Figure 4.

Adenosine uptake (A) and release (B) from cultured cortical neurons from wild type (Wt) or hENT1 transgenic (Tg) mice. (A) [³H]Adenosine (1 μmol/L; 0.1 μCi) uptake was performed at 22 °C using primary cultures of neurons (11–15 d in vitro) using an assay volume of 0.5 mL and an uptake interval of 1 min. Non-specific uptake was determined with dipyridamole (Dpr; 30 μmol/L). For further details on methods, see³⁶. ^cP<0.01; t-test. (B) Neurons were incubated for 30 min at 37 °C with [³H]adenine to radiolabel intracellular ATP, then treated with buffer, Dpr (30 μmol/L), N-methyl-D-aspartate (NMDA; 100 μmol/L) or the combination of Dpr and NMDA for 30 min at 37 °C to evoke release of purines. [³H]Purines were separated by thin layer chromatography and quantified by scintillation spectroscopy. For additional details on methods, see²². Data are mean±SEM (n≥8) and are expressed as pmol/mg protein (A) or fold change, relative to wild type controls (B). Data were analyzed by two-way ANOVA. ^cP<0.01 vs controls. ^fP<0.01 vs NMDA. ⁱP<0.01 vs wild type NMDA.