5 mM KCl, 1 mM NaH2PO4, and 0.1 mM CaCl2), or low-sodium, low-calcium, bicarbonate-buffered cutting solution (85 mM NaCl,

75 mM Sucrose, 25 mM D-(+)-glucose, 4 mM MgSO4, 2.5 mM KCl, 1.25 mM Na2HPO4·H2O, 0.5 mM ascorbic acid, 25 mM NaHCO3, and 0.5 mM CaCl2). Cortical slices (400 μm thickness) were cut from the left hemisphere in the “across-row plane” and oriented 50° toward coronal from the midsagittal plane. These slices contain one barrel column from each whisker row (A–E) (Allen et al., 2003 and Finnerty et al., 1999). Slices Inhibitor Library were transferred to normal Ringer’s solution and incubated for 30 min at 30°C and 1–6 hr at room temperature before recording. Barrels were visualized by transillumination. Recordings were made at room temperature (22°C–24°C) with 3–6 MΩ pipettes using Multiclamp 700A, 700B, or Axopatch 200B amplifiers (Molecular Devices, Sunnyvale, CA). All recordings were made using normal Ringer’s solution except for input-output experiments that were recorded in high divalent Ringer’s (116 mM NaCl, 26.2 mM Selleckchem PF-2341066 NaHCO3, 8 mM D-(+)-Glucose, 4 mM MgSO4, 2.5 mM KCl, 1 mM NaH2PO4, and 4 mM CaCl2). A bipolar stimulating electrode (FHC, Bowdoin, ME) was placed

in the center of a L4 barrel. L2/3 neurons in the same radial column were selected for recording. Current-clamp recordings were made using K gluconate internal (116 mM K gluconate, 20 mM HEPES, 6 mM KCl, 2 mM NaCl, 0.5 mM EGTA, 4 mM MgATP, 0.3 mM NaGTP, 5 mM Na2phosphocreatine [pH 7.2], and 295 mOsm). In a subset of cells, biocytin (0.26%) replaced phosphocreatine to allow morphological reconstruction. Input resistance (Rinput) was measured with a 120 ms hyperpolarizing-current injection Amisulpride in each sweep. Series resistance (Rseries) was compensated by bridge balance. Cells were excluded if initial Rseries was >20 MΩ or if Rinput or Rseries changed by >30% during recording. Sweeps were collected at 10 s intervals. Voltage-clamp recordings were made using Cs gluconate internal (108 mM D-gluconic acid, 108 mM CsOH,

20 mM HEPES, 5 mM tetraethylammonium-Cl, 2.8 mM NaCl, 0.4 mM EGTA, 4 mM MgATP, 0.3 mM NaGTP, 5 mM BAPTA [pH 7.2], and 295 mOsm). Rinput and Rseries were monitored in each sweep in response to a −5mV test pulse. Rseries was not compensated. Pyramidal cells were excluded if Vm at break in was >−68mV, Rseries > 25 MΩ, or Rinput < 100 MΩ. Vm values for voltage-clamp recordings were corrected for the measured liquid junction potential (10–12mV), whereas those for current-clamp recordings were not. Data acquisition and analysis used custom software in IGOR Pro (Wavemetrics, Portland, OR). For L4 stimulation, excitatory-response threshold was defined as the L4 stimulation intensity that elicited EPSCs with no failures at ECl (−68mV).