Dorsal root ganglion neuron cultures
Lumbar DRGs were dissected from 100 g female Sprague-Dawley rats using procedures as described previously (Gomez et al., 2022). DRGs were excised and placed in sterile DMEM (Cat# 11965; Thermo Fisher Scientific, Waltham, MA). The ganglia were dissociated enzymatically with collagenase type I (5 mg/mL, Cat# LS004194; Worthington) and neutral protease (3.125 mg/mL, Cat# LS02104; Worthington, Lakewood, NJ) for 50 minutes at 37°C under gentle agitation. The dissociated cells were then centrifuged (800 rpm for 5 min) and resuspended in DMEM containing 1% penicillin/streptomycin sulfate (Cat# 15140, Life Technologies, Carlsbad, CA), 10% fetal bovine serum [HyClone]) and 30 ng/mL nerve growth factor (Cat# N2513, Millipore Sigma, St. Louis, MO). The cells were seeded on poly-D-lysine (Cat# P6407, Millipore Sigma, St. Louis, MO) and laminin (Cat#sc-29012, Santa Cruz Biotechnology, Dallas, TX) -coated 12- or 15-mm glass coverslips and incubated at 37°C. All cultures were used within 48 hours.