J Neurophysiol (January 1, 2003). 10.1152/jn.00559.2002
Submitted on Submitted 15 July 2002; accepted in final form 12 September 2002

Dynamic Recording of Cell Death in the In Vitro Dorsal Vagal Nucleus of Rats in Response to Metabolic Arrest

 ¹II. Physiologisches Institut, Georg-August-Universität Göttingen, D-37073 Göttingen, Germany; and  ²Perinatal Research Centre, Departments of Physiology and Pediatrics, University of Edmonton, Edmonton, Alberta T6G 2S2, Canada

Müller, Michael and Klaus Ballanyi. Dynamic Recording of Cell Death in the In Vitro Dorsal Vagal Nucleus of Rats in Response to Metabolic Arrest. J. Neurophysiol. 89: 551-561, 2003. Anoxic/ischemic neuronal death is usually assessed in cell cultures or in vivo within a time window of 24 h to several days using the nucleic acid stain propidium iodide or histological techniques. Accordingly, there is limited information on the time course of such neuronal death. We loaded acute rat brain stem slices with propidium iodide for dynamic fluorometric recording of metabolic arrest-related cell death in the dorsal vagal nucleus. This model was chosen because dorsal vagal neurons show a graded response to metabolic inhibition: anoxia and aglycemia cause a sustained hyperpolarization, whereas ischemia induces a glutamate-mediated, irreversible depolarization. We found that the number of propidium iodide-labeled cells increased from 27% to 43% of total cell count within 1-7 h after preparation of slices. Compared with these untreated control slices, cyanide-induced anoxia (30 min) or aglycemia (1 h) did not cause further cell death, whereas 3-h aglycemia destroyed an additional 13% of cells. Ischemia (1 h) due to cyanide plus iodoacetate immediately labeled an additional 20% of cells, and an additional 48% of cells were destroyed within the following 3 h of postischemia. Continuous recording of propidium iodide fluorescence showed that loss of membrane integrity started within 25 min after onset of the ischemic depolarization and the concomitant intracellular Ca²⁺ rise. The results show that propidium iodide can be used to monitor cell death in acute brain slices. Our findings suggest that pronounced cell death occurs within a period of 1-4 h after onset of metabolic arrest and is apparently due to necrotic/oncotic mechanisms.