Methods: Male 9-month-old SAMP8 mice were anesthetized with sodium pentobarbital and dental resin was applied to the upper molars to raise the vertical dimension of the bite (bite-raised condition). For in situ hybridization analysis, 14 d after surgery mice were placed in acrylic restrainers, and after a 1-h exposure to restraint stress, the mice were returned to their home cages for 1 h and then transcardially perfused with saline and 100 ml of 4% formaldehyde in 0.1 M phosphate buffer. The brains were removed and tissue sections were prepared on a sliding microtome. CRH cRNA probes labeled with digoxigenin were used. To amplify the digoxigenin signals, the probes were detected using an antidigoxigenin antibody conjugated to alkaline phosphatase, and diaminobenzidine was used as the chromagen. The sense controls showed no hybridization signals. All CRH mRNA signals were counted unilaterally.
Results: Restraint stress significantly increased CRH mRNA signals in the PVN of both control mice and bite-raised mice compared to non-restraint stress control mice. Furthermore, bite-raised mice had significantly more CRH mRNA signals than non-bite raised control mice.
Conclusions: This study suggested that long-term occlusal-disharmony increased the stress response to novel-acute stress in the PVN in aged SAMP8 mice. Occlusal disharmony might increase susceptibility to novel-acute stress.