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БИОМЕДИЦИНА | JOURNAL BIOMED | 2019 | Toм 15 | № 2 | 69–74 69
Е. И. Бонь, Н. Е. Максимович
«Методы оценки неврологических нарушений при экспериментальной церебральной ишемии»
https://doi.org/10.33647/2074-5982-15-2-69-74
METHODS FOR ESTIMATING NEUROLOGICAL DISTURBANCES
IN EXPERIMENTAL CEREBRAL ISCHEMIA
Elizaveta I. Bon*, Nataliya Ye. Maksimovich
Grodno State Medical University
230009, Republic of Belarus, Grodno, Gorkogo str., 80
Investigation of the brain pathology in experimental ischemia requires adequate methods for assessing the
neurological decit that occurs in laboratory animals, including sensory-based and behavioural disorders.
In this research, we aimed to compare motor and behavioural disorders in rats with partial and subtotal
experimental cerebral ischemia. The rats modelled with cerebral ischemia are found to exhibit a decrease in
muscle strength, resistance to hypoxia, motor and emotional activity. The animals with incomplete cerebral
ischemia demonstrated more pronounced sensory-based motor and behavioural disorders compared both
with those modelled with partial cerebral ischemia and, in particular, with the control group.
Keywords: cerebral ischemia, sensomotor and behavioural disorders, rats
Conict of interest: the authors declare no conict of interest.
Funding: The study was funded by the Belarusian Foundation for Basic Research (contract No. М18М-
036) grant for reagents and animals.
For citation: Bon E.I., Maksimovich N.Ye. Methods for Estimating Neurological Disturbances in Experimen-
tal Cerebral Ischemia. Journal Biomed. 2019;15(2):69–74. https://doi.org/10.33647/2074-5982-15-2-69-74
Submitted 08.10.2018
Revised 25.02.2019
Published 10.06.2019
МЕТОДЫ ОЦЕНКИ НЕВРОЛОГИЧЕСКИХ НАРУШЕНИЙ
ПРИ ЭКСПЕРИМЕНТАЛЬНОЙ ЦЕРЕБРАЛЬНОЙ ИШЕМИИ
Е. И. Бонь*, Н. Е. Максимович
УО «Гродненский государственный медицинский университет»
230009, Республика Беларусь, Гродно, ул. Горького, д. 80
Изучение патологии головного мозга при экспериментальной ишемии обусловливает потребность в
адекватных способах оценки возникающего у лабораторных животных неврологического дефицита,
включающего сенсомоторные и поведенческие нарушения. Целью работы явилось сравнительное
изучение двигательных и поведенческих нарушений у крыс с частичной и субтотальной экспери-
ментальной церебральной ишемией. Установлено, что крысы после экспериментальной церебраль-
ной ишемии обладали меньшей мышечной силой, были менее устойчивы к гипоксии, проявляли
меньшую двигательную и эмоциональную активность. У животных с субтотальной церебральной
ишемией наблюдались более выраженные сенсомоторные и поведенческие нарушения по сравне-
нию с крысами, которым моделировали частичную церебральную ишемию и, особенно, по сравне-
нию с контрольными животными.
4.0
МЕТОДЫ БИОМЕДИЦИНСКИХ ИССЛЕДОВАНИЙ | BIOMEDICAL METHODS
БИОМЕДИЦИНА | JOURNAL BIOMED | 2019 | Toм 15 | № 2 | 69–74
70
Ключевые слова: церебральная ишемия, сенсомоторные и поведенческие нарушения, крысы
Конфликт интересов: авторы заявили об отсутствии конфликта интересов.
Финансирование: Работа выполнена при поддержке БРФФИ (проект М18М-036).
Для цитирования: Бонь Е.И., Максимович Н.Е. Методы оценки неврологических нару-
шений при экспериментальной церебральной ишемии. Биомедицина. 2019;15(2):69–74.
https://doi.org/10.33647/2074-5982-15-2-69-74
Поступила 08.10.2018
Принята после доработки 25.02.2019
Опубликована 10.06.2019
Introduction
The expediency of studying the pathology of
the brain in experimental ischemia necessitates
adequate methods for assessing the neurological
decit that occurs in laboratory animals, includ-
ing various sensory and behavioural disorders.
A number of methods can be used to study the
degree of sensory-based motor and behavioural
disorders in animals, such as the forced swim-
ming, muscle strength, open eld tests, as well
as those for evaluating the modied indicators
of the depth of a neurological decit [3]. Among
other popular methods are the Bederson, gas ex-
posure, angular, paw- pulling tests.
This article sets out to generalize the avail-
able literature data on contemporary ap-
proaches to assessing sensory-based motor
reexes, learning and memory abilities in
experimental animals at different ages. In the
early postnatal period, reexes are assessed
using such methods as the slip on the surface,
negative geotaxis, avoidance of falling, reac-
tion to acoustic stimuli, olfactory reaction and
muscle strength tests.
Sensory-based motor and behavioural dis-
orders have been extensively studied using
various models of cerebral ischemia (CI) [7–
10]; however, the data obtained in these works
has not thus far been properly generalized.
Research works devoted to the investiga-
tion of CI effects have identied a number of
sensory-based motor disorders, including a de-
crease in motor activity when an animal is sus-
pended by the tail, walking along a circle on a
horizontal surface, dis-coordination of move-
ments during walking along the bar, a decrease
in the expression of unconditioned reexes,
animals’ inability to navigate through space.
Local ischemic damage to the anterior parts
of the frontal cortex of the rat brain leads to
disruption in the production, preservation and
reproduction of conditioned reexes, while
damage to the posterior parts of the frontal
cortex is accompanied by a loss of the ability
to navigate in T-shaped labyrinths [8–10].
The purpose of this research was a compar-
ative study of motor and behavioural disorders
in rats with partial and subtotal experimental
cerebral ischemia.
Tests for studying the maturation of the
nervous system in newborns
In new-born rat pups, methods for assess-
ing the development of sensory-based motor
reexes comprise the slip on the surface, neg-
ative geotaxis, avoidance of falling, reaction to
acoustic stimuli, olfactory reaction and muscle
strength tests. These measurements can be car-
ried out both multiple times to trace dynamic
changes and on a single occasion, e.g. on the
supposed day of maturation of the studied re-
ex in intact animals [1, 26–28].
Tests for studying sensory-based
motor disorders in adult animals
The degree of sensory-based motor disorders
in adult rats is identied using a diversity of
methods, including the Bederson, Garcia, angu-
lar, pulling-the-paw, open-eld tests, as well as
that for assessing the modied depth indices of a
neurological decit [3, 5–7, 11, 14, 15, 17–25].
Materials and methods
The experiments were performed on
30 female non-native white rats weighing
БИОМЕДИЦИНА | JOURNAL BIOMED | 2019 | Toм 15 | № 2 | 69–74 71
Е. И. Бонь, Н. Е. Максимович
«Методы оценки неврологических нарушений при экспериментальной церебральной ишемии»
230±20 g. All the requirements of the 2010/63/
EU Directive of the European Parliament and
the Council of 22.09.2010 on the protection
of animals used for scientic purposes were
observed during the experiments [16]. The
animals were kept in an air-conditioned room
(22°C) with mixed lighting under standard
vivarium conditions implying free access to
feed and water. One vivarium cage housed no
more than 5 animals. The choice of experi-
mental animals was determined by the simil-
arity between rats and humans in terms of the
brain angioarchitectonics. Before the research,
all the necessary conditions were met. Thus,
60 minutes before testing, the animals were
kept in a quiet, poorly lit place, without any
regrouping and changes in feeding and other
conditions [4]. Subtotal cerebral ischemia
(SCI) was modelled by a stepwise ligation of
a carotid artery, which manipulation promoted
the survival of animals compared to those
with simultaneous bandaging. Partial cerebral
ischemia (PCI) was modelled by ligation of
one common carotid artery (CCA) under intra-
venous thiopental anaesthesia (40–50 mg/kg).
The second group consisted of rats, which
were simulated with SCI 7 days following PCI
by ligation of the second (right) CCA [4]. The
control group (control) comprised false-oper-
ated animals. The tests were performed 5 days
following the surgery.
The ischemic damage to the brain was es-
timated by assessing the animals’ emotional
state, behaviour and motor activity. To this
end, the open eld, muscle strength and forced
swimming tests were applied.
Muscular strength was assessed by placing a
rat on a metal mesh with a length of 60 cm and
a centimetre scale, and determining the time
when the animal falls off after lifting the mesh
to a horizontal position (by 90°).
In the forced swimming test, the animals
were placed in a glass tank lled with water
(21°C) for determining the time during which
the animal can maintain swimming and oat-
ing behaviour.
Statistical processing of the obtained data
was carried out using the Statistica 10.0 pro-
gram for Windows (StatSoft, Inc., USA).
The values obtained were analysed by non-
parametric statistics. The quantitative data
were presented as Me (LQ; UQ), where
Me was the median, LQ — the value of the
lower quartile; UQ — the value of the up-
per quartile. Differences between the control
and experimental groups were considered
signicant at p<0.05 (the Kruskal — Wallis
and Mann — Whitney tests with the Bonfer-
oni correction) [1].
Results
A signicant decrease in muscle strength
was observed in both groups of the animals
with CI compared to the control group. Thus,
this indicator decreased by 75% (p<0.05) and
95% (p<0.05) in the PCI and SCI groups, re-
spectively. Muscular strength in rats with PCI
was 5 times more pronounced than in those
with SCI (Table).
In addition, the rats with PCI showed
a greater resistance to the load-induced hyp-
Fig. Muscle strength evaluation.
Рис. Оценка мышечной силы.
МЕТОДЫ БИОМЕДИЦИНСКИХ ИССЛЕДОВАНИЙ | BIOMEDICAL METHODS
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72
oxia in the forced swimming test. The time of
their oating on the surface exceeded that in
the animals with SCI by 58% (p<0.05). Com-
pared to the control group, the oating time
was by 57% (p<0.05) and 76% (p<0.05) lower
in the PCI and SCI groups, respectively.
The assessment of motor activity by the open
eld test also revealed the presence of motor
deciency in rats with CI. In comparison with
the control group, the rats with PCI and SCI
demonstrated a decrease in crossed squares by
20% (p<0.05) and 64% (p<0.05), respectively.
The motor activity in the horizontal plane test
was higher in rats with PCI by 44% (p<0.05)
than in the SCI group. In comparison with the
control group, the number of short washings in
the rats with PCI and SCI was 33% (p<0.05)
and 67% (p<0.05) lower, respectively. The rats
with PCI performed 50% (p<0.05) more wash-
ings compared to the SCI group.
Compared to the control group, the weight
loss in the rats with PCI and SCI was 33%
(p<0.05) and 67% (p<0.05) lower. The rats
with PCI made 50% more racks in comparison
with the SCI group (p<0.05).
The rats with PCI and SCI demonstrated a
40% (p<0.05) and 60% (p<0.05) decrease in
the number of defecation and urination acts
compared to the control. In terms of this indic-
ator, the rats with PCI differed from animals
with SCI by 33% (p<0.05).
Long-term washings and rearing posts were
observed only in intact animals.
Conclusions
The rats after experimental CI have demon-
strated a decrease in muscle strength, resist-
ance to hypoxia, motor and emotional activ-
ity. In animals with SCI, more pronounced
sensorimotor and behavioural disorders have
been observed compared to those modelled
with PCI. The morphological basis of the re-
vealed disorders is found to be damage to the
neurons of the brain, leading both to the de-
struction of physiological functional connec-
tions and physiological systems, as well as to
the emergence of pathogens. The latter causes
the destabilization of nervous processes (co-
ordinated activity of excitation and inhibition),
which subsequently results in the disruption of
brain cognitive functions. Another reason for
these disorders is an imbalance between the
levels of biogenic amines and neuromediators
in brain structures, which act as endogenous
pathogenic factors and determine the nature
and severity of ischemic damage [2, 12, 13].
Table. Indicators of sensory-based motor and behavioural tests. The quantitative data are presented in the form of
Me (LQ; UQ)
Таблица. Показатели сенсомоторных и поведенческих тестов. Количественные данные представлены в виде
медиана (нижний квартиль; верхний квартиль)
Group
Indicators
Muscle
strength
Forced
swimming
test
Open eld test
Number of
crossed squares
Number of
short washings Climbing Number of defecation
and urination acts
Control 20 (15; 24) 21 (18; 23) 67 (64; 72) 6 (5.1; 7.2) 9 (8; 9.4) 5 (4.6; 6.5)
PCI 5 (4.7; 5.9)*12 (15; 12.3)*54 (52; 59)*4 (3.5; 4.3)*6 (5.8; 6.4)*3 (2.5; 3.6)*
SCI 1 (0.5; 1.2)* # 5 (4.5; 5.7)* # 24 (22; 28)* # 2 (1.6; 2.4)* # 3 (2.6; 3.2)* # 2 (1.2; 2.3)* #
Note: * — p<0.05 compared to the control, # — p<0.05 compared to the PCI.
Примечание: * — p<0,05 по сравнению с контролем, # — p<0,05 по сравнению с частичной церебральной ишемией.
БИОМЕДИЦИНА | JOURNAL BIOMED | 2019 | Toм 15 | № 2 | 69–74 73
Е. И. Бонь, Н. Е. Максимович
«Методы оценки неврологических нарушений при экспериментальной церебральной ишемии»
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INFORMATION ABOUT THE AUTHORS | СВЕДЕНИЯ ОБ АВТОРАХ
Elizaveta I. Bon*, Cand. Sci. (Biol.), Grodno State
Medical University;
e-mail: asphodela@list.ru
Nataliya Ye. Maksimovich, Dr. Sci. (Med.), Prof.,
Grodno State Medical University;
e-mail: mne@grsmu.by
Бонь Елизавета Игоревна*, к.б.н., УО «Грод-
ненский государственный медицинский универ-
ситет»;
e-mail: asphodela@list.ru
Максимович Наталия Евгеньевна, д.м.н.,
проф., УО «Гродненский государственный меди-
цинский университет»;
e-mail: mne@grsmu.by
* Corresponding author / Автор, ответственный за переписку
