Hypoxic insult increases the level of extracellular glutamate, which leads to the influx of toxic Ca++ through the activation of N-methyl-D-aspartate(NMDA) receptors. The neuroprotective action of NMDA antagonist against hypoxic insult has been demonstrated in vitro. It has been demonstrated that the concentration of 50hydrox-ytryptamine(5-HT) also increased after ischemia in rat hippocampus. However, there is paucity of studies concerning the funtional relationships between the spontaneous release of 5-HT and NMDA receptor activity during hypoxia in vitro. Therefore, the present study was aimed to investigate whether hypoxia and/or NMDA was able to stimulate the release of 5-HT from the hypoxia-sensitive rat hippocampl slices.The hippocampus was obtained from the rat brain and sliced 400um thickness with manual chorpper. After 30 min's preincubation in the normal buffer, the slices were incubated for 20 min in a buffer containing 3H-5HT(0.1uM, 74uCi) for uptake, and washed. To measure the release of 3H-5-HT into the buffer, the incubation medium was drained off and refilled every ten minutes through a sequence of 14 tubes. Administration of NMDA or induction of hypoxia (gassing it with 95% N2/5% CO2) was done in the 6th and 7th tube, and APV was added 10 minutes prior to these manipulations. The radioactivities in each buffer and the tissue were counted using liquid scintillation counter and the results were expressed as a percentage of the total radioactivity. When slices were exposed to hypoxia for 20min, 3H-5-HT release was markedly decreased and a rebound release of 3H-5-HT was observed on the post-hypoxic period. NMDA(1mM) incereased 3H-5-HT release in the control group. NMDA also incereased rebound release of 3H-5-Htrelease. When 2-amino-5-hposphonovaleric acid (APV, 30uM or 60 uM) were added to the incubation media, NMDA-induced increase of 3H-5-HT release were blocked does-dependently. The rebound release of 3H-5-HT during post-hypoxic period was also blocked by APV. These results suggest that the spontaneous release of 3H-5HT decreases during hypoxic period, but 20min hypoxic exposure causes rebound increase of 3H-5-HT release during post-hypoxic period which is mediated by the increased activity of the NMDA receptor.
Animals ; Anoxia* ; Brain ; Glutamic Acid ; Hippocampus ; Ischemia ; N-Methylaspartate* ; Radioactivity ; Rats* ; Scintillation Counting ; Serotonin*