Short-Term Cortical Electrical Stimulation during the Acute Stage of Traumatic Brain Injury Improves Functional Recovery

Abstract

Functional restoration is an important issue in the treatment of traumatic brain injury (TBI). Various electrical stimulation devices and protocols have been applied in preclinical studies and have shown therapeutic potential for brain trauma. Short-term invasive cortical electrical stimulation during the acute stage of TBI might be a feasible adjuvant therapy for patients with moderate-to-severe brain injury receiving neurosurgical treatment in the intensive care unit. However, the therapeutic effects of short-term multisession cortical electrical stimulation for brain trauma are not clear. This study explored the therapeutic effects of acute-stage short-term cortical electrical stimulation on TBI. We conducted seven sessions of one-hour cortical electrical stimulation from day 0 to day 6 in rats after brain trauma by controlled cortical impact and then evaluated the functional outcome and histopathological changes. Our data showed that short-term cortical electrical stimulation improved motor coordination, short-term memory, and learning ability and attenuated neurological severity after brain trauma. Lesion volume, apoptosis, and gliosis after brain trauma were reduced, and trauma-induced neurogenesis in the hippocampus for the innate neural reparative response was increased. Our study demonstrated that short-term cortical electrical stimulation applied in the acute stage of traumatic brain injury is a potential adjuvant therapy to improve the recovery of neurological deficits.

Keywords: cortical electrical stimulation; functional recovery; neural stem cells; traumatic brain injury.

Figure 1

Schematic diagram of the experimental design: (A) Schedule of experiments, including traumatic brain injury (TBI) induction, cortical electrical stimulation (ES), modified neurological severity score (mNSS), rotarod test, Morris water maze (MWM) test, and novel object recognition (NOR) test. (B) Parameters of control cortical impaction and coordinates of craniotomy. (C) Parameters of CES. (D) Neurological test. (E) Histology and molecular analysis. (F) Demonstrating picture of the electrode device.

Figure 2

The neurological evaluation showed that CES improves functional recovery after TBI. (A) The modified neurological severity score (mNSS) evaluated on the third day after TBI showed no difference in terms of neurological severity between the TBI group (TBI + sham stimulation) and the TBI + ES group (TBI + cortical electrical stimulation). (B) On the seventh day after TBI, there was a significant improvement in mNSS in the TBI + ES group compared to the TBI group (p < 0.005). (C) The rotarod test showed a significantly prolonged latency to fall in the TBI + ES group (p < 0.0001). (D) The novel object recognition (NOR) test showed a significantly improved decimation index in the TBI + ES group (p < 0.001), indicating improved short-term memory by CES after TBI. (E) The Morris water maze (MWM) task showed that the escape latency in the sham group was significantly shorter than that in the TBI group from day 2 to day 5. The escape latency was significantly shorter in the TBI + ES group than in the TBI group, and there was no difference between the sham and the TBI + ES groups. Values are expressed as the mean ± SEM. **** p < 0.0001, *** p < 0.001, ** p < 0.005, * p < 0.05, ## p < 0.005, # p < 0.05.

Figure 3

CES decreased the lesion volume of cortical contusion. (A) Representative pictures of cresyl violet staining. (B) Quantification of lesion volume. The data showed significantly decreased lesion volume in the TBI + ES group, suggesting neuroprotection by CES. Values are expressed as the mean ± SEM. **** p < 0.0001, * p < 0.05.

Figure 4

CES reduced cleaved caspase 3 (cCas-3) immunoreactive cells in the perilesional cortex. (A) Representative picture illustrating the sample region for the perilesional cortex. The cortical specimen was obtained within 1 mm from the wall of the lesion cavity. (B,C) Representative pictures of immunofluorescent for cCas-3 (green). (D,E) Representative pictures of merged immunofluorescent of DAPI (blue) with cCas-3. (F) Quantitative analysis showed a significantly decreased number of cCas-3 immunoreactive cells. Values are expressed as the mean ± SEM. ** p < 0.001.

Figure 5

CES decreased the expression of Glial fibrillary acidic protein (GFAP) in the perilesional cortex. (A) Representative fluorescence images of GFAP (green), DAPI (blue), and merged picture. (B) Representative pictures of western blot for GFAP. (C) Quantitative analysis of western blot showed significantly decreased GFAP expression in the TBI + ES group compared to the TBI group. Values are expressed as the mean ± SEM. * p < 0.05, *** p < 0.001.

Figure 6

CES stimulates the proliferation of cells and increases the number of mature newborn neurons in the dentate gyrus of the hippocampus. (A) Representative fluorescence images of NeuN (green), BrdU (red), and merged picture. (B) Quantitative analysis showed significantly increased BrdU immunoreactive cells in the dentate gyrus. (C) Quantitative analysis demonstrated that BrdU/NeuN co-stained cells, indicating mature newborn neurons, are significantly increased in the TBI + ES group. Values are expressed as the mean ± SEM. **** p < 0.0001, ** p < 0.005.

Figure 7

CES stimulates migration from the subgranular zone (SGZ) to the granular cell layer (GCL). (A) Representative pictures of the DCX immunofluorescence stain (green) of the dentate gyrus. (B) The quantification of DCX⁺ cells in the dentate gyrus showed no significant difference between groups. (C) Highly magnified fluorescence pictures of DCX (green) and DAPI (blue) in the junctional zone between SGZ and GCL. (D) Quantitative analysis showed no significant difference in the number of DCX⁺ cells in the junctional zone between the sham and TBI groups. In the TBI + ES group, there was a significant increase in DCX⁺ cells in the junctional zone between SGZ and GCL, indicating that CES stimulates the migration of immature neurons from SGZ to GCL. Values are expressed as the mean ± SEM. ns, no significance; *** p < 0.001; ** p < 0.005.

Figure 8

CES upregulates PI-3 kinase/Akt, MEK/ERK, and Wnt/β-catenin signaling pathways. (A) Representative fluorescence images of Beta-catenin (green), DCX (red), DAPI (blue), and merged picture. (B) Representative image of western blot. (C) Quantification of Akt phosphorylation presented as the ratio of phosphorylated Akt (phospho-Akt) to total Akt. The data showed significantly downregulated phospho-Akt on the 18th day post-TBI, whereas CES treatment restored the downregulation. (D) The quantification of Erk phosphorylation (phospho-Erk) demonstrated significantly upregulated ERK phosphorylation, but CES did not further increase the upregulation. (E) The quantification of β-catenin showed the significant downregulation of β-catenin expression after TBI and CES treatment returned the expression level. Values are expressed as the mean ± SEM. ns, no significance; ** p < 0.005; * p < 0.05.