Isolation and Culture of Spiral Ganglion Cells

SGCs, including the SGN, were harvested from early postnatal (2–5 days) Sprague Dawley rats. The tissue preparation was performed in accordance with the German “Law on protecting animals” and with the Directive 86/609/EEC of the European Communities Council for protection of animals used for experimental purposes. Preparation and dissection of the spiral ganglia were previously described in detail.^{17 (link)} In brief, animals were decapitated, spiral ganglia prepared, collected, and the cells subsequently dissociated. Cell number was determined by Trypan blue exclusion assay in a Neubauer chamber. A cell number of 20 × 10⁴ SGC/mL was adjusted in SGC-medium. This SGC-medium based on Panserin 401 (Pan-Biotech) supplemented with insulin (8.7 µg/mL; Biochrom Ltd), penicillin (30 U/mL; Biochrom Ltd), glucose (0.15%; B. Braun Melsungen AG), PBS (Ca²⁺/Mg²⁺-free PBS; 0.172 mg/mL; Gibco by Life Technologies), HEPES-buffer solution (23.43 µM; Invit-rogen), and N2-supplement (0.1 µL/mL; Invitrogen).