Transcranial burst electrical stimulation contributes to neuromodulatory effects in the rat motor cortex

Abstract

Background and objective: Transcranial Burst Electrical Stimulation (tBES) is an innovative non-invasive brain stimulation technique that combines direct current (DC) and theta burst stimulation (TBS) for brain neuromodulation. It has been suggested that the tBES protocol may efficiently induce neuroplasticity. However, few studies have systematically tested neuromodulatory effects and underlying neurophysiological mechanisms by manipulating the polarity of DC and TBS patterns. This study aimed to develop the platform and assess neuromodulatory effects and neuronal activity changes following tBES.

Methods: Five groups of rats were exposed to anodal DC combined with intermittent TBS (tBES+), cathodal DC combined with continuous TBS (tBES-), anodal and cathodal transcranial direct current stimulation (tDCS+ and tDCS-), and sham groups. The neuromodulatory effects of each stimulation on motor cortical excitability were analyzed by motor-evoked potentials (MEPs) changes. We also investigated the effects of tBES on both excitatory and inhibitory neural biomarkers. We specifically examined c-Fos and glutamic acid decarboxylase (GAD-65) using immunohistochemistry staining techniques. Additionally, we evaluated the safety of tBES by analyzing glial fibrillary acidic protein (GFAP) expression.

Results: Our findings demonstrated significant impacts of tBES on motor cortical excitability up to 30 min post-stimulation. Specifically, MEPs significantly increased after tBES (+) compared to pre-stimulation (p = 0.026) and sham condition (p = 0.025). Conversely, tBES (-) led to a notable decrease in MEPs relative to baseline (p = 0.04) and sham condition (p = 0.048). Although tBES showed a more favorable neuromodulatory effect than tDCS, statistical analysis revealed no significant differences between these two groups (p > 0.05). Additionally, tBES (+) exhibited a significant activation of excitatory neurons, indicated by increased c-Fos expression (p < 0.05), and a reduction in GAD-65 density (p < 0.05). tBES (-) promoted GAD-65 expression (p < 0.05) while inhibiting c-Fos activation (p < 0.05), suggesting the involvement of cortical inhibition with tBES (-). The expression of GFAP showed no significant difference between tBES and sham conditions (p > 0.05), indicating that tBES did not induce neural injury in the stimulated regions.

Conclusion: Our study indicates that tBES effectively modulates motor cortical excitability. This research significantly contributes to a better understanding of the neuromodulatory effects of tBES, and could provide valuable evidence for its potential clinical applications in treating neurological disorders.

Keywords: motor evoked potential; neuromodulation; neuroplasticity; rats; transcranial burst electrical stimulation.

Figure 1

Schematic diagram of the experimental design for testing changes in cortical excitability after transcranial burst electrical stimulation (tBES), transcranial direct current stimulation (tDCS) or sham stimulation in anesthetized rats. In this representative experiment, anesthetized rats received intervention protocols for a duration of 20 min. Motor-evoked potentials (MEPs) were measured at baseline and at 0, 10, 20, and 30 min following the intervention. Subsequently, the rat brains were removed and subjected to immunohistochemistry for further investigation. DC, direct current; iTBS, intermittent theta burst stimulation; cTBS, continuous theta burst stimulation.

Figure 2

Placement and assembly of stimulation and recording electrodes. Throughout the entire experimental period, the rats were securely mounted on a stereotactic apparatus. Stimulation protocols were administered using an active electrode positioned 2.5 mm laterally and 1.5 mm anterior to the bregma, while a reference electrode was placed in the abdominal region. The same electrode configurations were employed to elicit motor-evoked potentials (MEPs). MEP data, recorded from the brachioradialis muscles, were subsequently analyzed to assess alterations in cortical excitability resulting from the intervention protocols.

Figure 3

Graph illustrating parameters of four stimulation protocols in the current study. The graph provides an overview of the parameters for the four stimulation protocols utilized in this study. tBES (+): This protocol combines tDCS (+) with intermittent theta burst (iTBS) mode. iTBS comprises 10 bursts of three pulses delivered at a frequency of 50 Hz, with each burst lasting 200 ms and repeated for 2 s, followed by an 8-s rest (A). tBES (−): In this protocol, tDCS (−) is paired with continuous theta burst (cTBS) mode, consisting of bursts of three pulses delivered at a frequency of 50 Hz, with each burst lasting 200 ms (B). tDCS (+): Anodal direct current stimulation is applied at an intensity of 0.3 mA (C). tDCS (−): Cathodal direct current stimulation is applied at an intensity of 0.3 mA (D). All stimulation protocols were administered for 20 min.

Figure 4

The time course changes in raw motor-evoked potential (MEP) signals for tBES (+), tDCS (+), tBES (−), tDCS (−), and the sham condition. MEP amplitudes increased after tBES (+) and tDCS (+), while they decreased following tBES (−) and tDCS (−) over the 30-min post-intervention period compared to baseline. No significant changes in MEP signals were observed under the sham condition.

Figure 5

The average normalized motor-evoked potential (MEP) amplitudes for the contralateral limb (A) and ipsilateral limb (B) across the five intervention protocols (tBES+, tDCS+, tBES−, tDCS−, and sham). The averaged responses of cortical excitability were calculated for each intervention group within 30 min following different stimulations on the contralateral limb (C). Asterisks (*) indicate statistically significant differences when comparing tBES (+), tBES (−), tDCS (+), and tDCS (−) with the sham group at the same time point. The data are presented as means, with error bars representing the standard error of the mean (SEM); ^*p ≤ 0.05.

Figure 6

Representative immunohistochemically stained slices with regions of interest (ROI) (A). Data for c-Fos are presented as the number of labeled cells within the ROI. The changes in cortical expression of c-Fos following tBES (+), tBES (−), and sham stimulation (B). Asterisks (*) indicate statistically significant differences, either between the right and left sides of the brain within a group or when compared to the sham group on the same side of the brain. The data are presented as means, with error bars representing the standard error of the mean (SEM); ^*p ≤ 0.05.

Figure 7

Representative images of GAD-65 immunostaining (A) and the average changes in GAD-65 expression following tBES (+), tBES (−), and sham stimulation (B). Asterisks (*) indicate statistically significant differences between the right and left sides of the brain within a group or when compared to the sham group on the same side of the brain. The data are presented as means, with error bars representing the standard error of the mean (SEM); ^**p ≤ 0.05, ^*p ≤ 0.01.

Figure 8

Representative images of glial fibrillary acidic protein (GFAP) immunostaining (A) and the average changes in GFAP expression following tBES (+), tBES (−), and sham stimulation (B). Asterisks (*) indicate statistically significant differences either between the right and left sides of the brain within a group or when compared to the sham group on the same side of the brain. Data are presented as means, with error bars representing the standard error of the mean (SEM).