Fig 2: Expression patterns of TRIM54, TTN, NCALD, and INA in the spiral ganglion neurons. (A) Expression pattern of TRIM54 in the spiral ganglion neuron. TRIM54 was observed in PRPH-positive Type II spiral ganglion neurons at P0 cochlea; no expression was detected in Type I spiral ganglion neurons. (B and C) Expression pattern of TTN in spiral ganglion neuron in E120 and P0 spiral ganglion neurons. In E120, TTN expression was broadly observed in spiral ganglion neurons. Its expression was detected in both Type I and II neurons. In P0 spiral ganglion neurons, still-board TTN expression could be detected in both Type I and II spiral ganglion neurons. (D) Expression pattern of NCALD in the spiral ganglion neuron. NCALD expression was broadly observed in Type I spiral ganglion neurons at P0 cochlea, and no expression was detected in PRPH-positive type II spiral ganglion neurons. (E) Expression pattern of INA in the spiral ganglion neuron. INA expression is observed in PRPH-positive Type II spiral ganglion neurons at P0 cochlea, and no expression was detected in Type I spiral ganglion neurons. The nuclei were counterstained with Hoechst (blue). Scale bar: 20 μm.

Fig 3: Cochlear neuronal development in E70 and E77. (A-E) Cochleovestibular ganglion in E70 cochlea. Expression of TUBB3, ISL1, and NEUROD1 was observed at E70 (A). Expression of POU4F1, SOX2, and PRPH was also observed in E70 cochleovestibular ganglion (B, C, and D). However, expression of MAFB and GATA3 was not detected in E70 cochleovestibular ganglion (D and E). (F-L) Spiral ganglion in E77 cochlea. Expression of TUBB3 and PRPH was observed in E77 spiral ganglion neurons (F). In E77, S100B-positive glial cells surrounding spiral ganglion neurons were observed (F). In E77, expression of NEUROD1 was observed in only an apical turn (asterisk in G), while RBFOX3 expression was observed in basal turns (H). No RBFOX3 expression was observed in the NEUROD1-positive spiral ganglion neurons (I). Expression of GATA3, ISL1, and MAFB was observed in the spiral ganglion neurons in E77 (J-L). The nuclei were counterstained with Hoechst (blue). Scale bar: 50 µm in A-E, G, and I, 100 µm in F, H, and J-L

Fig 4: Myelination of spiral ganglion neurons at W22.(A–E) Confocal images of a spiral ganglion in the middle turn of a cochlea at W22 showing DAPI (A), TUBB3 (B), PRPH (C), MBP (D) and the merged image (E). The spiral ganglion is delineated by the dotted line. (F–J) Confocal images of an axial transection of the cochlear nerve at W22 showing DAPI (F), TUBB3 (G), PRPH (H), MBP (I) and the merged image (J). (K–O) Deconvoluted confocal images of an axial transection of the cochlear nerve at W22 showing DAPI (K), TUBB3 (L), PRPH (M), MBP (N) and the merged image (O). Insets show TUBB3 (left), PRPH (middle) and the merge with MBP and DAPI (right) in high-magnifications examples of PRPH−/TUBB3+/MBP+ cochlear nerve fibers (upper inset) and PRPH+/TUBB3+/MBP+ cochlear nerve fibers (lower inset). Scale bar = 20 µm or 1 µm (insets in O).

Fig 5: Neonatal spiral ganglion neurons in the common marmoset can be divided into subtypes. (A) Low-magnification image of P0 cochlea. (B–D) CALB2 expression in the organ of Corti of the common marmoset. High expression of CALB2 was detected in the Type I spiral ganglion neurons located on the pillar side (Arrowhead in C); low expression of CALB2 was observed in the neurite located in the modiolar side neurite, which showed no CALB2 expression and pillar side CALB2-high neurites (Arrow in C). A higher magnification image of the bracket in A' was shown in B. No expression was observed in the PRPH-positive type II neurons (D). (E) CALB2 expression in the spiral ganglion neurons in the Rosenthal canal. At this level, CALB2 expression was observed in the part of the spiral ganglion neurons, and neurons could be subdivided into four types: CALB2 High neurons (Arrowhead in E'), CALB2 Low neurons, RBFOX3 positive neurons (Arrows in E), and PRPH positive neurons (* in E"’’). (F) A schema of Type I spiral ganglion neuron subtypes. Spiral ganglion neurons in the common marmoset could be subdivided into Type IA, Type IB, Type IC, and Type II. The nuclei were counterstained with Hoechst (blue). Scale Bar: 100 μm in A, 10 μm in B, and C, 20 μm in D and E. SGN: spiral ganglion neurons, OC: organ of Corti.