Limits...
Adenovector GAD65 gene delivery into the rat trigeminal ganglion produces orofacial analgesia.

Vit JP, Ohara PT, Sundberg C, Rubi B, Maechler P, Liu C, Puntel M, Lowenstein P, Castro M, Jasmin L - Mol Pain (2009)

Bottom Line: We postulated that GABA produced from the expression of GAD would reduce pain behavior by acting on GABA receptors on neurons within the ganglion.GABA receptors were not found on SGCs.Six days after injection of AdGAD65 into the trigeminal ganglion, there was a statistically significant decrease of pain behavior in the orofacial formalin test, a model of inflammatory pain.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anatomy, University of California San Francisco, San Francisco, CA 94143, USA. Jeanpvit@gmail.com

ABSTRACT

Background: Our goal is to use gene therapy to alleviate pain by targeting glial cells. In an animal model of facial pain we tested the effect of transfecting the glutamic acid decarboxylase (GAD) gene into satellite glial cells (SGCs) of the trigeminal ganglion by using a serotype 5 adenovector with high tropisms for glial cells. We postulated that GABA produced from the expression of GAD would reduce pain behavior by acting on GABA receptors on neurons within the ganglion.

Results: Injection of adenoviral vectors (AdGAD65) directly into the trigeminal ganglion leads to sustained expression of the GAD65 isoform over the 4 weeks observation period. Immunohistochemical analysis showed that adenovirus-mediated GAD65 expression and GABA synthesis were mainly in SGCs. GABAA and GABAB receptors were both seen in sensory neurons, yet only GABAA receptors decorated the neuronal surface. GABA receptors were not found on SGCs. Six days after injection of AdGAD65 into the trigeminal ganglion, there was a statistically significant decrease of pain behavior in the orofacial formalin test, a model of inflammatory pain. Rats injected with control virus (AdGFP or AdLacZ) had no reduction in their pain behavior. AdGAD65-dependent analgesia was blocked by bicuculline, a selective GABAA receptor antagonist, but not by CGP46381, a selective GABAB receptor antagonist.

Conclusion: Transfection of glial cells in the trigeminal ganglion with the GAD gene blocks pain behavior by acting on GABAA receptors on neuronal perikarya.

Show MeSH

Related in: MedlinePlus

GAD65-mediated analgesia is reversed by the GABAA receptor antagonist bicuculline. (A) Six days after injection of AdGAD65 into the trigeminal ganglion, a second injection was performed with saline, bicuculline or CGP46381 (a GABAB receptor antagonist), and the orofacial formalin test was carried out 10 minutes later. Bicuculline, but not CGP46381 reversed the AdGAD65-mediated analgesia in the second phase of the formalin test (RM ANOVA: F = 2.7; P < 0.001). AdGAD65 + bicuculline vs AdGAD65 + saline: **P < 0.01, ***P < 0.001. (B) Comparison of the averaged pain behavior in the second phase of the formalin test shows a significant difference between groups (one-way ANOVA: F = 15.6; P < 0.001). AdGAD65 produced a significany decrease in pain behavior. Bicuculline reverses AdGAD65-mediated analgesia. Bicuculline alone does not affect pain behavior. AdGAD65 or AdGAD65 + saline vs. AdGFP: ***P < 0.001; AdGAD65 or AdGAD65 + saline vs AdGFP: ###P < 0.001; AdGAD65 + saline vs. AdGAD65 + bicuculline: *P < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2734545&req=5

Figure 5: GAD65-mediated analgesia is reversed by the GABAA receptor antagonist bicuculline. (A) Six days after injection of AdGAD65 into the trigeminal ganglion, a second injection was performed with saline, bicuculline or CGP46381 (a GABAB receptor antagonist), and the orofacial formalin test was carried out 10 minutes later. Bicuculline, but not CGP46381 reversed the AdGAD65-mediated analgesia in the second phase of the formalin test (RM ANOVA: F = 2.7; P < 0.001). AdGAD65 + bicuculline vs AdGAD65 + saline: **P < 0.01, ***P < 0.001. (B) Comparison of the averaged pain behavior in the second phase of the formalin test shows a significant difference between groups (one-way ANOVA: F = 15.6; P < 0.001). AdGAD65 produced a significany decrease in pain behavior. Bicuculline reverses AdGAD65-mediated analgesia. Bicuculline alone does not affect pain behavior. AdGAD65 or AdGAD65 + saline vs. AdGFP: ***P < 0.001; AdGAD65 or AdGAD65 + saline vs AdGFP: ###P < 0.001; AdGAD65 + saline vs. AdGAD65 + bicuculline: *P < 0.05.

Mentions: The finding that adenovirus-mediated expression of GAD65 occurred principally in SGCs together with the presence of GABA receptor in the cell bodies of sensory neurons suggested that GABA might produce its antinociceptive effect inside the trigeminal ganglion in a paracrine manner. To test this possibility, we injected rats with AdGAD65, and 6 days later made a second injection into the trigeminal ganglion with either saline, bicuculline (a GABAA receptor antagonist) or CGP46381 (a GABAB receptor antagonist) followed 10 minutes later by the orofacial formalin test. When bicuculline (200 pmol in 2 μl) was injected into the trigeminal ganglion, GAD65-produced analgesia was reversed in the formalin test (Figure 5). The pain behavior observed from 12 to 28 min post-injection of formalin was higher after injection of bicuculline when compared to injection of saline (Figure 5A). On the contrary, the injection of CGP46381 (10 nmol in 2 μl) into the trigeminal ganglion had no effect on GAD65-mediated analgesia in the orofacial formalin test (Figure 5A). The reversion of GAD65-mediated analgesia by bicuculline was maximal as shown by the total pain behavior in the second phase of the formalin test (Figure 5B). In rats receiving AdGAD65 and bicuculline, the total pain behavior in the second phase of the formalin test (from 12 to 44 min) was similar to that from rats injected with either AdGFP or AdLacZ control adenoviruses (Figure 5B). Bicuculline alone in normal rats had no effect on the formalin test and the pain behavior in the second phase of the test was identical to that of control animals (Figure 5B).


Adenovector GAD65 gene delivery into the rat trigeminal ganglion produces orofacial analgesia.

Vit JP, Ohara PT, Sundberg C, Rubi B, Maechler P, Liu C, Puntel M, Lowenstein P, Castro M, Jasmin L - Mol Pain (2009)

GAD65-mediated analgesia is reversed by the GABAA receptor antagonist bicuculline. (A) Six days after injection of AdGAD65 into the trigeminal ganglion, a second injection was performed with saline, bicuculline or CGP46381 (a GABAB receptor antagonist), and the orofacial formalin test was carried out 10 minutes later. Bicuculline, but not CGP46381 reversed the AdGAD65-mediated analgesia in the second phase of the formalin test (RM ANOVA: F = 2.7; P < 0.001). AdGAD65 + bicuculline vs AdGAD65 + saline: **P < 0.01, ***P < 0.001. (B) Comparison of the averaged pain behavior in the second phase of the formalin test shows a significant difference between groups (one-way ANOVA: F = 15.6; P < 0.001). AdGAD65 produced a significany decrease in pain behavior. Bicuculline reverses AdGAD65-mediated analgesia. Bicuculline alone does not affect pain behavior. AdGAD65 or AdGAD65 + saline vs. AdGFP: ***P < 0.001; AdGAD65 or AdGAD65 + saline vs AdGFP: ###P < 0.001; AdGAD65 + saline vs. AdGAD65 + bicuculline: *P < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2734545&req=5

Figure 5: GAD65-mediated analgesia is reversed by the GABAA receptor antagonist bicuculline. (A) Six days after injection of AdGAD65 into the trigeminal ganglion, a second injection was performed with saline, bicuculline or CGP46381 (a GABAB receptor antagonist), and the orofacial formalin test was carried out 10 minutes later. Bicuculline, but not CGP46381 reversed the AdGAD65-mediated analgesia in the second phase of the formalin test (RM ANOVA: F = 2.7; P < 0.001). AdGAD65 + bicuculline vs AdGAD65 + saline: **P < 0.01, ***P < 0.001. (B) Comparison of the averaged pain behavior in the second phase of the formalin test shows a significant difference between groups (one-way ANOVA: F = 15.6; P < 0.001). AdGAD65 produced a significany decrease in pain behavior. Bicuculline reverses AdGAD65-mediated analgesia. Bicuculline alone does not affect pain behavior. AdGAD65 or AdGAD65 + saline vs. AdGFP: ***P < 0.001; AdGAD65 or AdGAD65 + saline vs AdGFP: ###P < 0.001; AdGAD65 + saline vs. AdGAD65 + bicuculline: *P < 0.05.
Mentions: The finding that adenovirus-mediated expression of GAD65 occurred principally in SGCs together with the presence of GABA receptor in the cell bodies of sensory neurons suggested that GABA might produce its antinociceptive effect inside the trigeminal ganglion in a paracrine manner. To test this possibility, we injected rats with AdGAD65, and 6 days later made a second injection into the trigeminal ganglion with either saline, bicuculline (a GABAA receptor antagonist) or CGP46381 (a GABAB receptor antagonist) followed 10 minutes later by the orofacial formalin test. When bicuculline (200 pmol in 2 μl) was injected into the trigeminal ganglion, GAD65-produced analgesia was reversed in the formalin test (Figure 5). The pain behavior observed from 12 to 28 min post-injection of formalin was higher after injection of bicuculline when compared to injection of saline (Figure 5A). On the contrary, the injection of CGP46381 (10 nmol in 2 μl) into the trigeminal ganglion had no effect on GAD65-mediated analgesia in the orofacial formalin test (Figure 5A). The reversion of GAD65-mediated analgesia by bicuculline was maximal as shown by the total pain behavior in the second phase of the formalin test (Figure 5B). In rats receiving AdGAD65 and bicuculline, the total pain behavior in the second phase of the formalin test (from 12 to 44 min) was similar to that from rats injected with either AdGFP or AdLacZ control adenoviruses (Figure 5B). Bicuculline alone in normal rats had no effect on the formalin test and the pain behavior in the second phase of the test was identical to that of control animals (Figure 5B).

Bottom Line: We postulated that GABA produced from the expression of GAD would reduce pain behavior by acting on GABA receptors on neurons within the ganglion.GABA receptors were not found on SGCs.Six days after injection of AdGAD65 into the trigeminal ganglion, there was a statistically significant decrease of pain behavior in the orofacial formalin test, a model of inflammatory pain.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anatomy, University of California San Francisco, San Francisco, CA 94143, USA. Jeanpvit@gmail.com

ABSTRACT

Background: Our goal is to use gene therapy to alleviate pain by targeting glial cells. In an animal model of facial pain we tested the effect of transfecting the glutamic acid decarboxylase (GAD) gene into satellite glial cells (SGCs) of the trigeminal ganglion by using a serotype 5 adenovector with high tropisms for glial cells. We postulated that GABA produced from the expression of GAD would reduce pain behavior by acting on GABA receptors on neurons within the ganglion.

Results: Injection of adenoviral vectors (AdGAD65) directly into the trigeminal ganglion leads to sustained expression of the GAD65 isoform over the 4 weeks observation period. Immunohistochemical analysis showed that adenovirus-mediated GAD65 expression and GABA synthesis were mainly in SGCs. GABAA and GABAB receptors were both seen in sensory neurons, yet only GABAA receptors decorated the neuronal surface. GABA receptors were not found on SGCs. Six days after injection of AdGAD65 into the trigeminal ganglion, there was a statistically significant decrease of pain behavior in the orofacial formalin test, a model of inflammatory pain. Rats injected with control virus (AdGFP or AdLacZ) had no reduction in their pain behavior. AdGAD65-dependent analgesia was blocked by bicuculline, a selective GABAA receptor antagonist, but not by CGP46381, a selective GABAB receptor antagonist.

Conclusion: Transfection of glial cells in the trigeminal ganglion with the GAD gene blocks pain behavior by acting on GABAA receptors on neuronal perikarya.

Show MeSH
Related in: MedlinePlus