| BackgroundMotion sickness (MS) is a common syndrome of dizziness, pale skin, cold sweat,salivation, abdominal discomfort, and even nausea, vomiting and other autonomic responsevestibular dysfunction, which is caused by abnormal acceleration stimulation.Seasickness,as one kind of motion sickness, is a common sailing health issues for ships members,which has a high incidence.The occurrence of seasickness brings crew varying degrees ofphysical discomfort, mental and psychological burden, and reducing the crew of fitness,efficiency and emergency response capabilities. Therefore, it is very important to study thereason and mechanism of seasickness, and to explore the new ways of preventionseasickness. And it has important practical significance to improve the efficiency andquality of life for maritime operations staff, to improve the combat effectiveness of thetroop ships member.Rat is one of common model for the research of motion sickness. However, rats don’thave the vomiting reflex. and conditions of anorexia nervosa(such as Kaolin pica) andspontaneous activity are used to determine the occurrence of motion sickness and severityindicators in rats at present. Recent years, scholars quantified piloerection, tremor,defecation, urine output of rodents,called motion sickness index(MSI),as a evaluationindicators to quantify the behavior of motion sickness in rats. MSI has a better stability andrepeatability compared with spontaneous activity and Kaolin pica.It is not a well recognized biological indicator to evaluate the susceptivity of motionsickness in rats. Some studies have shown that serum arginine vasopressin (AVP),adrenocorticotropic hormone (ACTH), glucocorticoids(GC), insulin, some traceelements(such as copper,iron,zinc),the protein expression of c-Fos in vestibule mayassociate with motion sickness.Research purposesMotion sickness index was used as a behavior evaluation of motion sickness in rats.Correlation analysis was used to analyse the relationship between motion sickness indexand the biological indicators in rats which may be related to motion sickness as reported.So as to provide experimental evidences for establishing a more consummate system of motion sickness comprehensive evaluation.Research MethodsSeveral possible biological markers associated with motion sickness are screenedby literature review. Motion sickness Index was used to evaluate motion sickness in rats.Correlation analysis was used to analyse the relationship between motion sickness indexand the change of serum AVP, ACTH, corticosteroids, insulin, trace elements copper,iron, zinc levels, brain vestibular nuclear c-Fos protein expression in rats after theseasickness stimulation exposure. Including the following sections:一. The correlation analysis between motion sickness index and thechanges of some serum hormone and element levels after seasicknessstimulation exposure(一) Experimental animals42adult male SD rat（sShanghai-BK Co., Ltd., Shanghai, China), weighing228±12g,were used in this animal study. All rats were caged under controlled conditions(temperature:22±2℃, humidity:60%-70%and lighting:8:00–20:00) and they had freeaccess to food and water.(二) Motion sickness-provocation simulation in ratsThe rotation device was used to provocate seasickness.The rats were put in therotation device freely before turning on the device. The rotation lasted for1hour. Therotatory device began to rotate at a constant angular acceleration of16°/s2in a clockwisedirection until the angular velocity reached120°/s2, then the box started to slow down at aconstant angular deceleration of48°/s2. After a1s-pause, the box continued to rotate incounter-clockwise direction in the same manner.(三) Motion sickness index scoreAfter acceleration stimulation, the piloerection, trembling and so on are recordedimmediately to calculate the scores of motion sickness index. motion sickness index of quantitative criteria for rats as follows: fecal pellets: Each1point, none0; urination: Yes,1.2points, none,0; piloerection: severe,1.2, mild,0.6none,0; tremble: yes,1.2, none,0.The final score result for the total points is motion sickness index.(四) Measurements and sampling1.The collection of bloodRats were decapitated immediately after acceleration stimulation and recordingmotion sickness index. Then all collected blood was centrifuged instantly at2,000rpm for30min at4°C. Supernate was isolated and stored at–80°C for subsequent analysis.2. Determination(1)The determination of serum hormone levelsSerum insulin, arginine vasopressin(AVP), corticosterone and adrenocorticotropichormone(ACTH) levels were measured by radioimmunoassay kits(RIA).(2)The determination of serum trace element levelsSerum copper(Cu)、iron(Fe)、zinc(Zn) levels were measured by inductively coupledargon plasma quantifier (IcAP).0.2ml serum samples were diluted with deionized water to5mL.二.The effects of the change of serum insulin levels on motionsickness index in rats(一) Experimental animals and grouping48healthy male SD rats，purchased from Sino-British SIPPR/BK Lab Animal Ltd(Shanghai, China), weighing259±32g, were used in this animal study. All rats werecaged under controlled conditions (temperature:22±2℃, humidity:50%-60%andlighting:8:00–20:00) and given standard rat chow and tap water.After feeding adaptation,the rats were randomly divided into control group (n=8), streptozotocin (STZ) group (n=8), insulin group (n=8), only rotation group (n=8), STZ+rotation group (n=8) and insulin+rotation group (n=8).(二) drug-induced animal model establishment for changing seruminsulin level in rats1. STZ administration for ratsSTZ group and STZ+rotation group rats were injected65mg/kg STZintraperitoneally after no food but water for12hours. All injection were finished in25minutes.2.Insulin administration for ratsInsulin and insulin+rotation group rats were injected2IU/kg insulinintraperitoneally in batches before30minutes at the beginning of rotation.(三) Motion sickness-provocation simulation in ratsThe rotation device and motion sickness activation condition were the same as theprevious description.After injected intraperitoneally2IU/kg insulin, the insulin+rotation rats rotated withonly rotation group and STZ+rotation group in batches. Rats in insulin group, controlgroup and STZ group did not rotate but place in the room of the rotation device for1h.(四) Motion sickness index scoreAfter seasickness stimulation, the performance characteristics such as defecation,piloerection, shivering were recorded to calculate motion sickness index. The method ofmotion sickness index score was were the same as the previous description.(五) Measurements and sampling1. The collection of bloodAfter seasickness stimulation, rats were immediately decapitated and take the trunkblood. Blood centrifuged for2000rpm×30min×4°C, the supernatant at-80°C for subsequent analysis of mineral elements. At the same time,baseline group rats weredecapitated and blood was obtained.2. DeterminationSerum insulin levels were measured by radioimmunoassay kits(RIA).三. The relationship study between motion sickness index andthe protein expression of c-Fos in rat vestibular nuclei afterseasickness stimulation exposure(一) Experimental animals30healthy male SD rats，purchased from Sino-British SIPPR/BK Lab Animal Ltd(Shanghai, China)，weighing264±15g，were used in this animal study. All rats werecaged under controlled conditions(temperature:22±2℃, humidity:50-60%and lighting:8:00–20:00) and given standard rat chow and tap water.(二)Motion sickness-provocation simulation in ratsThe rotation device and motion sickness activation condition were the same as theprevious description. The rotation lasted for1hour.(三)Motion sickness index score and groupingAfter rotation stimulation, the motion sickness index scores were recorded. Themethod of computing motion sickness index was the same as the previous description.According to interquartile method,taking the upper quartile and lower quartile level ofmotion sickness index scores,6rats in each group were decided,and the rest of therotation rats were removed.(四) Measurements and sampling1. The collection of tissueAfter motion sickness stimulation, the rats were immediately decapitated and taken the vestibular nuclei on ice.2. DeterminationWestern blot was used to measure the protein expression of c-Fos in rat vestibularnuclei.四. Statistical analysisAll analysis was performed using the SPSS16.0. The data were expressed as mean±standard error(SE). P<0.05were considered as statistically significant.The indicators inthree groups were compared using one way analysis of variance (one-way ANOVA),significant differences between the two groups were compared with Least SignificantDifference (LSD) method. Simple linear regression was used to analyze the relationshipbetween motion sickness index(Y)and each hormone and element(X), p=0.5was consideredas criterion for entering the indicator, p=1.0was considered as criterion for removing theindicator. Using multiple linear regression analyzed the relationship between the scores ofmotion sickness index and the entered indicators(p=0.05for entering,p=0.10forremovement).Results一.The correlation analysis between motion sickness index andthe changes of some serum hormone and element levels afterseasickness stimulation exposureSimple linear regression analysis was used to choice out Cu, Zn, AVP, insulin andcorticosterone(p=0.5for the entering criteria, p=1.0for removing criteria), motion sicknessindex as the variable Y, serum insulin and corticosterone, ACTH, AVP, Cu, Fe and Zn asvariable X respective.The five indicators were as X1, X2, X3,X4,X5, in multiple stepwiselinear regression and MSI was as Y(P=0.05for the entering criteria,and p=0.10for theremoving criteria. The establishment of the regression model (p <0.05). After seasicknessstimulation,serum insulin levels is negative with motion sickness index. 二.The effects of the change of serum insulin levels on motionsickness index in ratsAfter STZ administration, serum insulin levels decreased; after insulin administration,serum insulin levels increased, and the difference was statistically significant (p <0.05).Compared with the control group, after seasickness stimulation,motion sickness indexin the simple rotation group increased significantly (p <0.05); compared with the controlgroup, the motion sickness index of STZ+rotation rats increased (p <0.05); comparedwith the control group, the motion sickness index of insulin+rotation group was notstatistically significant (p>0.05)Compared with the simple rotation group, the motion sickness index of insulin+rotation group rats decreased, the difference was statistically significant (p <0.05);compared with the simple rotation group, the motion sickness index of STZ+rotationgroup was not statistically significant (p>0.05).Motion sickness index in insulin+rotation group was lower than that in STZ+rotation group, and the difference was statistically significant (p <0.05).三. The relationship study between motion sickness index andthe protein expression of c-Fos in rat vestibular nuclei afterseasickness stimulation exposureAfter1h stimulation, the protein expression of c-Fos in rat vestibular nuclei weredifferent in control,high-and low-MSI group(p<0.05). The protein expression of c-Fos inthe high-MSI group was significantly higher than that in control group(p<0.05), and wassignificantly higher than that in low-MSI group(p<0.05)ConclusionsIn this experiment, the motion sickness index was as behavioral evaluation index forrats, and the indicators, serum hormone insulin, corticosteroids, ACTH and argininevasopressin levels, serum trace elements copper, iron, zinc level and vestibular c-Fosprotein expression levels,which may be related to motion sickness,were measured, thecorrelations between motion sickness index and those indicators were analyzed. We foundthat serum insulin levels associated with motion sickness index.To further investigate therelationship between the serum insulin concentration and motion sickness index, we reduced serum insulin levels by treating STZ to the adult male SD rats to demolish the isletbeta cells of them and increased serum insulin levels by administration of insulin.Theresults showed that motion sickness index decreased in insulin＋rotation group afterseasickness stimulation. After seasickness stimulation, the protein expression of c-Fos inrat vestibular nuclei were different in control,high-and low-MSI group. In all, seruminsulin concentration and vestibular c-Fos have the potential indicator to become abiological evaluation of motion sickness, but the specific application needs to be identified. |
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