Pci 6251 interface

A larva was embedded in 2% low melting point agarose (Invitrogen). Recording electrodes were pulled from soda lime glass capillaries (1412227, Hilgenberg, Germany) on a DMZ Universal Puller (Zeitz, Germany) to a diameter of ~20 microns. It was filled with artificial cerebrospinal fluid (ACSF, 124 mM, NaCl, 2 mM KCl, 2 mM MgSO₄, 2 mM CaCl₂, 1.25 mM KH₂PO₄, 26 mM NaHCO₃, and 10 mM glucose) and placed on larva’s head above the optic tectum. The recordings were performed using WinEDR (John Dempster, University of Strathclyde, UK). Differential signal was amplified 10,000 times by DAGAN 2400 amplifier (Minnesota, USA), band pass filtered at 0.3–300 Hz and digitized at 2 kHz via a PCI-6251 interface (National Instruments, UK).
All the larvae used for LFPs displayed touch response. The duration of each recording was 10 min. An electrical discharge was classified as a positive event when its amplitude was at least three times the amplitude of the baseline, and had a duration of at least 100 ms. The analysis of the epileptiform events was done using automated detection software that was previously developed and validated by our group^{46 (link)}.

Larvae (6 dpf) in 100 µl VHC were arrayed individually in a 96-well plate and kept for 18 h in the dark at 28 °C. An equal volume of 800 μM EKP (2x solution) or VHC (control) was added to each well and hence larvae were incubated with 400 μM EKP for 15 min. Next, the treated larva was embedded ventral side down in 2% low melting point agarose bathed in artificial cerebrospinal fluid (ACSF, 124 mM NaCl, 2 mM KCl, 2 mM MgSO₄, 2 mM CaCl₂, 1.25 mM KH₂PO₄, 26 mM NaHCO₃, and 10 mM glucose). A blunt glass electrode (soda lime glass, Hilgenberg, Germany) was pulled with DMZ Universal Puller (Zeitz, Germany) to an opening of 15–20 microns, filled with ACSF and placed on the skin above the optic tectum. The recordings were performed using WinEDR (John Dempster, University of Strathclyde, UK). Differential signal was band pass filtered at 0.3–300 Hz and digitized at 2 kHz via a PCI-6251 interface (National Instruments, UK). LFP recordings started each time exactly 2 min after the removal of the larva from the EKP solution (or VHC) and were continued for 10 min at room temperature (24 °C).
EKP-induced events were considered as epileptiform activity when their signal exceeded three times the baseline and lasted for minimum 100 msec, as described previously^{44 (link)}. Electrophysiological recordings were analyzed in a blinded way using Clampfit 10.2 software (Molecular Devices, USA).