE

OUTER MITDCHONDRIAL MEMBRANE CHANNEL,VDAC. aT.Mirzabekov, aC.Ballarin, aM.C.Soraato. bM.Nicotini and cP. Zatta. aDip. Chimica Biologica e Centro CNR di Fisiologia Mitooondriale, Universita’ di Padova(ltaiy),bDip. Scienze FarmaceuticheUniversita di Padova (Italy), CCentro CNR Emocianine-metalloproteine, Padova (Italy).

ANIMAL AND CELLULAR MODELS

391 MOLECULARGENETIC STUDIES OF PRION DISEASESAND TRANSGENICMOUSE MODELSOF CNB DEGENERATttX Stanley B. Prusiner. Departments of Neurology and of Biochemistry and Biophysics, University of California, San Francisco, CA 94143, USA. Many studies of the infectious pathogens causing scrapie and other transmissible neurodegenerative diseases support the hypothesis that these pathogens are novel and different from both viroids and viruses. After convincing evidence was obtained showing that scrapie infectivity depends upon a protein component, the term “prion” was introduced to distinguish these infectious pathogens from others including viroids and viruses, Enriching fractions from Syrian hamster (SHa) brain for scrapie prion infectivity led to the discovery of the prion protein (PrP). Prion diseases include scrapie of sheep, bovine spongiform encephalopathy of cattle, as well as Creutzleldt-Jakob disease (CJD) and Gerstmann-Straussler-Scheinker syndrome (GSS) of humans. Transgenic (Tg) mice expressing both SHa and mouse (MO) PrP genes were used to probe the molecular basis of the species barrier and the mechanism of scrapie prion replication. Bioassays of brain extracts from two scrapieinlected Tg lines showed that the prion inoculum dictates which prions are synthesized de nova. even though the cells express both PrP genes. Our results argue that the species barrier for scrapie prions resides in the primary structure of PrP and formation of infectious prions is initiated by a species-specific interaction between PrPSc in the inoculum and homologous PrPC. Diswvery of mutations in the PrP gene from humans with GSS and familial CJD established that prion diseases are unique among human illnesses - they are both genetic and infectious. Tg mice expressing MoPrP with the GSS point mutation spontaneously develop neurologic dysfunction, spongiform degeneration and astrocytic gliosis. Inoculation of brain extracts prepared from these Tg(GSSMoPrP) mice into Syrian hamsters and Tg mice expressing wild-type PrP transgenes has produced neurodegeneration in recipient animals after prolonged incubation times, If wnvincing data on serial passage of prions from the inoculated recipients can bs obtained. then these results will argue that prions are devoid of foreign nucleic acid. Although it seems likely that transmissible prions are composed only of PrPSc molecules, a hypothetical second component such as a small polynucleotide remains a formal possibility. Studies on the structure of PrPSC and PrPc have been unsuccessful in defining a posttranslational chemical modification that distinguishes one PrP isoform from the other. These findings suggest that the difference between PrPSC and PrPC may be conformational. Whether distinct prion isolates or “strains” with different properties result from multiple conformers of PrPSc remains to be established,