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Biomedical subjects

Vicente Felipo

Publications and source records attributed to Vicente Felipo.

At least 37 records · Page 2Linked to original sources
PubMed2005

Limited capacity for ammonia removal by brain in chronic liver failure: potential role of nitric oxide.

Chronic liver failure leads to hyperammonemia and consequently increased brain ammonia concentrations, resulting in hepatic encephalopathy. When the liver fails to regulate ammonia concentrations, the brain, devoid of a urea cycle, relies solely on the amidation of glutamate to glutamine through glutamine synthetase, to efficiently clear ammonia. Surprisingly, under hyperammonemic conditions, the brain is not capable of increasing its capacity to remove ammonia, which even decreases in some regions of the brain. This non-induction of glutamine synthetase in astrocytes could result from possible limiting substrates or cofactors for the enzyme, or an indirect effect of ammonia on glutamine synthetase expression. In addition, there is evidence that nitration of the enzyme resulting from exposure to nitric oxide could also be implicated. The present review summarizes these possible factors involved in limiting the increase in capacity of glutamine synthetase in brain, in chronic liver failure.

Ammonia↗
PubMed2005

Altered modulation of motor activity by group I metabotropic glutamate receptors in the nucleus accumbens in hyperammonemic rats.

One of the neurological complications in hepatic encephalopathy is the impairment of motor coordination and function. Clinical signs of basal ganglia, cortico-spinal and cerebellar dysfunction have been commonly detected in these patients. We are studying the molecular bases of the alterations in motor coordination and function in hepatic encephalopathy. Hyperammonemia is considered the main factor responsible for the neurological alterations in patients with hepatic encephalopathy. Activation of metabotropic glutamate receptors (mGluRs) in the nucleus accumbens (NAcc) induces locomotion in rats. Asa first step in our studies on the alterations in motor co-ordination and function in hyperammonemia and hepatic encephalopathy we studied whether the control of motor function by mGluRs in the NAcc is altered in hyperammonemic rats. The locomotor activity induced by injection into the nucleus accumbens (NAcc) of DHPG, an agonist of group I mGluRs was significantly increased in hyperammonemic rats. Injection of DHPG increased extracellular dopamine but not glutamate in the NAcc of control rats. In hyperammonemic rats DHPG-induced increase in dopamine was significantly reduced, and extracellular glutamate increased 6-fold. The content of mGluR 1 but not mGluR 5, is increased in the NAcc of hyperammonemic rats. Blockade of mGluR 1 completely prevented motor and neurochemical effects induced by DHPG. These results show that modulation of both motor function and extracellular concentration of neurotransmitters by mGluRs in the NAcc is altered in hyperammonemia. This may contribute to the alterations in motor function in hepatic encephalopathy.

Animals↗
PubMed2005

Animal models in the study of episodic hepatic encephalopathy in cirrhosis.

The availability of an animal model is crucial in studying the pathophysiological mechanisms of disease and to test possible therapies. Now, there are several models for the study of liver diseases, but there still remains a lack of a satisfactory animal model of chronic liver disease with hepatic encephalopathy (HE) and abnormalities in nitrogen metabolism, as seen in humans. In rats, two models of chronic HE are widely used: rats after portacaval anastomosis (PCA) and rats with chronic hyperammonemia. The first one mimics the situation induced in cirrhosis by collateral circulation, and has the problem of the absence of hepatocellular injury. The model of hyperammonemia is useful to study the effect of ammonia as a brain toxic substance, but also lacks liver failure. Bile-duct ligation has been used to induce cirrhosis and could also be a model of HE, probably with the addition of a precipitant factor. An ideal model of HE in chronic liver disease must have liver cirrhosis and a precipitant factor of HE; it must also show neuropathological characteristic findings of HE, neurochemical alterations in the main pathways impaired in these complications of cirrhosis, and low-grade brain edema.

Animals↗
PubMed2005

Serines 890 and 896 of the NMDA receptor subunit NR1 are differentially phosphorylated by protein kinase C isoforms.

The NR1 subunit of the NMDA receptor has two serines (S890 and S896) whose phosphorylation by protein kinase C (PKC) differentially modulates NMDA receptor trafficking and clustering. It is not known which PKC isoforms phosphorylate these serines. In primary cultures of cerebellar neurons, we examined which PKC isoforms are responsible for the phosphorylation S890 and S896. We used specific inhibitors of PKC isoforms and antibodies recognizing specifically phosphorylated S890 or S896. The results show that PKC alpha phosphorylates preferentially S896 and PKC gamma preferentially S890. Activation of type I metabotropic glutamate receptors (mGluRs) with DHPG (3,5-dihyidroxy-phenylglycine) activates PKC gamma but not PKC alpha or beta. We found that activation of mGluRs by DHPG increases S890 but not S896 phosphorylation, supporting a role for PKC gamma in the physiological modulation of S890 phosphorylation. It is also shown that the pool of NR1 subunits present in the membrane surface contains phosphorylated S890 but not phosphorylated S896. This supports that differential phosphorylation of S890 and S896 by different PKC isoforms modulates cellular distribution of NMDA receptors and may also contribute to the selective modulation of NMDA receptor function and intracellular localization.

Amino Acid Sequence↗
PubMed2005

Modulation of NMDA receptors in the cerebellum. 1. Properties of the NMDA receptor that modulate its function.

NMDA receptors modulate important cerebral processes such as synaptic plasticity, long-term potentiation, learning and memory, etc. NMDA receptors in cerebellum have specific characteristics that make their function and modulation different from those of NMDA receptors in other brain areas. In this and the accompanying review we summarize the information available on the modulation of NMDA receptors in cerebellum. We review the properties of the NMDA receptor that modulate its function: subunit composition, post-translational modifications and synaptic localization. NMDA receptors are heteromeric ligand-gated ion channels assembled from two families of subunits, NR1 and NR2. There are at least eight splicing variant isoforms of the NR1 subunit and four types of NR2 subunits: NR2A, NR2B, NR2C and NR2D. NMDA receptors with different subunit composition or different splice variants of NR1 subunit have different properties. The expression of the different subunits and splicing variants varies during development. Two special characteristics of NMDA receptors in cerebellum that do not occur in other brain areas are the enrichment in the NR2C subunit and in the splice variant NR1b. As a consequence of these and other factors the pharmacology of NMDA receptors is also different in cerebellum than in other brain areas. The function and localization of NMDA receptors is also modulated by postranslational modifications including phosphorylation, glycosylation and nytrosylation. NMDA receptors are phosphorylated in serines of both NR1 and NR2 subunits and in tyrosines of NR2 subunits. Another factor modulating NMDA receptors function is the synaptic localization. The trafficking and clustering of NMDA receptors is modulated by phosphorylation and by interaction with other proteins. The signaling pathways and physiological modulators regulating NMDA receptor function as well as the role of these receptors in motor learning and coordination are reviewed in an accompanying article.

Alternative Splicing↗
PubMed2005

Modulation of NMDA receptors in the cerebellum. II. Signaling pathways and physiological modulators regulating NMDA receptor function.

NMDA receptors in cerebellum have specific characteristics that make their function and modulation different from those of NMDA receptors in other brain areas. The properties of the NMDA receptor that modulate its function: Subunit composition, post-translational modifications and synaptic localization are summarized in an accompanying article. In this review we summarize how different signaling molecules modulate the function of NMDA receptors. The function of the receptors is modulated by the co-agonists glycine and serine and this modulation is different in cerebellum than in other areas. The NMDA receptor also has binding sites for polyamines that regulate its function. Other signaling molecules that modulate NMDA receptors function are: cAMP, neurotrophic factors such as BDNF, FGF-2 or neuregulins. These and other molecules allow an interplay between NMDA receptors and other receptors for neurotransmitters that may in this way modulate NMDA receptor function. This has been reported, for example, for metabotropic glutamate receptors. The expression and function of NMDA receptor is also modulated by synaptic activity, allowing an adaptation of the receptors function to the external inputs. NMDA receptors modulate important cerebral processes. NMDA receptors in different brain areas seem to modulate different processes. Cerebellar NMDA receptors play a special role in the modulation of motor learning and coordination. This is also briefly reviewed.

Animals↗
PubMed2005

Chronic exposure to ammonia induces isoform-selective alterations in the intracellular distribution and NMDA receptor-mediated translocation of protein kinase C in cerebellar neurons in culture.

Hyperammonemia is responsible for most neurological alterations in patients with hepatic encephalopathy by mechanisms that remain unclear. Hyperammonemia alters phosphorylation of neuronal protein kinase C (PKC) substrates and impairs NMDA receptor-associated signal transduction. The aim of this work was to analyse the effects of hyperammonemia on the amount and intracellular distribution of PKC isoforms and on translocation of each isoform induced by NMDA receptor activation in cerebellar neurons. Chronic hyperammonemia alters differentially the intracellular distribution of PKC isoforms. The amount of all isoforms (except PKC zeta) was reduced (17-50%) in the particulate fraction. The contents of alpha, beta1, and epsilon isoforms decreased similarly in cytosol (65-78%) and membranes (66-83%), whereas gamma, delta, and theta; isoforms increased in cytosol but decreased in membranes, and zeta isoform increased in membranes and decreased in cytosol. Chronic hyperammonemia also affects differentially NMDA-induced translocation of PKC isoforms. NMDA-induced translocation of PKC alpha and beta is prevented by ammonia, whereas PKC gamma, delta, epsilon, or theta; translocation is not affected. Inhibition of phospholipase C did not affect PKC alpha translocation but reduced significantly PKC gamma translocation, indicating that NMDA-induced translocation of PKC alpha is mediated by Ca2+, whereas PKC gamma translocation is mediated by diacylglycerol. Chronic hyperammonemia reduces Ca+2-mediated but not diacylglycerol-mediated translocation of PKC isoforms induced by NMDA.

Ammonia↗
PubMed2005

Chronic hyperammonemia in vivo impairs long-term potentiation in hippocampus by altering activation of cyclic GMP-dependent-protein kinase and of phosphodiesterase 5.

Long-term potentiation (LTP) is impaired in the CA1 area of hippocampal slices from rats with chronic moderate hyperammonemia. We studied the mechanisms by which hyperammonemia in vivo impairs LTP. This process requires sequential activation of soluble guanylate cyclase, cyclic GMP-dependent protein kinase (PKG) and cyclic GMP-degrading phosphodiesterase. Application of the tetanus induced a rapid increase of cyclic GMP in slices from control or hyperammonemic rats, which is followed in control slices by a sustained decrease in cyclic GMP due to sustained activation of cyclic GMP-degrading phosphodiesterase, which in turn is due to sustained activation of PKG. In slices from rats with chronic hyperammonemia tetanus-induced decrease in cyclic GMP was delayed and transient due to lower and transient activation of PKG and of the phosphodiesterase. Hyperammonemia-induced impairment of LTP may be involved in the alterations of cognitive function in patients with hepatic encephalopathy.

3',5'-Cyclic-GMP Phosphodiesterases↗
PubMed2005

Correlation of nitric oxide and atrial natriuretic peptide changes with altered cGMP homeostasis in liver cirrhosis.

BACKGROUND: Cyclic GMP (cGMP) concentration is increased in plasma of patients with liver cirrhosis. Three possible mechanisms may contribute: increased cGMP synthesis by soluble (activated by nitric oxide), or particulate (activated by atrial natriuretic peptide (ANP)) guanylate cyclase or increased release from cells. AIM: The aim of this work was to analyze the possible contributors to increased plasma cGMP and to assess whether changes in the parameters of the system vary with the degree of liver disease (Child Pugh score) or by the presence of ascites. METHODS: We measured cGMP in plasma and lymphocytes, soluble guanylate cyclase activation by nitric oxide in lymphocytes, nitrates and nitrites and ANPs (activator of particulate guanylate cyclase) in plasma. We analyzed the correlation between changes in different parameters to discern which parameters contribute to increased plasma cGMP. RESULTS: The plasma content of nitrates+nitrites, ANP and cGMP are increased. Activation of soluble guanylate cyclase by nitric oxide is increased in patients while basal cGMP in lymphocytes is decreased. CONCLUSIONS: Both increased ANP and increased activation of soluble guanylate cyclase by nitric oxide contribute to increased plasma cGMP in patients. The concentrations of ANP and cGMP in plasma increase with the degree of disease and are higher in patients with ascites.

Adult↗
PubMed2004

Increased protein kinase A regulatory subunit content and cGMP binding in erythrocyte membranes in liver cirrhosis.

BACKGROUND/AIMS: Patients with liver disease show increased plasma cGMP and decreased intracellular cGMP in lymphocytes. The initial aim of this work was to assess whether decreased intracellular cGMP and increased plasma cGMP may be due to increased ATP-dependent release of cGMP from cells. The results obtained led to a new aim: to identify and quantify a protein responsible for the increased cGMP binding found in erythrocyte membranes from patients with liver disease. METHODS: ATP-dependent cGMP transport was determined in inside-out vesicles from erythrocyte membranes. cGMP-binding proteins were isolated from the membranes and identified by MALDI-TOF peptide mass fingerprint. Protein kinase A was quantified by immunoblotting. RESULTS: ATP-independent cGMP binding is increased in erythrocyte membranes from patients. There is a significant increase in the membrane content of a cGMP-binding protein with Mr 48,000, which was identified as the regulatory subunit of protein kinase A. CONCLUSIONS: The content of the regulatory subunit of protein kinase A is significantly increased (twice) in erythrocyte membranes from patients with liver cirrhosis. This protein binds cGMP strongly and may be responsible for the decrease in intracellular cGMP in liver disease.

Adult↗
PubMed2004

Hyperammonemia impairs long-term potentiation in hippocampus by altering the modulation of cGMP-degrading phosphodiesterase by protein kinase G.

Hyperammonemia impairs long-term potentiation (LTP) in hippocampus, by an unknown mechanism. LTP in hippocampal slices requires activation of the soluble guanylate cyclase (sGC)-protein kinase G (PKG)-cGMP-degrading phosphodiesterase pathway. The aim of this work was to assess whether hyperammonemia impairs LTP by impairing the tetanus-induced activation of this pathway. The tetanus induced a rapid cGMP rise, reaching a maximum at 10 s, both in the absence or presence of ammonia. The increase in cGMP is followed in control slices by a sustained decrease in cGMP due to PKG-mediated activation of cGMP-degrading phosphodiesterase, which is required for maintenance of LTP. Hyperammonemia prevents completely tetanus-induced cGMP decrease by impairing PKG-mediated activation of cGMP-degrading phosphodiesterase. Addition of 8Br-cGMP to slices treated with ammonia restores both phosphodiesterase activation and maintenance of LTP. Impairment of LTP in hyperammonemia may be involved in the impairment of the cognitive function in patients with hepatic encephalopathy.

3',5'-Cyclic-GMP Phosphodiesterases↗
PubMed2004

Sequential activation of soluble guanylate cyclase, protein kinase G and cGMP-degrading phosphodiesterase is necessary for proper induction of long-term potentiation in CA1 of hippocampus. Alterations in hyperammonemia.

Long-term potentiation (LTP) is a long-lasting enhancement of synaptic transmission efficacy and is considered the base for some forms of learning and memory. Nitric oxide (NO)-induced formation of cGMP is involved in hippocampal LTP. We have studied in hippocampal slices the effects of application of a tetanus to induce LTP on cGMP metabolism and the mechanisms by which cGMP modulates LTP. Tetanus application induced a transient rise in cGMP, reaching a maximum at 10s and decreasing below basal levels 5 min after the tetanus, remaining below basal levels after 60 min. Soluble guanylate cyclase (sGC) activity increased 5 min after tetanus and returned to basal levels at 60 min. The decrease in cGMP was due to sustained tetanus-induced increase in cGMP-degrading phosphodiesterase activity, which remained activated 60 min after tetanus. Tetanus-induced activation of PDE and decrease of cGMP were prevented by inhibiting protein kinase G (PKG). This indicates that the initial increase in cGMP activates PKG that phosphorylates (and activates) cGMP-degrading PDE, which, in turn, degrades cGMP. Inhibition of sGC, of PKG or of cGMP-degrading phosphodiesterase impairs LTP, indicating that proper induction of LTP involves transient activation of sGC and increase in cGMP, followed by activation of cGMP-dependent protein kinase, which, in turn, activates cGMP-degrading phosphodiesterase, resulting in long-lasting reduction of cGMP content. Hyperammonemia is the main responsible for the neurological alterations found in liver disease and hepatic encephalopathy, including impaired intellectual function. Hyperammonemia impairs LTP in hippocampus by altering the modulation of this sGC-PKG-cGMP-degrading PDE pathway. Exposure of hippocampal slices to 1 mM ammonia completely prevents tetanus-induced decrease of cGMP by impairing PKG-mediated activation of cGMP-degrading phosphodiesterase. This impairment is responsible for the loss of the maintenance of LTP in hyperammonemia, and may be also involved in the cognitive impairment in patients with hyperammonemia and hepatic encephalopathy.

3',5'-Cyclic-GMP Phosphodiesterases↗
PubMed2004

Alterations in soluble guanylate cyclase content and modulation by nitric oxide in liver disease.

Hyperammonemia is the main responsible for the neurological alterations in hepatic encephalopathy in patients with liver failure. We studied the function of the glutamate-nitric oxide (NO)-cGMP pathway in brain in animal models of hyperammonemia and liver failure and in patients died with liver cirrhosis. Activation of glutamate receptors increases intracellular calcium that binds to calmodulin and activates neuronal nitric oxide synthase, increasing nitric oxide, which activates soluble guanylate cyclase (sGC), increasing cGMP. This glutamate-NO-cGMP pathway modulates cerebral processes such as circadian rhythms, the sleep-waking cycle, and some forms of learning and memory. These processes are impaired in patients with hepatic encephalopathy. Activation of sGC by NO is significantly increased in cerebral cortex and significantly reduced in cerebellum from cirrhotic patients died in hepatic coma. Portacaval anastomosis in rats, an animal model of liver failure, reproduces the effects of liver failure on modulation of sGC by NO both in cerebral cortex and cerebellum. In vivo brain microdialisis studies showed that sGC activation by NO is also reduced in vivo in cerebellum in hyperammonemic rats with or without liver failure. The content of alpha but not beta subunits of sGC are increased both in frontal cortex and cerebellum from patients died due to liver disease and from rats with portacaval anastomosis. We assessed whether determination of activation of sGC by NO-generating agent SNAP in lymphocytes could serve as a peripheral marker for the impairment of sGC activation by NO in brain. Chronic hyperammonemia and liver failure also alter sGC activation by NO in lymphocytes from rats or patients. These findings show that the content and modulation by NO of sGC are strongly altered in brain of patients with liver disease. These alterations could be responsible for some of the neurological alterations in hepatic encephalopathy such as sleep disturbances and cognitive impairment.

Animals↗
PubMed2004

In vivo exposure to carbon monoxide causes delayed impairment of activation of soluble guanylate cyclase by nitric oxide in rat brain cortex and cerebellum.

Carbon monoxide induces delayed neurological and neuropathological alterations, including memory loss and cognitive impairment. The bases for the delay remain unknown. Activation of soluble guanylate cyclase by nitric oxide modulates some forms of learning and memory. Carbon monoxide binds to soluble guanylate cyclase, activating it but interfering with its activation by nitric oxide. The aim of this work was to assess whether exposure of rats to carbon monoxide alters the activity of soluble guanylate cyclase or its modulation by nitric oxide in cerebellum or cerebral cortex. Rats exposed chronically or acutely to carbon monoxide were killed 24 h or 7 days later. Acute carbon monoxide exposure decreased cyclic guanosine monophosphate (cGMP) content and reduced activation of soluble guanylate cyclase by nitric oxide. Cortex was more sensitive than cerebellum to chronic exposure, which reduced activation of soluble guanylate cyclase by nitric oxide in cortex. In cerebellum, chronic exposure induced delayed impairment of soluble guanylate cyclase activation by nitric oxide. Acute exposure effects were also stronger at 7 days than at 24 h after exposure. This delayed impaired modulation of soluble guanylate cyclase by nitric oxide may contribute to delayed memory loss and cognitive impairment in humans exposed to carbon monoxide.

Animals↗
PubMed2004

Acute ammonia intoxication induces an NMDA receptor-mediated increase in poly(ADP-ribose) polymerase level and NAD metabolism in nuclei of rat brain cells.

Acute ammonia toxicity is mediated by excessive activation of NMDA receptors. Activation of NMDA receptors leads to activation of poly(ADP-ribose) polymerase (PARP) which mediates NMDA excitotoxicity. PARP is activated following DNA damage and may lead to cell death via NAD+ and ATP depletion. The aim of the present work was to assess whether acute ammonia intoxication in vivo leads to increased PARP in brain cells nuclei and to altered NAD+ and superoxide metabolism and the contribution of NMDA receptors to these alterations. Acute ammonia intoxication increases PARP content twofold in brain cells nuclei.NAD+ content decreased by 55% in rats injected with ammonia. This was not due to decreased NAD+ synthetase nor increased NAD+ hydrolase activities and would be due to increased NAD+ consumption by PARP. Superoxide radical formation increased by 75% in nuclei of brains of rats injected with ammonia, that also induced protein nitrotyrosylation and DNA damage. Blocking NMDA receptors prevented ammonia-induced PARP, superoxide and nitrotyrosylation increase, DNA damage and NAD+ decrease. These results show that acute ammonia intoxication in vivo leads to activation of NMDA receptors, leading to increased superoxide formation and PARP content and depletion of NAD+ in brain cells nuclei that contribute to ammonia toxicity.

Amide Synthases↗
PubMed2004

Modulation of NMDA receptor function by cyclic AMP in cerebellar neurones in culture.

The signal transduction pathways involved in NMDA receptor modulation by other receptors remain unclear. cAMP could be involved in this modulation. The aim of this work was to analyse the contribution of cAMP to NMDA receptor modulation in cerebellar neurones in culture. Forskolin increases cAMP and results in increased intracellular calcium and cGMP that are prevented by blocking NMDA receptors. Similar effects were induced by two cAMP analogues, indicating that cAMP leads to NMDA receptor activation. It has been reported that phosphorylation of Ser897 of the NR1 subunit of NMDA receptors by cAMP-dependent protein kinase (PKA) activates the receptors. Forskolin increases Ser897 phosphorylation. Neither Ser897 phosphorylation nor cGMP increase induced by forskolin are prevented by four inhibitors of PKA, suggesting that NMDA receptor activation is dependent on cAMP but not on PKA. Inhibition of Akt prevents forskolin-induced phosphorylation of Ser897, suggesting a role for Akt in the mediation of the modulation of NMDA receptors by cAMP. Pituitary adenylate cyclase-activating polypeptide (PACAP) activates its receptors, increasing cAMP and also leading to phosphorylation of Ser897 of NR1 and activation of NMDA receptors. These results indicate that cAMP modulates NMDA receptor in cerebellar neurones and may play a role in NMDA receptor modulation by other receptors.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed2003

Sources of oxygen radicals in brain in acute ammonia intoxication in vivo.

The effects of acute ammonia intoxication on reactive oxygen species production by different sources in rat brain were studied. Ammonia intoxication in vivo leads to reduced activity of superoxide dismutase (SOD), catalase and glutathione peroxidase in brain nonsynaptic mitochondria and increased formation of O(2)(-) by submitochondrial particles. It also results in increased xanthine oxidase (XO) activity and decreased xanthine dehydrogenase (XDH)/XO activity ratio indicating conversion of XDH to XO and also increases monoamine oxidase A (MAO-A) activity but not of MAO-B. Blocking NMDA receptors with MK-801 prevents ammonia-induced oxidative stress, XDH to XO conversion and MAO-A activation. Ammonia intoxication did not lead to H(2)O(2) formation by mitochondria, in spite of increased O(2)(-) generation. The main source of H(2)O(2) in the mitochondrial matrix was Mn-SOD. Ammonia intoxication in vivo leads to increased superoxide and decreased hydrogen peroxide in nonsynaptic brain mitochondria. Increased superoxide is due to increased formation by the respiratory chain and by xanthine and aldehyde oxidases and decreased elimination by antioxidant enzymes. The reduced formation of hydrogen peroxide is due to the reduced activity of Mn-SOD. Prevention of ammonia-induced production of reactive oxygen species by MK-801 supports the idea that it is mediated by activation of NMDA receptors.

Aldehyde Oxidase↗