In this article, an overview of the technique and examples of our data demonstrate that our protocols produce excellent, reliable in situ hybridization of mRNAs and immunohistochemical localization of mRNA translation products in the temporal bone and brain stem. This in situ hybridization protocol has been used to localize eight different mRNAs, including four types of nicotinic acetylcholine receptor subunit mRNAs, actin mRNA, CGRP mRNA, and two mRNAs coding for gap junction proteins, in both paraffin-embedded and sectioned (6 microns) and cryostat sectioned (15 microns) temporal bones (unpublished data). In conclusion, in situ hybridization of mRNAs with single-stranded RNA probes allows screening of brain stem and temporal bone sections for the expression of specific genes, with cellular resolution, and this protocol gives reliable results for all mRNAs studied. Of great potential significance is the ability to assess variations in gene expression by in situ hybridization of specific mRNAs. It is expected that such variations accompany changes in the physiologic state of an organism, such as developmental, pathologic or experimentally induced, and that these variations in gene expression are important in understanding basic auditory processes at the cellular level.