Grass sd9k stimulator

Individual hippocampal neurons were isolated from the CA1 region using an enzyme-free mechanical dissociation procedure, as described previously (Akaike and Moorhouse, 2003; Jun et al., 2011; Vorobjev, 1991) . Briefly, slices including hippocampus were transferred into a 35 mm culture dish containing an external recording buffer with the following composition (in mM): 150 NaCl, 5 KCl, 2.5 CaCl 2 , 1 MgCl 2 , 10 HEPES, and 10 D-Glucose, pH adjusted to 7.4 with NaOH, osmolarity adjusted to 320 mOsm with sucrose. A firepolished glass micropipette was placed on the surface of the CA1 region. The pipette tip was vibrated horizontally within the CA1 region at 30Hz over a distance of 100-200 μm for ~2 min using a piezoelectric manipulator (LSS-3000, Burleigh/Exfo, Quebec, Canada) triggered by a Grass SD9K stimulator (Grass Technologies, West Warwick, RI). After the slice was removed, the isolated neurons were allowed to settle on the bottom of the dish for 10-15 min. For purely electrophysiological experiments, cells were isolated/examined in standard 35 mm cell culture dishes, (BD Biosciences, San Jose, CA). For experiments involving imaging, cells were isolated/examined in 35 mm cell culture dishes containing a glass coverslip insert (WillCo Wells B.V., Amsterdam, Netherlands). Cover-slips were coated with poly-l-lysine prior to use.