OBJECTIVES: The purpose of this study was to identify and characterize Na+-dependent, high affinity glutamate transporter (GLUT) activity in the hypothalamic paraventricular nucleus (PVN) and to compare GLUT activity in PVN of euhydrated versus water-deprived rats.
METHODS: Sprague-Dawley rats were deprived of water for two days before sacrifice. Control rats received water ad libitum. After sacrifice, PVN and cerebrum were removed and synaptosomes were prepared using standard techniques. Glutamate uptake was measured using [3H]-glutamate as substrate, physiological buffer, approximately 100 µg of synaptosomal tissue per assay and a Brandel cell harvester.
RESULTS: Glutamate uptake was saturable in PVN synaptosomes from euhydrated, control rats with a Vmax of 541 ± 22 pmol/min-mg protein (SEM) and Km of 17.6 ± 3.8 µM (SEM). In contrast, Vmax of glutamate uptake was 808 ± 58 pmol/min-mg protein in PVN of rats deprived of water for 2 days. This was significantly higher than controls (p<0.001). Km was 21.2 ± 7.3 µM and not significantly different from controls (NS).
CONCLUSIONS: Our results suggest that water deprivation of rats results in significantly higher synaptosomal glutamate uptake in PVN. Although the exact mechanism is unknown, increased transcription of the GLUT gene and/or increased cell surface expression of GLUT may contribute to the observed increase of glutamate uptake in dehydrated rats. Increased glutamate uptake may serve to restrict dehydration-induced activation of PVN efferent pathways specifically involved in release of neurohypophysial hormones and activation of sympathetic outflow that operate to maintain body fluid balance and cardiovascular function.