Anesthesia was induced in mice by intraperitoneal injection of 20% isoflurane. Decapitate the animal and scoop out the intact brain into a breaker of pre-chilled and oxygen-saturated cutting buffer (in mM: 228 sucrose, 26 NaHCO
3, 11 glucose, 2.5 KCl, 7 MgSO
4, 1 NaH
2PO
4, and 0.5 CaCl
2, pH 7.25, 330 to 340 mOsm), allowing the brain to cool evenly for ~1 min [
35]. The brain was trimmed and affixed on the specimen holder using adhesive glue. The reservoir was filled with the above cutting buffer. Mouse brain was sliced (300 μm) in a vibratome with the remaining pre-chilled cutting buffer. Subsequently, the slices were transformed to oxygenated artificial cerebrospinal fluid (ACSF [in mM]: 125 NaCl, 26 NaHCO
3, 2.5 KCl, 1.3 MgSO
4, 10 glucose, 1 NaH
2PO
4, and 2.5 CaCl
2, pH 7.25, 300 to 310 mOsm) at 28 °C for 1 h before transferring to the recording chamber at room temperature [
35]. The entire procedure was continuously supplied with mixed gas (5% CO
2 and 95% O
2) to maintain oxygen saturation. During recording, slices were immersed in a chamber with oxygenated ACSF at a rate of 2 ml/min in room temperature. Acute brain slices were imaged using a 40× Olympus water-immersion objective, differential interference contrast (DIC) optics, and a charge-coupled device camera. Patch pipettes were fabricated from borosilicate glass capillaries using a PC-10 pipette puller [
33]. To record the general postsynaptic current of PSCs (voltage clamp), pipettes were filled with a low Cl
− internal solution (in mM: 126 CsMeSO
3, 10 HEPES, 1 EGTA, 1 CaCl
2, 1 Na-GTP, 4 Mg-ATP, and 2 QX314, pH 7.4, 290 to 300 mOsm). EPSCs were recorded at a holding potential of −70 mV, while IPSCs were recorded at 0 mV. Similarly, to record AMPA and NMDAR-mediated EPSCs, pipettes were filled with another internal solution (in mM: 117.5 CsMeSO
3, 15.5 CsCl, 10 TEA-Cl, 10 HEPES, 10 EGTA, 8 NaCl, 1 MgCl
2, 0.3 Na-GTP, 4 Mg-ATP, 1 QX314, and 10 Na phosphocreatine, pH 7.4, 290 to 300 mOsm). AMPA receptor-mediated EPSCs were recorded at a holding potential of −60 mV, while NMDA receptor-mediated EPSCs were recorded at +40 mV. PSCs were evoked by 100-μA stimulus intensity delivering from the stimulating electrode (FHC bipolar platinum), which positioned over patched S1BF neurons retrogradely labeled with EGFP around ~100 μm away. AMPAR- and NMDAR-mediated PSCs were recorded in the presence of PTX (50 μM) for blocking the GABA receptors. Light-evoked PSCs from ChR2-mCherry
+ SC were triggered by single light pulses (473 nm, 2 ms, 20 mW) in the presence of 4-aminopyridine (20 μM) and tetrodotoxin (1 μM). Antagonists of GABAa receptors (PTX) and glutamate receptors (D-AP5/CNQX) were perfused with ACSF to examine the neurotransmitter receptor type of optically evoked postsynaptic currents. The resistance of pipettes varied between 4.0 and 5.5 MΩ, and the recorded data series resistance value is less than 15 MΩ. Recording data were filtered at 1 to 2 kHz and digitized at 10 kHz via a Digidata 1440A. Data acquisition was performed with Clampex 10, and subsequent analysis was conducted using Clampfit 10. The E/I ratios were determined by dividing the EPSC charge by IPSC charge.