DTIC ADA627842: Sympathetic Nerve Activity and Heart Rate Variability During Severe Hemorrhagic Shock in Sheep

0.3 MB·English

by  Defense Technical Information Center

AutonomicNeuroscience:BasicandClinical136(2007)43–51
www.elsevier.com/locate/autneu
Sympathetic nerve activity and heart rate variability during severe
☆
hemorrhagic shock in sheep
⁎
Andriy I. Batchinskya, , William H. Cookeb, Tom A. Kuuselac, Bryan S. Jordana,
Jing Jing Wanga, Leopoldo C. Cancioa
a U.S.ArmyInstituteofSurgicalResearch,3400RawleyE.ChambersAvenue,Building3611,FortSamHouston,Texas,78234-6315,UnitedStates
bLaboratoryforAppliedAutonomicNeurophysiology,DepartmentofHealthandKinesiology,UniversityofTexas,SanAntonio,
6900NorthLoop1604WestSanAntonio,Texas,78249,UnitedStates
c DepartmentofPhysics,UniversityofTurku,Turku,Finland
Received30January2007;receivedinrevisedform28March2007;accepted30March2007
Abstract
Introduction: In this study we explored direct and indirect measures of autonomic nervous system function, as well as changes in
cardiovascular complexity,during hemorrhagic shock(HS).
Methods: HSwasinducedinanesthetizedsheep(n=8)byremoving40ml/kgofbloodinfour10ml/kgstepsover40min.Resuscitationwas
performedwithlactatedRinger'sandre-infusionofshedblood.Renalsympatheticnerveactivity(RSNA)wasmeasuredbymicroneurography.
Spectralanalysisofheartratevariability(HRV)employedfast-FouriertransformationoftheR-to-Rinterval(RRI)oftheEKG.Thisgeneratedthe
normalizedhigh-frequency(HFnu)andlow-frequency(LFnu)powersoftheRRI,andtheirratio(LFnu/HFnu,aproposedindexofsympatho-vagal
balance).Additionally,non-linearmethodswereapplied:RRIcomplexitywasmeasuredbyapproximate(ApEn)andsample(SampEn)entropy
methods;RRIfractaldimensionwasmeasuredbycurvelengths(FDCL).PlasmacatecholaminesweredeterminedbyHPLC.
Results: The model caused profound HS; 2/8 animals survived till the end of resuscitation. RSNA increased in 7/8 sheep and, as HS
progressed,multipleburstcomplexeswereidentifiedfollowedbysympatheticwithdrawal.ConcomitantdecreasesinHFnuandincreasesin
LFnu/HFnu occurred after 20 ml/kg blood loss. ApEn and FDCL decreased after withdrawal of 40 ml/kg of blood. Catecholamine
concentrations increased throughout HS.LFnu/HFnuandRSNAwere notlinearly correlated.
Conclusions: HS led to an increase in RSNA with subsequent withdrawal. LFnu/HFnu increased during HS in association with vagal
withdrawalandlossofRRIcomplexity.RRIcomplexitymayinpartreflectvagalmodulationoftheheartrate.Changesindirectlymeasured
tonic sympathetictrafficdonotcorrelate with non-invasive measures of autonomic modulationof the heart.
©2007Elsevier B.V. All rightsreserved.
Keywords:Autonomicnervoussystem;Renalsympatheticnerveactivity;Hemorrhagicshock;Heartratevariability;Spectrumanalysis;Non-linearanalysis;
Complexity;Entropy
1. Introduction
The autonomic response to hemorrhagic shock (HS) is
Abbreviations:HS,Hemorrhagicshock;HF,high-frequencypower;LF,
complex,under-investigatedandinfluencedbythedynamics
low-frequencypower;RSNA,renalsympatheticnerveactivity;RRI,R-to-R
interval of the EKG; LR, lactated Ringer's; ApEn, approximate entropy; andmagnitudeofvolumedepletion(Secheretal.,1992;Evans
SampEn,sampleentropy;FDCL,fractaldimensionbycurvelengths. et al., 2001). Animal studies involving direct renal neural
☆ Theopinionsorassertionscontainedhereinaretheprivateviewsofthe recordings during HS showed increased renal sympathetic
authorsandarenottobeconstruedasofficialorasreflectingtheviewsofthe
nerve activity(RSNA) as the initial response tovolumeloss
DepartmentoftheArmyortheDepartmentofDefense.
⁎ (Skoogetal.,1985;Koyamaetal.,1988,1992;Malpasetal.,
Correspondingauthor.
E-mailaddress:[email protected](A.I.Batchinsky). 1998). With progression ofshock, a decrease inRSNAmay
1566-0702/$-seefrontmatter©2007ElsevierB.V.Allrightsreserved.
doi:10.1016/j.autneu.2007.03.004
Report Documentation Page Form Approved
OMB No. 0704-0188
Public reporting burden for the collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and
maintaining the data needed, and completing and reviewing the collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information,
including suggestions for reducing this burden, to Washington Headquarters Services, Directorate for Information Operations and Reports, 1215 Jefferson Davis Highway, Suite 1204, Arlington
VA 22202-4302. Respondents should be aware that notwithstanding any other provision of law, no person shall be subject to a penalty for failing to comply with a collection of information if it

does not display a currently valid OMB control number.
1. REPORT DATE 2. REPORT TYPE 3. DATES COVERED
30 OCT 2007 N/A -
4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER
Sympathetic nerve activity and heart rate variability during severe
5b. GRANT NUMBER
hemorrhagic shock in sheep
5c. PROGRAM ELEMENT NUMBER
6. AUTHOR(S) 5d. PROJECT NUMBER
Batchinsky A. I., Cooke W. H., Kuusela T. A., Jordan B. S., Wang J. J.,
5e. TASK NUMBER
Cancio L. C.,
5f. WORK UNIT NUMBER
7. PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) 8. PERFORMING ORGANIZATION
United States Army Institute of Surgical Research, JBSA Fort Sam REPORT NUMBER
Houston, TX 78234
9. SPONSORING/MONITORING AGENCY NAME(S) AND ADDRESS(ES) 10. SPONSOR/MONITOR’S ACRONYM(S)
11. SPONSOR/MONITOR’S REPORT
NUMBER(S)
12. DISTRIBUTION/AVAILABILITY STATEMENT
Approved for public release, distribution unlimited
13. SUPPLEMENTARY NOTES
14. ABSTRACT
15. SUBJECT TERMS
16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF 18. NUMBER 19a. NAME OF
ABSTRACT OF PAGES RESPONSIBLE PERSON

a. REPORT b. ABSTRACT c. THIS PAGE UU 9
unclassified unclassified unclassified
Standard Form 298 (Rev. 8-98)
Prescribed by ANSI Std Z39-18
44 A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51
occur(Skoogetal.,1985;Koyamaetal.,1988,1992)asasign 2. Materials and methods
ofirreversibleshock(Koyamaetal.,1988).
Because direct tonic nerve activity measurements are This study was approved by the U.S. Army Institute of
invasive, they are not usable as clinical monitoring tools. SurgicalResearchAnimalCareandUseCommitteeandwas
Alternatively, analysis of the R-to-R-interval (RRI) by fast- carriedoutinaccordancewiththeguidelinessetforthbythe
Fouriertransformhasbeenproposedasanindirectindexof AnimalWelfareActandotherfederalstatutesandregulations
cardiacautonomicmodulation(Akselrodetal.,1981;Pagani relatingtoanimals and studies involving animals and bythe
et al., 1986). The low-frequency component of the RRI 1996GuidefortheCareandUseofLaboratoryAnimalsofthe
powerspectrum,orlow-frequency(LF)power,isinfluenced NationalResearchCouncil.
by both sympathetic and parasympathetic activity. High-
frequency power (HF), by contrast, is influenced by 2.1. Animal preparation
parasympathetic activity alone. The ratio of LF to HF (LF/
HF) has been proposed as an index of “sympatho-vagal Neuteredmalesheep(n=8),weighing35.9±1.9kgwere
balance” (Pagani et al., 1986). The concept of sympatho- fastedfor24hpriortothestudy,premedicatedbyketamine,
vagalbalance,however,hasbeencriticallyappraised(Parati induced with isoflurane via mask and endotracheally
et al., 1995; Eckberg, 1997; Notarius and Floras, 2001). At intubated. With the animal in dorsal recumbency, surgical
present,theindirect heartrate variability(HRV) metrics are anesthesia was carried out with a combination of isoflurane
said to reflect cardiac autonomic influences in a qualitative (0.47±0.03%)andintravenousketamine(468±3.7μg/kg/h)
manner,andtheirfullmeaningremainscontroversial(Parati (KetaVed, Ketamine HCL, Phoenix, Scientific, Inc., St.
etal., 2006). Joseph, MO). Animals were ventilated in volume-control
Despitethiscontroversy,HRVmetricsarewidelyusedfor mode with a tidal volume of 13 ml/kg, respiratory rate of
obtaining indirect non-invasive insight into cardiovascular 12 per min and fraction of inspired oxygen of 100%. Tidal
autonomic modulation (Anonymous, 1996). Association of volume and respiratory rate were adjusted to maintain the
HRV metrics with volume status (Butler et al., 1994; endtidallevelofCO between30and40mmHg.Bilateral
2
Palazzolo et al., 1998; Goldstein et al., 1999; Batchinsky surgicalcatheterizationsoftherightexternaljugularveinand
etal.,2007)andwithmortalityinICU(WinchellandHoyt, bothfemoralarteriesandveinswereperformedfollowedby
1996)andtraumapatients(Cookeetal.,2006)hassuggested a midline laparotomy and splenectomy to exclude the
their potential utility as clinical monitoring tools (Winchell possibility of splenic auto-resuscitation during hemorrhage.
and Hoyt, 1996; Grossman etal., 2004; Cooke etal., 2006; Each animal then received a 1-l bolus of lactated Ringer's
Parati et al.,2006). (LR) solution followed by a maintenance infusion of LR at
Sauletal.(1990)reportedapositivecorrelationbetween 2ml/kg/h.Next,theleftkidneywasexposed,therenalnerve
the LF/HF ratio and directly measured sympathetic nerve dissectedandaunipolarTungstenmetalmicroelectrodewith
activity in humans, while others have not (Kingwell et al., a 75-μm shaft (FHC, Bowdoinham, ME) inserted into the
1994; Pagani et al., 1997; Cooke et al., 1999; Cooke and nerve. After the bursts were visually verified the electrode
Dowlyn,2000).Kingwelletal.(1994)reporteddissociation was glued in place with cyanoacrylate adhesive (Vetbond,
between the LF and cardiac sympathetic activity evaluated 3M, Saint Paul,MN).
by norepinephrine spillover. In animal studies that did not
involve direct RSNA recordings, RRI LF increased during 2.2. Physiologic measurements
hemorrhageinconsciousdogs(MadwedandCohen,1991),
decreasedduringhypovolemicshockinanesthetizedrabbits Arterial plasma lactate was measured with an i-STAT
(Goldstein et al., 1999), and showed a differential response bloodgasanalyzer(AbbotLaboratories,EastWindsor,NJ).
in tachycardic and bradycardic hemorrhaged dogs (Kawase Standardfour-leadEKGwasobtainedfromsurfaceelectro-
etal., 2000). desgluedtothepreviouslydepilatedskin intheaxillaeand
In this study we sought to characterize the changes in outerthighs.Acontinuouscardiacoutputpulmonaryarterial
RSNA and its morphology occurring in response to severe catheter(Model746HF8,EdwardsLifesciencesLLC,Irvine,
HS in sheep, and to determine the relationship, if any, CA) was placed via the left external jugular vein. Arterial
between theindirectlyassessedLF/HF ratioofthe RRI and pressure was transduced (Transpac® IV, Abbott, North
directly measured tonic RSNA. We hypothesized that HS Chicago,IL)atheartlevel.Allphysiologicwaveformswere
would cause concurrent increases in both the LF/HF ratio displayed continuously using a clinical monitor (Viridia
and RSNA, possibly followed by withdrawal of RSNA in CMS 2000,Boebingen,Germany).
pre-terminal subjects. We also hypothesized that HS would
be associated with loss of RRI complexity, a new metric 2.3. Experimental protocol
derived from non-linear biosignal analysis methods, which
quantifies RRI irregularity by assessment of structural Afterasteadynervesignalwasobtained(verifiedvisually
changes within the signal (Pincus, 1995; Porta et al., 1998; and by auscultation) the experimental protocol was started.
Richman andMoorman, 2000). Animalswerebledinfour10-mincycles,eachfollowedbya
A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51 45
15-min observation period. Blood was manually withdrawn 4, after LR resuscitation and after blood re-infusion) were
viaasyringefromthefemoralarteriallineduringeach10-min analyzed for every subject. Next, off-line, 800-beat data
bleeding period at a constant rate of 1 ml/kg/min for a total sectionsfromwithineachobservationperiodwereimported
bleedvolume(allfourcycles)of40ml/kg.Theremovedblood into the WinCPRS software. Automatic identification of R-
wascollectedintocommercialblood-collectionbagscontain- waves was carried outby means of an iso-electric line-shift
ing CPD anticoagulant (Terumo Corp., Tokyo, Japan). After algorithm.WeverifiedcorrectidentificationofeachR-wave
thefourthhemorrhagesegmentandobservationperiod,sheep ineverydataset.TheanalyzedRRIdatawasfreeofectopy.
wereresuscitatedwith3timestheshedbloodvolumewithLR ThesoftwaregeneratedtheinstantaneousRRIandSAPtime
overaperiodof25min,wereobservedfor15min,andwere series. Before spectral calculations, signals were windowed
given their shed blood back via infusion over 25 min. with Hanning function and re-sampled with a 5 samples/
ResuscitationwasaccomplishedwiththeModel245Ranger secondrate.There-sampledsignalswerezero-paddedtothe
(Blood/Fluid)Warmingsystem(AugustineMedicalInc,Eden nextpowerof2beforetheFFTroutinewasusedtocalculate
Prairie, MN). Following a final 15-min period after re- the power spectra. Next, the low-frequency (LF) power
infusion, surviving animals were euthanized by means of an (frequency range: 0.04–0.15 Hz), and high-frequency (HF)
overdoseofsodiumpentobarbital(Fatal-Plus,Dearborn,MI). power (frequency range: 0.15–0.4 Hz), components of the
RRI were calculated for each data set with baseline
2.4. Nerve signal analysis normalization. Because of the large inter-animal variability
in absolute values of the spectral data we normalized the
RSNAsignalswereamplified(90,000times)andfiltered spectraldatatothetotalpower(Anonymous,1996).Alldata
at low- and high-frequency cut-offs of 0.7 and 2 kHz reportedinthisstudy arenormalizedspectral data.Normal-
respectively using a Nerve Traffic Analysis System (Model ization was achieved by dividing the integrated LF and HF
662C-4;UniversityofIowaBioengineering,IowaCity,IA). spectra by the total power (minus oscillations occurring
The presence of bursts was verified both visually and by below 0.05 Hz) and multiplying this value by 100. The
auscultation. Automatic amplitude-based detection of sym- normalized,integratedareaswithintheLFbandweredivided
pathetic bursts was performed with WinCPRS software bynormalizedareasintheHFbandtoderivethenormalized
(AbsoluteAliensOy,Turku,Finland).Off-line,thesignalin ratio (LFnu/HFnu). This ratio was taken as an index of
all files was brought to a common baseline via global sympatho-vagalbalance(Paganietal.,1986).
polynomial fitting with the order of 4 and the trends were For non-linear analysis, approximate entropy (ApEn),
removed using the Butterforth high pass filter with the cut- sample entropy (SampEn) and fractal dimension by curve
off frequency of 0.005. RSNA was calculated with an lengths (FDCL) were calculated for the same 800-beat
averagingwindowwidthof5sandaveragewindowstepof segments using the software. Eight hundred beats were
1susingmeannormalization.DiscreteRSNAdatafileswere selectedasanoptimaldatasizeforanalysisinordertoprevent
selected during the observation periods, and were normal- anybiasinApEnwhichispronetochangesinthenumberof
ized with the baseline activity as the reference value. A data points (Pincus, 1995). Before entropy calculations,
scalingfactorwasapplied,calculatedastheratioofthemean linear trends were removed in all segments of the analyzed
areaunderthecurveforoneburstinagiventimepointdata data.ForbothApEnandSampEncalculationsthedimension
file,tothemeanareaunderoneburstatbaselineasshownin parametermwas2andthefilterparameterrwas20%ofthe
theformulabelow: standarddeviation.Analysisalgorithmsareidenticaltothose
SF¼ðarea TP=count TPÞ=ðarea BL=count BLÞ; usedbyKuuselaetal.(2002).
where SF is scaling factor, area is area under each burst, 2.6. Catecholamine assays
countisburstcountintheanalyzedsegment,TPistimepoint
ofexperiment,andBLisbaseline. Next,time pointactivity At each time point, 5 ml of arterial blood was collected
was multiplied by the scaling factor. RSNA is reported as intovacutainertubescontainingEDTA,andwascentrifuged
arbitrary units. at 1000 g for 10 min. The obtained plasma was stored in
polypropylene tubes at −80 °C until the day of the assay.
2.5. Waveform analysis Plasma samples were then thawed in the refrigerator and
assayed according to He et al. (1997) with modifications.
EKG and arterial blood pressure waveforms were con- Epinine (25 μl of a 100 ng/ml solution) was added to each
tinuouslymonitoredthroughouttheexperimentandrecorded 1 ml plasma sample and served as the internal standard.
at 500 Hz using an in-house digital data acquisition system Next,1mlofTRISbuffer(pH9)and50mgofacid-washed
during 15-min-long observation periods that followed each alumina was added, followed by shaking for 15 min. Then,
intervention(i.e.episodesofbloodwithdrawal,resuscitation the samples were centrifuged at 1000 g for 5 min to
or blood re-infusion). This allowed usto do theanalysis on precipitate the alumina, which was washed twice with 1 ml
the most stable discrete data sets. Thus 7 time points ofTRISbuffer.Thealuminawasmixedwith150μlof0.1M
(baseline,afterbleed1,afterbleed2,afterbleed3,afterbleed perchloric acid solution and vortexed for 1 min, than the
46 A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51
Table1
Hemodynamicandcatecholaminedata
Timepoint 0 1 2 3 4 5 6 p1 p2 p3
Event Baseline 10ml/kg 20ml/kg 30ml/kg 40ml/kg LR Re-infusion (0vs.2) (0vs.4) (2vs.4)
N 8 8 8 8 5 4 2
HR 86±5 82±3 85±2 117±12 138±12 114±11 109±17 NS b.0001 b.0001
SAP 101±6 74±5 48±5 38±3 29±3 99±18 109±34 b.0001 b.0001 b.0001
DAP 85±8 59±6 32±5 19±3 14±2 59±13 72±28 b.0001 b.0001 b.0001
CO 2.7±0.1 1.8±0.1 1.2±0.1 0.8±0.1 0.5±0.1 7.6±2.2 6.8±0.7 b.0001 b.0001 b.0001
Lactate 1.5±0.3 1.6±0.3 3.0±0.5 7.5±1.1 11.1±0.8 15.6±1.1 14.4±1.3 .041 b.001 b.001
Dop 74±37 85±47 53±12 118±23 265±77 164±31 119±53 NS .008 .003
Epi 64±36 45±8 118±23 3652±137 14595±6149 611±222 262±106 NS .002 .002
Norepi 382±97 429±88 501±63 1383±290 6901±3052 1095±210 1298±374 NS .009 .011
Timepoint:baseline(0),afterwithdrawalof10(1),20(2),30(3)and40ml/kg(4)ofblood;afterresuscitationwithlactatedRinger's(5);afterre-infusionofshed
blood(6).p1,significancelevelwhenbaselinecomparedwithtimepoint2;p2,significancelevelwhenbaselinecomparedwithtimepoint4;p3,significance
levelwhentimepoint2comparedwithtimepoint4.N,samplesizeateachtimepoint;HR,heartrate;SAP,systolicarterialpressure,mmHg;DAP,diastolic
arterialpressure,mmHg;CO,cardiacoutput,l/min;lactate,plasmalactatelevelsinmmol/l;Dop,plasmadopamine,ng/ml;Epi,plasmaepinephrine,ng/ml;
Norepi,plasmanorepinephrine,ng/ml.Dataaremeans±SEM.
samplewascentrifugedat1000gfor5min.Thesupernatant effect. The covariance structure for the mixed model was
was placed in microfilterfuge tubes and centrifuged for determined using the Bayesian Information Criterion.
2min,and100μlofthesamplewasinjectedintotheHPLC. Hochberg's step-up Bonferroni method was used for p
For calculation of norepinephrine, epinephrine, and dopa- value adjustment for multiple comparisons among time
mine levels in plasma samples, ratios of peak areas of each points. Log transformation was used when appropriate.
catecholamine to the peak area of epinine were compared Statistical significance was determined as pb.05.
againstthoseofaseriesofcalibrators.Theconcentrationsof
the catecholamines were expressed in ng/ml. The HPLC 3. Results
system consisted of a Waters 515 pump, Waters 717
autosampler, Coulochem 2 electrochemical detector Hemorrhageresultedinsevereshock,suchthatonly25%
equipped with a 5011 detector cell, and an Alltima C18 of animals survived until the end of the experiment. All
column (5 μm, 4.6 (ID)×150 (length) mm). The mobile animalssurvivedremovalof30ml/kgofbloodattimepoint
phase was 75 mM phosphate, 1.7 mM octanesulfonic acid, 3. Three animals died after bleed 3; one animal died after
25 μM EDTA, and 5% (v/v) acetonitrile in Milli-Q water. bleed 4; and two animals died after LR resuscitation. The
The flow rate of themobilephase was 1.5 ml/min. physiologic data are summarized in Table 1. Heart rate did
notchangebetweenbaseline(86±5bpm)andafterbleeds1
2.7. Statistics (82±30) or 2 (85±2), but peaked at 138±12 after bleed 4
(pb.0001). SAP decreased from 101±6 mm Hg to 48±5
Data analyses were carried out using SAS, version 8.1 afterbleed2(pb.0001)andreacheditsnadirafterbleed4at
(SAS Institute Inc. 1999). Variables measured at different 29±3 mm Hg (pb.0001). Diastolic arterial pressure
timeswereanalyzedusingamixedmodelANOVAallowing decreased progressively. Cardiac output decreased from
for time as a fixed effect and replicate subject as a random 2.7±0.1 l/min at baseline, to 0.5 l/min after bleed 4
Table2
Renalsympatheticnerveactivity,linearandnon-linearanalysisdata
Timepoint 0 1 2 3 4 5 6 p1 p2 p3
Event Baseline 10ml/kg 20ml/kg 30ml/kg 40ml/kg LR Re-infusion (0vs.2) (0vs.4) (2vs.4)
N 8 8 8 8 5 4 2
RSNA 586±61 848±163 1060±201 1014±312 216±91 1866±787 1435±1305 NS .006 .001
LFnu 0.59±0.08 0.80±0.05 0.88±0.04 0.80±0.10 0.87±0.06 0.40±0.06 0.57±0.13 .009 .001 NS
HFnu 0.35±0.07 0.17±0.03 0.11±0.03 0.16±0.07 0.12±0.05 0.52±0.06 0.36±0.08 .003 .004 NS
LFnu/HFnu 3.25±1.37 6.57±1.74 11.99±2.42 9.31±1.72 20.07±8.50 0.83±0.17 1.75±0.77 .006 .007 NS
ApEn 0.91±0.11 0.84±0.13 0.82±0.14 0.62±0.11 0.59±0.09 0.90±0.16 0.99±0.03 NS .033 NS
SampEn 0.90±0.14 0.85±0.17 0.79±0.15 0.57±0.11 0.52±0.09 0.83±0.22 0.96±0.04 NS NS NS
FDCL 1.72±0.04 1.70±0.03 1.65±0.03 1.50±0.05 1.49±0.06 1.53±0.02 1.75±0.06 NS b.001 .003
Timepoint:baseline(0),afterwithdrawalof10(1),20(2),30(3)and40ml/kg(4)ofblood;afterresuscitationwithlactatedRinger's(5);afterre-infusionofshed
blood(6).p1,significancelevelwhenbaselinecomparedwithtimepoint2;p2,significancelevelwhenbaselinecomparedwithtimepoint4;p3,significance
levelwhentimepoint2comparedwithtimepoint4.N,samplesizeateachtimepoint;RSNA,renalsympatheticnerveactivity;LFnu,normalizedlow-frequency
poweroftheRRI;HFnu,normalizedhigh-frequencypoweroftheRRI;LFnu/HFnu,ratioofthenormalizedlow-frequencypowertonormalizedhigh-frequency
poweroftheRRI;ApEn,approximateentropy;SampEn,sampleentropy;FDCL,RRIfractaldimensionbycurvelengths.Dataaremeans±SEM.
A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51 47
Fig.1.ExampleofRSNAactivityovertime:A.baseline;B.duringfirstbleed(500mlofbloodremoved).Noteasomewhatincreasedburstamplitudeandarea
whencomparedtoA;C.after2ndbleed(980mlofbloodremoved).Areaundereachburstincreasesfurther.Hyperpolarizationandrepetitivefiringsareevident;
D.duringthe3rdbleed(1150mlofbloodremoved).Formationofhyperpolarizedburstconglomerates;E.afterthe4thandfinalbleed(1304mlofblood
removed).RSNAsignificantlydecreased;F.6minaftercompletionof4thbleed.Sympatheticwithdrawal;G.duringresuscitation(1900mlLRhavebeen
infused).PartialreturnofRSNA;H.aftercompletionofresuscitationwithLR,3450mlinfused).ReturnoftonicRSNA.
(pb.0001). Lactate increased significantly from 1.5±0.3 at Fig.1showsthemorphologyofRSNAateachtimepoint
baseline, to 11.1±0.8 mmol/l after bleed 4 (pb.001). in one of the survivors of the full protocol. The pattern of
Changesin baseexcess were similar(data notshown). multiple burst complexes depicted in Fig. 3D is one of the
Plasmacatecholaminesincreasedsignificantlyafterbleed mostpronouncedofallseeninthestudyandwaspresentin
4 (Table 1). the two animals that survived the entire study. Similar
changes—featuring grouping of consecutive bursts into
3.1. RSNA morphology doublets(Fig.3B),triplets,andmultiplebursts(Fig.3C)—
wereobservedasafunctionofprogressivevolumedepletion
RSNA increased in 7/8 animals during HS, although the in all animals. In the 4 animals surviving past the LR
time point at which the highest value was reached varied resuscitation, activity increased to more than 3 times the
among animals. Thus, it increased numerically but not baselinevalue(1866±787)andattheendofthestudy,inthe
significantly from a baseline value of 586±61 to a peak eventual2fullsurvivorsoftheprotocoltheRSNAremained
value of 1060±201 after withdrawal of 20 ml/kg of blood almost 300% ofthe baselinevalue.
(Table 2). RSNA then decreased in all animals, reaching a
nadir of 216±91 after bleed 4 when compared to both post 3.2. Spectral analysis of EKG
bleed 2(p=.006), andto thebaseline(p=.001)(Table 2).
RRILFnuincreasedsignificantlyfromabaselinevalueof
0.59±0.08 to 0.88±0.04 after bleed 2 (p=.009) and was
⁎
Fig.2.ChangesinthenormalizedRRIhigh-frequencypower. ,p=.003,
baselinevs.postbleed2.†,p=.004,baselinevs.postbleed4.Seetextfor Fig.3.Changesinapproximateentropy.†,pb.033,baselinevs.postbleed
details. 4.Seetextfordetails.
48 A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51
sustained at a higher level after bleed 4 when compared to Withrepetitiveandsuperimposedfiringshyperpolarizationmay
baseline(Table2).RRIHFnudecreasedfrombaselinevalues then take place, which may lead to neuronal hyperexcitability
(0.35±0.07) to a nadir (0.11±0.03) after bleed 2 (p=.003, (Kandel et al., 1991) and the development of multiple burst
Table2,Fig.2).Afterbleed4,HFnuremainedlowcompared complexes(Fig.3C–D).Developmentofthesecomplexesmay
tobaseline(0.35±0.07atbaselinevs.0.11±0.03afterbleed bea“lastresort”compensatorymechanismduringpre-terminal
2, p=.004) but was not different from the nadir point after shock, and may be caused by altered neuronal recruitment,
bleed 2 (0.12±0.05, NS) (Fig. 2). The LFnu/HFnu ratio alteredfiringpatterns,and/orprolongedneurotransmitterrelease
increasedsignificantlyafterbleed2(p=.006)andremained eventually leading to receptor saturation. Such multiple burst
elevated throughout the HS period and significantly higher complexes, seen in the two full survivors, may characterize
than baseline after bleed 4 (p=.007) (Table 2). With the subjects with adequate autonomic reserves to withstand near-
LFnu/HFnu remaining elevated, and RSNA showing lethalhypotension.
withdrawal after bleed 4, a significant relationship between Sympathetic withdrawaloccurredin all surviving subjects
theLFnu/HFnuratioandRSNAusingalldatapointswasnot afterbloodlossof40ml/kg.Skoogetal.(1985)foundshort-
revealedby linearregression (p=.121, r2=0.048, n=51). lasting sympathetic activation followed by powerful sympa-
thetic inhibition as assessed by RSNA measurements in
3.3. Non-linear analysis of EKG anesthetizedhemorrhagedrats.Theauthorsproposedthatthe
observedsympatheticinhibitionwasmediatedbymechanically
Both RRI ApEn and SampEn (Table 2) followed similar sensitive cardiac vagal afferents triggered by markedly
trends in this study, decreasing numerically after each blood decreased cardiac filling (Skoog et al., 1985). Koyama et al.
withdrawal and reaching the lowest values after bleed 4 at (1988)reporteddecreasedsympatheticactivityintheirstudies
which point changes in ApEn were significant (ApEn: 0.91± on anesthetized dogs subjected to hemorrhage, and proposed
0.11atbaselinevs.0.59±0.09,p=.033,Table2,Fig.3).The thatprolongedbrainischemiacausesdepressionofsympathetic
fractal dimension by curve lengths (FDCL) progressively outflow in shock. Sympathetic withdrawal was suggested to
decreasedwitheachbloodwithdrawalfromabaselineof1.72± represent a fundamental mechanism for the development of
0.04toanadirof1.49±0.06followingbleed4(pb.001).This circulatoryshockinhumanssubjectedtolower-bodynegative
FDCL value after bleed 4 was also significantly lower when pressure (Convertino et al., 2004). Our findings support
comparedtopostbleed2(p=.003)(Table2). observations in the literature that severe hemorrhage may
causeafailureofcompensationcharacterizedbyadecreasein
4. Discussion sympatheticnerveactivity.
A decrease inRRI HFnu occurred during HS, consistent
We examined changes in RSNA in response to a highly withwithdrawalofvagalmodulationtotheheart(Palazzolo
lethalmodelofhemorrhagicshock(HS)inanesthetizedsheep, etal.,1998).Previously,weidentifiedadecreaseinRRIHF
andassessedtherelationshipbetweennon-invasiveindexesof in anesthetized swine subjected to isovolemic hemorrhagic
cardiac autonomic regulation and RSNA. There were 4 shock (Batchinsky et al., 2007). Similarly, Palazzolo et al.
principalfindings:(1)moderatebloodlossof20ml/kgcaused (1998) documented loss of vagally mediated RRI oscilla-
hypotension, morphologic evidence of RSNA activation, and tionsduring hypotension inconscious dogs.
increasesinLFnu,LFnu/HFnuratioandplasmacatecholamine RSNAandLFnu/HFnuratiodidnotcorrelateinourstudy.
levels; (2) severe blood loss of 40 ml/kg led to RSNA This dissociation between direct microneurographic measure-
withdrawal, sustained LFnu and LFnu/HFnu ratio, and mentsofsympathetictoneandindirectnon-invasiveestimates
continued increases in catecholamine levels; (3) all stages of of autonomic modulation derived from heart rate variability
blood withdrawal were associated with a decrease in the RRI analysishasbeenreportedinothersettingsaswell.Forexample,
HFnu; and (4) blood withdrawal led to decreases in RRI Paganietal.foundagoodcorrelationbetweenLFnuofHRVand
complexity,measuredbyApEnandFDCL. LFnuofmusclesympatheticnerveactivity(MSNA)variability,
RSNAmorphologychangedasafunctionofvolumestatus. but not between LFnu (or LFnu/HFnu) of HRVand MSNA
Despite some degree of inter-animal variability in RSNA res- burstrateinhumans(Paganietal.,1997).Florasetal.(2001)did
ponse to hemorrhage, 7 out of 8 animals demonstrated notfindwithin-subjectcorrelationsbetweenMSNAandLF/HF
increasingburstfrequencywithvolumedepletion.Theampli- ratioduringlower-bodynegativepressure(Florasetal.,2001).
tudeandareaundereachburstalsoincreasedandgroupingof CookeandDowlyn(2000)foundthatRRIspectralanalysisdid
burstsintodoubletsandtripletstookplaceasmorebloodwas not adequately reveal subtle changes in sympathetic traffic
withdrawn.Eventually,burstconglomeratesformedbeforethe duringhead-downtiltasassessedbymusclesympatheticnerve
onset of sympathetic withdrawal in survivors. To our know- activity. As shown by Kingwell et al. (1994) in patients with
ledge, this is the first report on RSNA burst conglomerate heart failure, cardiac sympathetic nerve firing measured with
formationduringseverehemorrhagicshock.Conglomerationof standard spillover techniques and LF changed in opposite
burstsmayindicatetheaccumulationofmultipleactionpoten- directions. Thus, the dissociation between direct tonic and
tialfirings(Tangetal.,2003),possiblyprovidingforastronger indirectHRVmeasuresofautonomicactivityisapredominant
sympathetic discharge in the face of progressive volume loss. findingintheliterature.Additionally,theconceptofdifferential
A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51 49
activationoftheautonomicnervoussysteminvariousvascular RRItimeseriesbyevaluatingthenumberofshortsegmentsor
bedsmaybeanadditionalreasonforthediscrepancybetween “sticks”necessarytofollowthesignalcurve(Chauetal.,1993,
the indirect measures of autonomic modulation of the heart Kuusela et al., 2002). A higher FDCL value indicates an
(HRV) and direct measures of sympathetic tone (Ninomiya exponential increase in the number of sticks required to
etal.,1971;Barmanetal.,1984).Forexample,Barmanetal. describethestructureofthesignal,implyinghighercomplex-
(1984)demonstratedthatincatsthereticularformationcontrols ity.Thedecreaseinfractaldimensionmetricsidentifiedinthis
thesympatheticdischargetocardiac,renalandcarotidnervesin study has been previously documented by us during
anon-uniformfashion.Directcardiacsympatheticnerveactivity isovolemic hemorrhagic shock in swine (Batchinsky et al.,
measurements (Jardine et al., 2005) will likely provide more 2007).ThesedataarealsoconsistentwiththefindingsbyWest
decisive evidence on the relationship between indirect HRV etal.,whoreportedadecreaseinthefractaldimensionofthe
metrics and tonic sympathetic traffic in the heart. Meanwhile, RRI during lower-body negative pressure in humans (West
however,ourfindingssupporttheevidenceintheliteraturethat etal.,2004).
changesinsurrogatemeasuresofcardiacautonomicmodulation Atthispoint,lossofcomplexityduringHSmayberelatedto
suchasLFnu/HFnuratio,donotcorrelatewithdirectmeasures decreasedvagalactivity,sinceitcorrelatedwithdecreasedHFnu
oftonicsympatheticdischargeintherenalvascularbed. inboththis and other studies (Palazzoloetal.,1998; Kuusela
In recent years, non-linear statistical methods have in- et al., 2002; Batchinsky et al., 2007). Alternatively, loss of
creasingly been applied to the analysis of the EKG. These complexity may also be viewed as a non-specific marker of
methods are designed to analyze dynamically changing non- physiologic deterioration under conditions of hypovolemia or
stationary data (Anonymous, 1996) such as those explored in criticalillness.
thisstudy.Complexityisafundamentalcharacteristicofnormal
physiologictimeseries,suchastheRRI.Ithasbeensuggested 5. Conclusions
thatlossofcomplexityor“decomplexification”isafeatureof
disease(GoldbergerandWest,1987)andofimpairedadaptation Inananesthetizedovinemodelofhighlylethalhemorrhagic
to stress (Lipsitz and Goldberger, 1992). Complexity can be shock, moderate hypotension (blood loss of 20 ml/kg) was
assessed by methods such as approximate entropy (ApEn). associatedwithanincreaseintheRRILF/HFratio(anindirect
ApEnquantifiestheprobabilityofencounteringasimilarpattern frequency-domain index of sympatho-vagal balance), and a
within the signal at the next incremental comparison (Pincus, decreaseintheRRIHF(anindexofvagalactivity).Directly
1995);itisaffectedbythenumberofdatapoints.SampEnisa measured tonic RSNA featured increases in the burst fre-
similarapproach,inwhichthevectorcomparisonwithitselfis quency,amplitude,andarea;couplingofburstsintodoublets
removed;thismakesthismethodindependent,intheory,ofthe and triplets; and, finally, formation of burst conglomerates.
numberofdatapoints(RichmanandMoorman,2000).Entropy Bloodlossof40ml/kgledtoRSNAwithdrawal.LF/HFdidnot
ishighinirregular,complexsignalsanddecreasesasthesignals correlate linearly with RSNA. RRI complexity, measured by
becomemoreregular.Inourstudy,bothentropymetrics(ApEn ApEn and fractal dimension methods, decreased after
and SampEn) showed similar trends decreasing numerically completion of the 40 ml/kg hemorrhage. Measures of RRI
with each blood withdrawal and reached a nadir after the 4th complexity may provide useful information about patient
bleed at which point the values for ApEn were significantly status during hypovolemia. Further work will be needed,
lower when compared to baseline (changes in SampEn were however, to understand the relationship between directly
non-significant).Thesefindingsareconsistentwithourprevious measured autonomic nerve activity and various indirect
work that identified a decrease in ApEn and SampEn during measuresofautonomicmodulationoftheheart.
isovolemic hemorrhagic shock in anesthetized swine (Batch-
inskyetal.,2007).Palazzoloetal.(1998)identifiedadecreasein Acknowledgments
ApEnduringhypotensioninconsciousdogs.Complexityinour
current study changed in opposite direction to the indirect WearegratefultoDr.MartyJavorsattheUniversityofTexas
measures of autonomic cardiac modulation, LFnu and LFnu/ Health Science Center at San Antonio for the analysis of the
HFnu. This is in agreement with Porta et al. who used a new plasmasamplesforcatecholaminelevels.Wearealsogratefulto
entropycalculationapproach,thecorrectedconditionalentropy Dr.CharlesWadeforhelpfulcommentsonthemanuscript.
method, and found loss of complexity in the presence of The study was funded by the Combat Casualty Care
increasedLFnuduringhead-uptiltinhumans(Portaetal.,2000) Research Program of the US Army Medical Research and
and during nitroprusside infusion or handgrip (Porta et al., Materiel Command, Ft.Detrick, Maryland.
2007).Correctedconditionalentropyisapromisingtoolwhich
allows for analysis of shorter data sequences without a-priori
References
selectionoftheembeddingdimension(Portaetal.,1998),unlike
ApEnandSampEn.Furtherstudieswillbeneededtodefinethe
optimalentropymethod.
Akselrod,S.,Gordon,D.,Ubel,F.A.,Shannon,D.C.,Berger,A.C.,Cohen,R.J.,
Anothermeasureofcomplexity,FDCL,isafractalimage
1981.Powerspectrumanalysisofheartratefluctuation:aquantitativeprobe
analysismethodthatdescribeschangesinthestructureofthe ofbeat-to-beatcardiovascularcontrol.Science213,220–222.
50 A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51
Anonymous, 1996. Heart rate variability: standards of measurement, Koyama,S.,Aibiki,M.,Kanai,K.,Fujita,T.,Miyakawa,K.,1988.Roleof
physiologicalinterpretationandclinicaluse.TaskForceoftheEuropean central nervous system in renal nerve activity during prolonged
SocietyofCardiologyandtheNorthAmericanSocietyofPacingand hemorrhagicshockindogs.Am.J.Physiol.254,R761–R769.
Electrophysiology.Circulation93,1043–1065. Koyama,S.,Sawano,F.,Matsuda,Y.,Saeki,Y.,Shibamoto,T.,HayashiJr.,T.,
Barman, S.M., Gebber, G.L., Calaresu, F.R., 1984. Differential control of Matsubayashi, Y., Kawamoto, M., 1992. Spatial and temporal differing
sympathetic nerve discharge by the brain stem. Am. J. Physiol. 247, controlofsympatheticactivitiesduringhemorrhage.Am.J.Physiol.262,
R513–R519. R579–R585.
Batchinsky, A.I., Cooke, W.H., Kuusela, T., Cancio, L.C., 2007. Loss of Kuusela, T.A., Jartti, T.T., Tahvanainen, K.U., Kaila, T.J., 2002. Nonlinear
complexity characterizes the heart-rate response to experimental methodsofbiosignalanalysisinassessingterbutaline-inducedheartrateand
hemorrhagicshockinswine.Crit.CareMed.35,519–525. blood pressure changes. Am. J. Physiol, Heart Circ. Physiol. 282,
Butler,G.C.,Yamamoto,Y.,Hughson,R.L.,1994.Fractalnatureofshort-term H773–H783.
systolicBPandHRvariabilityduringlowerbodynegativepressure.Am.J. Lipsitz,L.A.,Goldberger,A.L.,1992.Lossof‘complexity’andaging.Potential
Physiol.267,R26–R33. applications of fractals and chaos theory to senescence. JAMA 267,
Chau,N.P.,Chanudet,X.,Bauduceau,B.,Gautier,D.,Larroque,P.,1993. 1806–1809.
Fractaldimensionofheartrateandbloodpressureinhealthysubjects Madwed,J.B.,Cohen,R.J.,1991.Heartrateresponsetohemorrhage-induced
andindiabeticsubjects.BloodPress2,101–107. 0.05-Hzoscillationsinarterialpressureinconsciousdogs.Am.J.Physiol.
Convertino, V.A., Ludwig, D.A., Cooke, W.H., 2004. Stroke volume and 260,H1248–H1253.
sympatheticresponsestolower-bodynegativepressurerevealnewinsight Malpas,S.C.,Evans,R.G.,Head,G.A.,Lukoshkova,E.V.,1998.Contribution
intocirculatoryshockinhumans.Auton.Neurosci.111,127–134. ofrenalnervestorenalbloodflowvariabilityduringhemorrhage.Am.J.
Cooke, W.H., Dowlyn, M.M., 2000. Power spectral analysis imperfectly Physiol.274,R1283–R1294.
informschangesinsympathetictrafficduringacutesimulatedmicrogravity. Ninomiya,I.,Nisimaru,N.,Irisawa,H.,1971.Sympatheticnerveactivityto
Aviat.SpaceEnviron.Med.71,1232–1238. the spleen, kidney, and heart in response to baroceptor input. Am. J.
Cooke,W.H.,Hoag,J.B.,Crossman,A.A.,Kuusela,T.A.,Tahvanainen,K.U., Physiol.221,1346–1351.
Eckberg,D.L.,1999.Humanresponsestouprighttilt:awindowoncentral Notarius,C.F.,Floras,J.S.,2001.Limitationsoftheuseofspectralanalysis
autonomicintegration.J.Physiol.517(Pt2),617–628. ofheartratevariabilityfortheestimationofcardiacsympatheticactivity
Cooke,W.H.,Salinas,J.,Convertino,V.A.,Ludwig,D.A.,Hinds,D.,Duke,J.H., inheartfailure.Europace3,29–38.
Moore,F.A.,Holcomb,J.B.,2006.Heartratevariabilityanditsassociation Pagani,M.,Lombardi,F.,Guzzetti,S.,Rimoldi,O.,Furlan,R.,Pizzinelli,P.,
withmortalityinprehospitaltraumapatients.J.Trauma60,363–370. Sandrone,G.,Malfatto,G.,Dell'Orto,S.,Piccaluga,E.,etal.,1986.Power
Eckberg, D.L., 1997. Sympathovagal balance: a critical appraisal. spectralanalysisofheartrateandarterialpressurevariabilitiesasamarkerof
Circulation96,3224–3232. sympatho-vagal interaction in man and conscious dog. Circ. Res. 59,
Evans, R.G., Ventura, S., Dampney, R.A., Ludbrook, J., 2001. Neural 178–193.
mechanismsinthecardiovascularresponsestoacutecentralhypovolaemia. Pagani,M.,Montano,N.,Porta,A.,Malliani,A.,Abboud,F.M.,Birkett,C.,
Clin.Exp.Pharmacol.Physiol.28,479–487. Somers, V.K., 1997. Relationship between spectral components of
Floras,J.S.,Butler,G.C.,Ando,S.I.,Brooks,S.C.,Pollard,M.J.,Picton,P., cardiovascularvariabilitiesanddirectmeasuresofmusclesympathetic
2001. Differential sympathetic nerve and heart rate spectral effects of nerveactivityinhumans.Circulation95,1441–1448.
nonhypotensive lower body negative pressure. Am. J. Physiol., Regul. Palazzolo,J.A.,Estafanous,F.G.,Murray,P.A.,1998.Entropymeasuresofheart
Integr.Comp.Physiol.281,R468–R475. ratevariationinconsciousdogs.Am.J.Physiol.274,H1099–H1105.
Goldberger,A.L.,West, B.J.,1987.Fractalsinphysiologyandmedicine. Parati,G.,Saul,J.P.,DiRienzo,M.,Mancia,G.,1995.Spectralanalysisof
YaleJ.Biol.Med.60,421–435. blood pressure and heart rate variability in evaluating cardiovascular
Goldstein,B.,Mickelsen,D.,Want,A.,Tipton,R.,Cox,C.,Woolf,P.D.,1999. regulation.Acriticalappraisal.Hypertension25,1276–1286.
EffectofN(G)-nitro-L-argininemethylesteronautonomicmodulationof Parati,G.,Mancia,G.,DiRienzo,M.,Castiglioni,P.,2006.Point:cardiovascular
heart rate variability during hypovolemic shock. Crit. Care Med. 27, variabilityis/isnotanindexofautonomiccontrolofcirculation.J.Appl.
2239–2245. Physiol.101,676–678(discussion681-672).
Grossman,P.,Wilhelm,F.H.,Spoerle,M.,2004.Respiratorysinusarrhythmia, Pincus,S.,1995.Approximate entropy(ApEn)as acomplexitymeasure.
cardiacvagalcontrol,anddailyactivity.Am.J.Physiol,HeartCirc.Physiol. Chaos5,110–117.
287,H728–H734. Porta, A., Baselli, G., Liberati, D., Montano, N., Cogliati, C., Gnecchi-
He, H., Stein, C.M., Christman, B., Wood, A.J., 1997. Determination of Ruscone, T., Malliani, A., Cerutti, S., 1998. Measuring regularity by
catecholaminesinsheepplasmabyhigh-performanceliquidchromatogra- meansofacorrectedconditionalentropyinsympatheticoutflow.Biol.
phywithelectrochemicaldetection:comparisonofdeoxyepinephrineand Cybern.78,71–78.
3,4-dihydroxybenzylamineasinternalstandard.J.Chromatogr.,B,Biomed. Porta,A.,Guzzetti,S.,Montano,N.,Pagani,M.,Somers,V.,Malliani,A.,
Sci.Appl.701,115–119. Baselli, G., Cerutti, S., 2000. Information domain analysis of
Jardine, D.L., Charles, C.J., Ashton, R.K., Bennett, S.I., Whitehead, M., cardiovascularvariabilitysignals:evaluationofregularity,synchronisa-
Frampton,C.M.,Nicholls,M.G.,2005.Increasedcardiacsympatheticnerve tionandco-ordination.Med.Biol.Eng.Comput.38,180–188.
activityfollowingacutemyocardialinfarctioninasheepmodel.J.Physiol. Porta, A., Guzzetti, S., Furlan, R., Gnecchi-Ruscone, T., Montano, N.,
565,325–333. Malliani, A., 2007. Complexity and nonlinearity in short-term heart
Kandel,E.R.,Schwartz,J.H.,Jessell,T.M.,1991.PrinciplesofNeuralScience. period variability: comparison of methods based on local nonlinear
AppletonandLange,EastNorwalk,CT. prediction.IEEETrans.Biomed.Eng.54,94–106.
Kawase, M., Komatsu, T., Nishiwaki, K., Kimura, T., Fujiwara, Y., Richman, J.S., Moorman, J.R., 2000. Physiological time-series analysis
Takahashi,T.,Shimada,Y.,2000.Heartratevariabilityduringmassive usingapproximateentropyandsampleentropy.Am.J.Physiol,Heart.
hemorrhageandprogressivehemorrhagicshockindogs.Can.J.Anaesth. Circ.Physiol.278,H2039–H2049.
47,807–814. Saul,J.P.,Rea,R.F.,Eckberg,D.L.,Berger,R.D.,Cohen,R.J.,1990.Heart
Kingwell,B.A.,Thompson,J.M.,Kaye,D.M.,McPherson,G.A.,Jennings, rateandmusclesympatheticnervevariabilityduringreflexchangesof
G.L., Esler, M.D., 1994. Heart rate spectral analysis, cardiac autonomicactivity.Am.J.Physiol.258,H713–H721.
norepinephrinespillover,andmusclesympatheticnerveactivityduring Secher, N.H., Jacobsen, J., Friedman, D.B., Matzen, S., 1992. Bradycardia
human sympathetic nervous activation and failure. Circulation 90, duringreversiblehypovolaemicshock:associatedneuralreflexmechanisms
234–240. andclinicalimplications.Clin.Exp.Pharmacol.Physiol.19,733–743.
A.I.Batchinskyetal./AutonomicNeuroscience:BasicandClinical136(2007)43–51 51
Skoog,P.,Mansson,J.,Thoren,P.,1985.Changesinrenalsympatheticoutflow West, B.J., Scafetta, N., Cooke, W.H., Balocchi, R., 2004. Influence of
duringhypotensivehaemorrhageinrats.ActaPhysiol.Scand.125,655–660. progressive central hypovolemia on Holder exponent distributions of
Tang,X.,Chander,A.R.,Schramm,L.P.,2003.Sympatheticactivityandthe cardiacinterbeatintervals.Ann.Biomed.Eng.32,1077–1087.
underlyingactionpotentialsinsympatheticnerves:asimulation.Am.J. Winchell,R.J.,Hoyt,D.B.,1996.Spectralanalysisofheartratevariabilityin
Physiol.,Regul.Integr.Comp.Physiol.285,R1504–R1513. theICU:ameasureofautonomicfunction.J.Surg.Res.63,11–16.