Treatment with Melatonin Improves Cognitive Behavior and Motor Skills in a Rat Model of Liver Fibrosis.

INTRODUCTION AND AIM: Patients with liver cirrhosis (LC) and minimal hepatic encephalopaty have a higher accident rate. LC impairs the normal sleep-awake cycle and produces disturbances in behavior, cognition and motor skills. Abnormal melatonin (MT) levels have also been identified in LC. Administration of MT may regulate circadian rhythms and prevent the oxidative damage. We studied the effects of MT on spatial memory acquisition (SMA) and motor skills in a liver fibrosis model (LF)s. MATERIALS AND METHODS: Forty-five rats, divided into 4 groups. [G1: LF; G2: LF + MT; G3: MT; G4: Healthy control (HC)]. LF was induced by carbon tetrachloride intraperitoneal injection (0.2 mL/kg) for 5 months. MT was administered during 5 weeks (0.4 mg/kg/day). SMA was evaluated by using the Morris Water Maze protocol where the escape latency (EL) and mean speed were measured. Data were registered by SMART®. RESULTS: The EL measurement analyzed by two way ANOVA: cirrhosis presented a higher EL than controls or those treated with MT suggesting impaired memory acquisition which is rescued by MT treatment. The mean speed analysis revealed that LF presented higher speed than LF+MT or HC, suggesting that LF affects motor skills, which are improved by MT. To discard whether EL is affected by altered motor skills in LF treated with MT, we compared the average EL and speed between days 2 and 6 of the training protocol. Speed was not improved during the trials unlike EL, suggesting that memory acquisition is independent of motor skills. CONCLUSION: These findings suggest that MT improves cognition and motor skills in the LF model.

Multiorgan chronic inflammatory hepatobiliary pancreatic murine model deficient in tumor necrosis factor receptors 1 and 2.

AIM: To provoke persistent/chronic multiorgan inflammatory response and to contribute to stones formation followed by fibrosis in hepatobiliary and pancreatic tissues. METHODS: Tumor necrosis factor receptors 1 and 2 (TNFR1/R2) deficient mice reared in-house were given dibutyltin dichloride (DBTC) twice within 10 d by oral gavage delivery. Sham control animals received vehicle treatment and naïve animals remained untreated throughout the study. Animals were monitored daily for symptoms of pain and discomfort. The abdominal and hindpaw hypersensitivity were assessed with von Frey microfilaments. Exploratory behaviors were recorded at the baseline, after initiation of treatment, and before study termination. Histopathological changes were examined postmortem in tissues. Collagen accumulation and fibrosis were confirmed with Sirius Red staining. RESULTS: Animals lost weight after oral administration of DBTC and developed persistent inflammatory abdominal and hindpaw hypersensitivity compared to sham-treated controls (P < 0.0001). These pain related secondary mechanical hypersensitivity responses increased more than 2-fold in DBTC-treated animals. The drastically diminished rearing and grooming rates persisted after DBTC administration throughout the study. Gross as well as micropathology at one month confirmed that animals treated with DBTC developed chronic hepatobiliary injuries evidenced with activation of stellate cells, multifocal necrosis, fatty degeneration of hepatocytes, periportal infiltration of inflammatory cells, and prominent biliary ductal dilation. The severity of hepatitis was scored 3.7 ± 0.2 (severe) in DBTC-treated animals vs score 0 (normal) in sham-treated animals. Fibrotic thickening was extensive around portal ducts, in hepatic parenchyma as well as in lobular pancreatic structures and confirmed with Sirius Red histopathology. In addition, pancreatic microarchitecture was presented with distortion of islets, and parenchyma, infiltration of inflammatory cells, degeneration, vacuolization, and necrosis of acinar cells and distention of pancreatic ducts. Extent of pancreatic damage and pancreatitis were scored 3.6 ± 0.4 (severe) for DBTC-treated in contrast to score 0 (normal) in sham-treated animals. The gall bladder became expanded with ductal distention, and occasional bile stones were detected along with microscopic hepatic lesions. DBTC-treated animals developed splenic hypertrophy with increased weight and length (P < 0.01) along with thymic atrophy (P < 0.001). Finally, colitic lesions and colitis were prominent in DBTC-treated animals and scored 3.4 ± 0.3 (moderately severe) vs 0 (normal) for the sham-treated animals. CONCLUSION: This is the first report of chronic inflammatory multiorgan hepatobiliary pancreatitis, along with fibrosis and calculi formation induced reliably utilizing oral DBTC administration in TNFR1/R2 deficient mice.

Low grade cirrhosis induces cognitive impairment and motor dysfunction in rats: could be a model for minimal hepatic encephalopathy.

Hepatic encephalopathy (HE) is a nervous system disorder developed in the patients with liver cirrhosis. The low grade cirrhosis is of common occurrence, however, whether and the extent to which it affects brain function is not clearly understood. The present article examines certain neurobehavioral parameters in the rats with low grade chronic liver failure (CLF) induced by intraperitoneal administration of 50mg/kgb.w. thioacetamide up to 14 days. During Morris Water Maze tasks, the CLF rats, as compared to the control, showed insignificant decline in the escape latency score to find out the hidden platform throughout the learning days and also stayed for a significantly declined (p<0.01) time period at the place of the hidden platform during the retrieval test. They also showed impairment in the conditional discrimination ability, reflected by a significant decline in the active avoidance score (p<0.05) and increment in the number of non-response (p<0.05) during shuttle box tests. On rotarod performance, they exhibited significant decline in their riding time (p<0.01-001) on the rotating rod as a function of increasing speed. The findings suggest a moderate level cognitive impairment and motor dysfunction in the low grade CLF rats. Since, these impairments correlate with the early stage manifestation of HE in the patients, these CLF rats could serve as a model to study the pathogenesis of minimal hepatic encephalopathy.

Brain cholinergic impairment in liver failure.

The cholinergic system is involved in specific behavioural responses and cognitive processes. Here, we examined potential alterations in the brain levels of key cholinergic enzymes in cirrhotic patients and animal models with liver failure. An increase (~30%) in the activity of the acetylcholine-hydrolyzing enzyme, acetylcholinesterase (AChE) is observed in the brain cortex from patients deceased from hepatic coma, while the activity of the acetylcholine-synthesizing enzyme, choline acetyltransferase, remains unaffected. In agreement with the human data, AChE activity in brain cortical extracts of bile duct ligated (BDL) rats was increased (~20%) compared to controls. A hyperammonemic diet did not result in any further increase of AChE levels in the BDL model, and no change was observed in hyperammonemic diet rats without liver disease. Portacaval shunted rats which display increased levels of cerebral ammonia did not show any brain cholinergic abnormalities, confirming that high ammonia levels do not play a role in brain AChE changes. A selective increase of tetrameric AChE, the major AChE species involved in hydrolysis of acetylcholine in the brain, was detected in both cirrhotic humans and BDL rats. Histological examination of BDL and non-ligated rat brains shows that the subcellular localization of both AChE and choline acetyltransferase, and thus the accessibility to their substrates, appears unaltered by the pathological condition. The BDL-induced increase in AChE activity was not parallelled by an increase in mRNA levels. Increased AChE in BDL cirrhotic rats leads to a pronounced decrease (~50-60%) in the levels of acetylcholine. Finally, we demonstrate that the AChE inhibitor rivastigmine is able to improve memory deficits in BDL rats. One week treatment with rivastigmine (0.6 mg/kg; once a day, orally, for a week) resulted in a 25% of inhibition in the enzymatic activity of AChE with no change in protein composition, as assessed by sucrose density gradient fractionation and western blotting analysis. In conclusion, this study is the first direct evidence of a cholinergic imbalance in the brain as a consequence of liver failure and points to the possible role of the cholinergic system in the pathogenesis of hepatic encephalopathy.

Brain serotonin metabolism and behavior in rats with carbon tetrachloride-induced liver cirrhosis.

Increased brain serotonin metabolism has been suggested as an etiologic factor in the development of portal-systemic encephalopathy (PSE) in connection with liver disease. We therefore investigated brain serotonin metabolism and open-field behavior (spontaneous activity and exploration) in rats with carbon tetrachloride (CCl4)-induced liver cirrhosis. Brain serotonin metabolism was evaluated in rats pretreated with an amino acid decarboxylase inhibitor. The 5-hydroxyindoles were analyzed by high-performance liquid chromatography (HPLC) with electrochemical detection. The results revealed an increased serotonin synthesis rate in all investigated brain regions in rats with histologically verified diffuse micronodular cirrhosis of the liver. Slightly impaired open-field behavior (i.e., decreased spontaneous activity) in the cirrhotic rats could not be excluded. However, the elevated brain serotonin synthesis rate could not be correlated to any abnormalities in open-field behavior.