Med64 system

The intracerebroventricular LTP was performed with 3 mice in each group as follows: the mice were anesthetized with 2–3% isoflurane, and whole brains were immediately resected and soaked in ice-cold artificial cerebrospinal fluid (aCSF) saturated with 95% O₂ and 5% CO₂. Following sectioning at 300 μm thickness, the slices were incubated in oxygenated aCSF at 32 °C to recover for 40 min and at 20–25 °C to recover for 1 h. Then, slices were transferred to a recording chamber and submerged in aCSF perfusion. Slices were laid in a chamber with an 8 × 8 microelectrode array (Parker Technology, Beijing, China) in the bottom plane (each 50 × 50 mm in size, with an interelectrode distance of 150 μm) and kept submerged in aCSF. Signals were acquired using the MED64 System (Alpha MED Sciences, Panasonic). The field excitatory postsynaptic potentials (fEPSPs) in CA1 neurons were obtained by stimulating CA3 neurons. LTP was induced by applying three trains of high-frequency stimulation (100 Hz for 1 s, delivered 30 s apart). The LTP magnitude was quantified as the percentage change in the fEPSPs slope (10–90%) taken during the 60-min interval after LTP induction [25 (link)].

Multi‐electrode array (MEA) analyses of neurons and cortical organoids were performed as described (Nageshappa et al, 2016; Trujillo et al, 2019). Briefly, neurospheres were plated on dual‐chamber MEA; spontaneous spike activity was evaluated with the MED64 System (Panasonic). Glutamatergic (AP5, NBQX), GABAergic (Gabazine), and gap channel (Mefloquine) antagonism were used to verify neuronal activity. Recorded spikes were analyzed using the Neuroexplorer software (Nex Technologies, Madison, AL, USA). For cortical organoids, one‐month‐old organoids were plated on MEA plates and recordings were collected with a Maestro MEA system and AxIS Software Spontaneous Neural Configuration (Axion Biosystems, Atlanta, GA, USA). The plate was incubated in the machine for three minutes before five minutes of recording. Axion Biosystems’s Neural Metrics Tool classified as “active” those electrodes with at least five spikes/minute. Bursts were identified using an inter‐spike interval (ISI) threshold requiring a 5‐spike minimum and 100ms maximum ISI. Network bursts required a minimum of 10 spikes under the same ISI and at least 25% active electrodes.

The mice were anesthetized by isoflurane and the brain was quickly cut into artificial cerebrospinal fluid (aCSF) containing 119 mM NaCl, 26.2 mM NaHCO₃, 2.5 mM KCl, 11 mM glucose,1 mM NaH₂PO₄, 2.5 mM CaCl_2, and 1.3 mM MgSO4 (pH 7.4). In ice-cold aCSF brain were sectioned into 350 μm thick slices using a vibrating microtome (Leica, VT1000S, Germany). The sections were transferred to the recovery chamber with oxidized aCSF for at least 1.5 h at room temperature.
Acute brain sections were transferred from the recovery chamber to a recording chamber and submerged in CSF. An 8 × 8 microelectrode array (Parker Technology, Beijing, China) were used to record the signals. The sections were laid down in the bottom plane (50 × 50 μm in size, with an interpolar distance of 150 μm) and kept submerged in the aCSF. The fEPSP in CA1 neurons was recorded by stimulating CA3 neurons using a MED64 system (Alpha MED Sciences, Panasonic). Baseline recordings were collected for a minimum of 30 min. Following baseline an induction protocol that evoked LTP was applied. LTP induction protocol consisted of 1 train of 100 Hz stimulus that lasted for 1 s, and the field potential response for 1 h after the tetanus was recorded. The LTP magnitude was quantified as the percentage change in the fEPSP slope (10–90%) taken during the 60 min interval after LTP induction.