Figure 1.

A–C, RT-PCR (A, B) and qRT-PCR (C) analysis of mAChR expression in wild-type mice and their compensatory upregulation/downregulation in knock-out mice (C ). M₂, M₄, and M₅ receptors were consistently expressed in the following: A, inner hair cells (IHCs) and spiral ganglion neurons (SGNs) but not outer hair cells (OHCs) harvested from immature cochleas (P11–P13); B, both organ of Corti and spiral ganglion from microdissected adult cochleas (4 – 6 weeks); and C, whole adult cochleas (2 years). M₃ receptors were present only in whole-cochlear digests (C ), and M₁ receptors were never amplified from wild types (A–C). All five mAChRs are expressed in adult brain (B, positive control). A, B, A glutamate receptor (GluR2), otoferlin, and oncomodulin (Oncomod.) primers were used as markers for spiral ganglion neurons, inner hair cells, and hair cells, respectively. All bands appear at the location expected for the size of the amplicon (see Table 1), as calibrated by the ladder lane in each gel: dashed lines are positioned at amplicon sizes of 100, 300, 500, and 700 bp. The 8-bit gel micrographs were adjusted digitally: after inversion, offset was set to 0 by subtracting the mean pixel value from an empty lane, and gain was optimized by setting to 256 the mean pixel value from the 500 bp ladder band. Primer bands (<100 bp) were cut from graphs. C, Mean expression levels for mAChR mRNA are normalized to 18S rRNA, after adjusting for primer efficiency, as described previously (Stankovic and Corfas, 2003). Error bars represent SEMs.