Effects of cPKCβII/γ membrane translocation ischemic/hypoxic tolerance induced by morphine postconditioning in hippocampal slices

To determine whether or not morphine postconditioning can induce ischemic/hypoxic tolerance in neurons subjected to reperfusion injury after oxygen-glucose deprivation (OGD). Hippocampal slices of 400 µm thickness were prepared from healthy adult male BALB/c mice. The slices were incubated in oxygen-saturated ACSF without or with calcium, then were subjected to OGD for 20 min. After recovery, the samples were immersed in oxygenated artificial fluid for 2 hours in the presence or absence of morphine postconditioning at 3 µmol/L during the first 5 - 60 min. The assessment of slices injury was performed by a determination of the intensity of slice stain incubated with TTC (2% 2, 3, 5-triphenyltetrazolium chloride) and the leakage rate of LDH also evaluated. At the designated periods during incubation, some slices were immersed into liquid nitrogen for a later analysis of Western blot. The frozen slices were homogenized, sonicated and centrifuged to separate soluble and particulate proteins. 10% SDS-PAGE Western