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Advanced Neurology Non-invasive electroencephalography in rats

that among alpha-2-agonists, xylazine (i.p., 2 mg/kg) was 2.3. ECoG
the most effective agent to induce high-voltage spike- The rats were permanently implanted with epidural screw
and-wave spindles (HVS) in Fischer 344 rats. Similar to electrodes to perform long-term recordings during periods
27
WAG/Rij rats, HVS is associated with absence seizures of free behavior.
and spontaneously occurring SWDs. 10,28 The WAG/Rij rat
strain is widely recognized as a valid and reliable genetic 2.3.1. Electrode setup and implantation procedure
model for investigating absence epilepsy. 8-10,29-31 WAG/Rij The ECoG electrodes were constructed using stainless steel
rats are particularly valuable in preclinical investigations screws with a shaft length of 2.0 mm, a head diameter of
due to their remarkable similarities to human patients with 2.0 mm, and a shaft diameter of 0.8 mm. Each electrode
absence epilepsy. 23,30,32 In the present study, we used xylazine was equipped with four screws that were affixed to
for its sedative effect and its strong ability to elicit SWDs in wires and 4-pin connectors. The surgery was conducted
WAG/Rij rats. A prominent absence seizure-inducing effect under isoflurane anesthesia using the RWD Stereotaxic
of xylazine provided the basis for developing a novel, non- Anesthesia Setup (RWD Life Sciences, China). The rat’s
invasive EEG technique for rapidly diagnosing absence head was secured in the stereotaxic apparatus (Standard
epilepsy in genetically predisposed rats. The non-invasive Stereotaxic Instrument, RWD, Life Sciences, China). After
EEG technique was effectively validated in a large cohort shaving the rat’s head, the skin and soft tissues were excised
of WAG/Rij rats aged 5 – 15 months (n = 65, including from the cranium (Figure 1A). To implant the electrodes,
32 females and 33 males). four holes were drilled into the cranium: two holes over

2. Methods the right and left frontal cortices (AP +2 mm and L ±
2.5 mm) for the active frontal electrodes, one hole over the
2.1. Animals occipital cortex (AP −6 mm and L 4 mm) for the active
This study included adult WAG/Rij rats of both sexes (body occipital electrode, and one hole over the right cerebellum
weight 300 – 500 g). The rats were bred and reared at the for the reference electrode (Figure 1B). The entire assembly
Institute of Higher Nervous Activity and Neurophysiology was permanently attached to the skull utilizing a methyl
of the Russian Academy of Sciences (RAS) in Moscow, methacrylate monomer (Figure 1C). Following the
Russia. The rats were housed in a vivarium and maintained surgical procedure, the rats were administered metamizole
under standard conditions with a 12/12-h light-dark cycle (produced by FSSCI Microgen, Russia) intramuscularly
and unrestricted access to food and water. All experiments (25 mg/kg) to alleviate pain. To prevent damage to the
were conducted in compliance with Directive 2010/63/EU electrode connectors, the rats were individually confined
on the protection of animals used for scientific purposes. in cages and allowed to recuperate for at least ten days.
All phases of this study were approved by the Ethics The pain, suffering, and distress experienced by the rats
during the experiment were minimized, in keeping with
Committee of the Institute of Higher Nervous Activity the experimental protocols approved by the Animal Ethics
and Neurophysiology of the Russian Academy of RAS. Committee of our institution.
Specifically, the protocols for ECoG examination in rats
(protocol #4, October 26, 2021, and protocol #4, December 2.3.2. ECoG recording in free behavior
13, 2022) and additions for non-invasive EEG examination
in rats (protocol #4, May 29, 2024) were approved. The rats were placed in Plexiglas cages (25 × 60 × 60 cm), and
the ECoG was recorded during free behavior. The four-pin
The first group of 16 rats (9 females and 7 males) connectors on the rats’ heads were connected to an amplifier
was subjected to invasive ECoG examination in free through a swivel contact. The ECoG signals were transmitted
behavior during baseline and after i.p. xylazine injection. to a multichannel amplifier (PowerLab 4/35, LabChart 8.0
Among them, three rats (two females and one male) were software, ADInstruments, Sydney, Australia). The signals
subjected to non-invasive EEG examination before ECoG were bandpass filtered between 0.5 and 200 Hz, digitized at
examination. The second group including 32 female 400 samples per second per channel, and stored on the hard
and 33 male rats was subjected to non-invasive EEG disk. The rats’ behavior was monitored employing a Genius
examination under the influence of xylazine. eFace 1325R video camera and recorded using the video
capture module in LabChart software.
2.2. Pharmacological provocation of SWDs
SWDs were induced through intraperitoneal 2.4. EEG examination
administration of xylazine hydrochloride (Xyla, 20 mg/mL The EEG signal was captured from the scalp using a
xylazine hydrochloride, Interchemie Werken De Adelaar, wireless recording system (Physiobelt, Neurobotics,
the Netherlands) in dosages of 2 – 8 mg/kg. Moscow, Zelenograd, Russia). Two Physiobelt sensors

Volume 3 Issue 4 (2024) 3 doi: 10.36922/an.4464