Dorsal root ganglia from male Sprague-Dawley rats were obtained by surgical resection from adults 225 to 250 g in weight and processed/prepared as we have described previously.8 (link) Individual sections from 3 different rats were stained with antibodies specific to SRF (1:400; Cell Signaling Technologies22 (link)), AKAP79/150 (1:100; Santa Cruz Biotechnology9 (link)), and TRPV1 (1:2000; Neuromics [GP14103]17 (link)). Species-appropriate Alexa Fluor secondary antibodies (Molecular Probes at 1:200 dilution) were used to visualize TRPV1 (Alexa Fluor 488), AKAP79/150 (Alexa Fluor 633), and SRF (Alexa Fluor 568). Immunofluorescence was evaluated, and images were obtained with the use of a Nikon 90i microscope equipped with a C1si laser scanning confocal imaging system (Nikon, Melville, NY). Images were processed for illustration purposes with Adobe Photoshop CS2 (Adobe, San Jose, CA) and are representative of 3 individual animals.
For quantification, 8 DRG images were chosen randomly, and an unbiased observer quantified positively stained cells. Image J (NIH) was used for identifying positive expression against a background-quantified low threshold.
For quantification, 8 DRG images were chosen randomly, and an unbiased observer quantified positively stained cells. Image J (NIH) was used for identifying positive expression against a background-quantified low threshold.