Fig 1: Spontaneous excitatory postsynaptic current frequency and amplitude is unchanged following 10 and 20 days of TDP-43ΔNLS expression in layer 5 pyramidal neurons. (A,B) Representative traces of spontaneous excitatory postsynaptic currents (sEPSCs) from CaMKIIa-tTA/+ and +/+ mice (Control) and mice expressing a nuclear localization sequence-deficient human TDP-43 (TDP-43ΔNLS) following 10 (A) and 20 (B) days of transgenic expression. (C,D) The frequency of sEPSCs is not different between control and TDP-43ΔNLS mice at 10 days post expression (C; P30 + 10) and 20 days post expression (D; P30 + 20). (E,F) The amplitude of sEPSCs is not different between control and TDP-43ΔNLS mice at 10 days post expression (E) and 20 days post expression (F). Experiments conducted in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes and approved by the Animal Ethics Committee of the University of Tasmania (A16593). Mice were group housed (2–5 per cage) in individually ventilated cages on a 12-h light–dark cycle. Female mice were excluded from this study. CaMKIIa-tTa (The Jackson Laboratory Stock # 003010) and tetO-TDP-43ΔNLS (The Jackson Laboratory Stock# 014650) were fully backcrossed to a C57Bl/6J background for a minimum of 10 generations (confirmed test of 99.8–100% C57Bl/6J background). To collect tissue for electrophysiological assessments, mice were sacrificed via intraperitoneal injection of with sodium pentobarbital (200 mg/kg) followed by trans-cardial perfusion with ice cold carbogenated (95% O2, 5% CO2) perfusion solution containing (in mM): 92 choline chloride (Sigma-Aldrich Cat# C7527), 2.5 KCl (Sigma-Aldrich Cat# S9541), 1.2 NaH2PO4 (Sigma-Aldrich Cat# S5011), 30 NaHCO3 (Sigma-Aldrich Cat# S5761), 20 HEPES (Sigma-Aldrich Cat# H3784), 25 Glucose (Sigma-Aldrich Cat# G7021), 0.5 CaCl2 (Sigma-Aldrich Cat# C3306), 10 MgSO4•7H2O (Sigma-Aldrich Cat# M2773), 5 sodium ascorbate (Sigma-Aldrich Cat# A4032), 2 Thiourea (Sigma-Aldrich Cat# T8656), 3 sodium pyruvate (Sigma-Aldrich Cat# P5280), 5 N-Acetyl-L-Cysteine (Sigma-Aldrich Cat# A9165). Brains were immediately sliced (coronal sections) at 400 μm on a Leica VT1200s vibratome. Acute slices were recovered in a holding solution containing (in mM): 92 NaCl, 2.5 KCl, 1.2 NaH2PO4, 30 NaHCO3, 20 HEPES, 2 CaCl2, 2 MgCl2 (Sigma-Aldrich Cat# M2392) and 25 Glucose. Borosilicate glass capillaries (Harvard Apparatus) were pulled to produce patch pipettes with a tip resistance of 3–5 MΩ (P1000 pipette puller, Sutter Instruments). The intracellular solution contained (in mM): 120 Cs-methanesulfunate (Sigma-Aldrich Cat# C1426), 20 CsCl; (Sigma Aldrich Cat# C4036), 10 HEPES, 10 EGTA (Sigma-Aldrich Cat# O3777), 2 MgCl2, 2 Na2ATP (Abcam Cat# ab120385) and 0.3 NaGTP (Abcam Cat# ab146528) and CsOH (Sigma Aldrich Cat# 232041), at pH 7.4 and 280–290 mOsm. Slices were continuously perfused (> 4 mL/min) using aCSF (warmed to 30°C) containing 100 μM picrotoxin (Abcam Cat# ab120315) to eliminate GABAergic currents. The motor cortex was identified using a brain atlas at low magnification and layer V was identified using a 40x objective based on the presence of very large, sparse pyramidal neurons. Whole-cell patch-clamp recordings of primary motor cortex layer V pyramidal neurons (uncompensated series resistance of <20 MΩ) were made using a Heka EPC10 USB amplifier via the Heka Elektronik Patchmaster software. Voltage clamp recordings of EPCSs were made at-70 mV for 2 min, were sampled at 20 kHz and filtered with a low pass filter at 3 kHz. All recordings had a series resistance <15 MΩ and noise <10 pA, only cells with a stable series resistance over the course of the 2 min recording were used. Blinding was carried out for electrophysiological experiments and analysis. Each data point represents an individual cell. Groups were compared using a student’s t-test with significance threshold set to p < 0.05.