Abstract : Objective To investigate the effects of different doses of vitamin E (Vitamin E, Vit E) on learning and memory ability and choline acetyltransferase (ChAT) activity in chronic episode hypoxia (EHYP) rats . . Methods Rat model EHYP, and given the large (50IUP250g weight Pd), small (5IUP250g weight Pd) doses of Vit E intervention. Passive dark avoidance reflex tests evaluated the ability of learning and memory, the longer the incubation period (the STL), the stronger the learning ability and memory; rat cortex was determined by radiochemical method, striatum and hippocampus ChAT activity. Results After EHYP treatment , the STL of the EHYP group was significantly shorter than that of the control group (P<0.01), and the ChAT activity of each brain area was significantly decreased (P < 0.05) . After drug intervention , compared with the EHYP group , the STL of the Vit E large and low dose groups was significantly prolonged (Vit E high dose group : P < 0.05, Vit E low dose group : P < 0.01), but STL large dose group was significantly shorter than the low dose group (P <0. 05); on ChAT activity, the low dose group were ChAT activity in areas of the brain were significantly higher (P <0. 05), and large In the dose group, only the hippocampus and striatum ChAT activity was significantly increased (P < 0.05) . Conclusion Vit E can improve the learning and memory ability of EHYP rats and increase the activity of ChAT in the brain , and the small dose is better than the large dose.

Key words : hypoxia ; passive avoidance avoidance reflex test ; choline acetyltransferase ; vitamin E

Obstructive sleep ap2nea syndrome (OSAS) can cause learning and memory disorders has become an indisputable fact , a large number of studies have shown that chronic intermittent hypoxia (chronic epi2)

Sodic hypoxia, EHYP) played an important role in it. At present , there are many treatments for OSAS , but no effective method has been found to prevent and improve learning and memory disorders caused by OSAS . Vitamin E (Vitamin E, Vit E) is a fat-soluble vitamin . It has been reported in the literature to improve cognitive impairment caused by AD and aging through its antioxidant properties.

hinder. We established that reflect characteristics OSAS portion (i.e.: chronic intermittent hypoxia) EHYP rat model, learning and memory observed Vit E and choline acetyltransferase enzyme (choline acetyltrans2ferase, ChAT) activity of rat EHYP, Provide experimental evidence for finding ways to improve learning and memory impairment in OSAS patients.

Materials and Method

First, the material

Vit E (T3634, 1000 IUPg, Sigma company. Diluted with soybean oil, and the two concentrations of formulated 50 IUPml 5 IUPml.), 3H2 acetyl CoA (185CiPmmol, Amersham Life Science, Inc.), often down oxygen chamber

( Institute of Pulmonary Heart Disease , Fuwai Hospital , Chinese Academy of Medical Sciences ), O2 , CO2 Concentration Tester ( Shanghai Jiading Xuelian Instrument Factory ), Program Control Dark Box ( Pharmaceutical Institute of Chinese Academy of Medical Sciences ), Liquid Spark Instrument ( Xi'an 262 Factory ) .

Second, the method

1. Experimental animals and grouping: 40 male SD rats, after 190 ~ 200g, were purchased from Animal Center of Beijing Union Medical College Hospital, the passive dark avoidance reflex test screening, to meet the conditions 34 rats were randomly divided into control group (n = 7) , EHYP group (n = 9) , Vit E high dose group (n = 9), and Vit E low dose group (n = 9) . Died 35 days after treatment EHYP 7, the final number of rats in each experimental group as follows: Control group 7, EHYP group 7, Vit E 6 high dose group, Vit E 7 low dose group. There was no significant difference in body weight before hypoxia, body weight after hypoxia and weight gain in each experimental group (P>0.05) .

2. Establishment of animal model : (1) EHYP group produced EHYP rat model with reference to Xue Quanfu et al . The rats were placed in a normal-pressure hypoxic chamber , and nitrogen was injected into the chamber . At the same time, the amount of soda lime and silica gel was amplified in the chamber to absorb CO2 and water vapor , so that the oxygen concentration in the cabin was continuously controlled at 10 ± 0.5%. The CO2 concentration is always < 3 % and the humidity is 50 ± 10 % . Bulkhead small and extravehicular communication gap, the cabin pressure and atmospheric balance. The time for the oxygen concentration to decrease from 23% to 10% per anoxic was 45 minutes. During hypoxia , rats ingested and drank water regularly. Directly intragastric administration of 1 ml of soybean oil 30 min before hypoxia . Daily hypoxia for 8 hours, continuous anoxic 35 days later. (2) In addition to the control group not hypoxia, the remaining conditions were the same EHYP group. (3) Large and small dose groups of Vit E 30 ml before hypoxia, 1 ml large (50 IUPml) and small (5 IUPml) doses of Vit E were used instead of soybean oil by direct gavage . The rest were treated with EHYP group. 3. Determination of learning and memory : Refer to Ando et al. to determine the learning and memory ability of rats by passive avoidance avoidance reflex test. The detection steps are as follows : (1) Before the hypoxic treatment in the initial screening period , the rat is placed in the bright room with the back of the hole , and the timer is started . The rats entering the dark room within 90 s are used for the experiment. (2) Training of hypoxia, rat into the light compartment facing away from the hole, after entering the dark chamber, the energization stimulation 2s, 5s after electroporation animals are removed, return home cage. (3) Determination of 24 hours after the training, and then placed in the hole facing away from the light compartment rats, while the timer is started, the recording time to enter the dark chamber for the first time in rats as latency (the STL), observed 5min, has not entered Rat STL is specified as 300 s .

4. drawn: the behavioral experiments, rats were decapitated, the whole brain was placed on ice tray taken out, sequentially retrieves the cortex, hippocampus and striatum, meninges and blood vessels removed, quickly weighed out into liquid nitrogen. In order to avoid the cyclical rhythm change of ChAT activity , the rats in each experimental group were sacrificed at about 9 Am , and the rats were alternately decapitated.

5. Determination of ChAT activity : Refer to the method of Fonnum . Brain tissue to be tested by 1: 20 (WPV) was added 10mmolPL EDTA in PB buffer (0. 05M, PH = 7. 4 ). Homogenate was prepared with a homogenizer speed, take 40ul blank to a tube and the reaction solution while adding 40ul 20ul 3H2 acetyl-CoA, were incubated 30min at 37 degrees. The reaction was stopped with a reaction stop solution. The incubation solution flashes into the cup, add 2ml sodium tetraphenylborate 2 and 10ml of acetonitrile toluene scintillation solution, jog 1min, so that the generated 3H2 acetylcholine extracted into the toluene phase (3H2 using acetyl-CoA is not left in the aqueous phase). After standing for 10 min , the Decay Per Minute (DPM) value was measured in a liquid scintillation meter . The amount of protein bound to the homogenate, the activity of ChAT is calculated to pmol · mg · min expressed protein. 6. Determination of protein: Lowry et al reference method, with the test protein concentration value measured absorbance after Brilliant Blue G2250 interaction with proteins in the sample. Bovine serum albumin was used as the standard protein.

7. Statistical analysis: All data were expressed as mean ± standard deviation, the comparison between the measurement data of a plurality of samples and Q ANOVA test for significance test, comparison between a plurality of sample data by nonparametric method as significant Nemenyi Test , the significance level of the test is P < 0.05 .