Hypothermia, Metabolic Stress, and NMDA-Mediated Excitotoxicity

Abstract: Isolated embryonic retinas were metabolically stressed by inhibition of glycolysis either with iodoacetate (IOA) or by glucose withdrawal plus 10 mM 2-deoxy-D-glucose, and the effects of hypothermia were examined. Incubation at 30 versus 37°C during 30 min of hypoglycemia with IOA completely reduced the rapid swelling-related GABA release 6 ± 2 vs. 68 ± 10 nmol/100 mg of protein (mean ± SEM) for 30 and 37°C, respectively]. Histology of the retina immediately following 30 min of metabolic stress at 30°C appeared normal, whereas that at 37°C showed a pattern of acute edema, characteristic of NMDA-mediated acute excitotoxicity. Coincubation with a competitive or noncompetitive NMDA antagonist, respectively, CGS-19755 (10 μM) or MK-801 (1 μM), during 30 min of hypoglycemia at 37°C completely prevented tissue swelling, whereas extracellular GABA content remained at basal levels, indicating that the cytotoxic effects of IOA treatment for 30 min at 37°C were NMDA receptor mediated. Longer periods of hypoglycemia at 37° C produced acute toxicity that was only partially NMDA receptor mediated. Hypothermia delayed the onset of NMDA-mediated toxicity by 30–60 min. At 30°C, the rate of loss of ATP was slowed during the first several minutes of hypoglycemia (82 and 58% of maximal tissue levels at 30 and 37° C, respectively, at 5 min), but by 10 min, ATP levels were comparably reduced. After a transient exposure of retina to 50 μM NMDA in Mg²⁺-free medium, hypothermia significantly attenuated acute GABA release by 30%. At 24 h of recovery, lactate dehydrogenase release was decreased by 37%. Hypothermia had no effect when the exposure was done in medium containing physiological concentrations of Mg²⁺. The above results suggest that the protective effect of hypothermia during the metabolic insult is predominately directed at the cellular events that lead up to NMDA receptor involvement. Reduction in the rate of loss of ATP, however, does not fully account for the delay in involvement of NMDA receptors during metabolic stress at 30°C. The attenuation of direct NMDA-mediated toxicity in Mg²⁺-free medium further suggests that decreased temperature may result in altered channel properties during situations when the Mg²⁺ block is lifted.