Application of the method of pharmacoencephalography for the assessment of neuroprotective drug activity

Central nervous system disorders resulting from traumatic or vascular injuries have a high social significance. In view of this, the search and development of effective methods for the experimental evaluation of the effec-tiveness of new neuroprotective agents is an urgent task of biomedical research. The most common experimental models for assessing neuroprotective activity are models of ischemic stroke (temporary or permanent occlusion of the middle cerebral artery circulation, photothrombosis, etc. [Li, Zhang , 2021]) and traumatic brain injury (model of controlled cortical impact, fluid-percussion impact, etc.) [Marklund, Hillered, 2011]) in rats. Since traumatic and vascular injuries of the brain are accompanied by the appearance of a focus of necrosis (dead cells) and penumbra (cells with pronounced functional and metabolic disorders), the classical version of the positive effect of the studied drug in the experimental conditions on rodents is the preservation of the viability of penumbra cells while reduc-ing the severity of neurological deficit, accompanied by motor, emotional-behavioral and cognitive impairments.

In view of this, to assess the effects of neuroprotective agents in small laboratory animals, as a rule, behavior al and functional tests are used [Schallert, 2006] to assess the degree of motor impairment (for example, the Cylinder and Beam walking test), emotional and behavioral changes (Open field, Elevated plus maze) or cognitive function (Morris water maze, T-maze, etc.). The results of these tests are usually verified by methods of assessing the volume and histomorphological pattern of the lesion [Berger et al., 2008], as well as by biochemical and molecular genetic methods [Iino et al., 2003]. The data obtained allow us to comprehensively assess the activity of the tested drugs and draw a conclusion about the appropriateness of the proposed pharmacotherapeutic approach. Howev-er, all of these methods have their limitations, which, in the future, may affect the objectivity of the conclusions drawn. For example, the results of behavioral and functional tests are often subjective and their results are most dependent on the "hands" of the experimenter. Compliance with testing conditions, such as time of day, lighting and room temperature, pre-handling, etc., is of great importance. Equally important is the sequence of tests, as well as the time intervals between them. The disadvantage of histomorphological, biochemical and molecular ge-netic methods may be the need to remove animals from the experiment in order to take the necessary material for research, which does not allow tracking the course of the pathology in dynamics within one particular animal.

Neurophysiological methods such as electroencephalography (EEG) or electrocorticography (ECoG) allow assessing the state of the brain of small laboratory animals in terms of bioelectric activity parameters (amplitude-spectral characteristics, coherence of lead pairs, etc.). Despite the need for surgical intervention (implantation of EEG or ECoG electrodes), they do not have a number of disadvantages of the above methods, allowing testing of one animal as often as necessary. In view of this, it is interesting to use EEG/ECoG methods to assess the neuro-protective activity of new drugs in models of vascular and traumatic injuries in small laboratory animals. This paper presents some examples of such studies, as well as our own experience in evaluating the neuroprotective activity of the alpha-2-adrenergic agonist mafedine in a controlled cortical impact model in rats.

Pharmaco-EEG as a tool for evaluating the effects of drugs in models of ischemic stroke and traumatic brain injury in rats

The possibility of using neurophysiological methods in in vivo experiments to assess the neuroprotective activity of new drugs is an important issue for biomedical research. In rats with TBI in the study by G.A. Volo-khova, the administration of deproteinized dialysate from the blood of calves in the post-traumatic period contributed to positive changes in the EEG pattern, which correlated with the normalization of vertical and horizontal motor activity in the Open Field test [Волохова, Стоянов, 2015]. In a study by other authors on a model of cerebral ischemia in rats (bilateral occlusion of the carotid arteries), was shown a positive effect of the preventive use of a combination of vinpocetine with melatonin on the parameters of EEG rhythms in surviving animals. At the same time, this combination of drugs increased the survival rate of rats after ischemia up to 80.0% (compared to 34.8% of the group of control animals) [Ганцгорн, Макляков, Хлопонин, 2015]. We have shown in a series of pilot studies that open severe TBI in rats is accompanied by persistent changes in the parameters of the bioelectrical activity of the brain, which are detected on the 3rd and 7th days after injury [Сысоев, Крошкина, Оковитый, 2019a; Сысоев и др., 2019, 2020, 2020a]. Among the key features of the ECoG signal of injured animals, a decrease in the amplitudes and indices of θ-, α-, and β-rhythms, as well as an increase in the activity of the slow-wave δ-rhythm were noted [Сысоев, Крошкина, Оковитый, 2019a]. These changes were accompanied by a disruption in the operation of interhemispheric and intrahemispheric connections, as evidenced by a drop in the cross-correlation coefficient [Сысоев и др., 2020a]. It should be noted that these changes were recorded not only in the impact area, but also in remote areas of the cortex. Another important feature of the bioelectrical activity of injured animals was the change in the reflex responses of the cerebral cortex to photo- and somatosensory stimulation. Therefore, in injured rats, the amplitude of the P2 peak of VEP curves on the 3rd day after TBI in areas of the cortex remote from the impact site was higher than in healthy animals, and on the 7th day in the area of injury, it was lower [Сысоев и др., 2020]. Also, in rats with injury, the amplitude of early (N1, P2) and late responses (N2, P2, N3) of the cortex to somatosensory stimulation was reduced, as well as the latency of early responses increased and late responses decreased compared to the conditionally healthy group of animals [Сысоев и др., 2020b].

In the course of further studies [Sysoev, 2021], it was found that the administration of the sodium salt of the alpha-2 adrenoreceptor agonist mafedine 1 hour after TBI and in the next 6 days led to the normalization of interhemispheric connections of brain regions remote from the area of damage and intrahemispheric connections of the healthy hemisphere by the 7th day after injury. In addition, positive changes in the responses of the cortex to photo- and somatosensory stimulation were noted in such animals. The results obtained confirmed the previously identified cerebroprotective activity of the sodium salt of mafedine [Sysoev, 2019], and the suitability of recording and analyzing ECoG to assess the effect of pharmacological agents on the course of TBI in laboratory animals.

Conclusions

Thus, based on the results of the work of other authors, as well as the data of our own studies, we can conclude that the recording and analysis of EEG/ECoG in rats is a sensitive tool for assessing the functional state of the brain after traumatic and vascular injuries. This makes it possible to use these methods to identify the neuro-protective effects of drugs, complementing traditional behavioral, functional, or molecular genetic research methods.