Neonatal (P7) brain hypoxia-ischemia (HI) induces intracellular Bax protein shifts to the nucleus, mitochondria, and endoplasmic reticulum (ER), where it triggers the activation of the respective cell death signaling cascades. When compared with HI-treated rat pups, 100% D2 resuscitation of HI-treated rat pups increases HI-induced ER Bax levels, ER-mediated cell death signaling, and resultant lesion volume and inflammation due to increased necrotic-like cell death. To better characterize the role of Bax intracellular shuttling ER cell death signaling and necrotic-like cell death, we used rotenone-treated P5 neuronal cortical cultures to increase ER Bax levels and subsequent cell death signaling. We treated P5 primary cortical neurons with 25 μM and 100 μM rotenone as an apoptotic or necrotic-like stimulus, respectively, and measured intracellular organelle Bax levels and the subsequent activation of ER/mitochondrial cell death signaling. The 25 μM rotenone treatment promptly increased nuclear Bax levels followed by a later increase in mitochondrial Bax levels and caspase-mediated cleavage of α-fodrin. The 100 μM rotenone treatment also resulted in an early increase in nuclear Bax levels followed by a subsequent increase in ER Bax levels and calpain-mediated cleavage of α-fodrin. After pretreatment with the immunosuppressive and neuroprotective FK506, there was a delay in Bax intracellular shifts and cell death signaling for both the 25 and 100 μM rotenone treatments. These results suggest that the different outcomes of apoptotic-like vs. necrotic-like cell death resulting from the treatment of neuronal cultures with rotenone at 25 and 100 μM rotenone reflect changes in the intracellular trafficking of Bax among different organelles.
ASJC Scopus subject areas
- Cellular and Molecular Neuroscience