Arrhythmia Susceptibility and Myocardial Composition in Diabetes

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Arrhythmia Susceptibility and Myocardial Composition in Diabetes
Influence of Physical Conditioning
Shamsher Bakth, Joseph Arena, William Lee, RemyTorres, Bunyad Haider, BalvantC. Patel,
Michael M. Lyons, and Timothy J. Regan
Division ofCardiology, Department ofMedicine, UniversityofMedicineandDentistryofNewJersey,
NewJerseyMedicalSchool, Newark, NewJersey07103
Abstract We have undertaken a study ofelectrical vulnerability in
themyocardiumofacaninemodelafter 1 yrofalloxan-induced
Abnormalmyocardialcompositionindiabetesmellitushasbeen diabetes. Since altered cardiac cell action potentials have been
described, but the effects on ventricular vulnerability have not described in experimental diabetes with substantial hypergly-
beendefined.Wehaveassessed thesusceptibilitytoarrhythmias cemia (5), the action potential characteristics in this model of
inacaninemodelafter 1 yrofmilddiabetesinducedbyalloxan. mild diabetes have also been examined. In addition, we have
Since physical conditioning can affect metabolic abnormalities assessedtheoccurrenceofspontaneousarrhythmiasduringacute
in diabetes, this intervention has also been evaluated. Group 1 myocardial ischemia.
servedascontrolsandgroups3and4werediabetic.Animalsin Chronicexercise isknowntoimproveglucosetolerance (6)
the lattergroup aswellas nondiabeticcontrols ofgroup 2were andhasbeen showntoincreasethefibrillation thresholdinthe
exercised on a treadmill for the last 8 mo of the experiment. basalstateaswellasduringischemiain nondiabetics(7). How-
After 1 yr, anesthesia was induced with chloralose for vulnera- ever, it is not known whether the cardiac complications ofdi-
bilitystudies. Theventricularfibrillation threshold of24.4±1.9 abetesareaffectedbyenduranceexercise. Intheseexperiments,
mAingroup3wassignificantlylessthaninnormals(45.1±2.2). alteredmyocardial composition hasbeenrelatedtoobservations
Spontaneous arrhythmias werealsomoreprevalentindiabetics onventricularvulnerability.
during acute ischemia (group 3-A). Increased ventricular vul-
nerabilityafterepinephrineinfusionwaspresentinthesedentary Methods
diabetes despite normal ventricular function responsiveness. In
asuperfused preparation ofmyocardium, restingmembrane po- Westudiedhealthymalemongreldogs, 2-4yrofage,weighing22-26
tential andaction potential amplitude werenormal indiabetics, kg.Thedogshadnoclinicalevidenceofdiseaseduringtheinitial8-wk
and beta-adrenergic stimulation shortened repolarization more period. Hematocrit, serumalbumin,andelectrolytesaswellasglucose
tolerancetestswere normal beforeadmission to the study groups. All
thanincontrols. Myocardial collagen concentrations, whichin-
theanimals were fed the same diet consistingof8% fat, 22% protein,
cludedaninterfibrillar distribution onmorphologic examination, 58%carbohydrate,9%ash,and3%crudefiber.Individualanimalswere
were increased in group 3. In the trained diabetics ofgroup 4 placed sequentially in the four groups up to the limit ofthe number
thebasal vulnerability thresholdsandresponses toepinephrine plannedforthegivengroup.
were normal. While myocardial collagen levels were normal, Forstudiesofventricularvulnerabilityandcomposition,thefollowing
cholesterol andtriglyceride increments persisted. Thus, inmild groupswereformed.Group1consistedof11 nonexercisedcontrols,and
experimental diabetes, enhanced susceptibility to arrhythmias group2hadfivenormalcontrolsthatwereexercisedfor8mo.Thelatter
exists;thissusceptibilitymaybebasedonacombinationofnon- group was limited in size dueto time required for daily exercise ofa
homogenouscollagenaccumulationaffectinglocalconductionand totalof14dogs. Diabeticsofgroup3 (n = 9, nonexercised) andgroup
increased electrophysiologic sensitivity tocatecholamines. 4(n = 9, exercised) received seriallowdosesofalloxan. Supplemental
studiesto furtherdefinethesedentarydiabeticswereperformedinad-
ditionalanimalsasindicatedbelowtoassessactionpotentialcharacter-
Introduction
isticsandarrhythmiasduringischemia.
Diabeticmodel.Toproducemilddiabetes,alloxanmonohydratein
Abnormalities ofthe myocardium have been described in ani- sterile salinewasadministered intravenously in adoseof20 mg/kgat
malswithspontaneous(1)orexperimentaldiabetes(2,3)aswell 3-mointervals. Largerdosesofalloxanwereavoidedtopreventketosis.
as in human diabetics (4). Under conditions ofmild diabetes Glucosetolerancewasmeasuredbeforeandat4-mointervalsafterthe
without ketoacidosis, modest impairment of left ventricular initialalloxan intheunanesthetizeddogandserialbloodsampleswere
function was present (2). Since myocardial composition was taken aspreviouslydescribed (2). Plasmaglucosewasanalyzed bythe
found to be abnormal particularly in terms ofcollagen accu- glucose-oxidasemethod(8)tocalculatethedisappearanceratefromthe
mulation, the question ofan associated alteration in electrical vascularcompartment.
Thisdiabetic model exhibitsreducedglucosetoleranceandplasma
propertieshasbeen examined.
insulin levels after intravenous glucose (2) as well as abnormal post-
prandialglucoseconcentrations(9).Reducedventricularcomplianceand
Address reprint requests to Dr. Regan, UMDNJ-New Jersey Medical histochemical changes in the renal mesangium (2) areconsistent with
School, 100BergenSt.,Newark, NJ07103. changes in humandiabetes(10, 4). Hemoglobin A1, levels(11)inthis
Receivedforpublication 11 February 1985 and in revisedform 6 model were found to risefrom 2.41±0.31% to 3.95±0.33 (P<0.004;
September 1985. n = 7)(unpublishedstudies).
Astothequestionofadirecttoxiceffectofalloxanonmyocardium,
J.Clin. Invest. we have previously observed that myocardial abnormalities were not
©TheAmericanSocietyforClinicalInvestigation, Inc. present when the pancreatic effects ofalloxan and resultant diabetes
0021-9738/86/02/0382/14 $1.00 wereprevented(2).Forthiscurrentstudy,threeanimalsseparatefrom
Volume77,February 1986, 382-395 groups 1-4receivedabolusinjectionofglucosetoblockthepancreatic
382 Bakthetal.
effects ofalloxan. Ventricular vulnerability was assessed initially and fibrillation,resultinginahighvariance.Inaddition,inrecentpublished
after 1 yrwhencollagen contentwasalsodetermined. Thesewerecom- studies,the 5-msdurationhasbeenincommon useindeterminations
pared with fourseparate normal controls 9-12 mo after the initial vul- offibrillation thresholdthat do not involve acute ischemia(17). Defi-
nerability assessment, which also provided information on long-term brillationwasaccomplishedwithadirectcurrentpulseof50-100W/s,
reproducibility. deliveredthrough apairofcopperplates(150cm-2). Thiswasusually
Physical conditioning consisted ofrunningon atreadmill (Warren effectivewithin 10-15sanda 1-hintervalwasallowedwhentheanimal
E.Collins,Inc.,Braintree, MA)atarateof8mi/honan 180 inclinefor wasretested.
30min/d. Theanimalsexercised 5d/wk for 32 wkafteraninitial 1-2- Afterestablishingthethresholds foreachanimal inthebasalstate,
wk period ofgraded exercise (12). Two dogs were excluded from the therepetitiveextrasystolicresponseto l-epinephrine wasassessedsince
projectduetoinadequateperformance inthisinitialperiod. Basalheart thishormone isknown toaffect vulnerability(18), andcan bepresent
ratewas measured on at leasttwo separate occasions before beginning inenhancedconcentrationsindiabeticsundersomecircumstances(19).
thestudyandtoward theend ofthetraining period with thedog lying The hormonewasinfused intravenously ata rate of0.05 to 0.1 tg/kg
inaquiet,relaxedstate.Atthesesameperiodsbloodwastakenthrough per min. Metabolic responses were compared by assay ofplasma free
intracath tubing for analyses ofplasma lactate (13), before and at the fattyacids(20)andK+concentrations.Attheendoftheabovestudies,
termination ofexercise toassessdevelopment oftheconditioned state. clotswereremoved,allvesselsweresutured,andtheskinwasclosed.
All dogs were housed in pens large enough to permit free movement, Conduction intervals. Anelectrode catheterwaspassed underfluo-
withambulation outside the pensfor 10-15 min/d. roscopy via a carotid artery to the aortic root immediately below the
Ventricularvulnerability. Animalsweretestedundersterileconditions valvetorecordaHisbundleelectrogrambeforethevulnerabilitystudies
for susceptibility to arrhythmias under chloralose anesthesia, 100 mg/ (21).Bipolarrecordingsweremadewithamultipleelectrodeswitchbox.
kg,withsupplementsasneeded. VentilationwasregulatedbyaHarvard TheoutputwasledintotheA-Cinputofanelectrocardiogram pream-
pump (Harvard Apparatus Co., Inc., S. Natick, MA) via a cuffed en- plifier, filteredat40-500cps,andrecordedonphotographicpaperata
dotrachealtube,and40%oxygenwasusedtopreventhypoxicepisodes. speed of200 mm/s with an Electronics for Medicine recorder (White
ArterialpHwasmaintainedwithintherangeof7.36-7.42byadjustment Plains,NY).ThefirstrapidcomponentoftheHisimpulsewastakenas
ofrespiratoryrate.Bodytemperaturewasmonitoredwitharectalthermo- itsonset.His-Qtimeshavebeenreproducibleover 1-2hofobservation
couple,andwhenbelow370C,wascorrectedbyuseofaninfraredlamp inthedog(21)andduringacuteatrialpacing(22).InadditiontotheP-
focusedontheventral surfaceoftheanimal. RandcorrectedQ-Ttimes,theQRSdurationwastakenasthemaximum
Alltheanimalsinthefourgroupshadstudiesoftherepetitive ven- observed value in the standard leads. Mean values were obtained by
tricularresponsethresholdasanindexofvulnerabilityafter 1 yr.When handtothenearest0.5 msfrom threerepresentativebeatsinthebasal
itbecameapparentthatthisvulnerabilityindexdoesnotalwaysparallel state.
thefibrillationthreshold(14),determinationsoftheventricularfibrillation Hemodynamics. 2wklater,theleftventricularmechanicalresponse
thresholdwerealsoperformed.Thismeasurementwasmadeineightof to catecholamines wasdetermined forcomparison with the prior vul-
11 animalsingroup 1,twooffiveingroup2,sevenofnineingroup3, nerabilitychangesduringarepeatinfusionofl-epinephrineintheanes-
andsevenofnineingroup4. thetized state. While vulnerability wastestedin therightventricle, we
Inadditiontointergroupcomparisons,controlmeasurementsbefore haveassumedthatthehemodynamic responsecomparedwithcontrols
inductionofdiabetesweremadeinsomeofthediabetics.Fouranimals wouldbesimilarinbothventriclesofthediabeticanimals, particularly
ofgroup 3 andthree ingroup4wereevaluated beforealloxanadmin- sincethecompositionalchangesvs.controlsdescribedbelowwerecom-
istrationand 1 yrlater. parable. Wehavepreviouslyfoundthatthe1-epinephrinedoserangeof
Atleast1 helapsedafteranestheticinductionbeforetheformalstudy 0.05-0.1 ag/kgperminelicitsaminimaltomoderatepositiveinotropic
began (15). Electrical testing was accomplished with an intracavitary response inthe left ventricle (23). Sincethepeak maximal rate ofrise
leadsystem,consistingofatransvenoustripolarcatheter(platinumelec- ofventricularpressure(dP/dt)' intherightventricle, incontrasttothe
trodewithan interelectrodedistanceof1 cm andpolewidthof3 mm; left, occursafterthe isovolumic period (24), the use ofdP/dt, isas-
N23-7466;Electro-CatheterCorp.,Rahway,NJ)andelectrocardiogram sociatedwithsomewhatgreatererrorintherightascomparedwiththe
recording probe (semifloating probe No. 567; Electro-CatheterCorp.). leftventricle.Thus,thelattersitewasconsideredpreferable,particularly
Thecathetersystemwaspositionedviajugularveinunderfluoroscopic whentestingatlowintropicdoses.
controlattheapexoftherightventricle,withthedistalcathodalelectrode No.8Goodale-Lubincatheters(U.S.CatheterDiv.,C.R.Bard,Inc.,
localizedinthetrabeculae.Apacemaker(Medtronic,Inc.,Minneapolis, Billerica,MA)wereplacedintheleftventricularchamberandproximal
MN)wasemployedtodeliverrectangularpulses, 2 msin duration, to aortafromthecarotidarteriesundersterileconditions. Stathamtrans-
maintainaconstantheart rateof200 beats/min, adjustingpacemaker ducerswereused forthemeasurement ofleftventricular(P23Gb)and
currenttotwicethemiddiastolicthreshold. Testpulsesweregenerated aortic(P23Db)pressures,whichwererecordedonanamplifier-recorder
withanelectricallyisolatedsquare-wavepulsegenerator(S44;GrassIn- system (Electronics forMedicine). Thetransducers were placed atthe
strumentCo.,Quincy,MA)andanoperationalpowersupplytoproduce midthoraciclevelandbalancedforequalsensitivity. Themaximalrate
constantstimuli. Theelectricaloutputofthisunitwasisolatedandcal- ofriseofleftventricularpressure(dP/dt)wasobtainedusingaresistance-
ibrated with an oscilloscope current probe (P6021; Tektronix, Wood- capacitance differentiating circuit. The system had a linear frequency
bridge,NJ).Thepulsegeneratorwasequippedwithappropriatecircuitry responsefrom0-30cycles/s.Systolicejectionperiodwasmeasuredfrom
toshutofftheoutputofthepacemakerfor3.0safterdeliveryofthetest superimposed aortic and left ventricular pressure pulse tracings. End-
stimulus. The test impulse was delivered after every lOth-l5th paced diastolicpressurewasmeasuredoveratleastthreerespiratorycyclesand
beat. averaged. These measurements havebeenfoundtocorrelatewellwith
A single bipolarstimulus ratherthan atrain ofpulses was used in thepressuremeasurementsobtainedusingamicromanometertip(Millar)
viewoftheevidencethatthelatterelicitslocalreleaseofnorepinephrine catheter(25).Asanindexofmyocardialcontractilityweusedthevelocity
(16). Electrical scanningwasconducted at 3 ms intervalsbeginningat ofcontractile element (Vce) at peak dP/dt, calculated as the ratio of
the end ofthe effective refractory period and terminating 10 ms after maximaldP/dtrelatedtothesimultaneousleftventricularpressure(26).
theT-wave.Usingelectricpulsesof2msdurationthestimulusintensity
was increased in 2 mA steps and scanning continued until repetitive
extrasystoles were elicited. The lowest stimulus intensity that evoked 1. Abbreviations usedin thispaper: APA, action potential amplitude;
repetitiveextrasystolicbeatsintwoofthreetrialsdefinedthisthreshold. APD5o, actionpotentialdurationat50%repolarization;APD80,action
Todeterminetheventricularfibrillationthresholdweusedateststimulus potentialduration at80%repolarization; dPdt, maximal rateofriseof
durationof5 ms,sincetheshorterpulsedurationinpreliminarystudies ventricularpressure;MDP, maximumdiastolicpotential;Vce,velocity
ofnormalsoccasionallyrequiredan unusuallyhighcurrenttoproduce ofcontractileelement.
Diabetes, CardiacComposition, andVulnerability 383
Myocardial composition. 2 wk were allowed for recovery and the endocardialtissuewaspinnedwithsurfaceupina30-mlchamberand
animals were reanesthetized. 1 h later the thorax was incised and the superfusedat 10ml/minofTyrode'sequilibratedwith95%02/5%CO2.
heartrapidlyarrestedwithiced Ringer'ssolution. Transmural samples Temperaturewasmaintainedat37.0±0.50CandthepHat7.3to7.4.
from both ventricles were placed in liquid nitrogen forglycogen assay Theisolatedmyocardiumwasstimulatedthroughbipolarsilver-wire
(27).Bothventriclesandseptumwereseparatelyweighed.Sampleswere electrodesatarateof60min-'beginning30minaftersuperfusionwas
takenfromtheinnerandouterhalvesofleftventricle,theseptum,and begun.Thestimulatingelectrodewaslocated5-10mmfromtherecording
rightventricleforhydroxyprolineanalysestoestimatecollagenconcen- electrode. Rectangularpulses, 1-5 msinduration andtwicethreshold
tration(28).Transmuralsamplesofleftventriclewereextractedforassay voltage,weregeneratedbyastimulator(GrassInstrumentCo.)isolated
ofcholesterol (29), phospholipid (30), andtriglyceride (31). Forcation fromground.Transmembraneactionpotentialswererecordedthrough
analysis the samples were homogenized and extracted for 48-72 h in glasscapillary microelectrodesfilledwith3MKCIatatipresistanceof
distilled water, sufficient time forcomplete extraction. Potassium and 5-15megohms.Microelectrodeswerecoupledbyasilver-silverchloride
sodium were determined on an Autoanalyzer system (Technicon In- wiretoahighinputimpedanceamplifierwithcapacitanceneutralization.
strumentsCorp.,Tarrytown,NY)withflameattachment.Wedetermined Thedepthofthefluidoverthepreparationwasmaintainedataminimal
water content ofthe myocardium by drying the tissue samples in an leveltoreducecapacitanceintherecordingsystem. Theoutputofthe
ovenat 1IO'Ctoconstantweight, amplifierwasdisplayed on an oscilloscope and photographed with an
Forahistochemical assessment, transmural sampleswere taken as oscilloscopecamera(C4;Grass Instrument Co.). Action potential am-
unknownsfromthemid-leftventricleofthreeanimalspergroup.These plitude(APA),maximumdiastolicpotential(MDP),andactionpotential
examinations included trichrome stain for connective tissue (32, 33). durationat50%(APDjo)and80%repolarization(APD80)wereanalyzed
Forelectron microscopy, transmuralsampleswerefixedina2.5%cold from this film. After an initial 30-min equilibration, action potential
glutaraldehyde bufferedwithphosphate, washed, postfixed in osmium, characteristicsweredocumentedbyrecordingPurkinjetypeactionpo-
exposedtoleadanduranylacetate, andembeddedinepon. Eight l- m tentialsfromtheendocardialsurfaceandventricularmusclepotentials
sections were prepared for each dog. Approximately ten micrographs frombeneaththesubendocardial layers;7-15 cellsweresampled from
wereobtainedfromeachsectiontoacquirealongitudinalorientationat eachanimal.
anappropriate magnification. Aftercontrol recordings, incrementaldosesoffreshlypreparediso-
Acutemyocardial ischemia. Aftercompletion oftheabovestudies, proterenolweresuperfused, 30minforeachdose,atwhichtimesteady
weproposedtodeterminewhetherspontaneousarrhythmiasduringacute state measurements were taken. Doses ranged from 10-8 to l0-s M.
myocardialischemiahadadifferentprevalenceindiabeticanimalsthan Subsequently,drug-freesolutionwassuperfusedtocheckforreversibility
controls. Eight male mongrel dogs received alloxan, as above. These ofisoproterenoleffects.
sedentary diabetics (group 3-A) were compared with normal controls Statistics. Thedataareexpressedasmean±SE.Whenonlyonesta-
(group 1-A), matchedforageandsex. tisticalcomparison wasemployedbetween acontrolandintervention,
After 11-12 mo the dogs were anesthetized with chloralose. Oxy- Student'sttestforpaireddatawasused.Toassesssignificancebetween
genation,bodytemperature,andbloodpHweremaintainedasdescribed groups,analysisofvariancewasperformedusingDuncan'smultiplerange
above.Withchestintact,adoublelumen'No. 5balloon-tippedcatheter test for comparisons when F values were significant, at an a level of
was positioned via the carotid artery. Coronary angiography was per- <0.05 (38). Fisher'sexacttestwasusedtoevaluatearrhythmiasinthe
forme4beforeischemiatodeterminethedistributionandcourseofthe acute myocardial ischemia study. Action potential duration responses
leftanteriordescendingcoronaryartery. Two normal dogsandonedi- togradeddosesofisoproterenolwerecomparedbyattestfordifferences
abeticdogwereexcludedbecausetheanteriordescendingarterydidnot inregressionlineelevation(38).
coursetotheapexoramajorobliquevesseloriginatedclosetotheorigin
oftheanteriordescendens,thusdiminishingtheareaatrisk.Afterballoon Results
inflationintheproximal 1.5cmoftheleftanteriordescendingcoronary
artery(34),transmural ischemiawasevidencedbyareductionofmean Bodyweightinanimalsofthefourgroupswasmaintainedover
peripheralcoronaryarterypressureto -25mmHgandtheappearance
the 12moofobservationwhileplasmaalbuminandelectrolytes
ofaninjury potential onstandardlead 1. Nocardioactive agentswere
remainedwithinthenormalrange(TableI).Diabeticsofgroups
given.
Theanimalswereobservedupto 15min,ariskperiodforarrhyth- 3and4exhibitedareductionofglucosetolerancethatwassimilar
mias.Thislimitwasimposedsincethisdiabeticmodelexhibitsagreater inextentat4moaftertheinitialalloxanwithaGkof1.91±0.27
degreeofleftventriculardysfunctionduringischemiaovertimethando and 1.82±0.14, respectively. Fasting normoglycemia was not
normals(34),whichmayrepresentanadditionalvariableaffectingout- affected by the exercise regimen in group 4 diabetics, but at 1
come.Arrhythmiaswererecordedata25mm/spaperspeedonarecorder yrtheglucoseclearanceconstantwasincreasedto2.40±0.15 (P
(ElectronicsforMedicine)withautomaticdevel6per. <0.05), without achange ingroup 3.
Ventriculartachycardia wasdefinedas'aseriesofsixormore con- Evaluation ofthemodels. Toexaminethepotentialcardio-
secutive ventricularectopiccomplexes(35). Weexaminedthetotal in-
toxiceffectsofalloxan, wepreventedthedevelopmentofglucose
cidence ofventriculartachycardia andfibrillation inthetwogroupsas
intoleranceinthreeanimalsafterpretreatmentwith50gofglu-
wellasthesumofventricularectopiccomplexes(35, 36).Inthosethat
survived 15min,theheartwascold-arrestedwithicedRinger'ssolution, cose before each alloxan dosage. The fibrillation thresholdwas
andEvan'sBluedyewassimultaneouslyadministeredviathecoronary unchangedafter 1 yrintheseanimalsaswellasinfournormals
arterycatheterataortic-diastolicpressurelevelstodeterminetheareaat followed for 9-12 months without such treatment (Table II).
risk. In animals that fibrillated, the injection pressure wasthat ofthe Collagenconcentrations intransmural samplesofleftventricle,
residualaorticpressure. as a marker of diabetic changes, were 2.14±0.19 ag/mg dry
Electrophysiologicalstudiesinvitro.Todeterminewhetherthismodel weight in the glucose-alloxan group vs. 2.09±0.11 in normals.
ofdiabeteswasassociatedwithalteredcellactionpotentials,werecorded Thus, acardiotoxic effectofthiscompound appears unlikely.
intracellularpotentials(37), andtheresponse ofthesepotentialstoiso-
Adaptation to chronic exercise was indicated in groups 2
proterenolinfournormals(group 1-C)andthreediabetics(group3-C)
and 4 by a reduction ofbasal heart rate in the conscious state
preparedasabove. After 1 yrtheheartwascold-arrested, andtrans-
and the failure ofblood lactate to rise during acute exercise
muralsampleswererapidlytakenfromthefreewalloftheleftventricle
2.5cmabovetheapexandplacedincooloxygenatedphysiologicTyrode's (Table I). In addition, left ventricular weight, as a measure of
solution with the followingmillimolar composition: NaCI, 135; KCl, hypertrophy, increased similarly in these two groups.
4.0; dextrose, 5; CaCl2, 2.0; MgCl2, 1.0; NaHCO3, 15; Na2HPO4, 1.0. Ventricularvulnerabilityindiabetics. Incontrolanimalsfor
After removal ofthe epicardial two-thirds ofthe ventricular slice, the thediabeticseries(group 1),therepetitiveextrasystolicthreshold
384 Bakthetal.
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Diabetes, CardiacComposition, and Vulnerability 385
TableII. Serial VentricularFibrillation ThresholdinNormals* TableIII. VentricularFibrillation Threshold*
Animalno. Initial 12mo Group 1 Group3 Group4
mA mA mA mA mA
1 48* 46 38 27 41
2 53* 55 42 31 38
3 42* 41 44 18 43
4 39 43 45 22 36
5 45 39 43 20 37
6 38 35 48 31 46
7 49 48 49 22 48
Mean±SE 44.85±2.09 43.86±2.47 52
45.1±1.6 24.4±1.9 41.3±1.7
* Indicatesanimalsthatreceivedglucose-alloxan aftertheinitialdeter-
minationoffibrillation threshold. Theotherfouranimalsdidnotre- * Threshold forindividualanimalsasdeterminedaftertherepetitive
ceiveglucoseoralloxan. ventricularresponse. Unpairedttest:group 1 vs. 3,P<0.005;group
1 vs.4,N.S.;group 3 vs.4,P< 0.01.
was 45.8±2.0 mA in the basal state after 1 yr (Fig. 1). The
threshold wassignificantly lowerinthe nontrained diabetics of
group 3 at a level of27.5±3.8 mA. By contrast, in the trained betweencontrolsanddiabetics(TableV). Basalplasmafreefatty
diabetics ofgroup 4, this threshold at 12 mo was significantly acid(FFA)levelswerealsosimilarbetweengroup 1 andthetwo
higherthaninthesedentarydiabetics,didnotdifferfromgroup diabetic groups, with similar elevations during the infusion of
1 controls,butwaslessthanthelevelintheconditioned normals epinephrine. Group 2 controls differed from group 1 with a
(Fig. 1). higher basal FFA level. Neither plasma K+ decreases nor ele-
Evaluationoftheventricularfibrillationthresholdinthebasal vationofplasmaFFAappearedtocontributetothevulnerability
staterevealedasignificantreductioninthenontraineddiabetics differences in untrained andtraineddiabetics.
ofgroup3comparedwithcontrolsofgroup 1 (TableIII). Inthe Duringthe hemodynamic study 2 wk later, left ventricular
traineddiabeticsofgroup4thethresholdwassignificantlyhigher performance during administration ofepinephrine was com-
than that ofthe sedentary diabetics and approximated that of paredintermsofvelocityofcontractileelement(Vce)andfilling
thenormalcontrols. Onlytwooftheexercisednormalsofgroup pressure (Table V). The responses in diabetics ofgroups 3 and
2 weretested and hadfibrillation thresholds of51 and 53 mA. 4did notdiffer from those ofthe normals.
Inthe subset ofgroup 3 animalswith measurements before Conduction times. In thebasal state, P-R andQRS on lead
induction ofdiabetes aswell as 1 yrlater, the extrasystolic and
fibrillationthresholdsweresignificantly reducedattheterminal
studies (Table IV). The group 4 subset showed no change in TableIV. Serial Vulnerability Thresholds
eitherparameter from the prediabetic period.
Toassesstheresponsetoadrenergicstimulation, epinephrine Group3 Group4
was infused ata rate of0.05 and0.1 og/kgpermin. Theextra-
Control 12mo Control 12mo
systolic threshold was reduced to a significantly greater degree
in the nontrained diabetics vs. group 1 controls (Fig. 2). In the mA mA mA mA
traineddiabetics, ontheotherhand, thethreshold atthehigher A. Repetitiveventricularextrasysoles
dose was significantly greater than in the nontrained diabetic,
didnotdifferfromthecontrolsofgroup 1,butwassignificantly 35 25 37 36
lessthanthetrained normals. 47 22 41 45
DuringepinephrineinfusionmeanplasmaK+levelsdeclined 43 27 46 44
10%fromtheinitialnormallevelsbuttherewasnodifference 36 28 _
-
Mean±SE 40.3±2.8 25.5±1.3t 41.3±2.6 41.7±2.8
70 CONTROLS DIABETICS
B.Fibrillationthreshold
60 _ P-0.001
Figure1. Thresholdsforrepeti-
50 tiveventricularectopicrespon- 48 22 41 37
X 40 ses.o,exercised.Group3,the 42 20 43 46
nonexerciseddiabetics(n = 9), 47 31 49 48
- 30 hadasignificantreductionof 38 22
20 _ 0 0 threshold comparedwithgroup Mean±SE 43.8±2.3 23.7±2.4t 44.3±2.4 43.7±3.4
1 controls(n = 11). Inthe
o0 traineddiabeticsofgroup4(n
GRP= 9),thethresholdwassignifi- * Individualanimalsthatwerestudiedbeforeinductionofdiabetes
GROUP GROUP GROUP GROUP cantly higherthaningroup3 and 1 yrlater.
102P-0.032 4X diabetics, lowerthangroup2(n tPairedttest,P<0.025 (one-tail): P<0.05 (two-tail).
N.S. = 5),andsimilartogroup 1. §Pairedttest,P<0.0025 (one-tail);P<0.005(two-tail).
386 Bakthetal.
60 withtrichromewerenegativeingroups 1 and2(Fig. 3A). How-
_-a ,- TRAINED ever,inthesedentarydiabeticsofgroup3,accumulation ofblue-
50 - 1I stained material waspresentaround intramural vesselsandac-
cretionswere presentthroughout interstitium (Fig. 3 B), which
en 40 wasnegligibleingroup4. Onelectron micrography, ultrastruc-
a.
ture ofcell organelles in the sedentary diabetic appeared to be
30
. - P<O.003 within normal limits(Fig. 4). Theinterstitium containedfibrils
j withtheperiodicityofcollagenandamorphousmaterialaround
20 I
the capillary and nerve endings, as well as between adjoining
cardiaccells.
BASAL 0.05 0.1 Left and right ventricularglycogen concentrations were el-
EPINEPHRINE (,ug/Kg/min)
evatedingroups2and4butnotinthesedentarydiabetic(Table
Figure2. Repetitive ventricularresponseafterepinephrine. Thenon- VIII). Ingroups3 and4,cholesterolandtriglycerideconcentra-
exercisedgroup3diabetics(o)hadalowerthresholdatthehigherepi-
tions were increased in both ventricles. Phospholipid concen-
nephrinedosethantheexerciseddiabeticsofgroup4(A)andgroup 1
trations in left ventricle for groups 1 through 4, respectively,
controls(o).Group2wassignificantly higherthangroup4.
were 16.8±0.7MM/gm, 17.2±0.9, 16.7±0.8,and 17.0+0.6.There
wasalsonosignificantdifferenceinrightventricularconcentra-
II ofthe EKG times did not differ in the diabetic groups vs. tionsbetween thesegroups. Sodium wasalsoincreased butpo-
control(TableVI). Intheexercisedcontrolsofgroup2,P-Rwas tassium concentrations were normal in both diabetic groups
significantly longer. The H-Q time was significantly prolonged (TableVIII). Watercontentaspercentofleftventricularweight
indiabeticsofgroup3comparedwiththenonexercisedcontrols did notdiffersignificantly between the fourgroups. Ingroup 1
ofgroup 1.ExerciseddiabeticshadH-Qtimesthatdidnotdiffer the value was 78.1±0.06%; group 2, 78.3+0.04; group 3,
from theexercised controlsofgroup 2. 79.8±0.07; group4, 78.5±0.05.
Myocardialcomposition. Theconcentrations ofcollagen in Myocardialischemia. Toassesstheincidenceofspontaneous
theleftand rightventriclesaswell asseptum were significantly ventriculararrhythmias, we prepared additional animalstode-
increased ingroup 3 diabetic animals(Table VII). These incre- terminetheresponsetoa 15-minperiodofmyocardialischemia.
mentsabovecontrolswereproportionatelysimilarineachtissue, Thesedentary diabetic animals hadaglucose clearance rate of
although concentrations were highest in the right ventricle of 2.05±0.19, comparable tothelevelsingroup 3 diabetics. After
controlsanddiabetics. Collagen incrementsintheleftventricle anestheticinduction,heartrateandaorticpressurewereatsimilar
were moreevident inthesubendocardium butthiswasnotsig- levels in the control and diabetic animals before the onset of
nificantly greater than in the outer layers. Similar increments ischemia, andplasmaelectrolyteswerenormal. Duringcoronary
werenotobservedintheexerciseddiabeticsofgroup4inwhich occlusion the aorta to peripheral coronary artery pressure gra-
collagenconcentrationsapproximate thenormalcontrollevels. dientwas 88±4.9 mmHginthecontrolsand91±6.4 mmHgin
Toassesstheeffects ofthe hypertrophy ofexercise on collagen thediabetics,andthesinusrateresponsewascomparable(Table
accumulation, thecontentpertotalventriclewascalculatedfrom IX).Thenumberofectopicbeatsasventriculartachycardiawas
muscleweightandcollagenconcentration (TableVII). Content significantly greater in the diabetic animals (Table IX). Fibril-
was significantly greater in both the left and right ventricles as lation occurred in fouroftheseven diabeticsand intwo outof
wellasseptumingroup3butgroup4didnotdifferfromcontrols. 12 inthenormalgroup. Whenthosewith fibrillation andthose
Morphologic studies were performed to determine the dis- with ventricular tachycardia alone were compared with nondi-
tributionofcollagen. Histologicsectionsofleftventriclestained abetics, the difference was significant at P < 0.05. The area of
Table V ResponsestoEpinephrineInfusion inDiabetics
Left
Aortic ventricular
Freefatty pressure end-diastolic
K+ acid Heartrate mean pressure Vces
mEqlliter mEqlliter beats/min mmHg mmHg
Group 1
Basal 3.9±0.16 776±86 122±7 138±8 5.1±0.6 6.8±0.7
Epinephrine(0.1 tg/kgpermin) 3.6±0.16 1,478±234 127±9 126±10 3.9±1.3 10.6±1.9
Group3
Basal 4.1±0.12 716±96 133±10 140±4 4.9±0.7 7.5±0.5
Epinephrine(0.1 tg/kgpermin) 3.9±0.32 1,331±232 145±7 136±9 2.6±0.5 9.9±1.2
Group4
Basal 4.1±0.17 762±82 110±12 138±9 3.4±0.7 7.7±0.4
Epinephrine(0.1 sg/kgpermin) 3.7±0.22 1,460±310 117±15 124±10 3.7±1.2 12.0±1.4
Unpairedttest. Nosignificantdifferencesinresponsetoepinephrinebetweengroup 1 vs.groups3and4. *Ratio ofmaximaldP/dttosimulta-
neousleftventricularpressureincyclelengths/s.
Diabetes, CardiacComposition, and Vulnerability 387
Table VI. Myocardial Conduction inNormals andDiabetics ening at ADP50 and APD80 after isoproterenol in the diabetic
group, whichdid notoccurinnormals.
Heart
rate PR QRS QTc HQ
Discussion
beats/
min ms ms ms ms
Theseexperimentswereundertakentodeterminewhetheramild
Group 1
form ofdiabetes, 1 yrinduration,wouldalterthesusceptibility
(n= 6) 128±11 71±7 65±2.6 324±13.7 25.0±0.8
ofthe myocardium toarrhythmias. Thiscanine model ischar-
Group2
acterized by normal coronary arteries (2) and collateral blood
(n = 5) 94±15 96±8* 68±1.1 310±6.5 28.5±0.7f
flowresponseduringcoronaryocclusion(34),aswellaspreserved
Group3
ultrastructure ofcardiac cells(8).
(n = 6) 133±10 67±6 63±0.78 312±7 29.7±0.98t
Sedentarydiabetics. Diabetic dogsthatwere notphysically
Group4
conditioned showedchangesinelectrical vulnerability, withre-
(n = 6) 100±12 84±6 64±3.3 307±10 31.6±1.6t
duction ofboth the thresholds for repetitive extrasystoles and
ventricular fibrillation. Moreover, agreater incidence ofspon-
Unpairedttestvs.Group7. *P<0.05. *P<0.01. taneousventriculararrhythmiaswasevidentduringacuteisch-
emia ofthe anterior left ventricle. That arrhythmogenesis was
affected in both ventricles with similar compositional abnor-
risk intheregion ofthe anteriordescending artery was 33±1.8 malities issupported bythisobservation.
ginthe normal vs. 31.7±2.1 ginthediabeticgroup. Reduction ofvulnerability thresholds in the sedentary dia-
Actionpotentials. Todetermine whether electrophysiologic beticsmayberelatedtoseveral factors. Connectivetissuesepta
abnormalities were present in the sedentary diabetics, group normally presentwithin individual musclebundlesinaninter-
3-A was compared with group 1-A normals (Table X). In the mittentdistributionareconsideredtobeassociatedwithlocalized
basal state there was no difference in the MDP orAPA in the dissociation ofexcitation (39).Accordingly, theincreaseofcol-
myocardial cell between normals and diabetics. However, the lagencontentinthediabeticmyocardiuminanonhomogeneous
duration ofrepolarization at 50 and 80% was slightly, but sig- distributionmayexaggeratethisphenomenon. Thisviewissup-
nificantly, longer inthediabetic group(P<0.02). ported by the local electrode study, which revealed a modest
After isoproterenol, normals exhibited a significant short- increaseofHis-Qtime,approximatingthatofanethanolmodel
eningoftheAPD50orAPD80withoutachangeinMDPorAPA of subclinical heart disesase (22). Assuming that this at least
(Table X). This response was significantly greater in diabetic partially reflects subendocardial collagen accumulation, alocal
animals.Aplotofthelogconcentrationoftheadrenergicagonist dispersionofrefractoryperiodswouldfacilitatereentrantactivity
vs. the action potential duration in myocardial cells reveals andarrhythmias(40).Thismechanismhasbeenconsideredthe
greater shortening at both repolarization durations (Fig. 5). In basis forarrhythmias duringthreshold stimulation (41) aswell
thepresence ofsimilarslopes the regression line levels differed asearly in acute ischemia (42) in nondiabetics. Further, under
indiabetics vs. normals (P<0.05). appropriate conditions, triggered activity maybeelicited inthe
In the Purkinje fibers ofthe diabetics the MDP, APA, and diabetic (43).
therepolarization durationswerenotsignificantly differentfrom Catecholamine responses. Several observations supportthe
normalinthebasalstate.Therewas,however, significantshort- view that catecholamines may influence arrhythmia suscepti-
bilityindiabetics. Exogenousepinephrinewasfoundtoreduce
thethreshold forrepetitive ectopic beats in sedentary diabetics
Table VII. OH-ProlineConcentrations* andto shortentherepolarization phases oftheactionpotential
relativetonormals. Duringthevulnerabilitytestthesympathetic
Leftventricle
system inthemyocardium appearstobeactivatedatthesiteof
Group Inner1/2 Outer1/2 Septum Rightventricle thestimulatingelectrode, evenbythesingle-impulse technique
(16).Thus,anenhancedsensitivitytocatecholaminesindiabetics
1 (n = 11) 2.15±0.11 2.09±0.15 1.72±0.09 2.88±0.10 may partially account for the reduced fibrillation threshold in
2(n = 5) 2.16±0.28 1.88±0.26 1.64±0.21 3.0±0.29 thebasal state.
3(n = 9) 3.14±0.24t 2.63±0.27§ 2.41±0.28§ 3.56±0.27§ During ischemia a neurohormonal contribution to arrhyth-
4(n = 9) 2.11±0.17 2.02±0.14 1.83±0.22 3.08±0.09 mogenesisisattestedbytheprevention oflethal ventricularar-
rhythmias after beta-blockade or chemical sympathectomy in
OH-ProlineContent, MilligramsperRegionofHeart nondiabetics (44). While a similar antiarrhythmic efficacy in
Group Leftventricle" Septum Rightventricle mild diabeteswouldsuggest adominant role forthis system, a
quantitativecomparison with normalswouldrequireassessment
1 40.9±2.9 15.4±0.7 23.3±1.2 ofthe dose-response relationship to beta-adrenergic blockade.
2 42.4±5.9 16.0±2.5 26.7±3.4 Althoughareductionofvagalactivitycancontributetoarrhyth-
3 59.8±3.3t 25.0±2.2t 30.9±3.0§ mogenesis (45, 46), in experimental diabetes the efferent limb
4 40.7±5.0 17.6±2.4 25.6±1.7 ofthis nerve has been reported to be unimpaired asjudged by
the heart rateresponseto vagal stimulation (47).
*Concentrations in ug/mgdryweight. tP<0.01. §P<0.02. Altered responsiveness to catecholamines appearsto be se-
Contentsininnerandouterhalvesofmyocardiumwereaveragedfor lectivesincethemyocardialhemodynamicresponsedidnotdiffer
eachanimal. between control and diabetic animals. In adiabetic rat model,
388 Bakthetal
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Diabetes, CardiacComposition, and Vulnerability 391