Simultaneous transcranial electrical and magnetic stimulation boost gamma oscillations in the dorsolateral prefrontal cortex

doi:10.21203/rs.3.rs-1550621/v1

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Simultaneous transcranial electrical and magnetic stimulation boost gamma oscillations in the dorsolateral prefrontal cortex

https://doi.org/10.21203/rs.3.rs-1550621/v1

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Neural oscillations in the gamma frequency band have been identified as a fundament for synaptic plasticity dynamics and its alterations are central in various psychiatric and neurological conditions. Transcranial magnetic stimulation and alternating electrical stimulation may have a strong therapeutic potential by promoting gamma oscillations and plasticity. Here we applied intermittent theta-burst stimulation (iTBS), an established TMS protocol known to induce cortical plasticity, simultaneously with transcranial alternating current stimulation (tACS) at either theta (θtACS) or gamma (γtACS) frequency on the dorsolateral prefrontal cortex (DLPFC). We used TMS in combination with electroencephalography (EEG) to evaluate changes in cortical activity on both left/right DLPFC and over the vertex. We found that simultaneous iTBS with γtACS but not with θtACS resulted in an enhancement of spectral gamma power, a shift of individual peak frequency towards faster oscillations and an increase of local connectivity in the gamma band. These results were specific to the DLPFC and confined locally to the site of stimulation, not being detectable in the contralateral DLPFC. We argue that the results described here could promote a new and effective method able to induce long-lasting changes in brain plasticity useful to be clinically applied to several psychiatric and neurological conditions.

Transcranial alternated current stimulation

Transcranial magnetic stimulation

Dorsolateral prefrontal cortex

Gamma oscillations

During the last decades, a growing body of neurophysiological studies has focused on the possibility to interfere with and modulating neural oscillations. Cortical oscillatory activity represents the rhythmic activity of a population of neurons within a given frequency band ¹, is crucial for brain networks dynamics ^2,3 and underlies cognitive processes in healthy ⁴ and pathological brains ⁵. Gamma oscillations in the prefrontal areas are involved in several cognitive functions including attention and memory ^6,7. Furthermore, several studies in human and animal models have suggested a role for γ oscillations in inducing and supporting synaptic plasticity mechanisms in cortical prefrontal ⁸ and motor areas ⁹.

Non-invasive brain stimulation (NIBS) techniques have been widely used to induce plastic changes in cortical areas, to modulate brain rhythms and influence the ongoing cortical activity. Intermittent theta-burst stimulation (iTBS) is a form of repetitive transcranial magnetic stimulation (rTMS) method, which consists of bursts of high-frequency stimulation (3 pulses at 50 Hz) repeated at intervals of 200 ms, that can induce robust long-lasting changes in the stimulated area ¹⁰. It was firstly developed in animal models to mimic the natural patterns that support synaptic long-term potentiation (LTP) mechanisms ^11,12. iTBS has been successfully employed in the clinical ground in several conditions ranging from depression ¹³ to stroke recovery ¹⁴.

These iTBS after-effects seem to be mediated by GABAergic interneurons activity, which is also crucial for oscillatory activity modulation ^15–17.

Transcranial alternating current stimulation (tACS) is another NIBS protocol delivering electrical stimulation with a sinusoid alternated current within a specific frequency. tACS can entrain ongoing brain oscillations activity and modulate brain areas in a frequency-dependent manner ¹⁸. γtACS has been shown capable to interact with γ oscillatory activity in the motor cortex (M1) ^19,20 as well as in the prefrontal cortex ²¹. Moreover, tACS gained attention given the therapeutic potential to induce long-lasting increases of gamma oscillations, since a decrease in gamma activity is central in various psychiatric and neurological conditions such as schizophrenia ²² and Alzheimer’s disease ²³. However, the therapeutic effect of NIBS protocols acting on gamma oscillations is currently limited by the fact that the after-effects are often variable, of small magnitude, and short-lasting ²⁴.

Recently the pioneering work of Guerra and others showed that contemporary electrical and magnetic stimulation can promote robust after-effects on cortical oscillations when applied over M1 ^25–27. This combination has not been assessed in areas involved in cognitive functions such as the dorsolateral prefrontal cortex (DLPFC) which is involved in the pathophysiology of a wide range of neurological diseases ^28–31.

Here, we used a novel approach by combining transcranial magnetic stimulation (TMS) and electroencephalography (EEG) to assess the potential after-effects of simultaneous iTBS and tACS on DLPFC cortical activity ³². ^33,34.

We hypothesize that entraining γ frequency by tACS during iTBS could boost the long-lasting plasticity after-effects of iTBS on oscillatory activity by inducing a synergistic effect on the underlying local networks.

All 13 participants completed successfully the three-session protocol planned. The different stimulation protocols were all well-tolerated, and subjects reported no side effects. Single-pulse TMS evoked on EEG signal a well-known sequence of positive and negative deflections with amplitude ranging from − 3 to 3 µV and lasting up to ~ 250 ms (Fig. 1, Fig. 2). TMS-evoked cortical reactivity (panel (a)) last around 150 ms and it is characterized by a series of peaks such as P1 from 15 to 25 ms, P2 from 26 to 47 ms, P3 from 48 to 65 ms, P4 from 66 to 75 ms, P5 from 76 to 115 ms, P6 from 116 to 145 ms. This temporal dynamic, in terms of waveform and amplitude, looks similar for all the sites at baseline. The three peaks were detectable over all the three stimulated areas as expected (Veniero et al., 2013, Rosanova et al., 2009; Casula et al., 2016). No differences in the general amplitude are detected between DLPFC and Vertex (p > .05). In the first two peaks window (i.e. 15–60 ms after TMS) a dipole was focused over the stimulated area; from ~ 65 to ~ 120 ms after TMS spread negativity observable over both the hemispheres, followed by a strong positivity centred over the frontocentral electrodes, ranging from ~ 120 to ~ 250 ms after TMS. Figures 1,2 and 3 show the TMS-evoked potential (TEPs) over the different stimulation sites (l-DLPFC; r-DLPFC; vertex) in “tACS” and “Time” conditions. iTBS + γtACS stimulation showed a neuromodulation effect evident on the third peak while stimulating l-DLPFC (Fig. 1, panel (a)). Figure 1 (b) depict the peaks analysis revealing a significant ANOVA “tACS” (γ, θ, sham) x “time” (T0, T1, T2) interaction [F(10, 120) = 1.066 ; p = 0.05.; η² = .082]. Post-hoc analysis showed that in the iTBS + γtACS stimulation condition the third peak significantly differed in t0 vs t1 times conditions (p = 0.05). No effects were found for r-DLPFC (Fig. 2) and vertex (Fig. 3) sites.

Figures 4, 5, and 6 show local TMS-evoked cortical oscillations. In detail, the figures represent the local oscillatory activity (Fig. 4l- DLPFC; Fig. 5r- DLPFC; Fig. 6 vertex). As depicted in panel (a), TMS-evoked cortical oscillations have similar baseline activity patterns characterized by a remarkable activation around 50 ms after TMS pulse in the frequencies between 20 to 30 Hz. A second activation can be identified from around 50 ms to 250 ms in lower frequencies such as approximately 6 Hz to 10 Hz. Cortical maps show the topographical cortical oscillations activation, which was stronger in areas around the stimulated site. Panel (b) represents the most expressed individual frequencies for all the stimulation conditions and during time. In baseline for all the conditions and areas, the most expressed individual frequencies were in a range between 22 Hz to 28 Hz. Figure 4 shows l- DLPFC cortical oscillations patterns dynamics before and after iTBS + tACS stimulation. We analyzed gamma oscillation (mean 30–50 Hz) power and individual frequency shifting in time. Gamma oscillation power analysis shows a significant difference between conditions (“Time”, “tACS”) [F(4, 48) = 2.801; p = .03; η² = .189]; post-hoc analysis reveals that iTBS + γtACS increase power in time between t0 vs t2 [p = .05]. No effects were originated by the other “tACS” conditions (iTBS + θtACS /sham tACS). A latter analysis was focused on the individual frequency shifting right after the iTBS + tACS stimulation; iTBS + γtACS stimulation modulates spectral shifting in the most expressed individual frequency [F(2, 24) = 3.335; p = .05; η² = .217]; post-hoc analysis reveal the difference between T0 vs T1 “Time” conditions [p = .04]. iTBS + θtACS /sham tACS stimulation conditions have no effects on individual frequency shifting. Figure 4 panel (b) shows these effects. No significant effects were reported for the same analysis in r- DLPFC and vertex (Figs. 5 and 6, panel (b)).

Figure 7 shows the individual frequency wavelet phase-locking value analysis (W-PLV). W-PLV was conducted on the data collected during l-DLPFC TMS-EEG recordings between F3-F5 and F3-F4 before and after iTBS + tACS stimulation for the three “tACS” conditions. Panel (a) represents W-PLV in F3-F5 and F3-F2 channels pairs for the “tACS” conditions in time. PLV values are higher in T0 for lower frequency bands and in a range between 22 to 28 Hz for both couples of electrodes. Figure 7 panel (b) represents histograms that show the neuromodulation iTBS + γtACS stimulation effects on W-PLV in l- DLPFC during time (T0; T1; T2) for a mean Gamma band range (30–50 Hz). W-PLV analysis shows a significant difference between conditions (“Electrode”, “Time”) [F(2, 22) = 20.817; p = .01; η² = .654] for the Gamma frequency (mean between 30–50 Hz); post-hoc analysis shows significant difference between T0 and T1 [F5 p = .05; F2 p = .01] and T0 and T2 [F5 p = .004; F2 p = .002] for both the F3-F5 and the F3-F2 pairs. These effects are depicted respectively in panel (b)’s left (F3-F5) and right (F3-F2) side histograms. No significant effects were reported for the same analysis in r- DLPFC and vertex.

Here we show that simultaneous transcranial electrical and magnetic stimulation exerts a robust long-lasting increase in gamma oscillations in the DLPFC. We found that combined DLPFC iTBS-γtACS resulted in an enhancement of spectral gamma power, a shift of individual peak frequency towards faster oscillations, and in an increase of local connectivity in the gamma band. These results were specific to the stimulated area and confined locally to the site of stimulation, not being detectable in the contralateral DLPFC. This study aimed to take advantage of the iTBS and tACS properties to boost the capability that these two NIBS methods have to produce long-lasting oscillatory brain changes on DLPFC and to evaluate whether a synergistic effect would arise from this combination. Although there is great interest in the application of TBS in several neurological and psychiatric disorders, its clinical impact is somewhat limited by the variability of after-effects that have been reported in healthy studies evaluating the effects on the amplitude of the MEP. These studies showed that only approximately 50% of subjects undergoing iTBS show the expected significant motor MEP long-lasting increase ^35–38. Inter-subject variability is considered the most relevant limitation of non-invasive brain stimulation ^39,40 since it affects the effectiveness of neuromodulation techniques and limits their clinical applications. Moreover, the magnitude of increase in MEPs amplitude after M1 iTBS and in TEPs amplitude after DLPFC iTBS is in the range of 20–50% as compared to baseline, again potentially limiting the clinical impact of this plasticity inducing protocol ⁴¹. While initially iTBS was thought to produce more powerful and reproducible effects than other rTMS methods, a claim that has not been fully confirmed, its main attraction still relies on the speed of application with protocols lasting a few hundreds of seconds instead of several minutes ⁴¹. Indeed iTBS through the high-frequency neuronal activation modulates cortical inhibition and GABA-ergic synaptic transmission, resulting in an enhancement of γ band expression ⁴². iTBS is thought to activate Ca2 + influx to the postsynaptic neuron. The property, including the amount and the rate of the increase, determines the amount of the build-up of subsequent facilitation processes that modify the synaptic strength ⁴³. This notion is supported by animal studies showing that dysfunction of Inositol 1,4,5-trisphosphate receptors (InsP3Rs) that is required for LTP results in a conversion of LTD to LTP, while partial blockade of NMDARs to reduce the rate of Ca2 + influx results in a conversion of LTP to LTD ⁴⁴.

On the other hand, tACS is a relatively new technique that can effectively modulate oscillatory brain activity through weak external alternating current at specific frequencies ¹⁸. This effect is supported by research in animals ⁴⁵ suggests that TES in phase with network-induced patterns can enhance neuronal discharge activity. Through the mechanism of stochastic resonance ^46,47, the electric field, which in itself may be subthreshold, can be effectively summed with otherwise subthreshold effects of network-induced membrane voltage fluctuations, and the combined effect can generate spikes in a fraction of the neuronal population. However, there is still a lack of understanding about the exact mechanisms that modulate cortical activity as a function of tACS administration.

We argue that the robust synergistic effects observed here are the consequence of the interplay among gamma oscillations and the formation of cortical plasticity. The role of gamma activity in synaptic plasticity has been then confirmed throughout the last two decades by numerous investigations using electrophysiological recordings in animals ⁴⁸ and humans ⁴⁹. Although the exact physiological mechanism is still a matter of debate, it has been suggested that local inhibitory interneurons play a key role in synchronizing gamma oscillations among large neuronal populations ^50–53. Thus, when depolarized, local interneuron populations tend to generate synchronized inhibitory postsynaptic potentials in thousands of cells, inducing an entraining in fast gamma oscillations not only in local but also in distant neurons ^33,51,54,55. This is relevant since gamma-frequency synchronization between the activity of distant neuronal cells has emerged as a marker of connectivity within large cortical networks, during learning or memory processing ^56,57. During the formation of plasticity, an increase in gamma-activity coherence represents enhanced connectivity between distant neuronal populations in forming a new memory ⁵⁸. This latter element is particularly relevant since cognitive dysfunction in AD has been recently linked to a disorder of gamma oscillations ²³. In AD animal models, local changes in gamma oscillatory activity affect multiple brain centres critical for learning and memory, and other higher-order brain functions, such as the hippocampus and the prefrontal cortex ^59–63. Hence we believe that our current findings may have broad implications for treating gamma dysregulation in neurodegenerative disorders such as AD.

While we found that combined iTBS-γtACS induced robust after-effect on DLPFC cortical activity both in terms of excitability and oscillations, combined iTBS-θtACS did not result in any significant change. This finding is in line with the pioneering work of Guerra and colleagues showing that tACS delivered at lower frequencies in the alpha band leaves the iTBS-related LTP-like plasticity unchanged (Guerra et al., 2018, 2019).

In the current study, we adopted a protocol for combined stimulation of tACS and iTBS based on a synchronous start triggered externally. This approach is supposed to drive oscillatory activity using tACS while iTBS exerts its effects on local LTP-like mechanisms. However, other approaches are trying to apply controllable phase-synchronized rTMS with tACS to induce and stabilize neuro-oscillatory resting-state activity at targeted frequencies. For instance, Hosseinian and coll. ⁶⁴ used a novel circuit to precisely synchronize rTMS pulses with the phase of tACS in the bilateral prefrontal cortex (PFC). They found that 10-Hz resting-state PFC power increased significantly with peak-synchronized rTMS + tACS, while rTMS timed to the negative tACS trough did not induce local or global changes in oscillations. Moreover, they also developed a novel stimulation protocol, where a single circuit precisely synchronizes rTMS pulses with the phase of tACS in the theta frequency band ⁶⁵. Similarly, Zrenner et al., ⁶⁶ hypothesized that triggering TMS synchronized with the negative peak of endogenous alpha oscillations in left DLPFC would more effectively increase cortical excitability (as measured with TMS-evoked potentials) than a non-alpha-synchronized stimulation protocol.

The study has some limitations. Some control conditions such as theta-tACS alone and gamma-tACS alone are lacking. However, it has been shown previously that a few seconds of tACS are not supposed to exert any after-effect (Lafleur et al., 2021) and thus we did not weigh down our experimental procedure that was already quite demanding for the healthy subjects recruited for the study. The TMS-EEG measurements did not include a sham stimulation condition to control for peripherally evoked potentials and muscle artefacts. In this regard, we adopted all the TMS that can result in non-specific effects, such as auditory and somatosensory stimulation that can affect the EEG response (Rocchi et al., 2021). We adopted several methodological precautions to avoid these artefacts. To reduce the auditory response, we used an ad-hoc masking noise; to reduce bone conduction of the TMS click and scalp sensation caused by coil vibration we placed a 0.5 cm foam layer underneath the coil. Finally, while our evidence seems to suggest that the synchronization of rTMS with peak oscillatory activity may have an impact on subsequent plasticity induction, this approach is technically limited to lower frequency bands in the theta-alpha range. Current methodological restraints do not allow to transfer of a similar approach towards higher frequencies such as those used in the gamma band in our case (70 Hz) since these cannot be reliably detected online with non-invasive scalp EEG recordings. Such hypothesis however could be tested in the future in patients with implanted electrodes.

In conclusion, we argue that the results described here could promote a new and effective method able to induce long-lasting changes in brain plasticity and connectivity, useful to be clinically applied to several psychiatric and neurological conditions.

Participants and procedure

13 healthy participants were enrolled in the study. Experimental within-subject design included three different randomized sessions every participant underwent to: (1) iTBS + γtACS stimulation; (2) iTBS + θtACS stimulation; (3) iTBS + sham tACS stimulation. During every session subject underwent TMS-EEG neurophysiological assessment on three testing times: before (T0) iTBS + tACS stimulation, 0 min (T1) and 20 min (T2) after the iTBS + tACS stimulation. Three cortical areas were studied using TMS-EEG: left dorsolateral prefrontal cortex (l-DLPFC), right dorsolateral prefrontal cortex (r- DLPFC) and vertex (Fig. 8). The study was approved by the local ethics committee (Ethical Committee Fondazione Santa Lucia, Via Ardeatina 306, 00179 Roma; Prot. CE/PROG.811; 19/02/2021)TMS-safety guidelines and medical regulations were fully followed by experimenters and written informed consent was obtained from each participant.

iTBS + tACS stimulation

The iTBS + tACS stimulation was applied over l- DLPFC and last for 190s, with the tACS electrode on the scalp and the iTBS coil just above it. A figure-of-eight coil with a diameter of 70 mm was used to deliver iTBS over the scalp site corresponding to the l- DLPFC (F3, according to 10–20 system). A MagStim Super Rapid magnetic stimulator (Magstim Company, Whitland, Wales, UK) was used to deliver the biphasic waveform pulse, with a pulse width of ∼0.1 ms. iTBS consist of a 2 seconds train of TBS that is repeated 20 times, every 10 seconds for a total of 190 seconds (600 pulses). Stimulation intensity was set at 80% of the active motor threshold (AMT), defined as the lowest intensity able to produce MEPs < 200 µV in at least five out of ten trials when the subject performed a 10% of maximum contraction using visual feedback ⁶⁷. AMT was tested over the motor cortex of the hemisphere target of the iTBS stimulation (i.e. left) with the same stimulation condition, such as with the tACS electrode behind the coil. Electromyographic activity was recorded from the contralateral FDI muscle, using two Ag-AgCl surface cup electrodes (9 mm) in a belly-tendon montage. Responses were amplified through a Digitimer D360 amplifier (Digitimer Ltd, Welwyn Garden City, Hertfordshire, UK): filters were set at 20 Hz and 2 kHz, with a sampling rate of 5 kHz; they were then recorded by a computer using SIGNAL software (Cambridge Electronic Devices, Cambridge, UK). During stimulation, the coil was oriented 45° with respect to l- DLPFC. A neuronavigation system (SofTaxic; E.M. S., Bologna s.r.l.) coupled with a Polaris Vicra infrared camera was used to ensure that in each subject iTBS was applied over the same spot across different sessions. Indeed stimulation point was set and memorized during the first session and imported at the beginning of the other two sessions, to make sure the stimulated area was constant across sessions. A Brainstim multifunctional system for low-intensity transcranial electrical stimulation (E.M. S., Bologna s.r.l.) was used to deliver the current stimulation using saline-soaked sponge electrodes (7 × 5 cm²). The active electrode (anode) was placed on the scalp over the left- DLPFC (l- DLPFC, F3) and the reference (cathode) over the right deltoid muscle. During real stimulations, the current was set to 1 mA and delivered for 190s. No ramp-up and no ramp down were programmed for the stimulation. For the sham condition, the electric current was not applied, but there were a 2 seconds 1 mA ramp up and 2 seconds 1 mA ramp down, to give the subject real stimulation feelings. For γtACS the sinusoid frequency wave was set at 70 Hz; for θtACS, the sinusoid frequency wave was set at 5 Hz. iTBS and tACS were synchronized using a BrainTrigger (E.M. S., Bologna s.r.l.) and SIGNAL Software: both stimulations started exactly at the same time.

TMS-EEG cortical assessment

During every session, participants underwent an electroencephalographic (EEG) recording of a series of TMS pulses delivered in specific areas of interest. Consequently, it was possible to evaluate cortical reactivity, connectivity and plasticity during time. TMS was carried out using a Magstim R² stimulator with a 50 mm figure-of‐eight coil (Magstim Company, Whitland, Wales, UK), which produces a biphasic waveform with a pulse width of ∼0.1 ms. TMS pulses were applied over the l- DLPFC, right- DLPFC (r- DLPFC) and vertex. According to scientific literature, the coil was differently oriented with respect to the mid-sagittal axis of the subject's head for each stimulation site: 45° over l- DLPFC and r- DLPFC, and parallel over the vertex, with the handle pointing backwards. The intensity of stimulation of single-pulse TMS was set at 110% of the resting motor threshold (RMT), defined as the lowest intensity producing MEPs of > 50 µV in at least five out of 10 trials in the relaxed first dorsal interosseous (FDI) muscle of the right hand ⁶⁸. The coil position was constantly monitored using a neuronavigation system (SofTaxic; E.M. S., Bologna s.r.l.), to ensure a high degree of reproducibility within the same session and across different sessions. Stimulation points were set and memorized at the beginning of the first session and imported at the beginning of the other two sessions, to make sure the stimulated areas were constant across times and sessions.

A TMS-compatible DC amplifier (BrainAmp, Brain Products GmbH, Munich, Germany) was used to record EEG activity from the scalp. The EEG was continuously recorded from 64 sites positioned according to the 10–20 International System, using TMS‐compatible Ag/AgCl pellet electrodes mounted on an elastic cap. Additional electrodes were used as ground and reference. The ground electrode was positioned in AFz, while the reference was positioned on the tip of the nose. EEG signals were digitized at a sampling rate of 5 kHz. Skin/electrode impedance was maintained below 5 kΩ. Horizontal and vertical eye movements were detected by recording the electrooculogram to offline reject the trials with ocular artefacts.

Each TMS-EEG session consisted of single pulses (100 for T0, 80 for T1 and T2) applied at a random inter-stimulus interval (ISI) of 2 seconds with a variation of 20%. During TMS-EEG assessment subjects passive listen to a white noise and wear ear protectors to ensure the environmental noise does not affect the EEG signal. A short break was run between TMS-EEG sessions of either site. During the entire session, subjects were seated on a dedicated, comfortable armchair in a soundproofed room.

TMS-EEG Analysis

TMS–EEG data were analyzed offline with Brain Vision Analyzer (Brain Products GmbH) and EEGLAB toolbox running in a MATLAB environment (MathWorks Inc., Natick, MA). As a first step, data were segmented into epochs starting 1 s before the TMS pulse and ending 1 s after it. We first removed and then replaced data, using a cubic interpolation, from 1 ms before to 10 ms after the TMS pulse from each trial. Afterwards, data were downsampled to 1,000 Hz and bandpass filtered between 1 and 80 Hz (Butterworth zero-phase filters). A 50 Hz notch filter was applied to reduce noise from electrical sources. Then, all the epochs were visually inspected and those with excessively noisy EEG were excluded from the analysis. Independent component analysis (INFOMAX-ICA) was applied to the EEG signal to identify and remove components reflecting muscle activity, eye movements, blink-related activity, and residual TMS-related artefacts based on previously established criteria ⁶⁹. Finally, the signal was re-referenced to the average signal of all the electrodes.

To evaluate changes in cortical excitability we evaluated local cortical responses evoked by TMS. Local response was assessed, for each subject and three stimulation sites (l- DLPFC, r- DLPFC, vertex), by measuring the first six peaks (P1 from 15 to 25 ms, P2 from 26 to 47 ms, P3 from 48 to 65 ms, P4 from 66 to 75 ms, P5 from 76 to 115 ms, P6 from 116 to 145 ms) measured by the TMS-evoked potentials (TEPs) waveform at each electrode within a pooling calculated around the stimulation site (F1, F3, FC1 for l- DLPFC; F2, F4, FC2 for r- DLPFC; C1, C2, CZ for vertex).

Cortical oscillations analysis was performed using a time/frequency decomposition based on Morlet wavelet (cycles = 3; frequency resolution = 1 Hz from 4 to 50 Hz; temporal resolution = 1 ms) and then by computing TMS-related spectral perturbation (TRSP) ^70,71.

To measure oscillations power and to perform the peak shifting analysis we assessed the local TRSP for each stimulated site and stimulation condition. For each TMS area, we considered a pooling computed around the stimulation site (Same as Cortical excitability analysis, see above), and averaged the TRSP values for each of the 23 frequency layers, between 10 and 250 ms for alpha (α) and θ bands, and between 10 and 100 ms for beta (β) and γ band. These time windows were chosen considering the meantime windows of activity.

Wavelet Phase-locking value analysis is a measure of the phase synchronization of a pair of channels. It computes the randomness of the phase-locking between two channels: the index is calculated in a range between 0 and 1, where 0 represents the complete absence of phase-locking and 1 is the total phase synchronization between channels. After the computation of the wavelets and TRSP (see the paragraphs above) we applied the formula to compute the Wavelet Phase Locking Value (W-PLV) as reported in previous studies ⁷².

Statistical analysis

To assess the effect of iTBS + tACS stimulation on cortical excitability different repeated-measures ANOVA with within-subject factors “tACS” (γ, θ, sham) and “time” (T0, T1, T2) were performed for each site of TMS-EEG recordings. The analyses on cortical oscillations were conducted in the following steps. We first assessed the gamma oscillation at baseline (T0) in terms of frequency and power. To test if the iTBS + tACS protocol could have induced any change in the gamma band in terms of power of oscillation, we used a repeated-measures ANOVA with within-subject factors “tACS” (γ, θ, sham) and “Time” (T0, T1, T2). Then we focused on the individual frequency shifting analysis; we calculated the individual frequency peaks (the most expressed frequency in the whole oscillation spectrum) and, equally to the gamma band power analysis, we used a repeated-measures ANOVA with within-subject factors “tACS” (γ, θ, sham) and “Time” (T0, T1, T2) to assess the changes in the band’s expression in terms of shifting. For the Wavelet Phase-locking value analysis, we performed the same ANOVA analysis with different within-subject factors such as “Electrode” (F3 vs. F5; F3 vs. F4) and “Time” (T0, T1, T2).

Intermittent theta burst stimulation (iTBS); Transcranial alternating current stimulation (tACS); Dorsolateral prefrontal cortex (DLPFC); TMS-evoked potential (TEPs); TMS-related spectral perturbation (TRSP); Wavelet Phase Locking Value (W-PLV).

Funding

This research was partially supported by the Alzheimer’s Drug Discovery Foundation (ADDF), Brightfocus Foundation, and the Italian Ministry of Health to GK.

Data availability statement

Data could be requested from the corresponding author upon a rational statement.

Declaration of competing interest

None.

Author contribution statement

MM conceived the experimental setup, collected and pre-processed the data, performed the analysis and prepared the manuscript; EPC conceived the experimental setup, performed the analysis and prepared the manuscript; IB, MA, AD, VP, LC, MCP collected. pre-processed the data and performed analysis, and GK supervised the work. All authors have reviewed the manuscript.

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Simultaneous transcranial electrical and magnetic stimulation boost gamma oscillations in the dorsolateral prefrontal cortex

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Abstract

Figures

Introduction

Results

Discussion

Methods

Participants and procedure

iTBS + tACS stimulation

TMS-EEG cortical assessment

TMS-EEG Analysis

Statistical analysis

Abbreviations

Declarations

References

Competing Interests

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