normalization data transformation protocol
The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. ID_REF = VALUE = Signal ABS_CALL = indicating whether the transcript was present (P), absent (A), or marginal (M) DETECTION P-VALUE =
array scanning protocol
GeneChips were scanned using Molecular Dynamicfs GenePix4000A Lucida Module1(Amersham parmacia biotech Ludica base).
Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on Rat Expression Array using Affymetrix genechip hybridization Oven 640. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
Female Sprague-Dawley rats weighing between 180 g and 210 g (8–10 weeks old) were purchased and bread in a SPF graded clean room.
nucleic acid extraction protocol
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
sample treatment protocol
After general anesthesia with pentobarbital (30–40 mg/kg, i.p.) and the local administration of lidocaine (1%), an incision was made posterior to the left pinna near the external meatus. The left otic bulla was opened and the round window niche was infused with 3-NP (500 mM or 300 mM) dissolved in saline (pH adjusted to 7.4 with NaOH). Infusion with saline alone was used as a control. Following treatment, and before the wound was closed, a small piece of gelatin was placed onto the niche to keep the solution in place and to allow for head movement after the animals awoke. The right cochlea was surgically destroyed to avoid cross-hearing during the recording of auditory brain-stem responses. The second turn of the lateral wall (excluding the stria vascularis) was harvested under the dissection microscope.