Ann Rehabil Med.  2020 Oct;44(5):343-352. 10.5535/arm.19202.

Sequential Activation of AMPA Receptors and Glial Cells in a Pain Model of Lumbar Spine Disc Herniation

• Kwak DG
• Lee DG

• KMID: 2508459
• DOI: http://doi.org/10.5535/arm.19202

Abstract

Objective
To investigate the glial cell and AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor activity after surgery for disc herniation pain model.
Methods
In total, 83 Sprague-Dawley rats were randomly assigned to the following groups: control (n=16), sham-operated (n=4), rats for pain behavior evaluation (n=3), nucleus pulposus-exposed groups for AMPA receptors (n=30), and glial cell (n=30). The rats were tested for mechanical allodynia; immunohistochemical staining for AMPA receptors (GluA1 and GluA2) and glial cells (OX-42 and glial fibrillary acid protein [GFAP]) in the spinal dorsal horn was performed on postoperative days 3, 7, and 14.
Results
Mechanical withdrawal thresholds decreased after surgery, and this effect was maintained for up to 14 days. Immunohistochemical expression of GluA1 and GluA2 in the spinal dorsal horn had increased quantitatively on postoperative days 3 and 7 (p<0.05) to levels similar to that of the controls on postoperative day 14. Moreover, immunohistochemical expression of OX-42 and GFAP showed similar changes to AMPA receptors after surgery. Although the activity of AMPA receptors and glial cells achieved normalcy, the mechanical withdrawal threshold of the hind paw remained decreased 38 days after surgery.
Conclusion
The rat model of lumbar disc herniation showed increased expression of AMPA receptor and glial cell activity in the spinal dorsal horn 3 and 7 days after surgery, which deceased to control levels at 14 days. The AMPA receptors and glial cell activations showed similar patterns after disc herniation surgery.

Keyword

AMPA receptors; Intervertebral disc displacement; Spinal cord dorsal horn; Inflammation; Synaptic plasticity

Figure

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  Fig. 1. Study flow schematic for immunohistochemical examination. AMPA, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; GFAP, glial fibrillary acidic protein; NP, nucleus pulposus; POD, postoperative day.

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  Fig. 2. In the nucleus pulposus-exposed group, the left L5 nerve roots were exposed after hemilaminectomy and facetectomy (A) and the nucleus pulposus harvested from coccygeal disc was implanted in the exposed nerve roots without mechanical compression (B).

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  Fig. 3. Time course of neuropathic pain behaviors. Hyperalgesia in a rat model of lumbar disc herniation induced by implantation of the nucleus pulposus to the left L5 nerve root. The withdrawal threshold in the ipsilateral paw is noted at 3 to 38 days following surgery. D, postoperative day. *p<0.05 versus the baseline.

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  Fig. 4. Immunohistochemical expression of GluA1 and GluA2 in the dorsal horn of the L5-level spinal cord before (control), 3, 7, and 14 days after surgery. GluA1 are GluA2 are intensely stained in the superficial lamina of the dorsal horn. POD, postoperative day.

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  Fig. 5. Semi-quantitative analysis of immunoreactivity of AMPA receptors and microglial cells in the dorsal horn of the spinal cord at postoperative day (POD) 3, 7, and 14. (A) Mean green value of GluA1 and GluA2 is significantly higher at POD3 and POD7, compared with control. (B) Immunoreactivity of OX-42 and GFAP are also increased and return to the control value at 14 days after surgery, with a pattern similar to that of AMPA receptors. For quantification, values and numbers were counted and averaged. Values are presented as mean±standard error. AMPA, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; GFAP, glial fibrillary acidic protein. *p<0.05 versus the control group.

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  Fig. 6. Immunohistochemical expression of OX-42 and glial fibrillary acidic protein (GFAP) in the dorsal horn of the L5-level spinal cord before (control), 3, 7, and 14 days after surgery. Increased OX-42-positive microglia are noted in superficial ipsilateral dorsal horns at 3 and 7 days following surgery compared with control. GFAP-positive astrocytes are also diffusely elevated in the spinal dorsal horn following surgery. POD, postoperative day.

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