Combined Transcranial Magnetic Stimulation and Electroencephalography of the Dorsolateral Prefrontal Cortex

Podsumowanie

The protocol presented here is for TMS-EEG studies utilizing intracortical excitability test-retest design paradigms. The intent of the protocol is to produce reliable and reproducible cortical excitability measures for assessing neurophysiological functioning related to therapeutic interventions in the treatment of neuropsychiatric diseases such as major depression.

Streszczenie

Transcranial magnetic stimulation (TMS) is a non-invasive method that produces neural excitation in the cortex by means of brief, time-varying magnetic field pulses. The initiation of cortical activation or its modulation depends on the background activation of the neurons of the cortical region activated, the characteristics of the coil, its position and its orientation with respect to the head. TMS combined with simultaneous electrocephalography (EEG) and neuronavigation (nTMS-EEG) allows for the assessment of cortico-cortical excitability and connectivity in almost all cortical areas in a reproducible manner. This advance makes nTMS-EEG a powerful tool that can accurately assess brain dynamics and neurophysiology in test-retest paradigms that are required for clinical trials. Limitations of this method include artifacts that cover the initial brain reactivity to stimulation. Thus, the process of removing artifacts may also extract valuable information. Moreover, the optimal parameters for dorsolateral prefrontal (DLPFC) stimulation are not fully known and current protocols utilize variations from the motor cortex (M1) stimulation paradigms. However, evolving nTMS-EEG designs hope to address these issues. The protocol presented here introduces some standard practices for assessing neurophysiological functioning from stimulation to the DLPFC that can be applied in patients with treatment resistant psychiatric disorders that receive treatment such as transcranial direct current stimulation (tDCS), repetitive transcranial magnetic stimulation (rTMS), magnetic seizure therapy (MST) or electroconvulsive therapy (ECT).

Wprowadzenie

Transcranial magnetic stimulation (TMS) is a neurophysiological tool that allows for the non-invasive assessment of cortical neuronal activity through the use of rapid, time-varying magnetic field pulses¹. These magnetic field pulses induce a weak current in the superficial cortex beneath the coil which results in membrane depolarization. The ensuing cortical activation or modulation is directly related to the characteristics of the coil, its angle and orientation to the skull². The waveform of the pulse discharged from the coil and the underlying state of the neurons also influence the resultant cortical activation³.

TMS enables the assessment of cortical functions by evoking behavioral or motor responses or through the interruption of task-related processing. The excitability of cortico-spinal processes can be evaluated through recording electromyographic (EMG) responses elicited from single TMS pulses over the motor cortex, whereas intracortical excitatory (intracortical facilitation; ICF) and inhibitory mechanisms (short and long intracortical inhibition; SICI and LICI) can be probed with paired-pulse TMS. Repetitive TMS can disturb various cognitive processes, but is primarily used as a therapeutic tool for a variety of neuropsychiatric disorders. Furthermore, the combination of TMS with simultaneous electroencephalography (TMS-EEG) can be used to assess cortico-cortical excitability and connectivity⁴. Finally, if the administration of TMS is delivered with neuronavigation (nTMS), it will allow for precise test-retest paradigms since the exact site of the stimulation can be recorded. Most of the cortical mantle can be targeted and stimulated (including those areas that do not produce measurable physical or behavioral responses) thus the cortex can be functionally mapped.

The EEG signal evoked from single or paired pulse TMS can facilitate the assessment of cortico-cortical connectivity⁵ and the current state of the brain. The TMS-induced electric current results in action potentials that can activate synapses. The distribution of the postsynaptic currents can be recorded through EEG⁶. The EEG signal can be used to quantify and locate synaptic current distributions through dipole modelling⁷ or minimum-norm estimation⁸, when multichannel EEG is employed, and with the conductivity structure of the head accounted for. Combined TMS-EEG can be employed to study cortical inhibitory processes⁹, oscillations¹⁰, cortico-cortical¹¹ and interhemispheric interactions¹², and cortical plasticity¹³. Most importantly, TMS-EEG can probe excitability changes during cognitive or motor tasks with good test-retest reliability^14,15. Importantly, TMS-EEG has the potential to determine neurophysiological signals that may serve as the predictors of response to therapeutic interventions (rTMS or pharmacological effects) in test-retest designs^16,17.

The principles of neuronavigation for TMS is based on the principles of frameless stereotaxy. The systems use an optical tracking system¹⁸ that employs a light-emitting camera that communicates with light-reflecting optical elements attached to both the head (via a reference tracker) and the TMS coil. Neuronavigation allows for coil localization on the 3-D MRI model with the aid of a digitizing reference tool or pen. The use of neuronavigation facilitates the capture of the coil orientation, location and alignment to the subject's head as well as the digitization of the EEG electrode positions. These features are essential for test-retest design experiments and for accurate stimulation of a specified location within dorsolateral prefrontal cortex.

In order to utilize a TMS-EEG protocol in a test-retest experiment, there needs to be accurate targeting and consistent stimulation of the cortical region to obtain reliable signals. TMS-EEG recording can be vulnerable to different artifacts. The TMS induced artifact on the EEG electrodes can be filtered with amplifiers that can recover after a delay^19,20 or with amplifiers that cannot be saturated²¹. However, other types of artifact generated by eye movements or blinks, cranial muscle activation in proximity to the EEG electrodes, random electrode movement and their polarization, and by the coil click or somatic sensation must be taken into consideration. Careful subject preparation that ensures electrode impedances below 5 kΩ, immobilization of the coil over the electrodes and a foam between coil and electrodes to reduce vibration (or a spacer to eliminate low frequency artifacts²²), earplugs and even auditory masking should be used to minimize these artifacts²³. The protocol presented here introduces a standard process for assessing neurophysiological functioning when the stimulation is applied over the dorsolateral prefrontal (DLPFC). The focus is on common paired-pulse paradigms that have been validated in the studies of M1^9,15,16.

Protokół

All the experimental procedures presented here have been approved by our Local Ethical Committee following guidelines of the Declaration of Helsinki.

1. Head Registration for Neuronavigated TMS — EEG

1. Obtain a high resolution whole head T1-weighted structural MRI for each participant. Scan according to the neuronavigation manufacturer guidelines.
2. Upload the images on the navigation system. Check if MRIs are correctly scanned. Choose the cardinal points (pre-auricular points, the nasion and the tip of the nose). Insert the stimulation targets (based on anatomy or based on the head coordinates, MNI, or Talairach coordinates).
3. Place the head tracker in such a way so that it will not move during the stimulation session and allow free moving of the TMS coil. Have the participant insert the earplugs before the registration starts.
4. Align the participant's head to the 3-D MRI model. Touch on the participant's head with the digitizing pen at the cardinal points that were selected on the images of the MRI stack. Select and mark additional points over the parietal, temporal and occipital areas of the head to reduce the registration error over those areas.
5. Validate the registration. Place the digitizing pen on the participant's head. Check its representation on the computer. If it is not on the corresponding point in the MR, repeat step 1.4.
6. Calibrate the TMS coil in use (in some systems this step is not needed).
   1. Attach the trackers to the coil.
   2. Place the coil on the calibration block so all trackers are visible from the camera.
   3. Press the calibration button on the computer screen and keep the coil in the calibration position for 5 s.

2. TMS-EEG Experiment

1. Place the EEG cap on the head and prepare the electrodes
   1. Choose a cap that fits the head well. Ensure that all electrodes are tightly touching the scalp and are functional. If more than 2 electrodes do not work, then use another cap of the same or smaller size.
   2. Place the Cz electrode at vertex, half way between the line connecting the nasion and inion and the Iz electrode over the inion.
      ​NOTE: Place the vertical (above and under the eye contralateral to the stimulation eye) and/or horizontal electrodes (left from the left eye and right from the right, a little above each zygomatic bone) for electrooculography (EOG).
   3. Adjust the blunt tip of the syringe and fill it with electroconductive gel. Place the tip inside the hole of the electrode, and then lightly press the plunger flange until there is some paste on the skin. Scrub the scalp lightly using cross-like moves with the blunt tip. Ensure that the paste is not spilling out over the top to avoid bridging (short-circuit between the electrodes).
2. Place the EMG electrodes. Place two disposable disc electrodes (diameter of about 30 mm) over the right abductor pollicis brevis muscle (APB) for a belly tendon montage. Place the ground according to the manufacturer's guidelines.
3. Start the head registration. Follow steps 1.3–1.6. Use the MNI or Talairach coordinates of the DLPFC.
4. Hot spot and motor threshold.
   1. Add a sponge (artificial fiber made from polyutherane) under the coil in order to minimize the coil vibration over the electrodes during the TMS pulses. Note that the foam should be about 10 mm thick.
   2. Instruct the participant to be at rest — comfortable and with relaxed hands, legs and spinal column.
   3. Find the hot spot. Target the motor knob²⁴ as the initial landmark of cortical representation of APB in M1 and move the coil until there is corresponding APB movement. Use TMS intensities evoking MEPs of around 500 µV over APB. Optimize the coil orientation by changing its angle and tilt to evoke the biggest response over the hot spot.
   4. Save the coil positioning in the neuronavigator software and reduce the output intensity in steps of 2–3%. Give 10 pulses and if more than 5 out of 10 MEP responses over 50 µV are obtained, then continue reducing the intensity.
   5. When less than 5 out of 10 responses are evoked, increase the intensity by steps of 1–2%. MT is represented as the intensity that produces MEPs larger than 50 µV 5 out of 10 times²⁵. The inter-stimulus interval (ISI) for MT should be longer than 1 s, usually set at 3, 4 or 5 s.
5. Adjust the intensity using the following steps:
   1. Start at 120% of MT intensity to produce MEPs over M1 from 500 to 1,500 µV. Record 10 pulses with this stimulator's output so the average response is 1 mV. Increase or decrease the intensity in steps of 1–2% until reaching an average of 1 mV.
   2. For stimulation intensity, choose the intensity as a percentage of stimulator's output, e.g., 110%, 120%, etc.
   3. Find the corresponding induced field in V/m (if the system allows). Place the coil over DLPFC; adjust the stimulator's output until the calculation of the induced field becomes the same as the one over M1 for the same cortical depth.
6. Digitize the EEG electrodes, so that their position is registered to the brain anatomy.
   NOTE: This is a very important step for locating the distribution of neuronal activation and for accurate repositioning of the electrodes in the follow-up session.
7. Record the TMS-EEG
   1. Replace the earplugs with the earplugs with air tubes to connect to audio masking (e.g., white noise) if available and add headphones over them. Play the audio masking only during TMS pulse delivery.
      NOTE: This step can be applied to step 2.4.2 without playing the audio masking and with care so the head trackers are not moved.
   2. Mount the coil on the coil holder and make sure that the coil does not move or press the electrodes under it. Make sure that the sponge is between the electrodes and the coil.
   3. Remove all active screens out of the sight of the participant. Give instructions to the participant to stare on a fixed point, not to change his/her head position during TMS delivery and not to blink between the TMS pulses.
   4. Switch off any fluorescent lights. Run single pulse TMS, SICI, ICF and LICI in a random order for each participant. Give 100 single and paired pulses. Use various ISI's of 3–4 s (±20%) or a constant of 3–5 s (see Note). Give a break of 3–5 min between each condition so the participant can relax and stretch.
      NOTE: SICI and ICF involve a paired-pulse TMS paradigm with a subthreshold conditioning stimulus (CS) and a suprathreshold test stimulus (TS). The CS used in this protocol is 80% of MT and the TS at the intensity evoking a 1 mV MEP peak-to-peak²⁶. The inter-pulse interval used for optimal SICI is at 2 ms and for ICF at 12–13²⁷. The LICI paradigm involves the pairing of a supra-threshold CS at the intensity evoking the 1 mV MEP peak-to-peak followed by another suprathreshold TS again using the intensity that evoked a 1 mV MEP peak-to-peak and at an inter-pulse interval of 100 ms. The ISI for both single and paired pulse paradigms is determined by the stimulator's charging time (our system can allow paired pulses every 4 s), the amount of sessions (longer experiments would require smaller ISI to not overburden the participants) and the analysis that is going to take place. In this study, we used a constant ISI of 5 s due to our stimulator's restrictions and also because we would need several cycles of low frequency band (theta rhythm) for the time-frequency and power spectrum analysis.

Wyniki

Figure 1A illustrates the TMSevoked potentials after DLPFC stimulation over the F3 electrode after averaging 100 epochs from each session for one healthy volunteer. In this illustration, we highlight the effect of the CS on the TS in comparison to the single pulse condition when TS is applied alone. The CS modulates the N100 deflection in a clear manner even in one subject. In the SICI and LICI sessions, N100 is usually increased and in ICF d...

Dyskusje

TMS-EEG enables the direct and noninvasive stimulation of most cortical areas and the acquisition of the resulting neuronal activity with very good spatio-temporal resolution³⁰, especially when neuronavigation is utilized. The benefit of this methodological advance is based on the fact that TMS-evoked EEG signals originate from the electrical neural activity and it is an index of cortico-cortical excitability. This has tremendous potential in neuropsychiatric patient populations where TMS-EEG can ...

Materiały

Name	Company	Catalog Number	Comments
CED Micro1401-3	Cambridge Electronic Design Limited	CED Micro1401-3	Digital Data Recocrder
BISTIM'2 Package Option 1	Magstim	3234-00	TMS paired pulse stimulator
Magstim 200'2 Unit (2 items)	Magstim	3010-00	TMS stimulators
UI controller	Magstim	3020-00	TMS controller
BISTIM'2 UI controller	Magstim	3021-00	TMS controller
BISTIM connecting module	Magstim	3330-00	TMS connecting module
D70 Alpha Coil - P/N 4150-00 (Alpha 70 mm double coil)	Magstim	4150-00	TMS coil
Brainsight	Rogue-Resolutions	Brainsight 2	Neuronavigator
Model 2024F	Intronix	2024F	Electromyograph
Neuroscan SynAmps RT 64 channel System	Compumedics Neuroscan	9032-0010-01	Electroencephalograph
Quick-Cap electrode system 64	Compumedics Neuroscan	96050255	EEG Cap