Changes of learning and memory ability associated with neuronal nitric oxide synthase in brain tissues of rats with acute alcoholism

BACKGROUD： Ethanol can influence neural development and the ability of leaming and memory, but its mechanism of the neural toxicity is not clear till now. Endogenous nitric oxide （NO） as a gaseous messenger is proved to play an important role in the formation of synaptic plasticity, transference of neuronal information and the neural development, but excessive nitro oxide can result in neurotoxicity. OBJECTIVE ： To observe the effects of acute alcoholism on the learning and memory ability and the content of neuronal nitric oxide synthase （nNOS） in brain tissue of rats. DESIGN ： A randomized controlled animal experiment. SETTING ： Department of Physiology, Xinxiang Medical College MATERIALS： Eighteen male clean-degree SD rats of 18-22 weeks were raised adaptively for 2 days, and then randomly divided into control group （n = 8） and experimental group （n = 10）. The nNOS immunohistochemical reagent was provided by Beijing Zhongshan Golden Bridge Biotechnology Co.,Ltd. Y-maze was produced by Suixi Zhenghua Apparatus Plant. METHODS ： The experiment was carded out in the laboratory of the Department of Physiology, Xinxiang Medical College from June to October in 2005. ① Rats in the experimental group were intraperitoneally injected with ethanol （2.5 g/kg） which was dissolved in normal saline （20%）. The loss of righting reflex and ataxia within 5 minutes indicated the successful model. Whereas rats in the control group were given saline of the same volume. ② Examinations of learning and memory ability： The Y-maze tests for learning and memory ability were performed at 6 hours after the models establishment. The rats were put into the Y-maze separately. The test was performed in a quiet and dark room. There was a lamp at the end of each of three pathways in Y-maze and the base of maze had electric net. All the lamps of the three pathways were turned on for 3 minutes and then turned off. One lamp was turned on randomly, and the other two delayed automatically. In 5 seconds after alternation, pulsating electric current presented in the base of unsafe area to stimulate rat＇s feet to run to the safe area. The lighting lasted for 15 seconds as one test. Running from unsafe area to safe area at one time in 10 seconds was justified as successful. Such test was repeated for 10 times for each rat and the successful frequency was recorded. The qualified standard of maze test was that the rat ardved in the safe area g times during 10 experiments. The number of trainings for the qualified standard was used to represent the result of spatial learning. ③ Determination of the content of nNOS in brain tissue： After the Y-maze test, the rats were anaesthetized, and blood was let from the incision on right auricle, transcardially perfused via the left ventricle with about 200 mL saline, then fixed by perfusion of 40 g/L paraformaldehyde. Hippocampal CA1 region, corpus striatum and cerebellum were taken to prepare serial freezing coronal sections. The nNOS contents in the brain regions were determined with the immunohistochemical methods to reflect the changes of nitdc oxide in brain tissue. MAIN OUTCOME MEASURES ： The changes of learning and memory ability and the changes of the nNOS contents in the brain tissue of rats with acute alcoholism were observed. RESULTS ： One rat in the experimental group was excluded due to its slow reaction to electdc stimulation in the Y-maze test, and the other 17 rats were involved in the analysis of results. ① The training times to reach qualifying standards of Y-maze in the expedmental group was more than that in the control group [（34.33 ±13.04）, （27.50±8.79） times, P〈 0.05]. ② Forms and numbers of nNOS positive neurons in brain tissue： It could be observed under light microscope that in the hippocampal CA1 region, there were fewer nNOS positive neurons, which were lightly stained, and the processes were not clear enough; But the numbers of the positive neurons which were deeply stained as huffy were obviously increased in the experimental group, the cell body and cyloplasm of process were evenly stained, but the nucleus was not stained. The nNOS positive neurons in corpus stdatum had similar forms and size in the experimental group and control group. The form of the nNOS positive neurons in cerebellum were similar between the two groups. The numbers of nNOS positive neurons in hippocampal CA1 region and corpus striatum in the expedmental group [（18.22±7.47）, （11.38±5.00） cells/high power field] were obviously higher than those in the control group [（10.15±4.24）, （6.15±3.69） cells/high power field. The number of nNOS positive neurons in cerebellum had no significant difference between the two groups [（49.56±18.84）, （44.43±15.42） cells/high power field, P〉 0.05]. CONCLUSION ： Acute alcoholism may impair learning and memory ability, and nitric oxide may be involved in mediating the neurotoxic role of ethanol.

Study on Protection Mechanism of Viscum coloratum (Kom.) Nakai f. lutescens kitag Fruit Polysaccharides on Mice with Acute Alcoholic Liver Injury

This study was conducted to investigate the protective mechanism of V. coloratum fruit polysaccharides（ VCFP） on mice with acute alcoholic liver injury. Acute liver injury model was built by one-time alcohol gavage. The mice were randomly divided into VCFP-treated groups（ low-,medium-,and highdose）,alcohol model group and normal control group at the same time. VCFP-treated groups were administrated polysaccharide intragastrically continuously for4 weeks; and the alcohol model group and normal control group were administrated intragastrically the same amount of normal saline. The general situation and liver tissue morphological structure changes were observed. The results showed that the levels of ALT,AST,TC,TG,MDA contents,and CAT and GSH-Px activity of mice in the model group increased significantly（ P 〈 0. 01）,while the activity of SOD and weight and liver index decreased significantly（ P 〈 0. 01） compared with the normal control group. After 4 weeks of gavage,VCFP could significantly improve weight,liver index and SOD activity,and significantly reduce ALT,AST,TC and TG levels,MDA content and CAT and GSH-Px activity. These results suggest that VCFP can improve antioxidant enzyme activity,remove oxygen free radical and reduce membrane lipid peroxidation reaction,thereby protecting liver cells.

Liver transplantation for alcoholic liver disease

Alcoholic liver disease(ALD) is the second commonest indication for liver transplantation after viral hepatitis in the United States and Europe.Controversies surround the indications and allocation of scarce and expensive resource for this so called self inflicted disease.Controversies stem from the apprehension that alcoholic recipients are likely to relapse and cause damage to the graft.There is a need to select those candidates with lower risk for relapse with the available predictive factors and scores.Substance abuse specialist and psychiatrists are mandatory in the pre-transplant evaluation and in the post-transplant follow-up.There is conflicting evidence to support a fixed period of pretransplant abstinence,although most units do follow this.Alcoholic hepatitis(AH) continues to be a contraindication for transplantation,however there is a need for further research in this f ield as a subset of patients with AH who do not respond to medical treatment,have high early mortality and could benefit from transplantation.One year,3-year,and 5-year survival post-transplant is similar for both ALD and non-ALD recipients.The incidence of post-transplant rejection and retransplantation is also similar to other recipients.ALD with viral hepatitis especially hepatitis C virus leads to a more aggressive liver disease with early presentation for transplantation.ALD patients are more prone to develop de-novo malignancy;this is attributed to the long term effect of alcohol,tobacco combined with immunosuppression.Post-transplant surveillance is important to detect early relapse to alcoholism,presence of de-novo malignancy and treat the same adequately.

Structural Shifts of Gut Flora in Rat Acute Alcoholic Liver Injury and Jianpi Huoxue Decoction's(健脾活血汤)Effect Displayed by ERIC-PCR Fingerprint

Objective： To study the structural shifts of gut flora in rats with acute alcoholic liver injury （AALI）, and the effect of Jianpi Huoxue Decoction （健脾活血汤, JPHXD） on the gut flora. Methods： Thirty-six Sprague- Dawley rats were randomly allocated to the control, AALI and JPHXD groups equally. The rats in the control group were given water and those in AALI and JPHXD groups were given ethanol by intragastric gavage for 5 days, while rats in the JPHXD group were administered JPHXD simultaneously. The blood and liver tissue were collected at the end of the experiment. The activities of serum alkaline aminotransferase （ALT）, aspartate aminotransferase （AST）, hepatic γ/-glutamyitranspetidase （γ-GT） and hepatic tdglyceride （TG） levels were determined. Plasma endotoxin level in the portal vein was measured. Pathological changes of liver tissues were determined by hematoxylin and Eosin （HE） staining and oil red O staining. The total DNA of gut flora were extracted from fecal samples by Bead-beating method and determined by ERIC-PCR fingerprint method. The similarity cluster analysis and principal component analysis were performed to analyze the ERIC-PCR fingerprint respectively. Results： In the AALI group, the ratio of liver/body weight, activities of ALT, AST and hepatic γ-GT, amount of hepatic TG were elevated significantly compared with those in the control group （all P〈0.01）. JPHXD decreased the ratio, activities of ALT, AST, γ-GT and TG significantly compared with those in the AALI group （P〈0.05 or P〈0.01）. HE and oil red O staining showed that fat deposited markedly in liver tissue, while JPHXD alleviated pathological changes markedly. Plasma LPS level in rat portal vein in the AALI group increased significantly （P〈0.01）, but it was decreased significantly in the JPHXD group （P〈0.01）. The cluster analysis and principal component analysis of ERIC-PCR fingerprint showed that gut flora in the hALl group changed markedly, and JPHXD could recover gut flora to some extent. Conclusion： The structure of gut flora shifted markedly during acute alcoholic liver injury, JPHXD had prevention effect through the modification of gut flora.

Forensic appraisal of death due to acute alcohol poisoning:three case reports and a literature review

Death due to acute alcohol poisoning lacks specific anatomical characteristics,compared with other deaths due to drug poisoning.We report three forensic cases of death from acute alcohol poisoning due to inhibition of the respiratory centre and eventual asphyxia.Blood alcohol concentrations in the three fatalities were 5.28,3.33 and 3.78mg/mL,respec-tively.Lethal doses and blood alcohol concentrations showed differences between individu-als.Detailed auxiliary tests besides autopsy were undertaken.These cases show that forensic scientists should exclude other causes of death,combine the autopsy with auxiliary tests,and then make an appraisal.