國立台灣大學醫學院物理治療學系暨研究所 碩士論文
School and Graduate Institute of Physical Therapy College of Medicine
National Taiwan University Master Thesis
經皮神經電刺激應用於主要動作皮質對於動作學習時 皮質興奮性與前額葉活化之影響：經顱磁刺激與近紅外
線吸收光譜研究
Effect of Transcutaneous Electrical Nerve Stimulation on Primary Motor Cortex to Modulate Cortical Excitability and Prefrontal Activation during Motor Learning: A TMS
and NIRS Study 施政楷 Jheng-Kai Shih
指導教授：陸哲駒博士、張雅如博士 Adviser: Jer-Juhn Luh, PhD, Ya-Ju Chang, PhD
中華民國 103 年 7 月
July, 2014
誌謝
碩士班兩年的生活對於我目前人生而言其實算是蠻大的轉折，碩士班這兩年 我不斷的再學習，不管是待人處世還是論文實驗，但是對我來說最困難的永遠是 人情世故這塊，我總是學習如何讓自己成為一個知情打理的人，而在過程中我也 發現，不論是臨床或是學術表現，待人處事永遠需要具備來襯托所自己所奉獻的 每一事物，而過程之中必然有許多的貴人，而要感謝的就是這些貴人所帶來的人 生體悟。
首先要感謝的是陸哲駒老師，是你引領我踏入這電生理魔幻的世界，探索並 發掘每一種可能，我們總是為了讓神經元產生我們想要的改變不斷的努力。從一 開始專任助理到後面的碩士論文，在摸索的過程中，總是可以發現在我即將迷失 時，有一雙溫暖的手輕輕的帶起了我，而老師您對於學生的體貼、體諒無比是學 生的動力來源，感謝陸老師讓我在碩士班這兩年，圓了我一回又一回的夢。而在 完成這份對我來說有點深奧的論文，雅如老師提供了我很多資源與環境，對於實 驗解釋也幫助了我度過重重難關，每次見到您的笑容總是那麼的正向與積極，在 研究上的不安與恐懼總是能帶來安定，而另外要感謝的就是邱醫師、黃正雅老師、
周立偉老師，如果沒有你們，我可能仍然會陷在解不開的泥淖中，而這本論文更 是因為你們才能如此的完備且完善。
碩士班兩年的過程中，大部分的日子總是特別的忙碌與緊繃，而過程中最重 要的就是陪我努力跨出每一步的同學，在自己墜落時，輕輕的將負荷接起，在整 個論文建構過程中，最要感謝的就是奕瑄學姊，走在你已經幫我開好的橋樑上，
總是會讓我想起你辛苦的付出才能讓我這後輩承接順利，對不起我總是在你打 TMS 時候睡著，但是每次和你到長庚作實驗都是快樂的回憶，也是我所有操作與 實驗的源頭，還是要再次感謝學姊的幫忙，沒有你的幫忙，政楷可能現在還在長 庚收案。接著是實驗室好夥伴燕玲，總是可以在生活中畫龍點睛，創造出歡樂排 解壓力，而你同時也是實驗室的好夥伴，也是一起同時踏出亞洲的伴遊，謝謝你
總是可以掏心掏肺的討論生活中面臨的危機。接著是林沂，處事圓融總是我借鏡 的對象，同時也扮演了很重要的人力仲介，我實驗龐大的收案量都是靠你如同棉 花糖般綿密的人脈所連接，另外也要感謝你這位知心好友很多事情的陪伴。另一 個要感謝的是在碩一時時時刻刻陪伴的好友梓聰，總是能在你身上感受到真誠的 友誼，謝謝你始終當我的第一號受試者，
最後要感謝的就是我的母親、父親、哥哥，哥哥總是在第一時間義不容辭的 幫助我解決許多關於研究的問題，而論文的測試工具程式也是哥哥一手幫忙寫出 來的，而父親和母親總是在我回家的那一刻，讓我感覺到被家人支持與鼓勵的感 覺，雖然我人在遙遠的台北，但是台中永遠是我的歸宿，因為爸媽我才能勇往直 前，挑戰我的自我潛能。
這一路走來嚐到人生酸甜苦辣，也腳踩過了地球的另外一端，而兩年的生涯 就在剎那間飛逝，卻留下久久不能忘卻的人生美酒甘醇，永遠的感謝所有在這過 程中曾經幫過政楷的朋友們。
中文摘要
前言：根據先前的實驗結果所發現，經皮神經電刺激對於主要動作皮質的活性所 造成的影響，無法與動作學習所產生的興奮性神經塑性有所區別，因此本實驗主 要是為了瞭解頭顱經皮神經電刺激以及動作學習所產生去抑制現象之間的交互關 係，而本實驗主要藉由量測在執行內隱式順序性動作學習的受試者主要動作皮質 興奮性的改變，而本實驗另外使用近紅外線吸收光譜來量測前額葉在動作學習的 過程中血液動力學反應的改變。方法：本實驗總共從社區以及大學徵招了48位年 輕受試者，而受試者將會被隨機分配到電刺激組與電刺激伴隨動作學習測試組，
而受試者需要完成兩次實驗，包含經皮神激電刺激和假刺激，而兩次測試則是隨 機分配，試驗的間隔則需要大於一周，而本實驗成果測試項目包括：動作誘發電 位、皮質內誘發、皮質內抑制、前額葉血液動力學變化以及順序性反應性動作測 試中的反應時間長度，綜合以上神經生理或是行為科學上的量測來觀察經皮神經 電刺激的效果。結果：在動作電位上則是發現有接受經皮神經電刺激的兩種試驗 顯著上升，包含單純接受電刺激介入以及電刺激介入伴隨動作表現量測之情形，
發現動作學習對於經皮神經電刺激所產生的效果產生協同作用，而在有動作執行 的兩組(動作執行伴隨電刺激或假刺激)則是發現皮質內抑制所產生的抑制量明顯 上升，而有電刺激伴隨動作執行量測的組別改變抑制的百分比顯著較假刺激明顯，
而皮質內誘發則是在有電刺激的兩組(有無動作執行)顯著下降，但是組間的比較 則是沒有明顯差異，而順序性反應時間測試中則是兩組(電刺激/假刺激)隨著練習 的次數增加時明顯的降低反應時間，但是電刺激介入並沒有顯著影響動作表現，
因此兩組間則是沒有顯著差異，而在前額葉血液動力學變化中則是只有在電刺激 介入的情形有顯著效應，而在初始練習時發現有活性明顯上升的情形，而在30分 鐘後測時發現顯著低於初始值的活性，因此代表電刺激的組別有一定的幫助學習 固化的現象。討論：經皮神經電刺激顯著對皮質脊髓神經元產生明顯的興奮性誘 發，但是同時間則發現皮質內抑制也有明顯增加的情形，而皮質內抑制上升則是
被認為會干擾皮質重組或是去覆蓋現象(unmasking)，而顯著動作的電位上升伴隨 皮質內誘發明顯的下降則是有可能在皮質內誘發的量測中發生天花板效應，而在 動作表現上則沒有發現明顯電刺激造成的效應，結論：本實驗證實經皮神經電刺 激明顯增加主要動作皮質內之興奮性，而且動作練習對於經皮神經電刺激所產生 的效果產生聯合反應，需要更進一步大於24小時的長期追蹤，或是增加電刺激介 入的劑量，來進一步討論經皮神經電刺激的效應。
關鍵字：經皮神經電刺激、經顱磁刺激、近紅外線吸收光譜、內隱式順序性動作 學習、神經塑性
ABSTRACT
AIM: The relationship between cranial transcutaneous electrical nerve stimulation
(TENS) stimulation and cortical excitability change during motor learning process is
unknown. This study aims to explore the effects of cranial TENS application on cortical
excitability of primary motor cortex (M1) during implicit sequential motor learning
process in normal subjects. Prefrontal activation pattern in learning process was also
monitored by Near-infrared spectroscopy (NIRS). METHODS: 48 volunteers were
recruited from colleges and communities. Subjects were randomized into TENS
stimulation group (Stimulus-TENS/Sham) and TENS stimulation with motor task group
(Motor-TENS/Sham). Subjects in both groups need to accomplish two trials (TENS or
sham stimulation), the interval between trials was more than 1 week. Motor evoked
potential (MEP), intracortical inhibition (ICI) and intracortical facilitation (ICF), Serial
reaction time task and NIRS were measured to monitor physiological and behavioral
change process in motor learning. RESULTS: MEPs amplitude in both Motor-TENS
and Stimulus-TENS group increased significantly. MEP amplitude of Motor-TENS
were significantly higher than Stimulus-TENS in followed up 60 mins. Motor task
induced synergistic effect on enhancement of MEP amplitude. Higher suppression
effects of ICI were also found in both Motor-TENS and Motor-Sham. Motor-TENS go
higher suppression of ICI than Motor-Sham which revealed synergistic effect of TENS
stimulation. The ICF was also decreased in Motor-TENS and Stimulus TENS. However,
between group comparison showed no significant different. In SRTT performance,
reaction times significantly improved both in Motor-TENS/Sham but no difference
between TENS and sham stimulation. Prefrontal activation showed significant time
effect in TENS-Motor only. Increment activation in initial learning and following
decrease activation in retention test was observed. Consolidation effect in Motor-TENS
than Motor-Sham was also noted. DISSCUSSION: TENS intervention increased
corticospinal neuron excitability. However, significant increase suppression induced by
ICI may indicate disruption of cortical representation. Increment of ICF concurrent with
increment MEP showed ceiling effect existed in ICF measurement. TENS intervention
showed weak effect to altered motor learning process. CONCLUSION: TENS
stimulation increase cortical excitability and inhibitory shift of intracortical circuits.
Motor practice played a facilitation role to altered cortical excitability which induced
synergistic effect on TENS intervention. Further study should be done to investigate
the effect of TENS with long-term (more than 24 hour) effect or increase times of
stimulus program.
Key words: Transcutaneous electrical nerve stimulation, transcranial magnetic
stimulation, implicit sequential motor learning, near-infrared spectroscopy,
neuroplasticity
LIST OF ABBREVIATIONS
tES Transcranial electrical stimulation
ES Electrosleep
EA Electroanesthesia
TCES Transcerebral electrotherapy stimulation
TCET Transcranial electrostimulation treatment
NET NeuroElectric therapy
tDCS Transcranial direct current stimulation
tACS Transcranial alternating stimulation
tPCS Transcranial pulsed current stimulation
CES Cranial electrotherapy stimulation
TENS Transcutaneous electrical nerve stimulation
TMS Transcranial magnetic stimulation
MT Motor threshold
MEP Motor evoked potential
ICI Intracortical inhibition
ICF Intracortical facilitation
fMRI Functional magnetic resonance image
BOLD Blood oxygen dependent level
PET Positron emission tomography
SRTT Serial reaction time task
NIRS Near infrared spectroscopy
[O2Hb] Oxygenated hemoglobin
[HHb] Deoxygenated hemoglobin
[tHb] Total hemoglobin
[Hbdiff] Hemoglobin difference
OD Optical density difference
SCD Scalp cortical distance
GABA Gamma-Amino butyric acid
NMDA N-methyl-D-aspartate
LTP Long-term potentiation
EEG Electrical encephalography
EMG Electromyogram
M1 Primary motor cortex
DLPFC Dorsal lateral prefrontal cortex
SMA Supplementary motor area
FDI First dorsal interosseous
ANOVA Analysis of variance
CONTENTS TABLE
口試委員審會定書 ... ⅰ 誌謝 ... ⅱ 中文摘要 ... ⅳ Abstract ... ⅵ List of abbreviations ... ⅷ Contents table ... ⅹ
Figures list ... xii
Tables list ... xiii
Chapter 1: Introduction ... 1
1.1 Background ... 1
1.2 Purpose ... 4
1.3 Question and hypothesis ... 6
1.4 Significance ... 7
Chapter 2: Literature review ... 9
2.1 Historical view of transcranial electrical stimulation ... 9
2.2 Comparison stimulation parameter of transcranial electricl stimution ... 11
2.3 Cranial electrotherapy stimulation (CES) ... 13
2.2.1 Research in animal model... 14
2.2.2 Rresearch in human model ... 15
2.4 Transcranial magnetic stimulation (TMS) ... 18
2.4.1 Motor threshold ... 20
2.4.2 Motor evoked potential ... 21
2.4.3 Intracortcial facilitation and intracortcial inhibition ... 22
2.5 Near infrared spectroscopy (NIRS) ... 23
2.5.1 Mechanism of NIRS ... 23
2.5.2 Psychometric studies of NIRS ... 26
2.6 Serial reaction time task (SRTT) ... 29
2.6.1 Implicit sequential motor learning process ... 29
2.6.2 Brain activation pattern during SRTT ... 30
Chapter 3: Methods ... 32
3.1 Participants ... 32
3.2 Study design ... 33
3.3 Experimental procedure ... 33
3.4 Experimental assessment ... 36
3.5 Statistical analysis ... 37
Chapter 4 : Results ... 39
4.1 Basic data and baseline measurements ... 39
4.2 Effect of cranial ten interventions ... 39
4.2.1 Results of motor evoked potential ... 39
4.2.2 Results of pair pulsed stimulation ... 40
4.2.3 Results of prefrontal hemodynamic response ... 41
4.2.4 Results of serial reaction time task ... 42
Chapter 5: Discussion ... 43
5.1 Neurophysiological outcomes after cranial tens intervention ... 43
5.1.1 Motor evoked potential ... 44
5.1.2 Intracortical inhibition ... 45
5.1.3 Intracortical facilitation ... 46
5.1.4 Prefrontal hemodynamic response... 47
5.1.5 Serial reaction time task ... 48
5.2 Possible clinical application of cranial tens intervention ... 48
5.3 Limitation ... 49
5.4 Future studies ... 50
Chapter 6: Conclusion ... 51
Reference ... 52
Figures ... 64
Tables ... 72
Appendix ... 78
FIGURES LIST
Figure 1. Study design and group allocation ... 64
Figure 2. Experimental Procedure ... 65
Figure 3. Results of motor evoked potential ... 66
Figure 4. Results of Intracortical inhibition ... 67
Figure 5. Results of intracortical facilitation ... 68
Figure 6. Prefrontal hemodynamic responses in motor learning process ... 69
Figure 7. Results of reaction time in block sequence practice ... 70
Figure 8. Results of reaction time in random number practice (transfer task) ... 71
TABLES LIST
Table 1. Basic data of participants... 72
Table 2. Baseline of all outcome measurements ... 73
Table 3. Results of MEP amplitude and between trial comparisons ... 74
Table 4. Results of ICI measurement and between trials comparison ... 75
Table 5. Results of ICF measurement and between trials comparison ... 76
Table 6. Results of prefrontal hemodynamic response and between trials comparison 77
Chapter 1: Introduction
1.1 Background
Non-invasive brain stimulation (NBS) was one kind of central nervous system
stimulation by small intensity of electrical current or magnetic field generated current
to stimulate brain tissue. NBS can be classified into several types dependent on
stimulation parameters and electrodes placements such as transcranial direct current
stimulation (tDCS), transcranial alternating stimulation (tACS), transcranial magnetic
stimulation (TMS) and cranial electrotherapy (CES)1.
tDCS had much of evidences of its effect on cortical excitability modulation.
Direct electrical current induced polarization effect on cortex tissue. Cathodal
stimulation induced inhibitory effect on cortical excitability. Anodal stimulation
induced excitatory effect on cortical excitability. Polarized electrical field would
increase the difference of intracellular and extracellular membrane potential. The
positive change of extracellular membrane potential induced by anodal stimulation
would facilitate NMDA pathway1. However, single direction current may easily induce
skin burn.
tACS was one kind of alternating current stimulation which commonly used
biphasic sinusoid wave with low(10-50 Hz) and high frequency (kHz). tACS was
believed to interrupt electroencephalic oscillation which may induce change of brain
observed. After Fast Fourier transform of the electrical signal, three level of frequency
were identified. α wave was consist of signals which frequency range from 8-13 Hz, β
wave was 13-30 Hz and γ was 30-100Hz. The Previous tACS study investigated the
effect on cortical excitability, but the results were still controversial. β frequency range
tACS (15-20Hz) was found more effective than other frequency band. Inhibitory effect
which reduced intracortical facilitation(ICF) was reported3. Other studies also
indicated that the stimulation would interrupt subject’s motor performance4,5. In
addition,, Kanai in 2010 applied 20 Hz tACS on visual cortex and found that the
phosphate threshold was decreased6. Another study also reported that tACS can induce
phosphate7.
Cranial electrotherapy (CES) was developed over 20th century. CES stimulation
can modulate neurotransmitter level which relieved insomnia, bipolar syndrome,
depression or anxiety8. The electrodes were placed on supra-auricular and
infra-auricular area where try to stimulate brain stem through cranial nerve or cervical
nerve. Previous study revealed that the CES would enhance dopamine and
norepinephrine level in rat’s hypothalamic area9. General brain activity was significant
decrease after CES stimulation also reported in fMRI study10. Connectivity of frontal
lobe and parietal lobe was significant decrease that indicated the stimulation effect
disrupted cortico-cortical network. CES also showed effect of interruption of neural
electrical oscillation status which lead to lower shift of median frequency.
Although the neurophysiological measurements reached statistical significant, but most of the studies didn’t reach significant in functional task. Transcranial pulse
current stimulation (tPCS) was another category of CES stimulation. This kind of
stimulation used unidirectional or bidirectional symmetric rectangular current, so
concept of stimulation was more related to tDCS.
One study reported the effect of tPCS on the gait pattern in Parkinson’s disease.
Significant improvements of strike length and gait velocity were noted11. However,
polarization effect induced by tPCS was less than tDCS. Abhihek in 2012 compared
different stimulation parameter 8. He indicated the biphasic current charge imbalance
current may have less effect on neuromudulation because of the symmetrically
bidirectional current.
As we know, CES stimulation can effectively influence brain stem
neurotransmitter level in animal study. Interruption of cortical connectivity also
reported in fMRI study. Recent years, integrated therapy was gradually be emphasized.
Combination of non-invasive brain stimulation with movement training would be more
efficient of training effect. Recently, study showed that combination of constraint
induce movement therapy with concurrent tDCS showed significant effect on
increasing cortical excitability of affected hemisphere and reducing excitability of
unaffected hemisphere in stroke patients12. tDCS combined robotic assisted therapy
also showed significant effect on spasticity control13.
TENS was commonly used in pain management. Few of studies reported that
cranial TENS intervention improve motivation of dementia population14. Parameter of
Transcutaneous electrical nerve stimulation (TENS) was similar as CES. The electrical
stimulation waveform was biphasic rectangular current with or without resting time
Our study investigate the effect of cranial TENS intervention on cortical excitability.
Because of feasible and easily approach of TENS, the effect of cranial TENS on
movement performance or cortical excitability are important for further clinical using.
Previous study showed increase cortical excitability after TENS intervention, but
motor learning process itself may increase cortical excitability. The relationship
between TENS intervention and motor learning process was unclear15.
1.2 Purpose
This study investigated the effect of cranial TENS on cortical excitability and implicit sequential motor learning process. Our previous study revealed the effect of
low frequency (15Hz) cranial TENS intervention. Significant increase of cortical
excitability was observed. Although significant change of cortical excitability, we
cannot clarify the excitatory effect was result of motor practice or TENS. We separated
the experiment into two groups. First group was cranial TENS stimulation group and
second group was cranial TENS combined with serial reaction time task (SRTT).
Independent measurement of cortical excitability help us compared the difference
induce by motor practice. In addition, previous study showed no significant difference
of thumb pinch accuracy task between TENS stimulation and sham stimulation. In this
experiment we choose SRTT as functional task which was common design in motor
learning studies. After repeated practice single number sequence, subjects would build
up the linkage of each number they taped. Prevention of any information about exist
sequence in the number they taped was important to ensure implicit leaning process.
We also measure cortical hemodynamic response of prefrontal cortex. Prefrontal
cortex increase activation in initial SRTT practice then decrease activation in retention
test was reported.
1.3 Question and Hypothesis
Question 1: Does cranial TENS intervention can modulate cortical excitability of
primary motor cortex?
Null hypothesis: There is no significant difference of cortical excitability between
cranial TENS trials and sham stimulation trials.
Alternating hypothesis: There are significant differences of cortical excitability
between cranial TENS trials and sham stimulation trials.
Question 2: Are there any neurophysiological differences between the subjects with
motor practice or not during cranial TENS intervention?
Null Hypothesis: After cranial TENS intervention, there are no significant differences
of cortical excitability between the subjects with motor practice or not.
Alternating Hypothesis: After cranial TENS intervention, there are significant
differences between the subject with motor practice or not.
Question 3: Is there any difference of prefrontal activity pattern during practice after
cranial TENS intervention?
Null hypothesis: cranial TENS treatment do not change activation pattern of prefrontal
cortex.
Alternating hypothesis: cranial TENS intervention significant decrease prefrontal
activation after motor practice.
Question 4: Does subjects would get significant more improvement of reaction time in
cranial TENS trial compare with sham stimulation trial in functional reaction time
task?
Null hypothesis: There is no significant different improvement of reaction time
between TENS trials and sham stimulation trials.
Alternating hypothesis: There is significant more improvement of reaction time in
cranial TENS trials compared with sham stimulation trials.
1.4 Significance
TENS were common used in pain management in Taiwan. TENS devices were
more feasible compared to tACS or tDCS in Taiwan. Cranial TENS intervention was
lack of evidence about the treatment effect. This study investigated the effect of cranial
TENS on cortical excitability of M1. Movement related disorder common change
excitability of primary motor cortex. In Parkinson’s disease, due to the impaired
dopamine pathway which lead to imbalance of facilitated circuit or inhibit circuit.in
basal ganglia16. Compensated change of increasing excitation of motor cortex was
observed. In stroke patient, the cortical excitability was unbalance between affected
hemisphere and unaffected hemisphere17. Unaffected hemisphere showed increasing
but affected hemisphere showed decreasing of cortical excitability.
The future goal is to identify the pattern of cortical excitability change after
cranial TENS and applied in neurological impaired population. For example, applied
the cranial TENS on Parkinson’s population or affected hemisphere of stroke patient to
restore balance of cortical excitability.
Chapter 2 Literature Review
2.1 Historical View of Transcranial Electrical Stimulation
Transcranial electrical stimulation(tES) had been developed for more than century
ago. The pioneer of transcranial electrical stimulation was Electrosleep (ES) and
Electroanethesia (EA). The concept of EA was applying high frequency (3500-10000
Hz) transcranial electrical stimulation to serve as substitution for chemical anesthesia.
Due to high frequency and high intensity of stimulation parameter, Electroanesthesia
was less related to our study. In 1902, Louise Robinivitch designed another type of the
transcranial electrical stimulation which was the first study of Electrosleep. The
frequency was set between 30-100Hz with biphasic rectangular current with pulse
which was more related to our experimental concept of stimulation parameter. In 1914,
Electrosleep first served as clinical use and published first clinical report at the same
time. Because of the relaxation effect, ES then be used as treatment of sleep disorder..
In 1950s, the Electrosleep reemerged to get public attention in Europe. In 1960, ES
evolved to other names such as Transcerebral electrotherapy (TCES) in 1960s,
NeuroElectric therapy (NET) in 1970s and Cranial electrotherapy (CES) in 2010. Such
kind of transcranial electrical stimulation was used as treatment for anxiety, insomnia
and depression18.
Polarization electrical stimulation was developed based on ES with DC offset.
Initial polarization electrical stimulation was TCES combined DC bias, but latter the
completely direct current developed in 1969 by Brown which stimulation parameter
was same as contemporary tDCS. Transcranial pulse current stimulation (tPCS) was
also based on polarization stimulation type. The stimulation parameter of tPCS was
using the monophasic pulse current. tPCS has significant effect on improving gait performance in Parkinson’s disease was reported in 201311.
Another contemporary type of tES was transcranial alternating stimulation (tACS).
In 2011, tACS was developed. The stimulation waveform was sinusoidal biphasic
current which was equal charge. Instead of polarization effect, tACS hypothesized that
sinusoidal wave can disrupt current brain electrical oscillation status and change the
brain oscillation pattern19. The effect of tACS on cortical excitability was still
controversial. Large clinical trial of tACS for neurological disorders still lack of
evidence. Even though, one fMRI study reported that β band frequency (15Hz)
tACS have inhibit effect on cortical abnormal activity in idiopathic cervical dystonia
patients20.
2.2 Comparison Stimulation Parameter of Transcranial Electricl Stimution
In 2013, Berkan comprehensively compared different type noninvasive brain
stimulation. Base on previous category of tES which included ES, EA, Neuroelectric
therapy (NET), CES, tACS, tDCS. The stimulation setting of Electrosleep was place
two active electrodes on bilateral frontal which ground placed on palm. Another
stimulation pattern which focus on optical nerve stimulation for relaxation effect. In
optical nerve stimulation condition, two active electrodes would place on bilateral eyes
area and ground place on mastoid process which intend to make current run through
eyes fossa. The intensity was between 0.1 to 0.5 mA.
NET was the precursor of CES. The electrodes setting shift from bilateral scalp in
Electrosleep to bilateral ears which current was more focus on brain stem stimulation
and tried to influence the pathway in brain stem including serotonergic and
noradrenergic systems21. Electrodes number decrease from 3 electrodes to 2 electrodes
that ground electrode was excluded. The intensity was smaller than CES which was
about 600μA. Stimulation frequency was larger range (0.5- 100Hz) which was same as
CES. In NET, electrodes placed on bilateral forehead which concept was not focus on
brain stem stimulation. CES stimulation generated fixed electromotive in 5V with
pulse current.
In Electroanesthesia(EA), four electrodes were placed on bilateral frontal and
occipital area. EA was more intensive type of electrical stimulation. Stimulation was
high which was about 40 mA with continue DC current. Another type of EA used
biphasic current, but the stimulation intensity was related lower ( about 10 mA) with
frequency near 10000 to 20000 Hz.
In tDCS, the electrode placement was changed from brainstem stimulation
concept to specific brain area stimulation. In M1 stimulation condition, the active
electrode was placed on hotspot of M1 defined by tanscranial magnetic stimulation
(TMS) mapping technique or placed on C3 and C4 in 10-20 EEG electrode system.
The passive electrode was placed on contralateral supraorbital area which current
flowed into unilateral motor cortex and premotor cortex. Stimulation intensity was set
at 1-2mA. Electrode placement of tACS was also similar as tDCS. Depending on
stimulation the target brain area, the tACS can alter subject’s visual, cognitive, motor,
sensory function. Previous montage study also show that the low frequency AC current
was more efficient to induce electrical field change in cortex than DC current22-24. The
electrical field induced in cortex by AC current was five times more than DC current in
realistic head model which designed based on different tissue conductivity22. In our
experiment, we combined the CES stimulation with contemporary setting of electrodes.
We use TENS as intervention tools to induce neurotransmitter change and further
change cortical excitability. The current generated by TensMed 931 was symmetric
biphasic current with pulse. The waveform of our stimulation tool was more close to
the concept of CES, NET and TCES. The electrodes were place on M1 and
contralateral supraorbital area.
2.3 Cranial Electrotherapy Stimulation
Cranial electrotherapy stimulation (CES) was developed from Electrsleep which
mechanism was based on peripheral nerve stimulation to induce neurotransmitter level change central nervous system. Instead of using “transcranial” as term of description,
CES used “cranial” as its term which emphasize the current not flow through the scalp1.
The peripheral stimulation mechanism focuses on the stimulus transmit into CNS with
change of antinociception reaction. The pathway of the stimulus may go from cranial
nerve I-III and VIII to activate of brainstem center. Some of the CES emphasize the
electrode need to place on bilateral eyes to induce peripheral pathway to influence
central nervous system.
Second potential mechanism was CES may induce release of neurotransmitter.
CES also have good effects on biological clock of brain which controlled in
hypothalamic suprachiasmatic nucleus (SCN) in brain stem21. The serotonin level and
noradrenaline level was also increase after CES intervention. Base on previous
physiological review, the serotonergic system was more response to stimulation with
10, 20 and 100Hz. TCET was one type of CES which was asymmetric biphasic current
with equal charge. When TCET applied on rat, the dopamine and norepinephrine level
were significant increase. The stimulation seems to induce synthesis of
neurotransmitter in midbrain or hypothalamus. No neurochemical response was found
in hindbrain neurotransmitter synthesis center which indicate the neurotransmitter
synthesis response was localized not whole brain.9
Third potential mechanism was alternating current may interruption or
disturbance ongoing cortical activity. Previous study also showed CES can alter EEG which showed lower shift median frequency of α wave. The frequency of CES was set
at 0.5Hz and 100Hz. higher frequency has more obvious effect on interruption of α
wave. Reduction median frequency of α wave was associated with more relaxation
status10. α wave was also related to awake and sleep status change. Significant desynchronizing of α wave was found in sleep status25.
2.3.1 Research in Animal Model
Some animal studies reveal how functional or structural change of the neural
tissue after stimulated by alternating current. In hippocampus cell, high frequency
electrical stimulation would induce long tern potentiation (LTP) which was long last
change of synaptic efficiency. Low frequency electrical stimulation do not induce LTP
liked response or even resulted in long term depression. Recent study overthrows this
concept and indicated that low frequency electrical stimulation (1Hz) would also
produce LTP response with specific afferent system. The 0.5 Hz alternating pulse
current induce CA1 LTP like response. However, low frequency stimulation on CA3
was ineffective26.
Whether the AC current actually can influence neuron is controversial especially
when charge was balance. Transcranial electrostimulation treatment (TCET) was
biphasic current with pulse which charge was balance. In the hypothalamic level of the
rat, TCET significantly enhance dopamine and norepinephrine synthesis. In mid-brain
of the rat, the serotonin and dopamine significant enhance synthesis. Although the
dopamine and norepinephrine synthesis increase, the turned over rate not change which
detected by measured the metabolite of neurotransmitter.
2.3.2 Research in Human Model
CES commonly used as treatment of depression, anxiety, insomnia, pain, migrant
headache, pain of fibromyalgia and sleep-awake cycle disorder. Few of clinical trials
focus on the rehabilitation effect of CES on movement disorder. Most of experiments
of CES were focus on cognitive, biological clock, psychological problems. Malden’s
study in 1985 indicated that the CES can enhance occupation therapy effect on gross
motor performance in severe cerebral palsy children. Actually, the evidence level of
motor function related study was not strong. CES also facilitates sensory motor
integration in children. In Okoyey’s in 1986, the motor accuracy and hand function
significantly improved in minimal cerebral palsy children27.
CES was also effective for pain relief. Most of the studies focus on how
neurotransmitter of nociception system affected by CES. In another point of view that
patients with chronic degenerative joint pain syndrome show abnormal peak and
unsmooth pattern in EEG spectrum curve in bilateral frontal area. In contrast, normal
healthy subject show relative more smooth of frequency spectrum curve than chronic
degenerative joint pain patient. Both patient with pain and healthy subject received 20
mins of microcurrent stimulation with 0.5 Hz biphasic rectangular pulse current. The
electrode placed on bilateral trapezius or earlobe. The electrode placed on trapezius
muscle which was trying to induce neurotransmitter level change through cervical
nerve. After CES intervention, the unsmooth pattern of EEG spectrum significant
improved which was near normal spectral curve. Furthermore, pain score of patient
also significant decrease28,29. Though the cortical electrical oscillation status change
after CES intervention, the effect still believed derived from neurotransmitter modulate
effect in subcortical area.
Another study compared the effects of high frequency CES and low frequency on
EEG spectral curve. Low frequency CES (0.5Hz) showed significantly downshift of mean frequency of α band. High frequency CES (100Hz) showed significant downshift
of mean and median α band frequency. α band frequency downshift was more related
to relaxation status of subject which indicate the CES facilitation relaxation state. After 100Hz CES intervention the power fraction of β band frequency significant decrease. β
band frequency was more related to stress, arousal, problem solving state. It seems that
higher frequency of CES was more effective in brain oscillation modulation.30 Another
fMRI study reveal that CES stimulation disrupted cortico-cortical network such as
connectivity between frontal lobe and parietal lobe10. General hemodynamic response
of specific task was less in CES stimulation condition. Based on these findings,
changes pattern of power spectral curve after CES was suspected the results of changes
of neurotransmitter level in subcortical area.
2.4 Transcranial Magnetic Stimulation
In 1838 Micheal Faraday discovered the phenomenon of the area near electrical
current would induce magnetic field change. Technique of transcranial magnetic
stimulation (TMS) was based the Faraday’s law of magnetic field. Based on concept of
magnetic field drive electrical current, the initial machine of transcranial magnetic
stimulation was developed by Barker in 1985.31-33 Using magnetic field pass through
the neuron membrane induced depolarization of motor cortex was more painless than
tES. The direction of electrical field induced by TMS was penetrated perpendicularly
to the coil. The current cause by TMS would parallel to the coil. Thus, TMS derived
current was more focus on the cortex. By contrast, because of anode and cathode
placement, the electrical field direction induced by tES would more focus on
subcortical area33. The penetrated magnetic field generated electrical current on motor
cortex which induce depolarization of the pyramidal neuron. The initial directly
descending action potential was defined as “Direct Wave” which also called D-wave.
Then follow late waves with interval 1.2-2.0 ms after D-wave were “Indirect Wave”
which also called I-wave31,34. I-wave was more related to polysynaptic or synaptic
recurrent network which represent excitability of motor neuron pool in cortical level or
spinal level. The descending several I wave reach neuromuscular junction with
temporal and spatial summation which reach threshold and leaded to muscle
contraction which was called motor evoked potential (MEP). MEP amplitude naturally
fluctuated over time and affected by the integrity and excitability of cortico-spinal
neuron. With voluntary contraction, the size of MEP increased which represented the
higher level of excitability corticospinal motor neuron pool.
Safety issue of TMS had been discussed for several years. Metal implantation in
brain or target stimulation area may potentially displaced or damaged under current
generate through magnetic field including mental clips, deep brain stimulator, pace
maker and cochlear implants. External body mental objects also need to be removed
for any possible interaction such as glasses, watch and necklace. Some of side effect of
TMS had been reported previously. Increase in auditory threshold due to the high
pressure level sound derived from coil. In case of side effect of transient heavy hearing,
earplug was suggested for auditory blocked. Another side effect may occur was seizure.
Due to the electrical current generated in gray matter would induce imbalance of
excitatory circuits and inhibitory circuit in some people which may leads to seizure
when received TMS stimulation. The population probably induced seizure included
subjects with multiple sclerosis, bipolar syndrome, major depression and have family
history of seizure. Other side effect may exist such as, vascular syncope, increase heart
rate, increase blood pressure, psychiatric change, interaction effect on drug of
neuro-system.34 Pregnancy should prevent from the stimulator coil more than 0.7 meter
for the purpose of that fetus may be affected by the magnetic field.35
2.4.1 Motor threshold
The definition of motor threshold (MT) or cortical motor threshold (CMT) was
the lowest TMS stimulation intensity applied on motor cortex to induce muscle
contraction of muscle which was defined by Rossini et al in 199436. Motor threshold
represents as the excitation status of pyramidal neuron and spinal neuron. Due to the
fluctuation status of excitability of motor neuron, estimation of motor threshold needs
validated method. The MT was defined as percentage of simulation intensity which
with 50% success rate (5 times out of 10 times) to induce MEP more than 50μV peak
to peak amplitude. Average 75 pulses needed to deliver to confirm the motor threshold
by Rossini’s method but relative shortness compare to other method37. Due to lack efficiency of Rossini’s method, modified Rossini method was developed which use 3
times out of 6 times method was considered. However, No validity or reliability report
of modified Rossini method was delivered.
2.4.2 Motor evoked potential
I wave with spatial and temporal summation would induce peripheral muscle
contraction, the signal received from electromyogram called “Motor evoked potential”.
Motor evoked potential not only reflect integrity of cortical spinal neuron, also
represent excitability of corticospinal neuron. The higher intensity TMS stimulation,
the more large size of motor evoked potential induced. While muscle slightly
contraction status, the same intensity of magnetic field would lead to larger size of
MEP than without muscle contraction. In slightly contraction status, the corticospinal
neuron would increase excitability that cause the size of MEP was increase. Single
pulse MEP also can be used as mapping technique to defined functional distribution of
motor cortex. To monitor the change of cortical excitability of target muscle with specific stimulation point called “hotspot”.
Motor learning was a process which included neuroplasticity which was
functional change or structure change of neural system. Motor learning also induce
long-term potentiation in motor cortex which cause increase of amplitude of MEP after
learning process.38,39
2.4.3 Intracortical facilitation and intracortical inhibition
Intracortcial faciulitation(ICF) and intracortical inhibition(ICF) was related the
techniques of pair pulse stimulation. Combination of different interval of sub-threshold
stimulation following supra-threshold stimulation would induce inhibition or
facilitation effect on motor cortex33. The first sub threshold stimulation was conditional
stimulus and second stimulation was testing stimulus. With lower interval between 4-7
ms belong to ICI which represent excitation of inhibitory circuits and have inhibitory
effect on cortical excitaiblity. Di Lazzaro et al. in 2000 revealed that ICI was more
related to a-GABAergic pathway which was the primary inhibitory neurotransmitters
in brain40. Excitatory circuitry can be evaluated effect in interstimulation interval
between 7-20 ms which called intracortical facilitation. However, mechanism of
intracortical facilitation was unclear. With intake of antagonist of
N-methyl-D-aspartate (NMDA), ICF was significant decrease41. The suppressive effect
of NMDA antagonist supported that the ICF was represent the glutamatergic
transmission. Long-term potentiation which involving in increasing calcium ions level
of dendritic spine was involved in NMDA pathway.42 Due to the different mechanism
of ICI and ICF, Some of the authors indicated that the circuitries involved may be
independent.42
2.5 Near-Infrared Spectroscopy
Near-infared spectroscopy was used to measurement of blood flow of different
tissue. When the blood flow increase, concentration of oxygenated hemoglobin [O2Hb]
would increase, the concentration deoxygenated hemoglobin [HHb] would decrease
and total hemoglobin would increase. With such phenomenon which called
hemodynamic response can be applied on different tissue such as cerebral cortex,
muscle belly and tendon. Near Infrared was mainly absorbed by melanin, so the hairy
skin should prevent in measurement. Other tissue such as scalp, bone, cerebrospinal
fluid would absorb infrared but related stable than cortex. Cortex would change blood
flow by the time due to the metabolic rate increase of local area.
2.5.1 Mechanism of NIRS
The early experiment of NIRS was done by Jobsis in 1997 who found myocardiac
and cortex had high penetration rate of near infrared.43 Until 1980s, the first study of
applied NIRS on human cortex to detected hemodynamic response was published. The
near infrared can partial penetrated through tissue and partial reflex or absorbed by the
tissue. Different kinds of molecules have different absorption spectra. For example,
H2O was highly be absorbed from 1050 nm to more than 1300 nm of wavelength. To
detect the hemodynamic response of cortex focus on concentration change of
oxygenated hemoglobin (∆[O2Hb]) and deoxygenated hemoglobin (∆[HHb])
between different events. HHb was high in absorption spectra when applied near
infrared’s wavelength between 600nm to 1000 nm. O2Hb was high absorption spectra
when applied near infrared’s wavelength between 700 nm to 1150 nm. 810 nm was
equal absorption spectra of O2Hb and HHb The NIRS device often project two
wavelength above and below 810 nm to calculus ∆[O2Hb] and ∆[HHb] by
mathematics method of first degree polynomial in two variable.44
The formula show how to calculate level of ∆[O2Hb] and ∆[HHb] through optical
density difference (∆OD). ∆OD is change of optical density from initial to final. IFinal
and IInitial are the measured density in initial and final. ∆C is the change of the concentration. L is distance from light source to detector. ε is the extinction
Coefficient. B is differential pathlength factor which may be influenced by age effect.
To calculus concentration change of ∆[O2Hb] and ∆[HHb], the formula also can
rewrite into below:
λ is particular wavelength. To measure the concentration change of ∆[O2Hb] and
∆[HHb] by this equation which need more than two wavelength light to determine the
concentration of hemoglobin.
Neuro-activation would increase metabolic rate of neural tissue which was
coupled with hemodynamic response. Increase metabolic demanding which drives
vascular response to provide more oxygen to neuron. When increase blood perfusion of
local neuron, [O2Hb] will increase and [HHb] will decrease. With change of
hemoglobin concentration level which indicate neural activation status. Total
hemoglobin concentration [tHb] also common measured in NIRS device which was
sum of ∆[O2Hb] and ∆[HHb]. [tHb] can be serve as an index of blood perfusion of
neural tissue.
The shape of infrared light penetrated area with perpendicular projection was
described as banana shape or ellipsoid path. Penetration depth of infrared was about
2-3 cm.45 Whether infrared can flow into cortex was depending on scalp cortical
distance (SCD). Target tissue to estimate hemodynamic response as index of activation
was focus on gray matter. By mathematic model of ellipsoid, Depends on different
SCD, the amount of gray matter volume which penetrated by infrared can be predict.
With increase of SCD, measured the gray matter volume would decrease. Such as area
near central sulcus where SCD would be too long to measure the cortical
hemodynamic response.45 For example, the area control lower extremity in primary
motor cortex was located beneath central sulcus was not suitable for NIRS
measurement. In this study, we measure activation of prefrontal cortex where SCD was
related less which was feasible for NIRS measurement.
2.5.2 Psychometric studies of NIRS
Hemodynamic response of prefrontal cortex during motor practice was one of
major outcome of this study. Reliability and validity of NIRS is important to explain
outcome between trails variability. Gold standard of hemodynamic response of neural
tissue was fMRI. Several psychometric studies have been published already. The
limitation of fMRI was poor in temporal resolution and suitable task in restrict space
when scanning was not functional. In contrast, NIRS is portable and high temporal
resolution. Limitation of NIRS was poor spatial resolution and movement artifact
signal may occur. Strangman in 2006 compare reliability of NIRS in different ways of
data analysis. 19 subjecs were recruited to evaluation cortical hemodynamic response
during complex thumb opposition task. Each trail consist of 16 sec moving time with
16 sec rest. Peak and trough of ∆[O2Hb] and ∆[HHb] were observed. Trial to trial
reliability showd morderate correlated while Peason’s correlation coeeficient was 0.33.
Pairwise comparison of trials show first trial and 16th trial was most poor correlated.
However, first trial was more correlated with second trial. The sesults indicated the
longer interval between events the less reliabiltiy.
Blocke mean analysis of data showed imrpove measurement reliability. for
example, average of 4 trials had better reliability than average with 2 trials. Based on
two example above showed time effect on reliability of NIRS was exist43.
Aging effect on decreasing hemodynamic response also be reported in 2002 by
Mehagnoul-Schipper. He compare hemodynamic response of young population with
elderly. Control event to measured activation was frequent finger tapping task.
Subjects need to tap as fast as possible within time. Significant hemodynamic response
was observed not only in young but also in elder, but significant lower response of
∆[O2Hb], ∆[HHb] and ∆[tHb] in elder population was observed. Based on the results,
responsibility of NIRS was lower in elder population which may due to aging effect on
poor vascular response.46
Validation study of NIRS common compared the blood oxygenate level
dependent(BOLD) in fMRI with ∆[O2Hb], ∆[HHb] and ∆[tHb]. MRI compatible
NIRS device was used to concurrently measure hemodynamic response during specific
event. Okamoto in 2004 compare correlation between ∆BOLD and ∆[O2Hb],
∆[HHb] ,∆[tHb] during subjects doing apple peeling task. In multiple channel NIRS
mapping technique found increase activation in M1, supplementary motor area (SMA)
and premotor cortex. The activation area detected by NIRS was similar as fMRI.
Compare ∆[O2Hb] with ∆BOLD by pearson’s correlation show poor but significant
correlated. (r= 0.2, P<0.05) Similar result also found when compared ∆[HHb] with
∆BOLD (r= - 0.19, P<0.05)47. Strangman et al in 2002 also compared NIRS and fMRI.
However, different result found when using reciprocal ∆BOLD compared with
∆[O2Hb], ∆[HHb] and ∆[tHb]. High correlation was found in ∆[O2Hb] with
1/∆BOLD and ∆[tHb] with 1/∆BOLD, r value were between 0.8 to 0.9. Compared
∆[HHb] with 1/∆BOLD showed poor to moderate correlation which r value were
between 0.14 to 0.58.44 The results support the relationship between concentration of Hb and ∆BOLD was non-linear.
2.6 Serial reaction time task
2.6.1 Implicit sequential motor learning process
Serial reaction time task(SRTT) had been developed by Nissen in 198648. SRTT
was used as tool to evaluation temporal organization of behavior by psychologist.
Subject need to recognize four visual cues showed on screen independently with 12
words sequence. Each visual cue can find correspnding key on the keyboard. Number
1234 were common used as four visual stimulation. Subjects need to tap correspond
key as fast as possible. Any information of exist seuquence in the number series they
taped was prevented. Follwing several trials of practice was random number trial
which indicated ability of trasfer skill to other condition. In random number trail, the
expectation of next number from previous number was violate to aquired sequence.
Subject can built up connection of each number by repeated practice. The process
was without awareness of the sequence that make sure learning type was implicit
learning. To make sure the implicit learning process not turn into explicit learning
pattern through recall test. Recall test often done in end of experiment. Subjects were ask “Whether any sequence exist in the number you taped ?”. If subjects answer that
the numbers were seuqencial, they were ask to recall the memory about the sequence
they tape.49
2.6.2 Brain Activiation pattern during SRTT
Berns in 1997 revealed the brain region where response for the novelty task with
awareness. Based on SRTT task, positron emission tomography (PET) was used as
monitoring brain functional activity tool. When subjects aquired new sequence,
increase activation found in isplateral premotor cortex, isplateral anterior cingulate
cortex and contralaeral venral straitum. Contralateral cerebellum and isplateral
prenmotor cortex were more responsible for novelty of motor task which constant
increas activation through all practice session. Dorsal lateral prefrontal cortex (DLPFC)
was more responsible for the sequencial memory mentainence which showed decrease
activation when switch from aquired sequence to novelty sequence and constant
increase activation in repeated practice.50
Sequenctial motor memory was affected by several neural curcuits not only
cortico-cortical curcuits but also subcortico-cortical curcuits. Activity of SMA was
decrease after repeated TMS applied on M1 to interfered motor memory modification.
Interfered motor memory also weaker correlation of activation between SMA, M1,
cerebellium, anterior cyngulate cortex and straitum.51
Contextual interference of motor task also affected activation level of DLPFC,
SMA and M1 in fMRI. Compare with repeatitive sequence(1112, 2223, 3334) or
interleaved seuqence (2134, 4312, 1423) practice in SRTT, interleaved sequence
(difficult sequence) showed poor performance initially than repeatitive sequence(easy
sequence). Related increase activation in more difficult condition were bilateral
occipital lobe, temporal cortex, sensorimotor and premotor areas, premotor area,
inderior and medial prefrontal area and medial temporal area.52 In rentention test after
practice session, interleaved sequecne seuqence showed significant better performance
in reaction time. Significant increase M1 excitabiltiy was found in interleaced prctice
which showed decrease of SICI and increase of ICF and control MEP. The neural
curcuit shifted to more excitatory status with more difficult condition.53 Excitation
changed of M1 indicated more efficient of motor memory retrieve after preactice
session. Region of increasing cerabral blood flow finally decreased in retention test
were isplateral prefrontal, premorot cortex and inferior frontal areas compared to
practice session52. Prefrontal cortex was more related to declarative memory system
which may interaction with procedural memory system. Inhibit dorsal lateral prefrontal
cortex (DLPFC) which would induce consolidation of procedural memory54. It seems
that declarative memory reduce or even complete inhibit procedural memory. Decrease
activation DLPFC, supplementary motor (SMA), medial frontal indicates more
efficient of motor memory retrieval52. In addition to neurophysiological change,
reaction time were significant improve compared to initial condition.
Chapter 3 Methods
3.1 Participants
This study was cross-over single blind design. Subject did not know which kind if
intervention they received. Participant were randomized into two group, stimulation
with motor execution group and isolately received stimulation group. Subject needed
to acomplish two trials of experiment which were true stimulation trial and sham
stimulation trial. Subjects were recruited from university or community.
Inclusion criteria of participants: (1) age between 20 to 40.
Exclusion criteria: (1) Psychological disorder; (2) Have history or family history of
epilepsy; (3) Have head trauma history, received brain surgery ,brain tumor, stroke and
head metal implant; (4) Implant of pacemaker or electrical stimuator; (5) Vascular
syncope or unknown reason syncope; (6) Intermittent headache; (7) Taking drug
related to cognitive or emotional status. (8) Poor skin status.
Volunteers fulfilled criteria of above would be enrolled into this study. All subjects
should sign on consent which was approved by Chang Gung Medical Foundation
Institutional Review Board (see appendix). Subjects were allowed to reject further
investigation without any reason. The experiment would immediately stop when
uncomfortable status appeared during TENS intervention or receiving TMS such as
headache, burn pain of skin, dizziness.
Sample size was estimated based on Liao’s study15. The effect size was 0.72
calculated by treatment effect of TENS on MEP. Alpha level set at 0.05 and power was
set at 80%. Sample size was 24 subjects for each groups which was estimated by
G*Power 3.1.3. Total number of subjects need to enroll was 48.
3.2 Study Design
This study was randomized cross-over design. The subjects were randomized into
TENS stimulation group and TENS stimulation with SRTT group. Participants needed
to accomplish two trials of experiments which include TENS stimulation and sham
stimulation with randomized order. Four conditions need to comparison included (1)
Motor practice with TENS stimulation (Motor-TENS); (2) Motor practice with sham
stimulation (Motor-Sham); (3) TENS stimulation (Stimulus-TENS) and (4) Sham
stimulation (Stimulus-Sham). Wash-out period must to be more than 1 week (figure 1).
Single blinded was designed in this study.
3.3 Experimental procedure
Basic data include age, gender, medical history, sleeping time were collected.
Subjects were randomized into two groups. Permute block randomization was
performed. Block number was four which include four conditions. Before start TENS
or sham stimulation, cortical excitability and prefrontal activation in SRTT were
measured in baseline. Subject need to tap 7 blocks of SRTT concurrent measured the
activation of prefrontal cortex after TENS applied on scalp for 5 minutes. After TENS
intervention, Immediate TENS effect on cortical excitability was measured. Cortical
excitability and prefrontal activation in SRTT was also measured in follow up test at 30
min and 60 min (Figure 2.).
During TMS assessment, subjects sit on comfortable seat with armrest to prevent
any movement artifact in TMS assessment. Hand held circular coil was place on head
which generated current in anterior-posterior direction on cortex. The TMS device used
in this experiment was MagStim 200 stimulator. Surface electromyogram (EMG) of
first dorsal interosseous was assessed by active electrode placed on muscle belly. To
ensure no deviation of coil during assessment, dermatograph was use to mark on the
hotspot.
In NIRS measurement, The PortaLite produced by Artinis Medical System was
use. Detection optodes was placed on FP1 or FP2 in 10-20 EEG electrode system. FP1
and Fp2 were area approximately 20 to 30 mm above midpoint of eyebrow. The
emission optodes were laterally placed which approximately F7 or F8. The average
photon path from detection to emission cover right superior and middle frontal gyrus
which mainly cover Brodmann’s area 1055,56. The detection optode and emission
optode was covered by goggles with low opacity. Before measuring the concentration
of hemoglobin, light leakage was test to ensure no light pass through goggles.
To perform SRTT, subjects need to place right hand on keyboard with finger
correspond to key one the key board. Index finger was corresponding to 1; middle
finger was 2; ring finger was 3; little finger was 4. A screen was place in front of
subject with distance about 1 meter. Head position was with slight flex when looking at
screen. Visual cue was at central of screen in identifiable size.
The TENS current was generated by “Enraf-Nonius Muscle stimulator, TensMed
931”. The current waveform was biphasic rectangular current with pulse duration was
200 μs. Current was deliver through pair of rubber electrodes (6X8 cm2) placed on
Hotspot and contralateral supraorbital area. Elastic bandages were used to make
electrode well contact on skin. Intensity was 2 mA which was below sensory threshold
stimulation. Frequency was set at 15Hz based on previous experiment15 The TENS device was controlled behind subject, so the subjects doesn’t know the device was on
or not. Any uncomfortable feeling reported by subject or adverse effect such as burn
pain of skin, would be recorded.
3.4 Experimental Assessments
Basic data were initially retrieved from subjects included age, gender, current or
previous medical history.
TMS stimulations were generated by MagStim 200. To locate hotspot of primary
motor cortex respond to FDI muscle, 40 to 50% of TMS intensity was used. The area
where induce most high amplitude of MEP was defined as hotspot. After locate hotspot,
TMS intensity was decrease in 2% gradient. MT was defined as the smallest intensity which can induce MEPs amplitude more than 50μV in 50% success rate (5 out of 10).
Single pulse MEP amplitude which represents cortical excitability was measured at
120% intensity. Pair-pulse stimulation was used to assess intracortical inhibition(ICI)
and intracortical facilitation(ICF). Condition stimulation’s intensity was set at 70% of MT which was subthreshold level. Following testing stimulation’s intensity was set at
120% of MT which was suprathreshold. To assess ICI in this study, inter-stimulation
interval was set at 2 and 3ms. To assess ICF, inter-stimulation interval was set at 7, 10,
15 ms. Due to the excitatory or inhibitory effect of pair-pulse stimulation, order of 2
ms, 3 ms, 7 ms, 10 ms and 15ms would be randomized. All TMS assessment was
measured at baseline, immediate after TENS stimulation and follow up 30, 60 min.
Motor performance at baseline was assessed by SRTT program generated in Matlab
8.0 version. Three sequence consist of 12 words was used in this study
