As the first neural element in the auditory pathway, neurons in the spiral ganglion shape the initial coding of sound stimuli for subsequent processing. Within the ganglion, type I and type II neurons form divergent and convergent innervation patterns, respectively, with their hair cell sensory receptors, indicating that very different information is gathered and conveyed. Layered onto these basic innervation patterns are structural and electrophysiological features that provide additional levels of processing multifaceted sound stimuli. To understand the nature of this additional complexity of signal coding, we characterized the distribution of calretinin and calbindin, two regulators of intracellular calcium that serve as markers for neuronal subpopulations. We showed in acute preparations and in vitro that calretinin and calbindin staining levels were heterogeneous. Immunocytochemical analysis of colocalization further showed that high levels of staining for the two molecules rarely overlapped. Although varied amounts of calbindin and calretinin were found within each tonotopic location and neuronal type, some distinct subdistributions were noted. For example, calretinin levels were highest in neurons innervating the midcochlea region, whereas calbindin levels were similar across the entire ganglion. Furthermore, we noted that apical type II neurons, identified by antiperipherin labeling, had significantly lower levels of calretinin and higher levels of calbindin. We also established that the endogenous firing feature of onset tau of the subthreshold response showed a pattern related to quantified calretinin and calbindin staining levels. Taken together, our results suggest an additional dimension of complexity within the spiral ganglion beyond that currently categorized. J. Comp. Neurol. 522:2299–2318, 2014. © 2014 Wiley Periodicals, Inc.