IADR Abstract Archives
Nociceptor Sensitization Association With SRF-Dependent Up-Regulation
Objectives: The objective of this project is to determine whether metabotropic glutamate receptor type 1/5 (mGluR1/5) activation up-regulates Serum Response Factor (SRF) phosphorylation in cultured sensory neurons. SRF is linked to the transcription of a number of neuronal genes implicit in primary afferent sensitivity. SRF phosphorylation by protein kinase C (PKC), a kinase downstream of mGluR1/5, drives its activation, translocation to the nucleus, and positive transcriptional effects. Interestingly, the gene for A-Kinase Anchoring Protein 79/150 (AKAP) is positively regulated by SRF. The AKAP scaffolding protein is necessary for sensitization of several ligand-gated ion channels responsible for transducing painful somatosensory information. Therefore, we evaluated whether the continued activation of mGluR1/5 could drive SRF phosphorylation, which in turn, would drive transcriptional up-regulation of AKAP to support chronic pain phenotypes.
Methods: Trigeminal Ganglia (TG) were resected form rats and cultured for experimental manipulation. Neuronal cultures were treated with either vehicle or the mGluR1/5 agonist DHPG, in the presence/ absence of the PKC inhibitor GFX. Cell lysates were collected and analyzed for protein expression by Western blot. Immunoreactive band densitometry was conducted on enhanced chemiluminescent images to quantify changed in expression, and values were averaged between 6 individual trails to ascertain a mean and SEM. Statistical significance was determined using on-way ANOVA with Bonferroni correction.
Results: DHPG application to cultured sensory neurons stimulated a significant increase in SRF phosphorylation, resulting in an increase in AKAP expression. There effects were blocked following PKC inhibition by GFX co-treatment.
Conclusions: In conclusion, our results support the downstream signaling hypothesis, and support additional experiments designed to determine how mGluR1/5 coordinates the maintenance of persistent pain phenotypes. The knowledge gained from this research will help lead to the development of improved therapeutic strategies to treat chronic pain.
Methods: Trigeminal Ganglia (TG) were resected form rats and cultured for experimental manipulation. Neuronal cultures were treated with either vehicle or the mGluR1/5 agonist DHPG, in the presence/ absence of the PKC inhibitor GFX. Cell lysates were collected and analyzed for protein expression by Western blot. Immunoreactive band densitometry was conducted on enhanced chemiluminescent images to quantify changed in expression, and values were averaged between 6 individual trails to ascertain a mean and SEM. Statistical significance was determined using on-way ANOVA with Bonferroni correction.
Results: DHPG application to cultured sensory neurons stimulated a significant increase in SRF phosphorylation, resulting in an increase in AKAP expression. There effects were blocked following PKC inhibition by GFX co-treatment.
Conclusions: In conclusion, our results support the downstream signaling hypothesis, and support additional experiments designed to determine how mGluR1/5 coordinates the maintenance of persistent pain phenotypes. The knowledge gained from this research will help lead to the development of improved therapeutic strategies to treat chronic pain.