\PERGAMON Journal of Psychiatric Research 21 "0887# 300Ð319

9911Ð2845:87:,08[99 Þ 0887 Elsevier Science Ltd[ All rights reserved[PII] S 9 9 1 1 Ð 2 8 4 5 " 8 7 # 9 9 9 2 2 Ð 7

Myo!inositol in depressive and healthy subjects determined byfrontal 0H!magnetic resonance spectroscopy at 0[4 tesla

R[ Freya\�\ D[ Metzlerb\ P[ Fischera\ A[ Heidena\ J[ Scharfettera\ E[ Moserb\c\ S[ Kaspera

a Department of General Psychiatry\ University Hospital of Psychiatry\ A!0989 Vienna\ Austriab NMR `roup\ Institute for Medical Physics\ University of Vienna\ Austria

c Clinical MR unit\ University Hospital of Vienna\ Austria

Received 01 December 0886^ revised 15 May 0887^ accepted 7 July 0887

Abstract

Myo!inositol "mI# as a precursor in the phosphatidylinositol second messenger system has been reported to be reduced indepression[ By means of proton!magnetic resonance spectroscopy "0H!MRS# the mI levels in the frontal brain were investigated invivo in the present study[ Twenty!two patients "mean age] 31[7209[6 years# with depressive episodes according to ICD 09 "HAMDscore × 06# were compared to 11 healthy subjects "17[924[2 years#[ Two voxels "29×19×19 mm2# in the frontal lobes were examinedin a Siemens Magnetom SP 3999 at 0[4 T "STEAM sequence] TR � 2499 ms\ TE � 44 ms#[ With the total creatine "Cr# as an internalstandard\ mI:Cr ratios were calculated to follow the mI levels[ In the left frontal lobe\ mI:Cr was 9[3229[95 in depressive patientsand 9[3529[96 in healthy subjects^ concerning the right frontal lobe\ mI:Cr was 9[3529[97 and 9[3729[95\ respectively[ There wereneither signi_cant di}erences between the two groups nor between the hemispheres[ Since there was a signi_cant positive correlation"R � 9[5# between the age and the mI:Cr in the right frontal lobe of depressed patients\ age matched pairs analysis was performed"n � 1×09\ in each group] nine females\ one male\ ³ 39 years#[ In the right frontal lobe\ the patients| mI:Cr of 9[3929[94 was nowsigni_cantly lower than the controls| mI:Cr of 9[3429[95[ However\ most of the patients were on antidepressive medication[Interestingly\ it was exactly this group of patients which showed signi_cantly lower mI levels[

We regard our investigation as a pilot study which suggests an in~uence of age and antidepressants on mI levels and should betaken into consideration in further investigations in depressive patients[ Þ 0887 Elsevier Science Ltd[ All rights reserved[

Key words] Myo!inositol^ 0H!magnetic resonance spectroscopy^ Depression^ Antidepressive medication

0[ Introduction

Phosphatidylinositol "PI# is a major component of neu!ronal cell membranes[ The phosphoinositide cycle "Fig[ 0#has been discovered as a major second messenger system"Berridge and Irvine\ 0878#[ Receptor stimulation byneurotransmitters activates a phospholipase C enzyme ina number of membrane receptor signalling pathways[Phospholipase C triggers the breakdown of pho!sphatidylinositol!bis!phosphate "PIP1# to inositol 0\3\4!trisphosphate "IP2#\ which releases calcium from internalstores[ A series of phosphatases remove the phosphategroups from IP2 sequentially\ releasing free inositol[Inositol is a 5!carbon cyclic polyol carbohydrate chemi!cally closely related to glucose[ The biologically activestereoisomer is called myo!inositol "mI#[ It is the theoriesabout the action of lithium which have aroused the inter!

� Corresponding author[ Tel[] ¦32039399!2426^ Fax] ¦32039399!2439

est in the phosphoinositide "PI# cycle "Fig[ 0# and its rolein a}ective disorders[ Lithium reduces the brain levels ofinositol by inhibiting inositol monophosphatase"Hallcher and Sherman\ 0879#[ Its therapeutic functionin mania and depression might be a result of the mIdepletion which slows down the PI cycle in overactivesynaptic systems "Berridge and Irvine\ 0878^ Snyder\0881^ Kofman and Belmaker\ 0882#[

In cerebrospinal ~uid\ a markedly reduced level of mIhas been reported in depressed patients with unipolarand bipolar a}ective disorder "Barkai et al[\ 0867#[ Myo!inositol levels in the human brain were analysed by gas!liquid chromatography post mortem "Shimon et al[\0886#[ These data revealed signi_cantly reduced mI levelsin the frontal cortex of bipolar a}ective disorder patients"n�7# and suicide victims "n�09# as compared withcontrols "n�09#[ Inositol levels were 7[621[3 mmol:kgin the normal sample\ but only 4[821[0 and 5[621[1mmol:kg in bipolar disorder and suicides\ respectively\i[e[\ reduced by 19Ð29)[ There were similar but stat!istically non!signi_cant intergroup di}erences in the

R[ Frey et al[:Journal of Psychiatric Research 21 "0887# 300Ð319301

Fig[ 0[ Phosphoinositide cycle] Phospholipase C triggers the breakdown of phosphatidylinositol!bis!phosphate "PIP1# to inositol 0\3\4!triphosphate"IP2#\ which has a function as a second messenger[ Lithium reduces brain levels of myo!inositol by inhibition of inositol!phosphatase[

occipital cortex[ The authors found no correlationbetween age and cerebral inositol levels and no signi_cantdi}erence between inositol levels in males and females[

High doses of exogenously administered inositol maypass the bloodÐbrain barrier su.ciently "Spector\ 0877^Levine et al[\ 0882#[ Since mI has been widely appreciatedas a key metabolic precursor of a second messenger sys!tem for numerous neurotransmitters\ controlled trials ofinositol treatment have been published "review paper]Levine\ 0886#[ Under double!blind conditions\ the intakeof inositol 01 g:day led to an improvement in depression\which was signi_cantly superior to placebo at week 3"Levine et al[\ 0884#[

Magnetic resonance spectroscopy "MRS# o}ers uniquepossibilities for non!invasive studies of biochemistry inthe human brain in vivo[ An atomic nucleus with a spindistinguished from zero has properties like a little mag!netic dipole[ In a strong external magnetic _eld\ due tothe quantum nature of matter\ each magnetic dipole orspin aligns parallel or antiparallel to this _eld[ As a resultthe spins are split into two distinct groups with di}erentenergy levels and slightly di}erent population densities"Nuclear Zeeman e}ect#[ Transitions between theseenergy levels may occur by absorption or emission ofelectromagnetic radiation\ in particular at the so calledLarmor or resonance frequency[ As the external magnetic_eld is shielded by the electron con_guration of eachmolecule\ the di}erences in energy\ or frequencyabsorbed\ depend not only on the external magnetic _eldstrength and the particular species of nucleus "e[g[\ 0H or20P# but also on the speci_c chemical compound[ Protonswithin di}erent molecular structures absorb and reemit

energy in the radio frequency range at slightly di}erentfrequencies because of small di}erences in the local mag!netic _eld induced by chemical binding[ Thus\ proton"0H#ÐMRS allows to detect biochemical compounds dueto substance speci_c di}erences in resonance frequency[This frequency shift is referred to as {chemical shift|[ Agraph of the magnitude of energy absorption as a func!tion of frequency is called MR spectrum "Fig[ 1#[ It iswell known\ that the main resonance of mI appears at achemical shift of 2[45 ppm "parts per million as a measureof normalized frequency# in human brain "Michaelis etal[\ 0880#[ Compounds must be present at concentrationsof approximately 9[4Ð0[9 mmol:kg or greater to be visiblewith MR techniques used in human subjects in vivo[ 0H!MRS studies published so far indicate that the free "MRSvisible# mI concentration ranges between 2[8Ð7[0mmol:kg in adults "review paper] Henriksen\ 0884#[ Incontrast to mI\ the isomer scyllo!inositol yields a res!onance at 2[24 ppm[ Scyllo!inositol concentrations ofabout 9[4 mmol:kg were identi_ed in 0H!MRS of humanbrain at 1[9 T "Michaelis et al[\ 0882#[

Phosphorous!20!MRS showed abnormal phos!phorous metabolism in bipolar disorder "Kato et al[\0880\ 0881\ 0882^ Deicken et al[\ 0884# in the frontal lobe\presumably due to deviations in the phosphatidylinositolcycle[ Moreover\ the most consistent "but not speci_c#SPECT and PET _ndings in depressives have beenregional de_cits in the cerebral blood ~ow and metab!olism of the frontal lobe "Hurwitz et al[\ 0889^ Lesser etal[\ 0883^ Kasper et al[\ 0883^ Mozley et al[\ 0885#[ Thus\the left and right frontal lobes are the regions of interestin this investigation[

R[ Frey et al[:Journal of Psychiatric Research 21 "0887# 300Ð319 302

Fig[ 1[ Proton "0H# spectrum of human brain tissue] The spectrum was obtained after water suppression\ and with a relatively short echo time "44ms#[ Resonances of cerebral metabolites are due to N!acetylaspartate "NAA#\ total creatine "Cr#\ choline "Cho# and myo!inositol "mI#[ Cr resonancesat 2[92 ppm\ mI resonances at 2[45 ppm "chemical shift in parts per million#[ Note that no _ltering whatsoever was applied to improve the appearanceof the spectrum[

We hypothesized that reduced mI concentrationsmight be detected by means of 0H!MRS in the frontallobe of depressive patients when compared to healthysubjects[

1[ Methods

1[0[ Subjects

0H!MRS was performed in 11 patients between 01:84Ð97:85 and compared to 11 healthy volunteers[ The groupsare described in Tables 0 and 1] females and males agedbetween 07Ð59 years[ The diagnostic entry criteria for thestudy were depressive episodes in unipolar or bipolardisorder according to ICD 09 "F21\ F22\ F20[2\ F20[3#[The Hamilton Depression Scale "HAM!D#\ 10 items"Arbeitsgemeinschaft fu�r Methodik und Dokumentationin der Psychiatrie\ AMDP\ 0875#\ scored higher than06^ case history and symptomatology was documented"AMDP\ 0884#[ In the present sample\ nine patients havea family history of a}ective disorder "Table 1] Nos 2\ 3\4\ 8\ 00\ 01\ 04\ 08\ 11#[ All patients were hospitalized[The current antidepressive medication was continued^tricyclic antidepressants "TCA#\ selective serotonin re!uptake inhibitors "SSRI#\ the reversible monoamine oxi!dase A inhibitor "RIMA# moclobemid as well as trazodonand mianserin are mentioned in Table 1[ Some patientsreceived benzodiazepines in addition[ Patients with lith!

Table 0mI:Cr!ratio and demography in healthy subjects

Subject mI:Cr left mI:Cr right Age Sex

0 9[285 9[264 17 f1 9[276 * 14 m2 9[280 9[492 29 m3 9[371 * 19 m4 9[421 * 15 m5 9[458 9[494 16 f6 9[331 9[494 36 f7 9[549 9[377 16 m8 9[322 9[366 15 m

09 9[327 9[356 12 f00 * 9[322 15 f01 9[310 9[429 20 m02 9[406 9[423 16 m03 9[353 9[372 17 f04 * 9[276 21 f05 9[262 9[319 18 f06 9[497 9[520 15 m07 9[341 9[448 11 f08 9[350 * 14 m19 9[308 9[383 20 m10 9[261 9[312 16 m11 9[341 9[304 21 fx 9[347 9[368 17[9S[D[ 9[969 9[953 4[2

ium or anticonvulsant!prophylactic therapy were notincluded[ Further exclusion criteria were dementia\ alco!

R[ Frey et al[:Journal of Psychiatric Research 21 "0887# 300Ð319303

Table 1mI:Cr!ratio and demography in depressive patients

mI:Cr m:Cr AntidepressivePatient left right Age Sex ICD!09 HAM!D medication

0 9[323 * 24 f F22[0 12 SSRI1 9[337 9[247 22 f F22[0 192 9[267 9[272 41 m F20[3 17 TCA3 9[256 9[335 36 m F22[0 07 RIMA4 9[346 9[352 49 f F22[0 11 RIMA5 9[405 9[389 48 m F20[2 12 SSRI¦Mianserin6 9[369 9[331 28 f F22[0 08 SSRI7 9[437 9[592 40 f F22[0 088 9[308 9[304 14 f F22[9 07 SSRI¦Trazodon

09 9[327 9[396 28 m F22[0 19 SSRI¦TCA00 9[399 9[285 14 f F20[2 1001 9[256 9[447 45 m F22[1 12 TCA02 9[321 9[335 26 f F22[1 15 SSRI¦Mianserin03 9[496 9[401 48 m F22[0 1904 9[280 9[326 25 f F20[2 0705 9[254 9[202 25 f F22[2 29 TCA06 9[235 9[233 29 f F21[0 07 Mianserin07 9[248 9[333 48 f F22[0 08 RIMA08 9[376 9[365 31 f F22[1 2119 9[383 9[432 33 f F22[0 1410 9[345 9[531 43 f F22[1 18 RIMA11 9[274 9[332 22 f F22[1 16 SSRIx 9[329 9[344 31[7 11[5S[D[ 9[947 9[971 09[6 3[2

hol or drug abuse\ as well as cranial trauma during thelast 07 months[ Exclusion criteria for MRS investigationwere claustrophobia\ pacemakers and metallic implants[The protocol was approved by the local Ethical andScienti_c Review Committee[ The investigation was car!ried out in accordance with the latest version of the Dec!laration of Helsinki[ A written informed consent wasobtained after the nature of the procedure had been fullyexplained\ and healthy subjects were paid[

1[1[ 0HÐma`netic resonance spectroscopy "0H!MRS#

Localized 0H!MRS was performed in a routine clinicalMR scanner "Siemens Magnetom SP 3999\ Erlangen\Germany# at 0[4 tesla "52 MHz# using a stimulated echoacquisition mode "STEAM# measurement "Frahm et al[\0878a# and eddy current correction[ The mI peak at 2[45ppm is close to the large H1O resonance at 3[62 ppm[ Thewater concentration in human brain is about 49 mol:kgand\ therefore\ the water signal has to be reduced sig!ni_cantly[ Water!suppression was accomplished by a sin!gle frequency!selective pulse preceding the localizing partof the STEAM sequence[ Since mI resonances exhibitrelatively short T1 relaxation times\ measurements shouldbe performed at short echo times "TE#[ As a compromisebetween signal loss at long TE "× 099 ms# and eddycurrent or baseline problems at very short TE "³ 19 ms#\a TE of 44 ms was chosen[ The acquisition of 145 scans

with a repetition time "TR# of 2499 ms yielded spectrawith adequate signal!to!noise ratios "SNR# from the 01ml voxel[ After Fourier transformation of the collectedsignal\ spectra were zero order phase corrected[ However\no line broadening whatsoever was applied to improvethe appearance of the spectrum "Fig[ 1#[ Reproducibilityof single voxel 0H!MRS at 0[4 T is in the order of 4Ð09) if systematic quality assessment measurements areroutinely performed "Radlbauer et al[\ 0883b^ Moser andRadlbauer\ 0884Ð0885^ Metzler\ 0886#[ Linearity of esti!mated mI!signals vs absolute concentrations was testedwith the actual measurement sequence and parameters ina series of _ve test objects\ containing mI from 0Ð19mmol:l with constant creatine "09 mmol:l# and choline "4mmol:l# concentrations[ Linear regression analysisresulted in a su.ciently high correlation "R1 �9[8887#for mI[ Relative metabolite concentrations were cal!culated from peak areas\ i[e[\ via peak amplitude and line!width\ assuming Lorentzian line shapes\ using a home!written GaussÐNewton algorithm "for details seeRadlbauer\ 0883a^ Metzler\ 0886#[ With the total creatinesignal "creatine¦phosphocreatine\ Cr# as an internalstandard\ mI:Cr!ratios were given to follow mI changesin the brain[

1[2[ Volumes of interest "voxels#

Localizer images in axial\ coronal and sagittal orien!tation were obtained _rst[ Two voxels with 29×19×19

R[ Frey et al[:Journal of Psychiatric Research 21 "0887# 300Ð319 304

mm2 "01 ml# were then placed in the right and left frontallobe\ respectively "Fig[ 2#[ To improve SNR by increasingthe local magnetic _eld homogeneity\ local shimming oneach individual voxel was performed after global shim!ming on the whole head[ Relative voxel position in thepatients| brains was kept as constant as possible to avoidvarying partial volume problems "i[e[\ grey vs whitematter vs CSF#[ Therefore\ also Cr!concentrationsshould not vary much\ although we are aware of regionalvariations in the healthy human brain "Pouwels andFrahm\ 0887#[

1[3[ Statistics

The statistical package SPSS for Windows 5[9 wasused[ We tested mI:Cr ratios for normal distribution bymeans of the KolmogorovÐSmirnov test[ The assumptionthat there might be no normal distribution could not beveri_ed] in patients\ the P!value was 9[82 left!frontallyand 9[52 right!frontally^ in healthy subjects it was 9[53left!frontally and 9[84 right!frontally[ Thus\ the t!tests forindependent samples or paired samples were calculated totest di}erences between groups[ The Spearman cor!relation coe.cient was calculated to test correlationbetween the mI:Cr ratio and the HAM!D score\ the Pear!son correlation coe.cient to test correlation betweenmI:Cr and age[ In the context of age dependency analysisof variance "ANOVA# was performed introducing age asa covariable[ For matched pairs analysis\ t!test for pairedsamples was applied[ In addition to individual values"Tables 0 and 1#\ mean values2one standard deviationare given in the results section[

2[ Results

Seven out of 77 spectra were discharged due to tech!nical problems like patient movements or termination ofthe MRS examination by the patient[

Fig[ 2[ The voxels "white boxes# were localized based on series of T0 weighted axial\ coronal and sagittal images[ Voxels "29×19×19 mm2\ 01 ml#were placed in the left and right frontal lobe\ respectively[ They contain predominantly white matter[

2[0[ Healthy subjects

In controls\ mI:Cr was 9[3529[96 left frontally"n�19# and 9[3729[95 right frontally "n�07# "Table0#[ The interhemispheric di}erence was not statisticallysigni_cant "P�9[00#[ In the left frontal lobe\ mI:Cr wasalmost equal in females "9[3429[92^ n�7# and males"9[3529[95^ n�01#\ while in the right frontal lobe therewas a trend "P�9[96# towards lower mI:Cr in females"9[3429[92^ n�09# than in males "9[4029[93^ n�7#[

2[1[ Depressive patients

In depressives\ mI:Cr was 9[3229[95 left frontally"n�11# and 9[3529[97 right frontally "n�10# "Table1#[ The interhemispheric di}erence was not statisticallysigni_cant "p�9[00#[ There were no di}erences betweenfemales and males[

No correlation could be found between mI:Cr andHAM!D score "Spearman\ left] R�9[03\ P�9[4^ right]R�9[08\ P�9[3#[

2[2[ Correlation between mI:Cr and a`e

Except for one woman\ all healthy subjects were agedbelow 39 years "mean 17[924[2 years#\ but patientsranged between 14Ð48 years "mean 31[7209[6 years#[In the latter\ there was a signi_cant positive correlationbetween mI:Cr and age in the right frontal lobe "Pearson\R�9[50\ P�9[991\ Fig[ 3#\ while in the left frontallobe\ the positive correlation did not reach the level ofsigni_cance "Pearson\ R�9[12\ P�9[04#[

2[3[ Total sample] inter`roup differences

The di}erence between mI:Cr in controls and depress!ives was neither signi_cant in the left "P�9[06# nor inthe right hemisphere "P�9[21#[

R[ Frey et al[:Journal of Psychiatric Research 21 "0887# 300Ð319305

Fig[ 3[ Positive correlation between age and mI:Cr ratio in the right frontal lobe of depressed patients "Pearson correlation coe.cient\ R � 9[50\P � 9[991#[

2[4[ Total sample] inter`roup differences\ analysis of vari!ance

When mI:Cr ratio\ left\ was compared between patientsand volunteers with age as a covariable\ signi_cance wasP�9[17 "main e}ect] P�9[01^ covariable] P�9[32# inthe ANOVA[

When mI:Cr ratio\ right\ was compared between pat!ients and volunteers with age as a covariable\ signi_cancewas P�9[906 "main e}ect] P�9[901^ covariable]P�9[997# in the ANOVA[

2[5[ A`e! and sex!matched subsamples] inter`roup dif!ferences

Matched pairs analysis was performed for age! andsex!dependence[ Ten young patients "Nos] 1\ 6\ 8\ 00\ 02\04\ 05\ 06\ 11\ 09^ nine females and one male^ mean age22[223[1 years# were compared to ten controls "Nos] 03\04\ 09\ 07\ 11\ 05\ 0\ 00\ 5\ 19^ nine females and one male^mean age 16[722[1 years#[ In the right frontal lobe\ thecontrols| mI:Cr ratio of 9[3429[95 was signi_cantlyhigher than the patients| mI:Cr ratio of 9[3929[94"P�9[926#[ In the left frontal lobe\ the di}erencebetween the controls| mI:Cr of 9[3429[95 and thepatients| mI:Cr of 9[3029[92 also showed atendency "P�9[05# towards lower mI:Cr in patients[

2[6[ Treated patients vs dru`!free

At the time of the examination\ 04 patients "mean age]32[3 years# were receiving antidepressants^ seven patientswere drug!free "mean age] 30[3 years#[ In the left hemi!sphere\ the treated patients showed a mI:Cr ratio of9[3029[94\ while that of the untreated patients was9[3629[95[ The observed di}erence was statistically sig!ni_cant "P�9[921#[ In the right hemisphere\ the treatedpatients showed a mI:Cr ratio of 9[3429[97\ while thatof the untreated patients was 9[3729[98[ This di}erencewas not signi_cant "P�9[34#[

When comparing the treated patient group "n�04#with healthy subjects "n�11#\ the former showed sig!ni_cantly lower mI:Cr than the latter in the left frontallobe "9[3029[94 vs 9[3529[96\ P�9[925#[ Right front!ally\ no signi_cant di}erences were found "9[3429[97 vs9[3729[95\ P�9[01#[ Comparison of untreated patients"n�6# and healthy subjects "n�11# did not reveal anydi}erences between the two groups[

A combined analysis was performed for both frontallobes\ i[e[\ the voxels of the left and right frontal lobeswere added[ The treated patients showed a trend towardslower mI:Cr ratios than the untreated patients"9[7529[00 vs 9[8329[03\ P�9[03#[ When comparingthe treated patients with the healthy subjects\ the formershowed signi_cantly lower mI:Cr than the latter"9[7529[00 vs 9[8329[00\ P�9[934#[ There was no

R[ Frey et al[:Journal of Psychiatric Research 21 "0887# 300Ð319 306

di}erence between untreated patients and healthysubjects[

3[ Discussion

In the present study\ it has been hypothesized thatreduced mI concentrations in the frontal lobe of depress!ives might be detected by means of 0H!MRS when com!pared to healthy subjects[ Indeed\ in the left as well as inthe right frontal lobe\ mI:Cr mean values were lower indepressives than in controls\ but the di}erences were notstatistically signi_cant[ Both groups showed lower mIin the left hemisphere\ but signi_cant interhemisphericdi}erences occurred neither in controls nor in patients[

In our total sample\ the control group is younger thanthe patient group and shows a di}erent male:female ratio[We did not use a control group in the strict sense of theword because the samples were asymmetric with respectto age and gender[ One reason for refraining from age!matching was that to our knowledge there is no literatureindicating any age!correlated di}erences in mI con!centrations in healthy adults[ Studies by two of our co!authors yielded the same results "Metzler\ 0886^ Moserand Metzler\ in preparation#[ However\ in the mentionedstudies\ metabolite concentrations were determined onthe basis of di}erent TE and TR values and in a di}erentlocation[ The other reason why we are not able to providea control sample is a practical one] in 0886 the 0[4 teslaclinical MR scanner "Siemens# was replaced by anotherroutine scanner which is not equipped for MRS[Recently\ for MRS a 2[9 tesla whole body scanner"Bruker# was installed[

Pearson coe.cients were calculated to test correlationsbetween mI:Cr and age in both the patients and thehealthy volunteers[ As expected\ the latter group pro!vided no signi_cant correlation\ since 10 subjects wereyoung people between 19Ð21 years[ In patients\ in theright frontal lobe\ the positive correlation between mI:Crand age was statistically signi_cant "Fig[ 3#\ while inthe left frontal lobe this was only a trend[ Consistently\ANOVA type analysis introducing age as a covariableshowed a signi_cant di}erence between mI:Cr in the rightfrontal lobe of patients and healthy subjects[ The rela!tively high mI concentrations in elderlies might be due tothe fact that the water content is being reduced as onegets older "Chang et al[\ 0885#[ In Alzheimer|s disease\increased levels of mI were found by means of 0H!MRSas compared with elderly controls "Shonk et al[\ 0884#[However\ the present study only includes patients below59 years and without relevant memory impairment[

As far as a potential gender di}erence is concerned\Pouwels and Frahm "0887# found no statistically sig!ni_cant di}erences between mI concentrations in thefrontal lobes of males and females[ These _ndings are inline with the present study\ which did not reveal any

signi_cant gender di}erences in healthy subjects anddepressive patients[ It remains unexplained why right!frontal mI:Cr tended to be higher in male than in femalecontrols[

Matched pairs analysis was now performed for age!and sex!dependence[ Ten young patients "nine femalesand one male# were compared with ten controls[ In theright frontal lobe\ the patients| mI:Cr ratio of 9[39 wassigni_cantly lower than the controls| mI:Cr ratio of 9[34[In the left frontal lobe\ the di}erence between thepatients| mI:Cr of 9[3029[92 and the controls| mI:Cr of9[34 was not signi_cant[

At the time of our examination\ 04 out of 11 patientswere receiving antidepressive medication[ Unfortunately\due to our time schedule we were not able to investigatemore than seven patients before they received any medi!cation[ A wash!out period for the sole reason of carryingout an MRS would not have been ethically acceptable[Generally speaking\ analyses in subsamples without age!and sex!matching are problematic[ However\ patientswith medication showed a signi_cantly lower mI:Cr ratioin the left hemisphere than healthy subjects[ Moreover\ asigni_cant di}erence between patients with and withoutmedication was found] the former showed signi_cantlylower levels than the latter[ As medication e}ects shouldnot be expected to show a lateralization e}ect\ a com!bined analysis was carried out for both frontal lobes\ i[e[\the voxels of the left and right frontal lobes were added[It showed a signi_cantly reduced mI:Cr ratio in treatedpatients as compared to healthy subjects[

Our _nding that mI was reduced in the medicationgroup draws one|s attention to lithium which is supposedto lower the mI level "see Introduction and Fig[ 0#[ AnMRS!study in healthy volunteers showed no reductionof mI after one week of lithium ingestion "Silverstone etal[\ 0885#\ while an MRS!study in bipolar a}ective dis!order patients indicated a decrease of mI after four weeksof lithium administration "Manji et al[\ 0886#[ Therelationship between inositol\ lithium and a}ective dis!order is complex "Belmaker et al[\ 0884^ Hertz et al[\0886#[ Since lithium and maybe also other antidepressantslower mI\ it appears to be a self!contradictory hypothesisthat depressed patients should have low mI in CSF "Bar!kai et al[\ 0867# and brain tissue "Shimon et al[\ 0886#[However\ in Barkai et al[|s study there was a drug!freeperiod of at least 09 days\ while\ unfortunately\ Shimonet al[ do not specify whether the patients had receivedany antidepressive medication before their death[

Since treatment with large doses of inositol has anti!depressant e}ects "Levine et al[\ 0884#\ it could beassumed that such treatment ameliorates a de_ciency ofinositol[ However\ chronic exposure of cultured astro!cytes to a high level of mI caused a signi_cant decrease inthe inositol uptake when the cultures were subsequentlyexposed to a control medium[ Thus\ it could be the down!regulation of the inositol transport into the brain cells

R[ Frey et al[:Journal of Psychiatric Research 21 "0887# 300Ð319307

which is therapeutically bene_cial[ If so\ chronic treat!ment with inositol might resemble treatment with lithiumor antidepressants in making inositol less available forre!synthesis of PIP1 "Hertz et al[\ 0886\ Calker et al[\0886#[

It is di.cult to tabulate neurotransmitter systemswhich act through adenylate cyclase versus those whichact through phosphoinositides because most neuro!transmitters act through both "Baraban et al[\ 0878#[ Con!trolled trials in psychiatry suggest that inositol 01 g:dayhas therapeutic e}ects in the spectrum of illnesses respon!sive to SSRI\ including depression\ anxiety disorders andobsessive!compulsive disorder\ and is not bene_cial inschizophrenia or Alzheimer|s disease "Levine\ 0886#[ Inregard to the present data\ no di}erences were foundbetween SSRI!patients "n�6^ mean mI:Cr left] 9[33\mean mI:Cr right] 9[33# and normal controls "Table 0^n�11#[

In agreement with several authors\ the total creatinesignal "Cr\ peak 2[92 ppm# has been used in the presentstudy as an endogenous reference for spectroscopy of thehuman brain in vivo[ Assuming a constant Cr con!centration "Frahm et al[\ 0878b^ Michaelis et al[\ 0880^Christiansen et al[\ 0882^ Shonk et al[\ 0884#\ it is widelyaccepted that the observed di}erences in mI:Cr ratiosrepresent changes in mI concentration[ In the presentstudy\ subjects showed mI:Cr ratios between 9[20Ð9[54[Standard deviations were about 04) of mean values[Assuming total creatine to be about 09 mmol:kg "Frahmet al[\ 0878b#\ the mI concentration was between 3Ð4mmol:kg[ This lies within the range of 2[8Ð7[0 mmol:kg\which is presumed to be the mI concentration in 0H!MRSstudies in human brain in vivo "Henriksen\ 0884#[ Thesignal at approximately 2[4Ð2[5 ppm "Fig[ 1# is expectedto be mainly caused by mI[ It is reasonable to assumethat 69) of the major inositol peak is made up by mI\04) by mI!monophosphate\ while 04) is attributableto glycine\ the singlet resonance "at 2[45 ppm# of whichpartially overlaps with that of mI "Ross\ 0880#[

Myo!inositol was detected in the left and right frontallobes of 11 controls and 11 depressives by 0H!MRS at 0[4tesla[ The echo time of 44 ms reduced broad backgroundsignals from large\ relatively immobile biomolecules\ andmI peaks could be quanti_ed without distortions byresidual water or eddy currents[ Only 7) of all spectracould not be evaluated due to gross head movements[Interestingly\ this occurred only in a single patient\ butin six healthy subjects[ The preparation "i[e[\ localizerimages\ voxel selection\ shimming# and examination oftwo voxels took 69 min during which the subject|s headhad to remain motionless[ Generally\ it was not necessaryto give tranquilizers^ however\ examining more than twovoxels would have been too stressful for the subjects[

Proton spectra could easily be recorded from frontalareas without contamination by non!brain tissue[ Thevoxel of 01 ml proved to be large enough to create

su.cient signals for 0H!MRS analyses\ but naturally itcontained di}erent and unknown amounts of grey andwhite matter[ In three autopsy specimens\ there wereno signi_cant di}erences between mI in grey and whitematter "Narumi et al[\ 0858#[ High!_eld 0H!MRS "01 T#showed mI concentrations of 5 mmol:kg in humanfrontal brain extracts of histologically normal specimens"Petro} et al[\ 0878#[ They consisted mainly of grey matterand were obtained during surgery for epilepsy[ In thesame study\ grey and white matter were examined sep!arately in temporal pole biopsies and they contained simi!lar mI concentrations in both[ However\ in a recent paper"Brand et al[\ 0886#\ the proton spectra of astrocytesshowed speci_c signal patterns of mI\ which did notappear in neuronal cell extract spectra[ Thus\ the authorsconsidered mI to be a marker in proton spectra of gliacells[ Inositol might be sequestered in various pools\ andone cannot exactly determine which percentage isinvolved in second messenger synthesis[ Inositol has afunction in osmoregulation in brain glia cells[ Plasmahypertonicity induces an intracellular accumulation ofmI to prevent alterations in brain volume "Paredes etal[\ 0881#[ Moreover\ the glia cells may have a storagefunction for mI "Hertz et al[\ 0886#\ which can then gradu!ally be passed on to the neurons where it becomes aprecursor in the PI cycle[

The phosphorous!20!MRS study "Deicken et al[\ 0884#and the post!mortem study "Shimon et al[\ 0886# men!tioned above focused on bipolar disorder[ In the presentstudy\ lithium treatment was an exclusion criterion[ Thisis the reason why only four depressed bipolar patientswere included[ The two drug!free bipolar patients "Table1^ Nos] 00\ 04# had relatively low mI levels[ Correlationanalysis between the severity of illness\ as rated by theHAM!D score\ and the mI level did not produce anysigni_cant _ndings[

The present study is in line with other studies showingreduced mI levels in depressive patients[ However\ thedata are quite preliminary with respect to the in~uenceof age and treatment[ Further group comparisons shouldbe carried out in age!matched and untreated samples[Moreover\ how far di}erent kinds of antidepressantsin~uence mI in the human brain will have to be deter!mined[ Since the intake of inositol led to an improvementin depression "Levine et al[\ 0884#\ future 0H!MRS inves!tigations could help to plan therapeutic intervention orpredict their outcome[ However\ the development ofcertain standards in MRS is imperative "Moser\ 0883^Moser and Radlbauer\ 0884Ð0885#[

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