Dnaase type 1

Rats were anesthetized with CO₂ and then euthanized by decapitation. The NG were dissected in ice-cold Hank’s balanced salt solution. Thereafter, the ganglia were enzymatically dissociated in Earle’s balanced salt solution containing 0.6 mg/ml collagenase (Roche Diagnostics Corp. Indianapolis, IN), 0.4 mg/ml trypsin (Worthington Biochemical Corp. Lakewood, NJ), 0.1 mg/ml DNAase Type I (Sigma-Aldrich, St. Louis, MO) in a shaking water bath at 35°C for 65 min. Afterward, the neurons were centrifuged twice and the dispersed neurons were resuspended in minimal essential medium supplemented with 10% fetal bovine serum (VWR), 1% penicillin-streptomycin (all from Thermo-Fisher Scientific). The dissociated NG neurons were plated on poly-L-lysine coated dishes or coverslips and incubated (5% CO₂/95% air) at 37 °C overnight until experimentation. Power analysis and sample size calculations were performed with Sigmaplot 12.5 (Systat Software, San Jose, Ca). Based upon a 50% ± 10% expected difference with a power of 0.9 and alpha 0.05, a minimum of three animals were required for all experiments.

Rats were first anesthetized with CO₂ and then quickly euthanized by decapitation. The ganglia were dissected in ice-cold Hank’s balanced salt solution (HBSS). Thereafter, the ganglia were enzymatically dissociated in Earle’s balanced salt solution containing 0.6 mg/ml collagenase (Roche Diagnostics Corp. Indianapolis, IN), 0.4 mg/ml trypsin (Worthington Biochemical Corp. Lakewood, NJ), 0.1 mg/ml DNAase Type I (Sigma, St. Louis, MO) in a shaking water bath at 35°C for 45-min. The neurons were then dispersed by vigorous shaking, centrifuged twice for 6-min at 500 rpm. The dispersed neurons were resuspended in minimal essential medium supplemented with 10% fetal bovine serum (VWR), 1% penicillin-streptomycin (all from Thermo-Fisher Scientific). The dissociated NG neurons were plated on multiple poly-L-lysine coated dishes or coverslips and incubated (5% CO₂/95% air) at 37°C overnight until experimentation.