5 mM Sr2+ and increasing Mg2+ to 3.3 mM. To minimize voltage-clamp errors, we recorded CF-PC EPSCs either between −65 mV and −70 mV in the presence of 600–800 nM NBQX or at depolarized potentials (−15 to −10 mV). Drugs were applied in the bath or via a flow pipe (ValveLink 8.2, Automate Scientific, Berkeley, CA). KYN and NBQX were purchased from Ascent Scientific (Princeton, NJ), TBOA, cyclothiazide (CTZ), and (2S,1′S,2′S)-2-(carboxycyclopropyl) glycine (L-CCG-I) were purchased

from C646 Tocris Bioscience (Ellisville, MO). Picrotoxin was purchased from Sigma (St. Louis, MO). Whole-cell recordings were made from visually identified PCs with a gradient contrast system by using a 60 × water-immersion objective on an upright microscope selleck products (Olympus BX51WI). Pipettes were pulled from either PG10165 glass (WPI,

Sarasota, FL) with resistances of 0.8–1.5 MΩ or BF150-110 borosilicate glass (Sutter Instrument Co., Novato, CA) with resistances of 1–1.5 MΩ. The series resistance (Rs), measured by the instantaneous current response to a 1–2 mV step with only the pipette capacitance cancelled, was <5 MΩ (usually <3 MΩ) and routinely compensated >80%. CFs were stimulated (2–10 V, 20–200 μs) with a theta glass electrode (BT-150 glass, Sutter Instrument Co., Novato, CA) filled with extracellular solution placed in the granule cell layer. The paired-pulse ratio (50 ms interstimulus interval) was determined after the stimulation train. Responses were recorded with a MultiClamp 700B amplifier (Molecular Devices, Sunnyvale, CA), filtered at 4–10 kHz, and digitized (Digidata 1440A, Molecular Devices) at 50–100 kHz by using Clampex 10 acquisition software (Molecular Devices). Pipette solutions PD184352 (CI-1040) for EPSC recordings contained 35 mM CsF, 100 mM CsCl, 10 mM EGTA, 10 mM HEPES,

and 5 mM QX314, adjusted to pH 7.2 with CsOH or 9 mM KCl, 10 mM KOH, 120 mM K gluconate, 3.48 mM MgCl2, 10 mM HEPES, 4 mM NaCl, 4 mM Na2ATP, 0.4 mM Na3GTP, and 17.5 mM sucrose (pH 7.25 with KOH) for current-clamp recordings. In current-clamp recordings, PCs were injected with a negative current (<500 pA) to maintain a membrane potential between −65 and −70 mV during synaptic stimulation (−66.9 ± 0.8 mV at 0.05 Hz and −68.4 ± 0.8 mV at 2 Hz; n = 26; p > 0.05). The frequency of synaptic stimulation did not alter the CpS plateau potential from which spikelets were generated (−41.0 ± 0.9 mV at 0.05 Hz and −44.0 ± 1.1 mV at 2 Hz; n = 26; p > 0.05). For experiments described in Figure 7, the membrane potential was also kept at approximately −70 mV. The peak amplitude of the injected current used to evoke complex-like spikes varied across cells (5–18 nA, corresponding to peak conductances of 70–250 nS). The maximal rate of spikelet rise was measured from differentiating the CpS waveform. Spikelets and their height were determined from trough to peak by setting the peak detection threshold to within 2%–10% of the maximum peak with a separating valley of adjacent peaks of <90%.