动物实验,甲状腺功能低下 z-bo 2020-08-13 462

　　【Abstract】Objective To investigate the effect of hypothyroidism (hypothyroidism) on learning and memory in young rats (offspring). Methods Pregnant SD female rats replicated hypothyroidism pups models were included in the study (n = 40), all males were randomly selected from the day of birth to intraperitoneal injection of T42μgP100g every day until weaned, as the T4 replacement treatment group, the remaining 20 hypothyroidism pups No special treatment was given to the mice, and 20 male pups were set up as a normal control group. The dark and eight-arm experiments were performed on the offspring of the 50-day-old hypothyroidism group, and compared with the normal control and treatment groups at the same age. Results The 50-day-old low A group of young mice made the most mistakes in the dark avoidance experiment, and the memory latency was shortened after 24 hours, and the difference was significant; there was no reference memory error in the 50-day-old low-A group of eight-arm experiment. Significant difference, working memory errors increase, the difference is significant, there is no significant difference in the time to run the maze.

　　Conclusion 　 Hypothyroidism causes a decline in the learning and memory ability of young mice.

　　Thyroid hormones play an important role in mammalian brain development. The lack of thyroid hormone in the perinatal period will cause irreversible brain damage, causing severe sensory, movement disorders and mental retardation, and the time of onset, severity, and the time to start thyroid hormone therapy, etc. The factors are closely related to the occurrence and recovery of brain damage. Based on previous studies of hippocampal neuronal apoptosis in perinatal hypothyroidism (hypothyroidism) pups, we further used dark and eight-arm mazes to detect the changes in learning and memory in developing hypothyroid pups.

　　Materials and Methods

　　Animal model preparation and group replication of hypothyroidism pups model, SD male pups were included in the study, numbered, randomly selected hypothyroidism group pups were intraperitoneally injected with thyroxine (T4) 2μgP100 g body weight every day from the day of birth until they were weaned at 21 days of age. T4 replacement treatment group (20); the pups of the low-yu hypothyroidism group were not given special treatment (20); another normal control group (20) was set up. The 21-day-old pups were weaned, with 3 to 4 male rats per cage until the experiment. Free water intake, 12 hours of light every day (lighting time: 8: 00 ~ 20: 00). Room temperature 21±2℃, relative humidity 40%～60%.

　　2　 Learning and Memory Research

　　211　darkness avoidance experiment 　tested 50-day-old mice in each experiment. The dark avoidance test box (SFK2 I) has two distinct bright and dark chambers, with a small door communicating between them; the dark avoidance habit makes the young mice instinctively choose to enter the dark room, and the young mice that enter the dark room are immediately shocked (35 mV) and passively escape Safe bright room. If you do not enter the dark room for 5 minutes, it is considered to have completed the study. Record the number of times that the offspring entered the dark room by mistake before completing the study, which mainly reflects the recent learning ability of the offspring; after 24h, put it back into the bright room again, and record the offspring entering the dark room. The time in the dark room is the memory latency, which mainly reflects the recent memory ability of the offspring.

　　212　eight-arm maze experiment 　test 50 (60) day old mice of each experiment. Before the eight-wall maze experiment, the offspring's diet was restricted to maintain their weight at 80%-85% of their free-feeding weight. Observation was carried out from 10:00 to 17:00. Use radial eight-arm (four arms to put food) maze. The central area of the labyrinth has a diameter of 30 cm, and eight arms (50 cm×12 cm, surrounded by a 415 cm high wall) are extended at equal angles and lengths all around. The entire maze is 40 cm above the ground. Before training, the young mice adapt to the maze for 2 d, once a day. During adaptation, 3 to 4 young mice were placed in the maze at the same time, free to move and eat food for 10 minutes. After the adaptation, the training was carried out once a day for a total of 10 days. In each training, only four of the eight arms were placed with food (Bio2Serv, Frenchtown, NJ, USA), namely arms 1, 3, 5 and 6; this order was maintained throughout the experiment. The young mice are placed in the central area of the maze. At this time, doors are closed around the central area. After 15 seconds, the door is opened. The young mice can choose to enter any arm to take food. The offspring entered the arm with food and ingested food for the correct choice once, otherwise it is a wrong choice. The number of times the parameter is incorrectly selected is recorded. Re-entering the food arm is called a working memory error (WME), and entering the food arm is called a reference memory error (RME).

　　3　Statistical processing 　Use SPSS 1010 for Windows statistical software for data analysis and processing. The data are all expressed as x±s. One2Way ANOVA or independent sample t test was used for comparison between groups and within groups.

　　结 　果

　　Compared with the normal control group and the treatment group in the dark avoidance experiment, the 50-day-old hypothyroidism group made more mistakes when they reached the level of learning, and the difference was very significant (P<01001), and="" the="" treatment="" group="" had="" more="" errors="" than="" normal="" control="" but="" there="" was="" no="" statistical="" difference="" p="">0105). After 24 hours, the memory latency of the offspring of the hypothyroidism group was the shortest. Compared with the normal control and the treatment group, the difference was very significant (P<01001) p="">0105).

　　2'' Compared with the normal control and the treatment group in the eight-arm maze experiment, there was no significant change in the RME of the offspring in the hypothyroidism group (P<0105); the="" wme="" was="" significantly="" increased="" p="">0105); the total number of errors was significantly increased (P<0105); there="" was="" no="" statistical="" difference="" in="" total="" maze="" running="" time="" p="">0105)

　　Discuss 　 Discuss

　　Learning and memory are one of the advanced functions of the brain. Learning refers to the acquisition or development of experience (behavioral habits, perception, thinking, etc.); memory is the preservation and reproduction of experience. They are two different and closely related neurobiological processes. On the one hand, they are the basis of thinking and cognition, and on the other hand, they are closely related to the sensory and motor functions of the brain; this complex process includes at least acquisition, consolidation and reproduction. Three main stages. Because of the comparability of rat's learning and memory ability and human learning and cognition, it is often used to study human learning and memory. At present, there are many kinds of experimental models for studying the learning and memory abilities of animals from the overall level. The shuttle box experiment is one of them. Because of its simple and fast operation, it has been widely used in learning and memory experiments. In order to eliminate the influence of corticosteroid secretion rhythm on learning and memory, daily experiments are conducted at the same time period. This experimental study found that the number of mistakes entering the darkroom in 50-day-old hypothyroidism rats increased during learning, and the memory latency after 24 hours was shortened, indicating that hypothyroidism in the perinatal period affects the short-term learning and memory ability of the rats. However, the electrical stimulation used in the experiment may cause excitement and restlessness in the animal. Moreover, non-learning and memory factors such as the animal's own exercise and sensitivity to electrical stimulation will also affect the results of the experiment. Animal spatial memory test is one of the most commonly used methods to study animal learning and memory. Since the water maze test can effectively separate animal spatial recognition and other abilities, this method has been used in previous studies on the effect of hypothyroidism on the spatial memory of young mice. However, the main indicator determined in this experiment is the time it takes for the animal to find a safe platform in the water maze (incubation period), which is closely related to the state of the animal's motor function. Any drug that can affect the animal's motor function and the state of the animal itself may affect The results of the experiment. Thyroid hormone not only affects the nervous system, but also affects the development of skeletal muscles, which may affect the results of the test. In addition, the nature of animals walking around walls also limits its applications.

　　The eight-arm maze test is an animal learning and remembering the relative position of itself and the bait (the white pill placed at the end of the maze arm) by observing some fixed reference objects around it. This method can effectively rule out the animal's movement due to its own motor dysfunction. It is also possible to separate and analyze animal WME and RME at the same time because of the influence of observation indicators on learning and memory. For this reason, this experiment used the eight-arm maze for the first time to measure the spatial memory reproduction of hypothyroid pups during perinatal period. The study found that compared with normal controls and treated pups, 50-day-old hypothyroid pups increased WME, but RME did not change significantly. This separation of spatial memory can also be seen in the eight-arm maze test in which anticholinergic drugs reduce the acetylcholine content of rat hippocampal neurons. However, the hippocampus function is significantly impaired and WME and RME are both increased. The phenomenon of spatial memory separation in the hypothyroidism and eight-arm test and the specific mechanism need further research. Learning and memory are closely related to the plasticity of the synaptic structure and function of the central nervous system. Thyroid hormones can affect the proliferation, migration, differentiation, synapse development and myelin formation of nerve cells by regulating the expression of a variety of brain structural proteins and signal proteins, affecting the transmission of information between neurons, leading to learning and memory Wait for brain function to be impaired. This study found that T4 replacement therapy pups have significantly improved learning and memory compared with hypothyroid pups after birth, but laboratory test results did not reach the level of normal pups of the same age, indicating that the state of fetal hypothyroidism has caused partial and irreversible brain damage.