The nature and time course of neuronal vacuolation induced by the N-methyl-D-aspartate antagonist MK-801

Abstract

N-Methyl-D-aspartate (NMDA) antagonists cause neuronal vacuolation in the posterior cingulate and retrosplenial cortex of the rat. Because the nature of neuronal pathologic changes due to NMDA antagonists may affect the potential clinical use of this class of drugs, we undertook experiments to define the nature and time course of the vacuolation caused by high-dose (5 mg/kg) MK-801 (dizocilpine, 5-methyl-10, 11-dihydro-5H-dibenzo [a,d]cyclohepten-5,10-imine). Ultrastructural examination revealed the vacuoles to be not a form of hydropic cellular degeneration, but rather a dilatation of several intracellular compartments, chiefly endoplasmic reticulum and mitochondria. Study of the time course of the alterations revealed no light or ultrastructural features of neuronal necrosis in over 1 thousand neurons examined in layers 3 and 4 of the cingulate and retrosplenial cortex, 153 of which were vacuolated. The vacuoles resolved over time by decreasing in magnitude. Oxalate-pyroantimonate methodology revealed no redistribution of cell calcium in either vacuolated or non-vacuolated neurons. At 6 h, when vacuoles were consistently prominent in glutaraldehyde-fixed plastic-embedded tissue, a separate series of experiments was undertaken to vary methods of tissue preparation, and determine conditions under which vacuolation occurs. Frozen sections revealed no vacuoles. Subsequent paraffin embedding of the previously frozen tissue revealed no vacuoles, but vacuoles were seen in paraffin after perfusion fixation. Immersion fixation with brain refrigeration for 12 h prior to fixation revealed no vacuoles. Alcohol fixation also led to no visible vacuoles. We conclude that the vacuolation induced by NMDA antagonists is a reaction to aldehyde fixation of perturbed but living neurons, resulting in artifactual distortion of multiple intracellular compartments.