Monoamines and metabolites in autopsied brain tissue from cirrhotic patients with hepatic encephalopathy

Alterations in the metabolism of monoamine neurotransmitters have been proposed to be involved in the development of the hepatic encephalopathy (HE) associated with experimental and human liver failure. In order to evaluate this hypothesis, the monoamines and some of their metabolites were measured in homogenates of caudate nucleus (CAU), prefrontal (PFCo) and frontal cortex (FCo) dissected from brains obtained at autopsy from nine cirrhotic patients who had died in hepatic coma and an equal number of control subjects, free from neurological, psychiatric and hepatic disorders, matched for age and time interval from death to freezing of autopsied brain samples. Monoamine measurements were performed by high-performance liquid chromatography with ion-pairing and electrochemical detection after a simple extraction procedure. In all three regions investigated, concentrations of dopamine (DA) were unchanged in cirrhotic patients vs controls while its metabolites, 3-methoxytyramine (3-MT) and homovanillic acid (HVA) were selectively affected i.e.3-MT was found to be increased in CAU, while HVA levels were increased in FCo and CAU. DOPAC was also found to be unchanged in CAU. Noradrenaline (NA) levels were greatly increased in PFCo and FCo of cirrhotic patients but remained unchanged in CAU. No significant differences in the concentrations of either serotonin (5-HT) or of its precursor 5-hydroxytryptophan (5-HTP) were found in any of the three regions studied. However, 5-hydroxyindoleacetic acid (5-HIAA), the major metabolite of 5-HT, was increased in PFCo and CAU of cirrhotic patients. These findings show that selective alterations of catecholamine and 5-HT systems are involved in human HE and therefore, they may play an important role in the pathogenesis of certain neurological symptoms associated with this encephalopathy.     

Reduced brain levels of DHEAS in hepatic coma patients: significance for increased GABAergic tone in hepatic encephalopathy

Increased neurosteroids with allosteric modulatory activity on GABA(A) receptors such as 3α-5α tertrahydroprogesterone; allopregnanolone (ALLO), are candidates to explain the phenomenon of "increased GABAergic tone" in hepatic encephalopathy (HE). However, it is not known how changes of other GABA(A) receptor modulators such as dehydroepiandrosterone sulfate (DHEAS) contribute to altered GABAergic tone in HE. Concentrations of DHEAS were measured by radioimmunoassay in frontal cortex samples obtained at autopsy from 11 cirrhotic patients who died in hepatic coma and from an equal number of controls matched for age, gender, and autopsy delay intervals free from hepatic or neurological diseases. To assess whether reduced brain DHEAS contributes to increased GABAergic tone, in vitro patch clamp recordings in rat prefrontal cortex neurons were performed. A significant reduction of DHEAS (5.81±0.88 ng/g tissue) compared to control values (9.70±0.79 ng/g, p<0.01) was found. Brain levels of DHEAS in patients with liver disease who died without HE (11.43±1.74 ng/g tissue), and in a patient who died in uremic coma (12.56 ng/g tissue) were within the control range. Increasing ALLO enhances GABAergic tonic currents concentration-dependently, but increasing DHEAS reduces these currents. High concentrations of DHEAS (50 μM) reduce GABAergic tonic currents in the presence of ALLO, whereas reduced concentrations of DHEAS (1 μM) further stimulate these currents. These findings demonstrate that decreased concentrations of DHEAS together with increased brain concentrations of ALLO increase GABAergic tonic currents synergistically; suggesting that reduced brain DHEAS could further increase GABAergic tone in human HE.     

Increased CSF levels of somatostatin in patients with brain tumours and intracranial hypertension

The cerebrospinal fluid (CSF) levels of somatostatin in patients with brain tumours, communicating hydrocephalus, lumbar-disc disease (treated as a control) were measured by specific radioimmunoassay. The somatostatin concentration in the patients with brain tumours and intracranial hypertension was significantly higher compared to those with brain tumours and normal CSF pressure. CSF somatostatin content in patients with communicating hydrocephalus, was similar to patients with brain tumours and normal CSF pressure, and did not show a significant difference from the control group. The authors discuss possible reasons for such results obtained in patients with brain tumours and intracranial hypertension.     

The effect of hepatic encephalopathy, hepatic failure, and portosystemic shunt on brain volume of cirrhotic patients: a voxel-based morphometry study

Purpose

To evaluate the effect of hepatic encephalopathy (HE), hepatic failure, and portosystemic shunt (PS) on the brain volume alteration in cirrhotic patients with MRI voxel-based morphometry (VBM).

Methods

Sixty cirrhotic patients (overt HE [OHE], n = 11; minimal HE [MHE], n = 19; non HE [nHE], n = 30) including 12 with pre- and post-transjugular intrahepatic portosystemic shunt (TIPS) scanning and 40 healthy controls were recruited. Neuropsychological and laboratory tests were performed in all patients. VBM was analyzed with ANOVA test among 4 groups, and t-tests for patients with different hepatic function, PS scores, and TIPS. Multiple linear regression was performed to investigate the effect of venous blood ammonia levels, Child-Pugh scores, and PS on the brain volumes in all patients.

Results

Cirrhotic patients exhibited decreased volume in many areas of gray matter (GM), increased volume in thalamus, and increased whiter matter (WM) volume, with the extent of affected brain volume greater in HE patients than nHE patients. Hepatic failure also resulted in decreased GM volume. Patients with high PS scores and TIPS displayed decreased GM and increased WM volume in some regions. Post-TIPS patients displayed increased GM volume in the thalamus. Multiple covariate regression results suggested that Child-Pugh score was a major factor to affect GM volume, while PS mainly affected WM volume.

Conclusion

Brain structure abnormalities appeared bilaterally symmetrical in cirrhotic patients, and the impairment was more extensive in HE patients than those without HE. Increased thalamus volume was not associated with HE progression. Hepatic failure and PS altered cirrhotic patients’ brain structure.     

Drug metabolism in human brain: high levels of cytochrome P4503A43 in brain and metabolism of anti-anxiety drug alprazolam to its active metabolite

Cytochrome P450 (P450) is a super-family of drug metabolizing enzymes. P450 enzymes have dual function; they can metabolize drugs to pharmacologically inactive metabolites facilitating their excretion or biotransform them to pharmacologically active metabolites which may have longer half-life than the parent drug. The variable pharmacological response to psychoactive drugs typically seen in population groups is often not accountable by considering dissimilarities in hepatic metabolism. Metabolism in brain specific nuclei may play a role in pharmacological modulation of drugs acting on the CNS and help explain some of the diverse response to these drugs seen in patient population. P450 enzymes are also present in brain where drug metabolism can take place and modify therapeutic action of drugs at the site of action. We have earlier demonstrated an intrinsic difference in the biotransformation of alprazolam (ALP) in brain and liver, relatively more alpha-hydroxy alprazolam (alpha-OHALP) is formed in brain as compared to liver. In the present study we show that recombinant CYP3A43 metabolizes ALP to both alpha-OHALP and 4-hydroxy alprazolam (4-OHALP) while CYP3A4 metabolizes ALP predominantly to its inactive metabolite, 4-OHALP. The expression of CYP3A43 mRNA in human brain samples correlates with formation of relatively higher levels of alpha-OH ALP indicating that individuals who express higher levels of CYP3A43 in the brain would generate larger amounts of alpha-OHALP. Further, the expression of CYP3A43 was relatively higher in brain as compared to liver across different ethnic populations. Since CYP3A enzymes play a prominent role in the metabolism of drugs, the higher expression of CYP3A43 would generate metabolite profile of drugs differentially in human brain and thus impact the pharmacodynamics of psychoactive drugs at the site of action.     

Increased levels of 4-hydroxynonenal and acrolein in the brain in preclinical Alzheimer disease

Previous studies demonstrate increased levels of 4-hydroxynonenal (HNE) and acrolein in vulnerable brain regions of subjects with mild cognitive impairment and late-stage Alzheimer disease (LAD). Recently preclinical AD (PCAD) subjects, who demonstrate normal antemortem neuropsychological test scores but abundant AD pathology at autopsy, have become the focus of increased study. Levels of extractable HNE and acrolein were quantified by gas chromatography–mass spectrometry with negative chemical ionization, and protein-bound HNE and acrolein were quantified by dot-blot immunohistochemistry in the hippocampus/parahippocampal gyrus (HPG), superior and middle temporal gyri (SMTG), and cerebellum (CER) of 10 PCAD and 10 age-matched normal control (NC) subjects. Results of the analyses show a significant (P < 0.05) increase in levels of extractable acrolein in the HPG of PCAD subjects compared to age-matched NC subjects and a significant decrease in extractable acrolein in PCAD CER. Significant increases in protein-bound HNE in HPG and a significant decrease in CER of PCAD subjects compared to NC subjects were observed. No significant alterations were observed in either extractable or protein-bound HNE or acrolein in the SMTG of PCAD subjects. Additionally, no significant differences in levels of protein carbonyls were observed in the HPG, SMTG, or CER of PCAD subjects compared to NC subjects.     

Parecoxib and its metabolite valdecoxib directly interact with cannabinoid binding sites in CB1-expressing HEK 293 cells and rat brain tissue

Cyclooxygenase 2 inhibitors (COX 2) such as parecoxib (par) and valdecoxib (val) are used in the treatment of neuropathic pain. Using the radioligand binding assay it was demonstrated that both the prodrug par as well as its active metabolite val have a specific affinity to the cannabinoid (CB) receptor measured in CB1-expressing HEK 293 cells and rat brain tissue. Agonist activity was detected by GTPγS assays, cAMP formation experiments and ex vivo modulation of glutamate and GABA release of the rat brain tissue. In comparison to the specific cannabinoid agonist, WIN 55,212-2, the two COX 2 inhibitors are about 2 orders of magnitude less potent. The data suggest that the analgesic effects of par and its metabolite val in Wistar rats may be at least partially mediated by a direct interaction with the CB1 receptors. The COX 2 inhibitors appear to be a hypothetically useful tool for add-on therapy of neuropathic pain.     

Increased plasma and tissue levels of vitamin E in the spontaneously diabetic BB rat

Increased plasma and tissue levels of vitamin E were found in spontaneously diabetic BB rats (D) as well as asymptomatic/diabetes-prone BB rats (AD) in comparison to levels in non-diabetic control rats (ND). Treatment of D rats with insulin for 30 days returned plasma and tissue values of vitamin E to control levels. The changes reported here could not be explained solely on the basis of variations in total lipid content of plasma. These data suggest the metabolism of vitamin E is altered in asymptomatic and spontaneously diabetic BB rats and this alteration returns to control values following insulin treatment. Furthermore, it might be speculated that these data indicate a relationship between vitamin E and insulin.     

Increased glucose metabolism and ATP level in brain tissue of Huntington's disease transgenic mice

Huntington's disease (HD) is a progressive neurodegenerative disorder characterized by multifarious dysfunctional alterations including mitochondrial impairment. In the present study, the formation of inclusions caused by the mutation of huntingtin protein and its relationship with changes in energy metabolism and with pathological alterations were investigated both in transgenic and 3-nitropropionic acid-treated mouse models for HD. The HD and normal mice were characterized clinically; the affected brain regions were identified by immunohistochemistry and used for biochemical analysis of the ATP-producing systems in the cytosolic and the mitochondrial compartments. In both HD models, the activities of some glycolytic enzymes were somewhat higher. By contrast, the activity of glyceraldehyde-3-phosphate dehydrogenase was much lower in the affected region of the brain compared to that of the control. Paradoxically, at the system level, glucose conversion into lactate was enhanced in cytosolic extracts from the HD brain tissue, and the level of ATP was higher in the tissue itself. The paradox could be resolved by taking all the observed changes in glycolytic enzymes into account, ensuing an experiment-based detailed mathematical model of the glycolytic pathway. The mathematical modelling using the experimentally determined kinetic parameters of the individual enzymes and the well-established rate equations predicted the measured flux and concentrations in the case of the control. The same mathematical model with the experimentally determined altered V(max) values of the enzymes did account for an increase of glycolytic flux in the HD sample, although the extent of the increase was not predicted quantitatively. This suggested a somewhat altered regulation of this major metabolic pathway in HD tissue. We then used the mathematical model to develop a hypothesis for a new regulatory interaction that might account for the observed changes; in HD, glyceraldehyde-3-phosphate dehydrogenase may be in closer proximity (perhaps because of the binding of glyceraldehyde-3-phosphate dehydrogenase to huntingtin) with aldolase and engage in channelling for glyceraldehyde-3-phosphate. By contrast to most of the speculation in the literature, our results suggest that the neuronal damage in HD tissue may be associated with increased energy metabolism at the tissue level leading to modified levels of various intermediary metabolites with pathological consequences.     

The effects of a single acute dose of dexamethasone on monoamine and metabolite levels in rat brain

Twenty male Sprague-Dawley rats were injected intraperitoneally with either 20 micrograms of dexamethasone or an equivalent volume of saline. The rats were then sacrificed at either one or four hours after the injections and their brains analyzed for monoamine and metabolite content using High Performance Liquid Chromatography with Electrochemical Detection. Significant effects were seen in dopaminergic and serotonergic systems, but these effects varied depending on the area of rat brain studied. Significant increases in dopamine (DA) levels were seen in the hypothalamus and nucleus accumbens of the dexamethasone treated rats when compared with saline treated rats. There was no significant effect of dexamethasone on DA levels in frontal or striatal brain areas. In the dexamethasone treated rats a significant increase in serotonin (5-HT) was observed in the hypothalamus; a significant decrease in 5-HT was observed in the frontal cortex. Biological and clinical implications of these findings are discussed.