Functional brain mapping using specific sensory-circuit stimulation and a theoretical graph network analysis in mice with neuropathic allodynia

Figure 1. Segregation of BOLD signals detected after forepaw and hindpaw stimulation. BOLD signals were detected in regions adjacent to the contralateral S1 cortex of adult mice following stimulation of the forepaw (cyan) and hindpaw (magenta). The brain regions activated by 2000 Hz electrical stimulation of Aβ fibers were clearly visualized on BOLD echo planar images. P < 0.05 voxel-wise FWE corrected; Scale bar, 2 mm. (B) The increase in MR signal in the contralateral S1 cortex was closely correlated with the stimulus performed between 20–30 s (gray). X axis, scan time [sec]; Y axis, signal intensity; Error bar, SE; Strength of stimulation, 500 μA at 2000 Hz.

Figure 2. BOLD mapping by the selective stimulation of intact peripheral nerves. (A) The BOLD signal (red) from 5 Hz stimulation of C-fibers (nociceptive and thermal sensations) was specifically located in the contralateral ACA and S1 forepaw region. After 250 Hz stimulation of Aδ-fibers (fast pain) and Aβ-fibers (touch sensation), the signal (blue) was observed in the contralateral ACA, S1, and thalamus. After 2,000 Hz stimulation of Aβ-fibers, the activated area (green) was detected only in the contralateral S1 (B). To obtain the signal change (%), the MR signal in the ACA, S1, and thalamus was analyzed quantitatively. The results indicated that a significantly higher BOLD signal was detected in the S1 region with all types of sensory input through C-, Aδ- and Aβ-fibers, whereas those in the ACA and thalamus were only observed after painful stimulation through C- and Aδ-fibers. Intensity of stimulation, 500 μA; forepaw, P < 0.05 voxel-wise FWE corrected.

Figure 3. Behavioral assessment of pain and touch sensation in peripheral nerve injury model mice. (A and B) Mechanical allodynia was evaluated using von Frey filaments (0.02 g, 0.16 g). Seven days after peripheral nerve injury, significantly higher scores were detected for both 0.02 and 0.16 g stimulation. (C) Thermal allodynia was evaluated, and significant hypersensitivity was observed at postoperative day 2 and later. (D–F). The sensory fiber-selective threshold change was evaluated in the EPW test with electrical stimulation at 2000 Hz (D), 250 Hz (E), and 5 Hz (F). A significant difference in withdrawal threshold was detected at postoperative day 7 after 2000 Hz stimulation. Behavioral analyses indicated that mice at postoperative day 7 showed the greatest allodynia. 2-sample t-test (*p < 0.05, **p < 0.001).

Figure 4. Visualization of allodynia in peripheral nerve injury model mice with fMRI. (A) BOLD signals in intact mice (green) were only observed in the contralateral S1 hindpaw region following 2000 Hz electrical stimulation. At postoperative day 7, an additional BOLD signal (magenta) was detected in the ACA, S1, and thalamus of the same animals. (B) For the quantitative assessment for allodynia, the signal change (%) was calculated by subtracting the MR signal in the ACA, S1, and thalamus, before and after peripheral nerve injury, from sham-operated mice. Significantly higher BOLD signals were observed in the ACA and thalamus after peripheral nerve injury. There were no significant differences between sham and SNL groups. Green, intact (preoperation); magenta, peripheral nerve injury; blue, sham-operated, MR signal threshold P < 0.05 voxel-wise FWE corrected, 2-sample t-test (*p < 0.05, **p < 0.005).

Figure 5. Visualization and graph theory analysis of brain networks in the peripheral nerve injury model mice with resting state functional connectivity MRI. (A) A connection matrix was expressed in terms of the temporal correlations between 576 architectonic subdivisions in the intact and spinal nerve ligation (SNL) mouse whole brain. Vertical and horizontal axes indicate the identification number of ROIs. The color bar shows the correlation between ROIs. (B) Brain networks were visualized in terms of 576 nodes and edges based on a connection matrix. (C) Characteristics of the brain network as calculated by a graph theory approach. Green, intact (preoperation); magenta, peripheral nerve injury. Paired t-test (*p < 0.05, **p < 0.001).