Written by - Sun Wei, Liang Ting

Editor-in-charge - Wang Sizhen

Editor — Binwei Yang

Stroke is one of the most important causes of death and disability worldwide

In view of the above background, the research group has used the rat thalamus hemorrhagic post-stroke pain model in recent years and found that: (1) the site of thalamus hemorrhage is closely related to the occurrence of pain, such as hemorrhage involvement of the spinal motha, thalamus ventral basal nucleus group and posterior nucleus group of the thalamus, which can cause pain sensitivity, and other parts are rare [8]; (2) Long-term administration of gabapentin or pregabalin and other anticonvulsants for the treatment of thalamus hemorrhagic post-stroke pain has drug resistance, the cause of drug resistance may be related to off-target effect, such as experiments have shown that the expression of gabapentin target molecule α2δ1 increased in the 1-2 weeks after thalamus hemorrhage, but the expression downs returned to normal levels at weeks 3-4, so the drug had the best effect in the 1-2 weeks after thalamus hemorrhage, but the efficacy weakened from the 3rd week, had to be increased, and the side effects such as large dizziness and drowsiness of the drug dose were significant.

Based on previous data analysis, the authors found that thalamus hemorrhage injury can not only upregulate the expression of α2δ1 molecular protein in the thalamus, but also upregulate the expression of α2δ1 in the dorsal horn of the bilateral spinal cord, suggesting secondary injury at the spinal cord level[9]; In addition, CPSP-associated pain sensitivity (i.

In order to explore the answers to these questions, the research group based on the theoretical knowledge of retrograde axonal degeneration (neural axonal degeneration), assuming that after thalamus hemorrhage, in addition to causing local bleeding foci and peripheral neuron death, it must also lead to injury to the spinothalamic tract (STT) neurons originating in the dorsal horn of the spinal cord (STT), then the injured axon terminal will undergo retrograde ulceration along the nerve fibers Eventually, secondary degenerative necrosis occurs in neuronal cells and protrusions in the dorsal horn of the spinal cord, and the neural tissues of transgender necrosis cause secondary neuroinflammation, and secondary nerve inflammation of the dorsal horn of the spinal cord sensitizes the surviving neurons involved in the dorsal horn of the spinal cord - the flexor circuit of the anterior horn of the spinal cord, which in turn causes pain sensitivity

On August 12, 2022, Professor Chen Jun's research group at the Second Affiliated Hospital of the Air Force Military Medical University published a paper entitled "Secondary damage and neuroinflammation in the spinal dorsal horn mediate post-thalamic haemorrhagic stroke" at Frontiers in Molecular Neuroscience Pain hypersensitivity: SDF1-CXCR4 signaling mediation" research paper proposes that secondary nerve injury and inflammation in the dorsal horn of the spinal cord are involved in regulating the occurrence

Previous studies have shown that region-specific hemorrhagic strokes produced by intrathalamus injection of collagenase type IV (ITC) can lead to bilateral mechanical hyperalgesia in rats, severe neuroinflammatory responses at the primary site of injury to the thalamus, and neuronal fatal injury [8-12].

Figure 1 Experimental flowchart and hypothesis diagram

(Source: Liang T.

Figure 2 The pro-apoptotic factor and TUNEL-labeled neurons in rats in the CPSP group were increased
.

(Source: Liang T.
et al.
, Front Mol Neurosci, 2022)

Using immunofluorescence labeling and the Western Blot method, the authors observed that 7 days after ITC injection, the apoptosis inhibitor Bcl-2-labeled neurons in the dorsal horn of the rat spinal cord decreased, while the pro-apoptotic factor Bax and TUNEL-labeled neurons increased significantly, while intrathalamus saline injection (ITS) did not have this effect, and protein quantification also verified these changes (Figure 2).


Electron microscopy observed that 7 days after ITC injection, there was significant neuronal degeneration and synaptic loss in the dorsal horn of the spinal cord, while there was no significant change in the ITS group (Figure 3
).

Multi-electrode arrays at the dorsal horn of the spinal cord (multi-electrode arrays containing 2 tetrode x 4 = 32 channels) were recorded and found that three different types of neurons recorded—low-threshold mechanoreceptivity (LTM), broad dynamic threshold (WDR), and injury-specific susceptibility (NS) neurons—were lost (Figure 4
).

These results suggest secondary death of primary thalamus neurons due to thalamus hemorrhagic stroke and neuroinflammation through retrograde ulceration of STT
.

Figure 3 The dorsal horn of the spinal cord of rats in the CPSP group had significant neuronal degeneration and synaptic structure loss
.

(Source: Liang T.
et al.
, Front Mol Neurosci, 2022)

Figure 4 The spinal cord dorsal horn LTM, WDR, and NS neurons in the CPSP group were all lost
.

(Source: Liang T.
et al.
, Front Mol Neurosci, 2022)

Then, the team tested whether retrograde ulceration caused by thalamus hemorrhagic stroke induced secondary neuroinflammation
of the dorsal horn of the spinal cord.

The results showed that after 7 days of ITC injection, Iba-1 (microglia biomarker), GFAP (astrocyte biomarker) and chemokinE SDF-1 and its receptor CXCR4-labeled cells increased significantly on the dorsal side of the spinal cord (Figure 5A-B), compared with no such response
in the ITS group.

CXCR4 co-localization with NeuN-, Iba1-, and GFAP-expression in the dorsal horn of the spinal cord is approximately 40%-50% (Figure 5C-D
).

Western blot analysis also showed a significant increase in protein expression levels in the bilateral dorsal horns of the spinal cord Iba1 and GFAP, as well as SDF1 and CXCR4 (Figure 5E-F
).

These results confirm secondary neuroinflammation
of the dorsal horn of the spinal cord after primary thalamus hemorrhagic stroke.

Figure 5 Significant secondary neuroinflammation
occurred in the dorsal horn of the spinal cord in rats in the CPSP group.

(Source: Liang T.
et al.
, Front Mol Neurosci, 2022)

To verify the presence of central sensitization in spinal cord dorsal horn neurons in living CPSP rats
.

MeA record analysis found that the three spinal cord dorsal horn units (LTM, WDR, NS) in the ITS and ITC groups can be divided into two types, namely self-generating (ITC vs.
ITS: 46 vs.
275) and non-self-generating (ITC vs.
ITS: 121 vs.
279) (Figure 6A-B).


In non-auto-emitting neurons in the IT or ITC group, the frequency distribution probability of the impulse response of LTM, WDR, and NS neurons to brush, pressure, and pinch stimulation is shown
in Figure 6C.

The peak response frequencies of LTM, WDR, and NS neurons were all related to the intensity of the stimulus, and under the stimulation of high-intensity mechanical stimulation (pressure and squeeze), the PEAK response frequency of the ITC group to WDR, NS and LTM was higher than that of the ITS group
.

These results suggest that there is a central sensitization effect in the surviving non-spontaneous spinal cord dorsal horn neurons under the neuroinflammatory response induced by secondary nerve injury
.

Figure 6 Superexcitation
occurs in residual neurons in the dorsal horn of the spinal cord in rats in the CPSP group.

(Source: Liang T.
et al.
, Front Mol Neurosci, 2022)

Finally, the group observed the pharmacological effects of intrathecal microglia inhibitors (MC), astrocyte inhibitors (luorocitrate, FC) and CXCR4 antagonists (AMD3100) on the expression levels of spinal dorsal horns Iba1, GFAP, SDF-1 and CXCR4 proteins by immunoprotechnic blotting and behavioral testing
.

The results showed that intrathecal injection of MC, FC, and AMD on the 10th day after ITC or ITS injection reversed the high expression of dorsal Iba1, GFAP, SDF1, and CXCR4 on the dorsal side of the spinal cord (Figure 7) and alleviated mechanical hyperalgesia caused by ITC (Figure 8
).

The same intrathecal injection treatment did not change the basal threshold of mechanical stimulation in the ITS group (Figure 8
).

Figure 7 Pharmacological effects
of intrathecal injection of MC, FC, and AMD3100 on the expression levels of Iba1, GFAP, SDF-1, and CXCR4 proteins in the dorsal horns of the spinal cord.

(Source: Liang T.
et al.
, Front Mol Neurosci, 2022)

Fig.
8 Effect of intrathecal injection of MC, FC and AMD3100 on the behavior of pain in rats
.

(Source: Liang T.
et al.
, Front Mol Neurosci, 2022)

In summary, the study combined with histoimity immunofluorescence, electron microscopy observation, immunoproteching, in vitro multi-electrode array extracellular recording, and intrathecal administration to explore the potential mechanism
of post-stroke pain (CPSP) C.

Found: in situ lesions of the thalamus can lead to secondary neuronal death and neuroinflammation in the dorsal horns of the bilateral spinal cord; And SDF1-CXCR4 signaling plays a key role in mediating secondary neuroinflammation at the spinal cord site; In addition, secondary neuronal injury also induces hyperexcitation of spinal cord dorsal horn neurons, thereby participating in the maintenance of CPSP mechanical pain sensitivity; Finally, it was confirmed that the mechanical pain sensitivity of CPSP can be alleviated by anti-neuroinflammation drug intervention, which provides theoretical support
for the primary sensory input segment as a new target for clinical treatment of CPSP.

Of course, in order to more intuitively observe the primary lesion of the thalamus can induce secondary injury of the spinal cord, the trace of axon retrograde ulcer between the thalamus and the spinal cord needs to be further verified
.

In summary, this series of experimental evidence suggests that primary injury to the thalamus can induce secondary neuronal death and neuroinflammation in the dorsal horn of the spinal cord through the retrograde degeneration process of the STT axon, which may be another important site for
the clinical treatment of CPSP.

Link to the original article: https://doi.
org/10.
3389/fnmol.
2022.
911476

The study is supported by the National Natural Science Foundation of China (31771159) and part of the Luzhou Municipal People's Government and the Southwest Medical University Science and Technology Strategic Cooperation Project (No
.
2019LZXNYDJ38).

Corresponding Author: Professor Chen Jun

(Photo courtesy of Professor Chen Jun's lab)

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[1] Mol Psychiatry-Wei Zheng/Fang Liao team collaborated to reveal vascular cognitive impairment and early events of white matter lesions in dementia

【2】Cell Rep | Xing Dajun's research group published a paper to reveal the differences between the primary visual cortex of macaque monkeys and the convolutional neural network integrating visual spatial information

[3] Neuron| Hu Yang's group revealed a large number of genes that promote optic nerve regeneration and their significant neuroprotective effect in glaucoma mice

[4] Cereb Cortex-Zhang Yuxuan research group reveals neurophysiological evidence for speech processing of task modulation

[5] Trends Cogn Sci—Qiu Jiang's team writes creative problem-solving perspective articles in knowledgeable fields

[6] Front Aging Neurosci—Yu Hongmei's team constructed a hierarchical multi-classification diagnostic framework for AD based on imaging features and clinical information

[7] The J Neurosci-Jin Mingyue/Guangchang Shinji team reveals that during brain development α-Syn and tau play an important synergistic physiological function

[8] Cell Death Differ — Xia Xiaobo's team revealed for the first time the correlation between "iron death" and the pathogenesis of glaucoma

[9] Sci Adv—Zeng Kewu/Tu Pengfei team revealed a new target for the regulation of neuroinflammation by the active ingredient of wild horses in Traditional Chinese medicine

[10] JNE—High-resolution time-frequency analysis estimates the functional connections of the brain in the EEG, helping to diagnose Alzheimer's disease

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References (swipe up and down to read)

(1)  GBD 2019 Stroke Collaborators (2021).
Global, regional, and national burden of stroke and its risk factors, 1990-2019: a systematic analysis for the global burden of disease study 2019.
Lancet Neurol.
20, 795-820.
doi:10.
1016/S1474-4422(21)00252-0.

(2)  Owolabi, M.
O.
, Thrift, A.
G.
, Mahal, A.
, Ishida, M.
, Martins, S.
, Johnson, W.
D.
, et al.
(2022).
Primary stroke prevention worldwide: translating evidence into action.
Lancet Public Health 7, e74-e85.
doi:10.
1016/S2468-2667(21)00230-9.

(3)  Scholz, J.
, Finnerup, N.
B.
, Attal, N.
, Aziz, Q.
, Baron, R.
, Bennett, M.
I.
, et al.
(2019).
The IASP classification of chronic pain for ICD-11: chronic neuropathic pain.
Pain 160, 53-59.
doi: 10.
1097/j.
pain.
0000000000001365.

(4)  Klit, H.
, Finnerup, N.
B.
, and Jensen, T.
S.
(2009).
Central post-stroke pain: clinical characteristics, pathophysiology, and management.
Lancet Neurol.
8, 857-868.
doi:10.
1016/S1474-4422(09)70176-0.

(5)   Kumar, B.
, Kalita, J.
, Kumar, G.
, and Misra, U.
K.
(2009a).
Central poststroke pain: a review of pathophysiology and treatment.
Anesth.
Analg.
108, 1645-1657.
doi:10.
1213/ane.
0b013e31819d644c.

(6)  Liampas A, Velidakis N, Georgiou T, et al.
Prevalence and Management Challenges in Central Post-Stroke Neuropathic Pain: A Systematic Review and Meta-analysis.
Adv Ther, 2020, 37:3278-3291.
doi: 10.
1007/s12325-020-01388-w

(7)  Choi, H.
R.
, Aktas, A.
, and Bottros, M.
M.
(2021a).
Pharmacotherapy to manage central post-stroke pain.
CNS Drugs 35, 151-160.
doi:10.
1007/s40263-021-00791-3.

(8)   Yang, F.
, Fu, H.
, Lu, Y.
F.
, Wang, X.
L.
, Yang, Y.
, Yang, F.
, et al.
(2014).
Post-stroke pain hypersensitivity induced by experimental thalamic hemorrhage in rats is region-specific and demonstrates limited efficacy of gabapentin.
Neurosci.
Bull.
30, 887-902.
doi:10.
1007/s12264-014-1477-5.

(9)   Yang, Y.
, Yang, F.
, Yang, F.
, Li, C.
L.
, Wang, Y.
, Li, Z.
, et al.
(2016).
Gabapentinoid insensitivity after repeated administration is associated with down-regulation of the α2δ-1 subunit in rats with central post-stroke pain hypersensitivity.
Neurosci.
Bull.
32, 41-50.
doi:10.
1007/s12264-015-0008-3.

(10)   Yang, F.
, Luo, W.
J.
, Sun, W.
, Wang, Y.
, Wang, J.
L.
, Yang, F.
, et al, (2017a).
SDF1-CXCR4 signaling maintains central post-stroke pain through mediation of glial-neuronal interactions.
Front.
Mol.
Neurosci.
10, 226.
doi:10.
3389/fnmol.
2017.
00226.

(11)   Cai, W.
, Wu, S.
, Pan, Z.
, Xiao, J.
, Li, F.
, Cao, J.
, et al.
(2018).
Disrupting interaction of PSD-95 with nNOS attenuates hemorrhage-induced thalamic pain.
Neuropharmacology 141, 238-248.
doi:10.
1016/j.
neuropharm.
2018.
09.
003.

(12)   Huang, T.
, Fu, G.
, Gao, J.
, Zhang, Y.
, Cai, W.
, Wu, S.
, et al.
(2020).
Fgr contributes to hemorrhage-induced thalamic pain by activating NF-κB/ERK1/2 pathways.
JCI Insight 5, e139987.
doi:10.
1172/jci.
insight.
139987.

End of this article