The Optimum Concentration of N-Methyl D-Aspartate to Induce Dorsal Root Ganglion Neuron Activation through the N-Methyl D-Aspartate Receptor Pathway: Creating a Neuron Model For the in-vitro Study of Pain

Background: In the in-vitro study on chronic pain, the N-methyl D-Aspartate receptor (NMDAR) activation in the dorsal root ganglion (DRG) neuron became one of the most important mechanisms to activate the chronic pain pathways. NMDAR activation can be induced using an NMDAR agonist. No guidelines explain the NMDA optimum concentration to induce DRG neuron activation through the NMDAR pathway. This study aims to find the optimum concentration of NMDA to induce DRG neuron activation through the NMDAR pathway.
Materials and Methods: We treat DRG neuron culture derived from the F11 cell line with 10, 20, 40, 60, 80, and 100 µM NMDA. Phosphorylated extracellular signal-regulated kinase (pERK), an activated neuron biomarker, is measured using an immunocytochemistry assay as a neuron activation biomarker. We validate the NMDA optimum concentration by measuring intracellular Ca2+ level, mitochondrial membrane potential (Δψm), and cytosolic adenosine triphosphate (ATP) in the activated neuron. Those parameters are the downstream process following NMDAR activation and are related to neuron activity. Statistical analysis was performed using the One-Way ANOVA test with α=5%.
Results: We found that NMDA 80 µM significantly had the highest pERK intensity and showed the most optimum neuron activation. Validation tests show an increase in intracellular Ca2+ influx and Δψm. NMDA 80 µM also causes significant depletion in the cytosolic ATP concentration related to neuron activation. NMDA 80 µM induces neuron activation by increasing pERK, Ca2+ influx, Δψm, and cytosolic ATP depletion.
Conclusion: NMDA 80 µM is the optimum concentration to induce DRG neuron activation through the NMDA receptor pathway.