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Changes in axonal excitability of primary sensory afferents withgeneral anaesthesia in humans

Maurer, K; Wacker, J; Vastani, N; Seifert, B; Spahn, D R

Maurer, K; Wacker, J; Vastani, N; Seifert, B; Spahn, D R (2010). Changes in axonal excitability of primary sensoryafferents with general anaesthesia in humans. British Journal of Anaesthesia, 105(5):648-656.Postprint available at:http://www.zora.uzh.ch

Posted at the Zurich Open Repository and Archive, University of Zurich.http://www.zora.uzh.ch

Originally published at:British Journal of Anaesthesia 2010, 105(5):648-656.

Maurer, K; Wacker, J; Vastani, N; Seifert, B; Spahn, D R (2010). Changes in axonal excitability of primary sensoryafferents with general anaesthesia in humans. British Journal of Anaesthesia, 105(5):648-656.Postprint available at:http://www.zora.uzh.ch

Posted at the Zurich Open Repository and Archive, University of Zurich.http://www.zora.uzh.ch

Originally published at:British Journal of Anaesthesia 2010, 105(5):648-656.

For Peer Review

1

Induction of general anaesthesia changes axonal excitability of primary sensory afferents in humans

K. Maurer, 1 J. Wacker,

1 N. Vastani,

1 B. Seifert,

2 D.R. Spahn

1

1 Institute of Anaesthesiology, University Hospital of Zurich, Switzerland

2 Biostatistics Unit, Institute of Social and Preventive Medicine, University of Zurich, Switzerland

Correspondence to:

Dr. Konrad Maurer

Head, Pain Research Unit

Institute of Anaesthesiology

University Hospital Zurich

Rämistrasse 100

8091 Zurich

Switzerland

Phone: +41 44 255 9379

Fax: +41 44 255 4409

Email: [email protected]

Short title: General anaesthesia changes nerve excitability.

Key words: Anaesthetics i.v., propofol

Anaesthetics volatile, sevoflurane

Nerve, membrane

Nerve, transmission

Ions, ion channels, voltage-gated

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Summary

Background

Intraoperative monitoring of neuronal functions is important in a variety of surgeries. The type of

general anaesthetic used may have an important influence on the interpretation and quality of the

obtained recordings. Although primary effects of general anaesthetics are synaptic-mediated, it

remains unclear to what extent they affect excitability of the peripheral nervous afferent system.

Methods

40 patients were randomised in a stratified manner into two equally sized groups. Induction was

performed either with propofol or sevoflurane. We used the technique of threshold tracking (QTRAC®)

to measure nerve excitability parameters of the sensory action potential of the median nerve before

and after induction of general anaesthesia.

Results

Only a small number of parameters of peripheral nerve excitability of sensory afferents changed after

induction of general anaesthesia and they were similar for both, propofol and sevoflurane. The

maximum amplitude of the sensory nerve action potential decreased in both groups significantly

(Propofol: 25.3 %; Sevoflurane: 29.5 %; both P < 0.01). The relative refractory period also decreased

similarly in both groups (Propofol: -0.6 (0.7) ms; Sevoflurane: -0.3 (0.5) ms; both P < 0.01). Skin

temperature at the stimulation site increased significantly in both groups (Propofol: +1.2 (1.0) °C;

Sevoflurane: +1.7 (1.4) °C; both P < 0.01).

Conclusions

Changes of excitability of primary sensory afferents after induction of anaesthesia with propofol and

sevoflurane were subtle and have been found under high concentrations of the anaesthetics. They

were non-specific and can potentially be explained by the temperature changes found.

Comment [km1]: Ref Question 1 & 2

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Intraoperative monitoring of neuronal function is important in a variety of surgeries. It is well known

that the type and concentration of general anaesthetics have an important influence on the quality of

the neurophysiological signals recorded and, hence their interpretation.1 All the electrophysiological

(clinical) investigations have so far considered the effect of general anaesthetics on afferent axons as

negligible, since axons are not usually considered as neural targets for these agents.2-4

In the

peripheral nervous system, a well organized interaction of different subtypes of voltage-gated ion

channels defines the size, frequency and the speed of an action potential. Even a slight shift in

membrane potential of a nerve membrane can lead to severely altered excitability properties, and

thereby modulate the information conveyed to the central nervous system.5

6 Propofol and

sevoflurane seem to affect human voltage-gated ion channels at clinically relevant concentrations and

therefore could contribute to the effects on peripheral afferent excitability. 7-10

The aims of our investigation were to test whether propofol and sevoflurane have different effects on

axonal excitability of peripheral sensory afferents after induction of general anaesthesia and how

sensory nerve monitoring could be influenced during anaesthesia.

We used the technique of threshold tracking, a diagnostic tool which assesses axonal nerve

excitability of myelinated peripheral nerves in a non-invasive manner.11-13

In contrast to conventional

nerve conduction studies threshold tracking technique uses subthreshold currents and provides

information about nerve excitability and ion channel function.

Comment [km2]: Ref. Question 7

Comment [km3]: Ref. To new reference 10

Comment [km4]: Ref. Clarification 3)


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Ref. Question 7

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Ref. To new reference 10

kmau

Ref. Clarification 3)

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Methods

Study subjects and randomisation

Study subjects: The study was performed at the University Hospital Zurich after approval by the local

ethics committee (University Hospital Zurich, StV. 5-2008) and in accordance with the Declaration of

Helsinki 2008 (registration number NCT00696254, www.clinicaltrials.gov). All study subjects gave

written informed consent after careful instructions concerning the study details, in particular they

agreed on the omission of premedication with a tranquiliser to exclude as many unknown influencing

factors as possible. Inclusion criteria were: German speaking patients scheduled for surgery under

general anaesthesia; age 18 – 70 years; weight 50 - 100 kg; signed informed consent. Exclusion

criteria were: Known peripheral neuropathy; diabetes mellitus; neuro-psychiatric diagnosis; pregnant/

breast-feeding women; congestive heart disease; more than 2 risk factors out of 4 for postoperative

nausea and vomiting (PONV): female, non-smoker, known PONV, planned opioids postoperatively;

participation in other studies; inability of verbal expression. 40 patients were randomised in a stratified

manner into two equally sized groups of 20 subjects (Figure 1). Induction was performed either with

propofol or sevoflurane.

Threshold tracking

We used a computer-assisted threshold tracking program (QTRAC ©

Institute of Neurology, Queen

Square, London, UK) to investigate the excitability parameters of myelinated axons and ion

conductances in the median nerve.11-14

This technique provides different information than

conventional nerve conduction studies which use supramaximal stimuli and provide information about

conduction velocity and amplitude. Threshold tracking technique uses subthreshold currents and

provides information about excitability and ion channel function. For more details about the technique

itself and the electric model which it is the base for the understanding and correct interpretation of the

data obtained refer to previous publications.11

13-16

In the current study we measured the excitability of the median nerve stimulated transcutaneously at

the wrist (Figure suppl. A-C). The sensory nerve action potential (SNAP) was recorded antidromically

at Dig. II with skin electrodes. The temperature at the stimulation site was measured at the end of

each measurement. We used a recording protocol comprising 5 main parts: Stimulus-response curve,

strength-duration relationship, recovery cycle, threshold electrotonus and current-threshold

relationship. Further information about the stimulation patterns in our protocols, how the


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measurements were plotted and interpreted, is provided in the supplemental method and suppl.

figure.

Excitability measurements during anaesthesia induction in patients

The study was performed in the anaesthesia induction room in the hospital’s OR facility. After the

completion of the first nerve excitability measurement immediately prior to induction, the anaesthesia

was administered exclusively with propofol or sevoflurane, respectively. Notably, no local

anaesthetics, muscle relaxants or opioids were used until the second measurement was completed.

Pulse oxymetry (Draeger Infinity Delta Systems, Draeger Medical Systems, Danvers, MA) was

attached on a finger on the same arm where we measured the nerve excitability. Subjects were

monitored with ECG, non-invasive blood pressure (NIBP) and and end-tidal gas analysis (FiO2 and

etCO2; volume %) using a KION anaesthesia workstation (KION workstation and Maquet SC 7000

screen, Maquet-Siemens, Rastatt, Germany). During the first measurement, baseline end-tidal CO2

values were determined during quiet spontaneous respiration using a tight face mask. In order to

reach a comparable depth of anaesthesia in both groups we aimed at a stable BIS value below 40

(BIS QuatroTM sensors with Infinity® BISx Pod for Draeger; Software version 1.03, Aspect Medical

Systems, Inc., Norwood, MA). According to this parameter we varied the target concentration of the

propofol infusion (using a software-controlled infusion pump with the pharmacological model of

Schnider et al. 17

programmed to plasma concentration) and the inspiratory concentration of

sevoflurane, respectively. To minimize burning pain at the i.v. cannulation site the intravenous

lactated Ringer’s infusion was infused at a high flow rate until the patient was deeply sedated. For the

induction of anaesthesia with sevoflurane we asked the subjects to breath 100 % oxygen for 3

minutes with a tightly sealed oxygen mask. To begin anaesthesia, we opened the sevoflurane

vaporizer maximally (8 %) and then decreased the concentration gradually over the next few minutes.

Controlled manual ventilation was used to stabilise ventilation, maintaining end-tidal CO2 values at

the baseline level

Before we recorded the second measurement we adjusted the concentrations of anaesthetics until

the patients were in a steady state which was defined as follows: end-tidal CO2 persistently at

baseline level, haemodynamic stability, BIS consistently < 40, constant calculated plasma

concentration of propofol or difference of inspired and expired concentration of sevoflurane < 0.3 %,

respectively.


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The second nerve excitability measurement was performed when a sustained steady state was

established (end-tidal CO2 persistently at baseline level, haemodynamic stability, BIS consistently <

40, difference of inspired and expired concentration of sevoflurane < 0.3 %). After the second

measurement was recorded, the study ended for the patient. Standard anaesthesia techniques were

carried out by addition of a muscle relaxant, opioid and benzodiazepine where indicated. The

anaesthesia was continued according to institutional standards for the respective procedures.

Statistical Analysis

All values are given as means and standard deviations (SD) except in figures 3, 4 and Suppl. Figures

2 and 3 where standard errors of the mean (SEM) were used to visualise the significance between

groups. To analyse data we used the software QtracP (Version 3/4/2009, ® Institute of Neurology,

University College London, UK). Data were tested for a normal distribution with Lilliefors test for

normality. Gender was compared between groups using Fisher’s exact test. Data before and after

induction of anaesthesia within the same group were analysed with a paired t-test. To compare

excitability changes between the two groups after induction of anaesthesia we used an unpaired t-

test. A Bonferroni correction was performed to address multiple comparisons for the same variable.

Thus, p-values of comparisons before and after induction of anaesthesia within the same group and

between groups after intervention were considered significant if p < 0.013.


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Results

Changes of general parameters during anaesthesia induction

We enrolled 17 female and 23 male patients in the investigation (Fig. 1). All 40 patients completed the

study. The demographic data of the studied population before starting anaesthesia are shown in table

1. After inducing anaesthesia, a comparable anaesthesia depth was achieved during the second

measurement in both groups (Propofol: BIS 25 (8). Sevoflurane: 27 (10); P = 0.42). The calculated

plasma concentration of propofol during the second measurement was 6.6 (1.3) µg ml-1. The end-

expiratory sevoflurane concentration was 5.5 (1.3) %. The concentration of end-expiratory CO2 during

and after the induction of anaesthesia was not significantly higher than the value during spontaneous

breathing before anaesthesia induction (Propofol spontaneous: 4.5 (0.5) kPa; anaesthetised: 4.6 (0.5)

kPa, P = 0.09. Sevoflurane spontaneous: 4.6 (0.4) kPa; anaesthetised: 4.6 (0.4) kPa, P = 0.72.).

There was no difference in the end-expiratory CO2 levels between the two groups before (P = 0.59) or

after induction of anaesthesia (P = 0.84) (Fig. 2 A). Skin temperature at the stimulation site increased

significantly in both groups, (Propofol before: 32.5 (0.9) °C; after: 33.7 (1.1) °C, P < 0.01. Sevoflurane

before: 32.4 (1.2) °C; after: 34.1 (1.0) °C, P < 0.01). No difference existed between the groups after

induction of anaesthesia (P = 0.17) (Fig. 2 C). Mean arterial pressure dropped significantly in both

groups (Propofol before: 101 (13) mmHg; after: 81 (12) mmHg, P < 0.01. Sevoflurane before: 101

(14) mmHg; after: 78 (12) mmHg, P < 0.01) but was similar when compared between the groups (P =

0.55) after the induction (Fig. 2 D). Blood pressure stabilised after reaching equilibrium of anaesthetic

depth, end-expiratory CO2 concentration, stable BIS value and stable SO2. We did not need to take

any pharmacological or other measures in any of the patients to stabilise blood pressure.

Changes of excitability parameters during anaesthesia induction

A summary of the excitability changes during the anaesthesia induction is given in table 2 and table 3.

We observed significant changes in only three excitability parameters following the induction of

anaesthesia.

First, the maximum amplitude of the SNAP decreased in both groups significantly (Before propofol:

39.9 (5.8) µV; after: 29.8 (4.4) µV, P < 0.01. Before sevoflurane: 47.1 (7.8) µV; after: 33.2 (5.3) µV, P

< 0.01) which corresponds to a decrease of 25.3 % and 29.5 %, respectively (Fig. 3). No difference

was observed between the two groups after induction of anaesthesia (P = 0.44). As a result, the

stimulus-response curve shifted downwards on the stimulus-response plot. In the group with

Comment [km5]: Ref. Comment nr. 8)


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sevoflurane, additionally a distinctive shift of the curve to the right was observed. For this group, this

implies that more current was needed to elicit the target action potential.

Second, during the early phase of the recovery cycle, the curve shifted to the left in both groups (Fig.

4). Consequently, the first intersection with the control threshold (the current that is normally needed

to elicit the targeted size of the action potential, plotted as a straight line at 0 % on the y-axis) and

representing the end of the relative refractory period (RRP, s. Suppl. Fig. 5 C.), occurred earlier. The

RRP decreased similarly in both groups (Before propofol: 4.4 (1.2) ms; after: 3.9 (1.1) ms, P = 0.01.

Before sevoflurane: 4.0 (1.1) ms; after: 3.7 (1.2) ms, P < 0.01). No difference was observed between

the two groups after induction of anaesthesia (P = 0.28).

Third, the overshoot after a hyperpolarising conditioning stimulus was less prominent after

anaesthesia induction (Before propofol: 20.3 (2.9) %; after: 16.1 (3.4) %, P = 0.01. Before

sevoflurane: 19.7 (6.2) %; after: 14.5 (5.6) %, P = 0.01; Table 3). ‘Overshooting’ represents an

increase in current required to reach the target threshold after the end of the hyperpolarising

conditioning stimulus.15

It corresponds to the activities of certain voltage gated membrane ion

channels (mainly slow potassium currents) which counteract the changes of the membrane potential

induced by the hyperpolarising conditioning stimulus. Consequently, we would also expect a

decrease of excitability at the end of the hyperpolarising stimulus (e.g. at TEh [90-100ms]). However,

this parameter remained stable after induction, which cannot be conclusively explained at present.


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Discussion

This study shows for the first time that general anaesthesia affects excitability of primary sensory

afferents significantly. Using subthreshold currents rather than supramaximal currents could detect

smaller changes of nerve excitability properties than conventional nerve conduction studies or SSEP.

Possible intrinsic effect of anaesthetics on excitability parameters

Previous studies have shown that both anaesthetics alter nerve excitability at clinically relevant

concentrations by modulating voltage-gated sodium and/or potassium channels in the central and

peripheral nervous system.7 8 10 12 18

In our investigation, parameters sensitive to changes in

membrane potential remained almost unchanged after the induction of anaesthesia. However, we

found three parameters which could be explained by an intrinsic effect of the general anaesthetics on

peripheral nerve excitability:

(i) The relative refractory period was shorter: Investigations with the same threshold-tracking

technique have shown that blocking of sodium channels results in a shift of the recovery

cycle curve to the left combined with a decrease of superexcitability in sensory afferents.

19 20

The latter parameter, however, remained unchanged in our investigation.

(ii) The amplitude of the maximum peak response decreased: The size of a compound

action potential decreases by temporal dispersion caused by different underlying

mechanisms: differential slowing of individual fibers, availability of the largest-diameter

fibers and changes in the amplitude of individual spikes.21 22

An intrinsic blocking of

voltage-gated sodium channels could have led to a reduced size of peak response.23

However, this conclusion could only be drawn in stable recording conditions. In our study,

the temperature at the stimulation site changed significantly and, therefore, was likely to

have an important influence on the response size (s. below).

(iii) We observed no change in latency: According to previously published data we would

expect a pronounced decrease in latency, hence increase in conduction velocity, of about

3 % caused by the extent of temperature change at the stimulation site.24

But the

latencies remained unchanged before and after the induction of anaesthesia. This could

imply that the faster kinetics of the voltage-gated ion channels induced by the higher

temperature (s. below) was counteracted by a partial block of sodium channels.


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None of the above findings, however, are specific for an intrinsic effect of the investigated general

anaesthetics on nerve excitability. Temperature change at the stimulation site increased significantly

and, therefore, has to be taken into account when interpreting these results.

Changes of excitability caused by an increase of skin temperature during anaesthesia induction.

Temperature variation results in several changes of peripheral nerve excitability. Although in our

study the temperature differences between the first and the second measurement were small, they

were significant. We can explain this by a decrease in sympathetic activity.25

Consequently, a change

in skin microcirculation occurred and resulted in a vasodilation with a substantial increase in skin

temperature.26

27

Temperature changes affect all nerve excitability indices measured with threshold

tracking to some extent.28

The most prominent effect is a decrease of the refractoriness with

increasing temperature.24 28

Our results are in line with this finding, reflected by the leftward shift of

the recovery cycle curve indicating a shortening of the relative refractory period. The underlying

mechanism is a faster recovery from inactivation of voltage-gated sodium channels due to the higher

temperature.29

An increase in temperature also leads to a linear increase in conduction velocity.30

31

According to

previously published data we would expect a decrease in latency of about 3 % caused by the extent

of temperature change measured in our study.24

However, latencies were not different and also, we

did not find a linear relationship of refractoriness and latency changes described by Burke et al.28

Therefore we might assume that the temperature change of around 1.5 °C was too small to affect

excitability. Also, temperature changes affect conditioned evoked potentials much more than

unconditioned potentials.28

Refractoriness – in contrast to conduction velocity - was measured with a

supramaximal conditioning stimulus, therefore, our findings would fit well with this theory.

In our investigation the size of the compound action potential decreased after induction of

anaesthesia with both anaesthetics. The effect of temperature on the size of the compound action

potential is more complex to explain. On one hand, the compound action potential is sensitive to

temporal dispersion of individual action potentials of nerve axons. Increasing the temperature,

therefore, decreases the amount of dispersion of the compound action potential and results in greater

amplitude.31

On the other hand, an increase of temperature decreases the duration of the action

potential and leads to a smaller action potential. Hence, the measured size of the action potential at

higher temperatures is a combination of both effects. In the study by Kiernan et al. who investigated


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motor nerve fibres, the peak amplitude increased only when rising the temperature between 32° C to

35° C, although the overall effect of temperature increase was a decrease of SNAP.24

Similar non-

linear functions between SNAP size and temperature changes have been described by Ludin et al.30

Anaesthesia depth, ventilation and haemodynamic changes did not affect the measurements

Our study endpoint was to compare the effect on peripheral nerve excitability of two different general

anaesthetics at an equipotent dosage. Clinically, at first glance these concentrations appear to be

high because propofol and sevoflurane are usually used in combination with co-anaesthetics (e.g.

opioids or benzodiazepines). In our study however, patients were not premedicated and until the end

of the second measurement we did not use any other co-medication. Consequently, the target

plasma concentrations of propofol and the end-expiratory sevoflurane concentrations had to be

chosen higher than normally to reach BIS values below 40.

Several parameters which could have influenced nerve excitability were controlled very closely. In our

study, anaesthesia induction was performed exclusively by one senior anaesthesiologist and

normoventilation was achieved throughout the recording period. We cannot fully exclude alveolar

hypercapnia. However, according to previously published data strength-duration time constant would

probably be the most sensitive parameter to detect hypercapnia.32

Since this parameter also

remained unchanged we may assume that alveolar pCO2 did not influence our measurements.

According to our measurements, depth of anaesthesia was equal in both groups and therefore

contributed equally – if at all – to the measured excitability changes.

Although induction of anaesthesia caused a significant drop in blood pressure values, they never

reached low physiological values. Furthermore, the oxygen saturation in the same arm remained

stable. Therefore, we are sure that the perfusion pressure in the neuronal tissue at the stimulation

site was high enough to prevent hypoperfusion or hypoxemia of the nerve and did not cause

previously described changes in excitability.33-35

Conclusion

Induction of general anaesthesia with propofol and sevoflurane resulted in a change of excitability of

primary sensory afferents. These changes were subtle and have been found under high

concentrations of the anaesthetics. A direct effect of general anaesthetics on excitability could not be

excluded and was minimal at most; it is more likely that the significant changes we found were

Comment [km6]: Ref. To Comment nr. 4)


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Ref. To Comment nr. 4)

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Ref. To Question 1) & 2)

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caused by an increase in temperature at the site of stimulation. Further investigations with specific

designs are needed to elucidate the differentiation between the two mechanisms. However, our

findings should raise the awareness of possible interference during intraoperative neuromonitoring

caused by general anaesthetics in the peripheral nervous system.

Comment [km7]: Ref. To Question 1) & 2)


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Acknowledgements:

Staff of the Institute of Anaesthesiology, University Hospital Zurich, Switzerland

Prof. Hugh Bostock, UCL Institute of Neurology, UCL, London, United Kingdom


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References:

1. Chen Z. The effects of isoflurane and propofol on intraoperative neurophysiological

monitoring during spinal surgery. J Clin Monit Comput 2004; 18: 303-8

2. Pathak KS, Amaddio MD, Scoles PV, Shaffer JW, Mackay W. Effects of halothane, enflurane,

and isoflurane in nitrous oxide on multilevel somatosensory evoked potentials.

Anesthesiology 1989; 70: 207-12

3. Raymond SA, Shin HC, Steffensen SC. A role for changes in axonal excitability in general

anesthesia. Ann N Y Acad Sci 1991; 625: 307-10

4. Sloan TB, Jäntti V. Anesthetic effects on evoked potentials. In: Nuwer MR, ed. Intraoperative

Monitoring of Neuronal Functions: Elsevier, 2008; 94-126

5. Raymond SA, Thalhammer JG, Popitz-Bergez F, Strichartz GR. Changes in axonal impulse

conduction correlate with sensory modality in primary afferent fibers in the rat. Brain Res

1990; 526: 318-21

6. Amir R, Michaelis M, Devor M. Membrane potential oscillations in dorsal root ganglion

neurons: role in normal electrogenesis and neuropathic pain. J Neurosci 1999; 19: 8589-96

7. Friederich P, Urban BW. Interaction of intravenous anesthetics with human neuronal

potassium currents in relation to clinical concentrations. Anesthesiology 1999; 91: 1853-60

8. Friederich P, Benzenberg D, Trellakis S, Urban BW. Interaction of volatile anesthetics with

human Kv channels in relation to clinical concentrations. Anesthesiology 2001; 95: 954-8

9. Friederich P, Benzenberg D, Urban BW. Ketamine and propofol differentially inhibit human

neuronal K(+) channels. Eur J Anaesthesiol 2001; 18: 177-83


123456789101112131415161718192021222324252627282930313233343536373839404142434445464748495051525354555657585960

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15

10. Herold KF, Nau C, OuYang W, Hemmings HC, Jr. Isoflurane inhibits the tetrodotoxin-resistant

voltage-gated sodium channel Nav1.8. Anesthesiology 2009; 111: 591-9

11. Bostock H, Cikurel K, Burke D. Threshold tracking techniques in the study of human

peripheral nerve. Muscle Nerve 1998; 21: 137-58

12. Burke D, Bartley K, Woodforth IJ, Yakoubi A, Stephen JP. The effects of a volatile

anaesthetic on the excitability of human corticospinal axons. Brain 2000; 123 ( Pt 5): 992-

1000

13. Kiernan MC, Lin CS, Andersen KV, Murray NM, Bostock H. Clinical evaluation of excitability

measures in sensory nerve. Muscle Nerve 2001; 24: 883-92

14. Z'graggen WJ, Bostock H. Nerve membrane excitability testing. Eur J Anaesthesiol Suppl

2008; 42: 68-72

15. Burke D, Kiernan MC, Bostock H. Excitability of human axons. Clin Neurophysiol 2001; 112:

1575-85

16. Maurer K, Bostock H, Koltzenburg M. A rat in vitro model for the measurement of multiple

excitability properties of cutaneous axons. Clin Neurophysiol 2007; 118: 2404-12

17. Schnider TW, Minto CF, Gambus PL, et al. The influence of method of administration and

covariates on the pharmacokinetics of propofol in adult volunteers. Anesthesiology 1998; 88:

1170-82

18. Frenkel C, Urban BW. Human brain sodium channels as one of the molecular target sites for

the new intravenous anaesthetic propofol (2,6-diisopropylphenol). Eur J Pharmacol 1991;

208: 75-9

Comment [km8]: Ref. To comment nr. 9)


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For Peer Review

16

19. Kiernan MC, Isbister GK, Lin CS, Burke D, Bostock H. Acute tetrodotoxin-induced

neurotoxicity after ingestion of puffer fish. Ann Neurol 2005; 57: 339-48

20. Kuwabara S, Misawa S, Tamura N, et al. The effects of mexiletine on excitability properties of

human median motor axons. Clin Neurophysiol 2005; 116: 284-9

21. Fink BR, Cairns AM. Differential slowing and block of conduction by lidocaine in individual

afferent myelinated and unmyelinated axons. Anesthesiology 1984; 60: 111-20

22. Gokin AP, Philip B, Strichartz GR. Preferential block of small myelinated sensory and motor

fibers by lidocaine: in vivo electrophysiology in the rat sciatic nerve. Anesthesiology 2001; 95:

1441-54

23. Butterworth JF, Strichartz GR. Molecular mechanisms of local anesthesia: a review.

Anesthesiology 1990; 72: 711-34

24. Kiernan MC, Cikurel K, Bostock H. Effects of temperature on the excitability properties of

human motor axons. Brain 2001; 124: 816-25

25. Landsverk SA, Kvandal P, Bernjak A, Stefanovska A, Kirkeboen KA. The effects of general

anesthesia on human skin microcirculation evaluated by wavelet transform. Anesth Analg

2007; 105: 1012-9

26. Matsukawa T, Sessler DI, Sessler AM, et al. Heat flow and distribution during induction of

general anesthesia. Anesthesiology 1995; 82: 662-73

27. Gonzalez de ZJ, Olmos A, Alvarez JC, Ruiz N, de AB, Gonzalez-Fajardo JA. [Core and

cutaneous thermal changes in the upper limb after anesthesia induction]. Rev Esp Anestesiol

Reanim 2000; 47: 287-92


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17

28. Burke D, Mogyoros I, Vagg R, Kiernan MC. Temperature dependence of excitability indices of

human cutaneous afferents. Muscle Nerve 1999; 22: 51-60

29. Louis AA, Hotson JR. Regional cooling of human nerve and slowed Na+ inactivation.

Electroencephalogr Clin Neurophysiol 1986; 63: 371-5

30. Ludin HP, Beyeler F. Temperature dependence of normal sensory nerve action potentials. J

Neurol 1977; 216: 173-80

31. Denys EH. AAEM minimonograph #14: The influence of temperature in clinical

neurophysiology. Muscle Nerve 1991; 14: 795-811

32. Mogyoros I, Kiernan MC, Burke D, Bostock H. Excitability changes in human sensory and

motor axons during hyperventilation and ischaemia. Brain 1997; 120 ( Pt 2): 317-25

33. Bostock H, Baker M, Grafe P, Reid G. Changes in excitability and accommodation of human

motor axons following brief periods of ischaemia. J Physiol 1991; 441: 513-35

34. Grosskreutz J, Lin C, Mogyoros I, Burke D. Changes in excitability indices of cutaneous

afferents produced by ischaemia in human subjects. J Physiol 1999; 518 ( Pt 1): 301-14

35. Krishnan AV, Lin CS, Kiernan MC. Excitability differences in lower-limb motor axons during

and after ischemia. Muscle Nerve 2005; 31: 205-13

36. Bostock H, Rothwell JC. Latent addition in motor and sensory fibres of human peripheral

nerve. J Physiol 1997; 498 ( Pt 1): 277-94

37. Weiss G. Sur la possibilité de render comparables entre eux les appareils servant

al'excitation électrique. Archives of Italian Biology 1901; 35: 413-47


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38. Bostock H. The strength-duration relationship for excitation of myelinated nerve: computed

dependence on membrane parameters. J Physiol 1983; 341: 59-74

39. Raymond SA. Effects of nerve impulses on threshold of frog sciatic nerve fibres. J Physiol

1979; 290: 273-303

40. Bostock H, Baker M. Evidence for two types of potassium channel in human motor axons in

vivo. Brain Res 1988; 462: 354-8


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Declaration of interests

None of the authors have a conflict of interest.


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Funding

This study was funded by the Swiss Foundation for Anaesthesia Research (SFAR), by the Swiss

National Science Foundation (Grant Nr. SPUM 33CM30_124117) and the Institute of

Anaesthesiology, University Hospital Zurich, Switzerland.


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Table 1. Demographic data.

PROPOFOL SEVOFLURANE P value

Age (y) 43 (14) 39 (12) 0.37

Weight (kg) 77 (10) 72 (15) 0.37

Height (cm) 177 (7) 173 (11) 0.30

Female/male 6/14 11/9 0.21

Values are presented as number (n) or as mean and (SD). P < 0.05 is considered statistically

significant.


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Table 2: Overview of measured parameters.

before

PROPOFOL

after

PROPOFOL P value

before

SEVOFLURANE

after

SEVOFLURANE P value P value*

Latency (ms) 4.9 (0.5) 4.9 (0.4) 0.17 4.9 (0.4) 4.9 (0.4) 0.16 (0.85)

Peak response

SNAP (µV) 39.9 (5.8) 29.8 (4.4) <0.<0.<0.<0.01010101 47.1 (7.8) 33.2 (5.3) <0.0<0.0<0.0<0.01111 (0.44)

Strength-duration

time constant (µS) 665 (135) 657 (134) 0.67 668 (184) 599 (265) 0.21 (0.40)

Rheobase (mA) 7.1 (1.6) 6.7 (1.5) 0.30 6.6 (1.6) 6.4 (1.6) 0.82 (0.73)

Relative refractory

period (ms) 4.4 (1.2) 3.9 (1.1) <0.0<0.0<0.0<0.01111 4.0 (1.1) 3.7 (1.2) <0.0<0.0<0.0<0.01111 (0.28)

Superexcitability

(%) -14.1 (5.8) -15.1 (5.0) 0.30 -18.1 (10.2) -15.1 (7.5) 0.14 (0.93)

Subexcitability (%) 13.6 (6.5) 12.0 (4.0) 0.17 12.3 (3.7) 15.7 (20.9) 0.52 (0.44)

Temperature (C) 32.5 (0.9) 33.7 (1.1) <0.0<0.0<0.0<0.01111 32.4 (1.2) 34.1 (1.0) <0.0<0.0<0.0<0.01111 (0.17)

CO2 (kPa) 4.5 (0.5) 4.6 (0.5) 0.09 4.6 (0.4) 4.6 (0.4) 0.72 (0.84)

BIS 98 (3) 25 (8) <0.0<0.0<0.0<0.01111 97 (2) 27 (10) <0.0<0.0<0.0<0.01111 (0.42)

SO2 (%) 99 (1) 99 (1) 0.32 99 (1) 99 (1) 0.41 (0.83)

MAP (mmHg) 101 (13) 81 (12) <0.<0.<0.<0.01010101 101 (14) 78 (12) <0.0<0.0<0.0<0.01111 (0.55)

Values are presented as mean and (SD). P < 0.013 is considered statistically significant. *P-values in

brackets represent comparison between ‘after PROPOFOL’ and ‘after SEVOFLURANE’.

Comment [km9]: Ref. Comment nr. 8)


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kmau

Ref. Comment nr. 8)

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Table 3 : Threshold electrotonus and current/threshold relationship

before

PROPOFOL

after

PROPOFOL P value

before

SEVOFLURANE

after

SEVOFLURANE P value P value*

TEh(90-100ms)

% 125.0 (20.8) 119.0 (16.3) 0.28 120.4 (19.7) 122.3 (22.8) 0.68 (0.66)

TEd(10-20ms) % 58.6 (7.8) 60.6 (5.2) 0.12 61.2 (6.1) 60.4 (8.9) 0.42 (0.89)

TEd(90-100ms)

% 42.7 (7.8) 44.6 (7.1) 0.17 44.6 (7.0) 45.6 (10.7) 0.37 (0.72)

TEh(10-20ms) % 69.6 (12.9) 67.7 (6.9) 0.70 70.9 (9.0) 73.8 (16.2) 0.94 (0.19)

TEd(undershoot)

% 21.9 (4.8) 21.9 (4.6) 0.88 21.1 (3.4) 19.0 (3.9) <0.01<0.01<0.01<0.01 (0.03)

TEh(overshoot) % 20.3 (2.9) 16.1 (3.4) <0.0<0.0<0.0<0.01111 19.7 (6.2) 14.5 (5.6) <0.<0.<0.<0.01010101 (0.36)

Resting I/V

slope 0.6 (0.1) 0.6 (0.1) 0.11 0.6 (0.2) 0.6 (0.2) 0.06 (0.50)

Minimum I/V

slope 0.3 (0.1) 0.3 (0.1) 0.65 0.3 (0.1) 0.3 (0.1) 0.70 (0.90)

Hyperpol. I/V

slope 0.7 (0.5) 2.2 (4.8) 0.84 0.4 (0.1) 0.6 (0.4) 0.29 (0.41)

Values are presented as mean and (SD). P < 0.013 is considered statistically significant. *P-values in

brackets represent comparison between ‘after PROPOFOL’ and ‘after SEVOFLURANE’.


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Legends to illustrations

Figure 1. Enrolment and randomisation of patients recruited to the study

Figure 2. Ventilation parameters and temperature during anaesthesia induction.

A: Endexpiratory CO2 remained stable after the induction of anaesthesia. We therefore can assume

that tissue pH was unaffected by the anaesthesia induction B: Peripheral oxygen saturation -

measured with pulsoxymetry on the arm where the recording was made - did not change after the

induction indicating that no tissue ischemia occurred at the stimulation and recording site. C: The

temperature at the stimulation site increased significantly during the induction with both general

anaesthetics. This increase was similar with both, propofol and sevoflurane. D: Mean arterial

pressure decreased significantly in both groups. The changes occurred in all subjects within a

physiological range. Solid lines indicate means; dashed lines indicate SEM. Empty circles indicate

values before induction, grey circles represent propofol, black circles sevoflurane after induction.

Figure 3. Peak response and latency changes.

A: The stimulus-response curve of the maximum sensory nerve action potential showed a downwards

shift on the y-axis after induction of anaesthesia with both general anaesthetics. No significant shift

occurred on the x-axis indicating that the current strength needed to elicit maximum response was not

affected. B: The size of the maximum peak response was significantly smaller for both anaesthetics

but no difference was found between the two. The latency of the peak response was not affected in

either group. Circles indicate the mean current needed to elicit a 50 % of the maximum peak

response. Empty circles indicate values before induction, grey circles represent propofol, black circles

sevoflurane after induction. Error bars SEM.

Figure 4. Recovery cycle.

A. The early recovery cycle curve was affected after induction with both anaesthetics. This is best

illustrated by the shift of the first intersection of the curve with the control threshold (end of the relative

refractory period). No significant shift on the y-axis at a given interval between the conditioning

stimulus and test stimulus (interstimulus interval) occurred at interstimulus intervals longer than the

relative refractory period. This indicates that neither the superexcitable nor the subexcitable periods


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were affected. B: The maximum extent of the test current changes was equally stable for both,

superexcitability and subexcitability after induction of anaesthesia with both anaesthetics. The relative

refractory period was significantly shorter in both groups but not different between the groups. Circles

represent mean threshold changes at a given interstimulus interval. Empty circles indicate values

before induction, grey circles represent propofol, black circles sevoflurane after induction. Error bars

are SEM.


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Suppl. Methods: Threshold tracking

The threshold-tracking program QTRAC (©

Institute of Neurology, Queen Square, London, UK)

adjusts the stimulus strength in a feed-back controlled manner for different test paradigms to produce

the target response using proportional tracking.36

In this study we used this technique to investigate

the excitability parameters of the median nerve by recording antidromically the sensory nerve action

potential (SNAP) the way it was described for the first time by Kiernan et al. (Suppl. Figure 2A).13

Throughout the recording protocol, a stimulus frequency of 1 Hz and a stimulus width of 0.5 ms were

chosen except otherwise stated.

As a first step, a stimulus-response curve was generated. The stimulus strength was gradually

stepped up until a maximum response of the sensory nerve action potential was reached (Suppl.

Figure 2B-1). An amplitude of about 40% size of the maximum amplitude was then defined as the

target amplitude and the stimulus current needed to reach this amplitude was called ‘threshold

current’. This target amplitude is chosen automatically by the program as the point with the maximal

slope between 30% and 50% of the stimulus response. The rationale behind the defined ‘threshold

current’ is that a small change in stimulus strength would have a largest change in amplitude of the

target response.16

In the following, changes of this ‘threshold current’ were measured continuously in

response to various test stimulus configurations and automatically adjusted.

Second, to record the strength duration relationship, stimuli of different widths (0.1 ms to 0.5 ms)

were applied (Suppl. Figure 2B-2). The strength-duration time constant (τSD) was calculated off-line

from thresholds measured according to Weiss’s formula.37 38

Third, the recovery of excitability following a single supramaximal 0.5-ms conditioning stimulus was

measured (Suppl. Figure 2B-3). Excitability changes were recorded at 18 different conditioning-test

intervals decreasing from 200 ms to 1 ms in an approximately geometric sequence. The resulting

recovery cycle curve allows to identify the different phases of the excitability changes of a nerve after

a supramaximal stimulus (e.g. relative refractory period, superexcitable period, subexcitable period)

as described by Raymond (Suppl. Figure 2C).39

Three stimulus conditions were tested in turn (control

test stimulus, supramaximal conditioning stimulus alone, and the combined conditioning and test


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stimuli) on three different channels. The response to the conditioning stimulus alone subtracted from

the response to the combined stimulus results in the effective response to the test stimulus.

Fourth, a threshold electrotonus was recorded (Suppl. Figure 2B-4). The excitability properties of the

nerve were altered by passing a polarising 100-ms subthreshold current through the whole nerve.

The polarising currents were set to + 40 % (depolarising) and - 40 % (hyperpolarising) of the

threshold current. Thresholds were then tested at defined delays between 0 ms and 200 ms during

and after the start of the polarising current (‘threshold-electrotonus’).40

During this section of the

protocol, the control, depolarised and hyperpolarised thresholds were tested in turn on three different

channels of the program.

As a last step the current-voltage relationship was recorded (Suppl. Figure 2B-5). Excitability of the

nerve was tested at a fix interval of 200 ms after the start of a polarising current. The polarising

currents were stepped down from + 50 % (depolarising) of the threshold current to -100 % (=

hyperpolarising current) in steps of 10 %. During this section of the protocol, the control and the

thresholds after polarisation were tested in turn on two different channels of the program.


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Suppl Fig 5: Experimental setup and stimulation protocol.

A: Setup for excitability measurements: The median nerve was stimulated with the cathode placed at

the wrist and the anode placed on the radial side of the forearm. A sensory nerve action potential

(SNAP) was recorded antidromically from Dig. II. The signal was amplified and data were then

digitized by a data acquisition unit. According to the signal size recorded a ‘threshold tracking’-

software (QTRAC©) adjusted the current applied by the stimulator for each stimulus. If the signal was

too small as for the defined response size (defined as 40% of the maximum peak response) the

current of the next stimulus was increased and vice versa. Skin temperature was measured at the

stimulation site. Oxygen saturation was measured by pulsoxymetry throughout the recording on the

same side where the recording was being performed.

B: Graphical depiction of the stimulation patterns used in the protocol. 1: A 0.5 ms rectangular

stimulus was steadily increased until the size of the SNAP was maximal. 2: To record the strength

duration relationship, stimuli of different widths (0.1 ms to 0.5 ms) were applied 3: After a

supramaximal conditioning stimulus, excitability changes were recorded at different conditioning-test

intervals between 200 ms and 2 ms in an approximately geometrical sequence. 4: To record a

threshold electrotonus, 100 ms subthreshold polarising currents set to + 40% (depolarising) and -

40% (hyperpolarising) of the target threshold current were applied. Thresholds were tested at defined

delays during and after the start of the polarising current. 5: To record the I/V-curve subthreshold

currents with different polarisation strengths were applied and threshold was tested at 200 ms after

the start of the polarising current.

C: The different periods during a recovery cycle of a sensory afferent nerve are illustrated. The

change of currents of the test stimuli to reach the threshold is plotted on the y-axis in a normalised

way whereby zero represents the unconditioned control threshold. Immediately after a supramaximal

conditioning stimulus the nerve enters the refractory period. The refractory period ends as soon as

the curve crosses the zero line for the first time (indicated with the dashed line). Thereafter, the nerve

is superexcitable until it becomes subexcitable (second intersection of the curve with the zero line).

Normal excitability is restored after 200 ms.

Suppl. Fig. 6. Threshold electrotonus.

A: On the y-axis the reduction of threshold induced by the polarising conditioning current is shown

(polarising current starts at 10 ms). Positive values imply that a weaker current was needed to reach


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the threshold and vice versa. Threshold changes were affected by neither of the anaesthetics,

propofol or sevoflurane, after induction of anaesthesia. B: The histograms of the threshold reduction

at 100 ms of the conditioning stimulus (indicated by the black arrow in A) illustrate the similarity of the

measured values before and after the induction of anaesthesia. Circles represent mean threshold

changes at a given time interval. Empty circles indicate values before induction, grey circles represent

propofol, black circles sevoflurane after induction. Error bars are SEM.

Suppl. Fig. 7. Current-threshold relationship.

A: Current-threshold relationship reflects the rectifying properties of the nerve membrane as a

response to long polarising conditioning currents. On the depolarising side (right from the zero-axis)

the decrease of excitability reflects outward rectification, on the hyperpolarising side (left from the

zero-axis) the increase of excitability reflects inward rectification illustrated by the open arrows. B:

The slope of the current-threshold relationship is the threshold analogue of the conductance. Both,

minimum slope of the curve on the hyperpolarising side and resting slope (intersection of the two zero

axis) were not altered by propofol and sevoflurane, respectively. Circles represent mean threshold

changes at a given polarisation defined as a percentage of the threshold current. Empty circles

indicate values before induction, grey circles represent propofol, black circles sevoflurane after

induction. Error bars are SEM.


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Figure 1

215x279mm (200 x 200 DPI)


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Figure 2

189x133mm (300 x 300 DPI)


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Figure 3 186x193mm (300 x 300 DPI)


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Figure 4 196x159mm (300 x 300 DPI)


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Supplemental figure 5 182x271mm (300 x 300 DPI)


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