Fig. 8.
Up-regulation of d-amino acid oxidase (DAAO) immunostaining expressed in astrocytes in the spinal cord after morphine tolerance. Rats were injected twice daily for 7 days either with morphine (10 mg/kg) or equal volume of saline. The spinal cords (L3–5 segments) were obtained, and double immunofluorescence was labeled with DAAO and the astroglial marker fibrillary acidic protein (GFAP). The measurement of GFAP (A)-, DAAO (B)-, and double DAAO/GFAP (C)-immunolabeled surface areas was quantified in the spinal dorsal horn (laminae I–V) using Image J computer program (National Institutes of Health, Bethesda, MD). The average percentage of immunolabeled surface area was the fraction of the positive immunofluorescent surface area of total measured area in the picture from three sections of each spinal cord (six animals in each group). Data are presented as means ± SEM. *Denotes statistically significant difference from the saline-treated group (P < 0.05 by one-way ANOVA followed by post hoc Student–Newman–Keuls test). Representative photomicrographs of GFAP (D, G)-, DAAO (E, H)-, and double GFAP/DAAO (F, I)-immunofluorescent staining of the left spinal cord were obtained from the naïve and morphine-tolerant rat. Scale bar: 400 μm.