| Motion sickness (MS) can occur when the sensory inputs about body position contradict what is expected. It can be provoked by abrupt changes in movement or when one is exposed to moving visual scenes while the body is in a relatively fixed state. Signs of motion sickness include pallor, restlessness, cold sweat, nausea, excessive salivating, and vomiting. In the serious conditions, it can cause dehydration and shock. MS is one of the main causes of decrease of battle ability. Therefore, the research for MS protection and therapy is an important topic in military medicine and maritime hygiene.Several effective treatments are available to prevent MS symptoms. A number of pharmacological agents, including antihistamines and anti-muscarinics, reduce some of the signs and symptoms of MS. But Pharmacological agents prescribed to alleviate the nausea of motion sickness have yielded less than satisfactory results, as they either do not sufficiently relive symptoms or the side-effects produced by the recommended dosages render the user functionally compromised. Thus, preparation of new medicine against MS is needed.It is required for research on MS to establish all experimental model. Vomiting is the criterion of gastrointestinal malaise in most studies dealing with MS. However, studies on the MS using common laboratory animals such as rats and mice are limited due to the lack of the ability to vomit. Pica, the consumption of a substance without nutritional value such as kaolin, can be induced in rats by various stimuli and in mice following exposure to rotation-induced hypergravity. So pica was used as an analogue to emesis in rats or mice in previous studies. But there exists dispute. In addition, it is an indirect observation and might be not sensitive enough. A score based on symptoms seen frequently in rats and mice during MS, such as piloerection, tremble, urinal and fecal incontinence, as an index for the severity of MS in rodents was usually used. Therefore, these symptoms may theoretically be taken as the parameters for MS in rodents in stead of emesis. Practically, the fecal granules of rats are easily to be scored and the existence of urinal incontinence, piloerection and tremble are easily determined. So, a new method for MS in rodents and to evaluate the anti-MS effects of a new combination drug in rats should be improved. In our preliminary study, glutamatergic vestibular neurons express Fos after vestibular stimulation, which provide molecular index for MS.The precise etiology of motion sickness is still unclear. Histamine are involved in the development of the symptoms and signs of motion sickness。Histamine H1-blockers are clinically effective in preventing motion sickness, suggesting that the histaminergic neuron system plays an important role in the development of motion sickness. In eukaryotes, the pyridoxal-requiring enzyme histidine decarboxylase (HDC) is the primary enzyme responsible for synthesizing histamine and is consequently a crucial regulatory step for histamine biosynthesis.Researches show that, dexamethasone can downregulate histidine decarboxylase mRNA and enzyme activity in rat lung and significantly reduced nasal allergy-like behaviors, up-regulation of HDC mRNA, HDC activity and histamine content, which suggest that Glucocorticoid hormones may be used for Motion Sickness Prophylaxis and therapy. The results of relation between sensitivity of MS and stress hormones in blood suggest that, MS associated with insufficient stress in stimulation of abnormal acceleration, and that increasing stress level could treat MS. Glucocorticoid hormones were used in spaceflight kinesia, which suggest it could be appled in MS prophylaxis and therapy. However, the applyment of glucocorticoid hormones in large dose long-termly can initiate a serial adverse effect, such as complication of digestive system, induceing even aggravating infection. Also large dose glucocorticoid hormones can downregulate the expression of GR and its effect.Ginsenoside is the main active component of ginseng and it can elevate the ability of physical stress through upregulating bind activity and transcription activity of GR. Ginsenoside can increase the effect of dexamethasone for relieving vomit of patient after radiotherapy and Chem. On the basis of above, we assume that, glucocorticoid hormones could relieve rat's MS symptom after stimulus of mimic MS by suppressing the activity of HDC and decreasing content of histamine and that combination with ginsenoside could decrease the applying dose of glucocorticoid hormones and not affect the pharmacodynamic action and lighten the side-effect.Part I Comparative study of MS indices of RatObjective:To compare the accuracy and reproducibility of commonly used three indices of rat motion sickness, we conducted repeated experiments under the same condition. To guide the choice of MS indices of Rat.1. Methods:1.1 Grouping of experimental animalMale SD rats(180±20)g were were purchased from Sino-British SIPPR/BK Lab Animal Ltd (Shanghai, China) and grouped to subjected to stimulation group with saline, stimulation group with scopolamine and control group. All the animals were caged under controlled conditions (temperature:22±2℃and lighting:8:00-20:00) and received standard rat/mice chow and tap water ad libtum. All experimental procedures were in accordance with institutional animal care guidelines. They received (i.g.) solvent, dose of scopolamine at 1.0 mg/kg with SCO or same dose of NS with CON and SAL respectively 1/d, 1day before and 3 days in rotating experiment.1.2 Modeling of MS ratsEach animal was placed in an individual centrifuge cage (diameter=20 cm, height=25 cm). Rats were accelerated on three successive days by Crampton model, one hour per day.1.3 Measurement of kaolin intakeKaolin (Sinopharm Chemical Reagent Co., Ltd) was mixed with 1% gum Arabic (Sinopharm Chemical Reagent Co., Ltd) to form a thick paste which was then made into pellets of the same length as the food pellets. Kaolin and food were provided in separate compartments in a divided food hopper. Each animal was placed in an individual cage for for the daily kaolin and food intake. Body weight, food and kaolin intake were monitored daily 1 day before and 3 days in rotating experiment.1.4 Investigation of motion sickness (MS) indexMS symptoms were observed and recorded. The score of MS symptoms was obtained according to evaluation criteria. This score defined as MS index is suitable for rats. Fecal granules None:0; 1 per fecal granule; Urination None:0; urination:1.2; Piloerection None: 0; light:0.6; severe:1.2; Tremor None:0; tremor:1.2.1.5 Investigation of spontaneous activity The spontaneous activity were tested by open-field test. One day before stimulation and one to three day during the stimulation, the rats were put into the open-field (high 40cm, width and long 100cm, undersurface divided into 25 equal square) and recorded the activity within the fist 3 min.1.6 Investigation of the protein expression of c-fos in rat vestibular nucleiThe protein expression of c-fos in rat vestibular nuclei was analysed by Western blot. The relative densities of bands were analyzed with Quantity One.2. Results:2.1 Compared with the control group, the stimulated groups presented the features that kaolin consumption increased, but data variated greatly in groups.2.2 MS index increased signifcantly, which could be attenuated by scopolamine.2.3 Results show that spontaneous activity of rats decreased following the repeating of experiments, which suggest that, exploratory behavior of rats decreased following the familiarity with environment. The results of spontaneous activity were not stastically signifcant between CON and SAL.2.4 The protein expression of c-fos in rat vestibular nuclei increased significantly in SAL(p>0.01) and SCO(p<0.01) compared with CON. Meanwhile, The protein expression of c-fos in rat vestibular nuclei is significantly lower in SCO than in SAL(p<0.01).PartⅡAntimotion sickness effects of with dexamethasone in ratsObjective:To investigate the antimotion sickness effects of with dexamethasone in rats and to observe the valid dose of dexamethasone for Antimotion sickness effects1.Methods1.1 Grouping of experimental animalMale SD rats(180±20)g were were purchased from Sino-British SIPPR/BK Lab Animal Ltd (Shanghai, China) and grouped to subjected to stimulation group with saline, stimulation group with scopolamine and control group. All the animals were caged under controlled conditions (temperature:22±2℃and lighting:8:00-20:00) and received standard rat/mice chow and tap water ad libtum. All experimental procedures were in accordance with institutional animal care guidelines. Male SD rats were randomly divided into 4 groups (AS, AD, SS, SD) or 8 groups(SAL, SCO, DEXL, DEXM, DEXH;). The rats were exposed to abnormal acceleration for one hour.1.2 Other methods:the same as mentioned above.2.Results2.1 Results shows that, the motion sickness index of AG, SS and SG decreased significantly compared with AS (P<0.05, P<0.01) and that SG decreased significantly compared with SS, but there is no notable difference between AG and SS.2.2 The protein expression of c-fos in rat vestibular nuclei increased significantly in AG, SS and SG decreased significantly compared with AS. but there is no notable difference between AG and SS. (P>0.05)2.3 The motion sickness index of SAL, SCO, DEXL, DEXM, DEXH were significantly higher than CON (P<0.05, P<0.01), and MS index of DEXL, DEXH were significantly higher than SCO (P<0.05, P<0.01), but there is no notable difference between DEXM and SCO (P>0.05)2.4 The protein expression of c-fos in rat vestibular nuclei increased significantly in SAL, SCO, DEXL, DEXM, DEXH (p<0.01) compared with CON. But there is no notable difference between DEXM and CON (P>0.05)2.5 During acceleration-exposition days, there are no significant difference rat body weight increment compared with before exposition in SAL, SCO. But the there are notable body weight decrease difference compared with before exposition in DEXL, DEXM and DEXL. Among all groups, the DEXH body weight decrease mostly.PartⅢAntimotion sickness effects of ginsenosides combined with dexamethasone in rats and it's possible mechanism.Objective:To investigate the antimotion sickness effects of ginsenosides combined with dexamethasone in rats and the possible mechanism.1.Method: 1.1 Grouping of experimental animal1.1.1 Antimotion sickness effects of ginsenosides combined with dexamethasone in rats:Fifty SD rats were randomly divided into 5 groups which were untreated (normal saline), scopolamine-treated, ginsenosides-treated, dexamethasone-treated and ginsenosides+dexamethasone-treated. The rats in each group were fed with corresponding ingredients respectively, and then the rats were exposed to abnormal acceleration for one hour. The motion sickness index, the level of kaolin consumption and the course and time of spontaneous activity and weight were observed.1.1.2 The mechanism of Antimotion sickness effects of ginsenosides combined with dexamethasone in rats:Sixty SD rats were randomly divided into 6 groups which were untreated (normal saline) (ASM), (SSM), ginsenosides+dexamethasone-treated(SGM). The rats in each group were fed with corresponding ingredients respectively, and then the rats were exposed to abnormal acceleration for one hour. (ASC), (SGC), (SSC).The motion sickness index, the level of kaolin consumption1.2 ELISA was used to detect the histamine and activity of HDC and Real-Time PCR were used to test the HDC mRNA expression1.3 Other methods:the same as mentioned above2.Results:2.1 The motion sickness index and the level of kaolin consumption of acceleration-exposed rats in the ginsenosides+dexamethasone-treated group was significantly lower than that in untreated group.2.2 The protein expression of c-fos in rat vestibular nuclei decreased significantly in SCO,DEX,DEX+GIN (p<0.01) compared with SAL. And it decreased significantly in DEX+GIN compared with SCO. But there is no notable difference between GIN and SAL (P>0.05),between SCO and DEX.2.3 And the course and the time of spontaneous activity of acceleration-exposed rats in the ginsenosides+dexamethasone-treated group was significantly higher than that in untreated group;2.4 The level of body weight increment of acceleration-exposed rats in the ginsenosides+dexamethasone-treated group was significantly higher than that in the dexamethasone-treated group.2.5 Compared with SSC, histamine content of ASM and SSM increased significantly (p<0.05), content of SGC decreased significantly (p<0.05), content of SGM, ASC didn't change significantly (p>0.05)2.6 Compared with SSC, the histidine decarboxylase (HDC) enzyme activity of ASM, ASC increased significantly (p<0.05), activity of SGM, SGC decreased significantly (p<0.05), activity of SSM didn't change significantly (p>0.05)2.7 Compared with ASC, levels of HDC mRNA of SSM increased significantly (p<0.05), levels of SGC, ASM and SGM decreased significantly (p<0.05)Statistical analysis:Statistical analysis was performed by using the SPSS13.0 statistical program. The differences between the drug-treated group and control group were analyzed by Repeated Measures, one—way ANOVA, the paired t test. Data are expressed as mean±SD. Statistical significance was judged at p<0.05 or P<0.01。Conclusion:1. MS index and c-fos protein in rat vestibular nuclei showed much better accuracy and reproducibility than kaolin intake and spontaneous activity under this experiment condition, which indicate that we can apply MS index and c-fos protein in rat vestibular nuclei as main indices, spontaneous activity as assistant index to evaluate the anti-MS effects of new drug in rats.2. Ginsenosides combined with dexamethasone in rats can decreased MS indices and the expression of c-fos protein in rat vestibular nuclei significantly, which indicate that the combination of ginsenosides and dexamethasone can relief the symptom of MS of rat.3. There is no significant difference in anti-MS effect between ginsenosides combined with low dose dexamethasone and single high dose dexamethasone. But the losing of weight was improved after applied dexamethasone, which proved that the combination is much better than single use of dexamethasone.4. Compared with control rats, MS rats treated with ginsenosides combined with dexamethasone, decreased, the histidine decarboxylase (HDC) enzyme activity of hypothalamus decreased, levels of HDC mRNA decreased. All indicate that ginsenosides combined with dexamethasone can relief the symptom of rat MS by downregulating the expression of HDC in hypothalamus and histamine content of vestibule and hypothalamus. |
