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OBJECTIVE To screen a mouse anxiety model with good reproducibility, little stimulation and user-friendliness by comparing the uncertain empty bottle drinking water stimulation with restraint stress to induce anxiety-like emotion in mice in order to find a suitable pharmacodynamic evaluation model for anti-anxiety drugs which provides an experimental basis. METHODS A mouse anxiety model was established using the uncertain empty bottle drinking water stimulation method. The mice were divided into freely drinking water group, regular drinking water group, physiological stress group and empty bottle stimulation group, with eight mice in each group. The freely drinking water group was free to eat and drink all day, the regular drinking water group mice were given free water for 2 periods per day, while the regular drinking water group mice were given water for 10 min at 2 fixed time with a 12 h interval every day. At the same time, physiological stress group mice had access to drinking water for 10 min once a day, but at another time they were given neither water nor empty bottles. The empty bottle stimulation group mice were given empty bottle stimulation randomly once or twice every day so that they drank water 12 times and empty bottle stimulation 16 times. The experiment lasted 14 d. The mouse anxiety model was established with restraint stress method. The experimental mice were divided into normal control group and restraint stress group, with eight mice in each group. The normal control group was normally reared, while the restraint stress group mice were confined to the tube at 9:00 every morning, headed towards the bottom of the centrifuge tube and were restrained for 2 h on the first day, and then the daily binding strength was extended for 2 h until 8 h every day for 21 d. The changes of body mass in mice after 7 and 14 d of empty bottle stimulation and after 7, 14, and 21 d of restraint stress were observed. At the end of the modeling, the mice were tested for autonomous activities and exploration behaviors through the open field test. The elevated plus maze test was used to detect the number of times they entered the open arms and the time spent in the open arms. The plasma corticosterone (CORT) content and the content of serotonin (5-HT) in the hippocampus of mice were determined by ELISA after 21 d of restraint stress. RESULTS Uncertain empty bottle drinking water stimulation method: compared with the freely drinking water group, the regular drinking water group showed a significant decrease in the residence time in the open arm (P<0.05). Compared with the regular drinking water group, the body mass of the mice decreased significantly after 7 d of empty bottle stimulation (P<0.05). There was no significant change in the behavioral indicators. Restraint stress method: compared with the normal control group, the body mass of mice was significantly decreased after 14 d of restraint stress (P<0.05). There was no significant change in the autonomous activity or exploration behavior of mice. The number of times the mice entered the open arms and the residence time in the open arms were significantly decreased (P<0.05). After 21 d of restraint stress, the body mass of the mice decreased significantly (P<0.05), and plasma CORT content (P<0.01) and 5-HT content in hippocampus (PO.01, P<0.05) of mice were significantly reduced. CONCLUSION The mouse anxiety model established with the uncertain empty bottle drinking water stimulation method results in poor stability and sensitivity, with more uncontrollable factors. The mouse anxiety model was not successfully established until 14 d of stimulation. The restraint stress for 14 d could successfully a establish mouse anxiety model, and the mechanism may be related to HPA axis dysfunction and 5-HT metabolic abnormalities.
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Objective To evaluate the effectiveness of a rat model for comorbidity of Tourette syndrome and anxiety with empty water bottle stimulation plus iminoodipropionitrile(IDPN) injection.Methods The 48 male SD rats were randomly divided into four groups:the blank control group,the TS group,the anxiety group and the comorbidity group.The blank control group was injected with saline for 7 days.The TS groop was injected with 3,3-iminodipropionitrile (IDPN) with 250 mg/kg once a day for 7 consecutive days.The anxiety group was given empty water bottle stimulation for 21 consecutive days.The comorbidity group was given empty water bottle stimulation plus IDPN injection.At the end of the 3rd week,the behavioral changes of the stereotyped movement,elevated plus-maze and open field of the rats in each group were measured,and the contents of monoamine neurotransmitters in striatum and hippocampus were determined by HPLC.Results The results of stereotyped movement showed that there was no significant difference between the groups except for the blank control group.The elevated plus-maze test showed that the 0E/TE values of the comorbidity group (21.33±11.35) ％ and the anxiety group (17.68±16.89) ％ were significantly decreased,lower than that of the blank control group (73.24± 19.33) ％ and TS group(61.43±21.84) ％.The results of open field test showed that the total scores of open field in the comorbidity group(15.22±9.87)and anxiety group (11.17±10.76) were lower than that of the blank control group (41.86±33.30) and TS group(48.83± 17.65) (P＜0.01).However,there was no significant difference between the comorbidity group and the anxiety group.The test of monoamine neurotransmitters in striatum showed that the content of HIAA in the comorbidity group(0.03±0.00) ng/mg was the highest,and that of the TS group and anxiety group (0.02±0.00) ng/mg was higher than that of the blank control group (0.01±0.00) ng/mg (P＜0.01).The DA test showed that the content of DA in the comorbidity group (0.03±0.00) ng/mg was the highest,and that of the comorbidity group,TS group(0.02±0.00) ng/mg and anxiety group was higher than that of the blank control group(0.01±0.00) ng/mg (P＜0.01).The expression of 5-HT was most significant among the groups (P＜0.01),and there was significant difference between the anxiety group ((0.011 ± 0.001) ng/mg)and the comorbidity group ((0.014±0.002) ng/mg) (P＜0.01).The expression of HVA in the three model groups ((0.05±0.00) ng/mg) was higher than that in the blank group ((0.02±0.00) ng/mg) (P＜ 0.01).The expression of DOPAC in the TS group ((0.23±0.02) ng/mg) was higher than that in the blank control group((0.16±0.01) ng/mg) and comorbidity group ((0.16±0.02) ng/mg) (P＜0.01).The test of monoamine neurotransmitters in hippocampus showed that the content of 5-HT in the comorbidity group ((0.14±0.02) ng/mg) was the highest,followed by the anxiety group ((0.1 ± 0.03) ng/mg) and the TS group ((0.07±0.04) ng/mg),which were all higher than the blank control group((0.04±0.03) ng/mg) (P＜0.05,P＜0.01),and there were significant differences between the comorbidity group and the TS group or anxiety group (P＜0.01).The expressions of HIAA and HVA were higher in the comorbidity group((0.44±0.04)ng/mg,(0.01±0.00) ng/mg),TS group ((0.46±0.15) ng/mg,(0.01 ±0.01) ng/mg) and anxiety group ((0.46±0.08)ng/mg,(0.01±0.00) ng/mg) than that in the blank control group((0.21±0.10)ng/mg,(0±0) ng/mg) (P＜0.05,P＜0.01).Conclusion This study confirms the reliability of the model and it is an ideal animal model for the study of TS with comorbidity of anxiety,which can be used for follow-up research.
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Objective To compare the biological characteristics of several different anxiety rat models established by different methods of stress at different time points and provide experimental basis for the most appropriate modeling methods .Methods 60 rats were randomly divided into normal , empty bottle stress , chronic emotional stress ( CES ) group, restraint stress for 3h, 6h, and modeling respectively .In the experimental 7 d, 14 d, 21 d, elevated plus maze and fear condition system was used to test anxiety-like behavior in rats , open field test to study anxiety or depression-like behavior , forced swimming test was used to detect depression-like behavior in rats , and using the Elisa test kit to detect the contents of 5-HT, DA in the hippocampus in rats .Results Anxiety-like behavioral test results showed that rats in empty bottle stress, CES, 6 h restraint stress group started to have anxiety-like behavior since 14 d, then anxiety-like behavior was becoming increasingly apparent .Forced swimming test results showed that immobility time in 6 h restraint rats was significantly increased in the first 7 d(P <0.05).Meanwhile, compared with control group, hippocampal 5-HT, DA contents in empty bottle stress and CES rats increased significantly since 14 d.Conclusions Among several stress methods established anxiety model , anxiety-like behavior in 3 h restraint stress was not obvious; 6 h restraint stress exhibited a depression-like behavior in the forced swimming test might be due to prolonged stress .Empty bottle stress and CES can successfully establish the anxiety rat model , and the anxiety behavior of the rats have some differences . Corresponding model methods can be selected according to different experimental purposes .
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Objective: To study the correlation between the empty bottle volume, negative pressure and gas production of the freeze-dried powder in the out-patient pharmacy intravenous admixture center of a children' s hospital in order to provide reference for the drug production. Methods:20 ml Syringes were used to measure the volume of empty bottles, negative pressure and produced gas. The relationship between the theoretical drug dissolution volume and the actual dissolution volume was compared, and the precautions for the drug production were put forward. Results:Among the tested 30 drugs, 6 ones were with the actual dissolution volume half of the theoretical dissolution volume, 8 ones were with negative pressure in the bottles, and 3 ones were with produced gas after dissol-ving. It was appropriate that the empty bottle volume be 4 ml larger than the theoretical dissolution volume, and it was appropriate that the negative pressure volume of drugs was slightly larger than the theoretical dissolution volume. Negative pressure should be still kept in the bottles after the gas production. Conclusion:The design of part of freeze-dried powder injection needle shows defects resulting in drug mixing difficulties to a certain extent.
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Objective To evaluate the effectiveness of a rat model for comorbidity of Tourette syndrome and anxiety with empty water bottle stimulation plus iminoodipropionitrile(IDPN) injection.Methods The 48 male SD rats were randomly divided into four groups:the blank control group,the TS group,the anxiety group and the comorbidity group.The blank control group was injected with saline for 7 days.The TS groop was injected with 3,3-iminodipropionitrile (IDPN) with 250 mg/kg once a day for 7 consecutive days.The anxiety group was given empty water bottle stimulation for 21 consecutive days.The comorbidity group was given empty water bottle stimulation plus IDPN injection.At the end of the 3rd week,the behavioral changes of the stereotyped movement,elevated plus-maze and open field of the rats in each group were measured,and the contents of monoamine neurotransmitters in striatum and hippocampus were determined by HPLC.Results The results of stereotyped movement showed that there was no significant difference between the groups except for the blank control group.The elevated plus-maze test showed that the 0E/TE values of the comorbidity group (21.33±11.35) ％ and the anxiety group (17.68±16.89) ％ were significantly decreased,lower than that of the blank control group (73.24± 19.33) ％ and TS group(61.43±21.84) ％.The results of open field test showed that the total scores of open field in the comorbidity group(15.22±9.87)and anxiety group (11.17±10.76) were lower than that of the blank control group (41.86±33.30) and TS group(48.83± 17.65) (P＜0.01).However,there was no significant difference between the comorbidity group and the anxiety group.The test of monoamine neurotransmitters in striatum showed that the content of HIAA in the comorbidity group(0.03±0.00) ng/mg was the highest,and that of the TS group and anxiety group (0.02±0.00) ng/mg was higher than that of the blank control group (0.01±0.00) ng/mg (P＜0.01).The DA test showed that the content of DA in the comorbidity group (0.03±0.00) ng/mg was the highest,and that of the comorbidity group,TS group(0.02±0.00) ng/mg and anxiety group was higher than that of the blank control group(0.01±0.00) ng/mg (P＜0.01).The expression of 5-HT was most significant among the groups (P＜0.01),and there was significant difference between the anxiety group ((0.011 ± 0.001) ng/mg)and the comorbidity group ((0.014±0.002) ng/mg) (P＜0.01).The expression of HVA in the three model groups ((0.05±0.00) ng/mg) was higher than that in the blank group ((0.02±0.00) ng/mg) (P＜ 0.01).The expression of DOPAC in the TS group ((0.23±0.02) ng/mg) was higher than that in the blank control group((0.16±0.01) ng/mg) and comorbidity group ((0.16±0.02) ng/mg) (P＜0.01).The test of monoamine neurotransmitters in hippocampus showed that the content of 5-HT in the comorbidity group ((0.14±0.02) ng/mg) was the highest,followed by the anxiety group ((0.1 ± 0.03) ng/mg) and the TS group ((0.07±0.04) ng/mg),which were all higher than the blank control group((0.04±0.03) ng/mg) (P＜0.05,P＜0.01),and there were significant differences between the comorbidity group and the TS group or anxiety group (P＜0.01).The expressions of HIAA and HVA were higher in the comorbidity group((0.44±0.04)ng/mg,(0.01±0.00) ng/mg),TS group ((0.46±0.15) ng/mg,(0.01 ±0.01) ng/mg) and anxiety group ((0.46±0.08)ng/mg,(0.01±0.00) ng/mg) than that in the blank control group((0.21±0.10)ng/mg,(0±0) ng/mg) (P＜0.05,P＜0.01).Conclusion This study confirms the reliability of the model and it is an ideal animal model for the study of TS with comorbidity of anxiety,which can be used for follow-up research.