Oxygen-sensing persistent sodium channels in rat hippocampus

1. Persistent sodium channel activity was recorded before and during hypoxi a from cell-attached and inside-out patches obtained from cultured hippocam pal neurons at a pipette potential (V-p) of +30 mV. Average mean current (I ') of these channels was very low under normoxic conditions and was similar in cell-attached and excised inside-out patches (-0.018 +/- 0.010 and -0.0 25 +/- 0.008 pA, respectively, n = 24). 2. Hypoxia increased the activity of persistent sodium channels in 10 cell- attached patches (I' increased from -0.026 +/- 0.016 pA in control to -0.15 6 +/- 0.034 pA during hypoxia, n = 4, P = 0.013). The increased persistent sodium channel activity was most prominent at a V-P between +70 and +30 mV (membrane potential, V-m = -70 to -30 mV) and could be blocked by lidocaine , TTX or R56865 (n = 5). Sodium cyanide (NaCN, 5 mM; 0.5-5 min) increased p ersistent sodium channel activity in cell-attached patches (n = 3) in a sim ilar manner. 3. Hypoxia also increased sodium channel activity in inside-out patches fro m hippocampal neurons. Within 2-4 min of exposure to hypoxia, I' had increa sed S-fold to -0.18 +/- 0.04 pA (n = 21, P = 0.001). Sodium channel activit y increased further with longer exposures to hypoxia. 4. The hypoxia-induced sodium channel activity in inside-out patches could be inhibited by exposure to 10-100 muM lidocaine applied via the bath solut ion (I' = -0.03 +/- 0.01 pA, n = 8) or by perfusion of the pipette tip with 1 muM TTX (I' = -0.01 +/- 0.01 pA, n = 3). 5. The reducing agent dithiothreitol (DTT, 2-5 mM) rapidly abolished the in crease in sodium channel activity caused by hypoxia in excised patches (I' = -0.01 +/- 0.01 pA, n = 4). Similarly, reduced glutathione (G-SH, 5-20 mM) also reversed the hypoxia-induced increase in sodium channel activity (I' = -0.02 +/- 0.02 pA, n = 5). 6. These results suggest that persistent sodium channels in neurons can sen se O-2 levels in excised patches of plasma membrane. Hypoxia triggers an in crease in sodium channel activity. The redox reaction involved in increasin g the sodium channel activity probably occurs in an auxiliary regulatory pr otein, co-localized in the plasma membrane.