In this review, we provide a description of the recent methods used for immunohistochemical staining of the human inner ear using formalin-fixed frozen, paraffin and celloidin-embedded sections. We also show the application of these immunohistochemical methods in auditory and vestibular endorgans microdissected from the human temporal bone. We compare the advantages and disadvantages of immunohistochemistry (IHC) in the different types of embedding media. IHC in frozen and paraffin-embedded sections yields a robust immunoreactive signal. Both frozen and paraffin sections would be the best alternative in the case where celloidin-embedding technique is not available. IHC in whole endorgans yields excellent results and can be used when desiring to detect regional variations of protein expression in the sensory epithelia. One advantage of microdissection is that the tissue is processed immediately and IHC can be made within 1 week of temporal bone collection. A second advantage of microdissection is the excellent preservation of both morphology and antigenicity. Using celloidin-embedded inner ear sections, we were able to detect several antigens by IHC and immunofluorescence using antigen retrieval methods. These techniques, previously applied only in animal models, allow for the study of numerous important proteins expressed in the human temporal bone potentially opening up a new field for future human inner ear research.

No abstract

No abstract

Glutamate plays an important role in the central nervous system as an excitatory neurotransmitter. However, its abundance can lead to excitotoxicity which necessitates the proper function of active glutamate transporters. The glutamate-aspartate transporter (GLAST) has been shown to exist and function within non-human cochlear specimens regulating the inner ear glutamate concentration. In this study, we examined micro-dissected human cochleas from formalin-fixed celloidin-embedded temporal bone specimens of three different types of patients (Meniere's disease, normal controls, and other otopathologic conditions) and examined the differential expression of GLAST in the spiral ligament of the basal, middle, and apical turns of the cochlea. Immunohistochemical staining was performed with polyclonal antibodies against GLAST and image analysis was carried out with the Image J analysis software. In contrast to other studies with non-human specimens, GLAST was expressed in the spiral ligament fibrocytes but was not detected in the satellite cells of the spiral ganglia or supporting cells of the organ of Corti in the human cochlea. Our data also showed that GLAST expression significantly differs in the basal and apical turns of the cochlea. Lastly, post-hoc analysis showed a difference in the GLAST immunoreactive area of patients with Meniere's disease when compared to that of patients with other otopathologic conditions—such as presbycusis or ototoxicity. These results may potentially lead to further understanding of different disease states that affect hearing.