Ravi Akundi - Academia.edu (original) (raw)

Papers by Ravi Akundi

Annals of the New York Academy of Sciences, 2003

The former popular hypnotic chloral hydrate has been known to spontaneously condense with tryptam... more The former popular hypnotic chloral hydrate has been known to spontaneously condense with tryptamine in the body to give rise to a highly unpolar 1-trichloromethyl-1,2,3,4-tetrahydro-beta-carboline (TaClo) derivative. Earlier studies have revealed the relative permeability of the molecule through the body, and its ability to induce Parkinson-like symptoms in rats. In this study, we report that TaClo is a highly cytotoxic agent that leads to apoptosis in human neuroblastoma cells involving caspase-3 activation.

Neurochemistry International, 2005

Adenosine binds to a class of G-protein coupled receptors, which are further distinguished as A(1... more Adenosine binds to a class of G-protein coupled receptors, which are further distinguished as A(1), A(2a), A(2b) and A(3) adenosine receptors. As we have shown earlier, the stable adenosine analogue NECA (N6-(R)-phenylisopropyladenosine) stimulates IL-6 expression in the human astrocytoma cell line U373 MG via the A(2b) receptor. The mechanism by which NECA promotes astrocytic IL-6 expression has not been identified. By using various inhibitors of signal transduction, we found that p38 mitogen-activated protein kinases (MAPK) activation (inhibitor SB202190), but not extracellular signal-regulated kinase (ERK) (PD98059) and c-jun N-terminal kinase (JNK)(SP600125), is essential in the NECA-induced signalling cascade that leads to the increase in IL-6 synthesis in U373 MG cells. Results obtained with protein kinase C (PKC) inhibitors that have different substrate specificities, indicated that the PKC delta and epsilon isoforms are also involved in adenosine receptor A(2b) dependent upregulation of IL-6 expression. This is supported by the fact that NECA induced the activation of PKC delta and epsilon in U373 MG cells.

Frontiers in Cellular Neuroscience, 2014

Brain inflammation is a common occurrence following responses to varied insults such as bacterial... more Brain inflammation is a common occurrence following responses to varied insults such as bacterial infections, stroke, traumatic brain injury and neurodegenerative disorders. A common mediator for these varied inflammatory responses is prostaglandin E 2 (PGE 2 ), produced by the enzymatic activity of cyclooxygenases (COX) 1 and 2. Previous attempts to reduce neuronal inflammation through COX inhibition, by use of nonsteroidal anti-inflammatory drugs (NSAIDs), have met with limited success. We are proposing the two-hit model for neuronal injury-an initial localized inflammation mediated by PGE 2 (first hit) and the simultaneous release of adenosine triphosphate (ATP) by injured cells (second hit), which significantly enhances the inflammatory response through increased synthesis of PGE 2 . Several evidences on the role of exogenous ATP in inflammation have been reported, including contrary instances where extracellular ATP reduces inflammatory events. In this review, we will examine the current literature on the role of P2 receptors, to which ATP binds, in modulating inflammatory reactions during neurodegeneration. Targeting the P2 receptors, therefore, provides a therapeutic alternative to reduce inflammation in the brain. P2 receptor-based anti-inflammatory drugs (PBAIDs) will retain the activities of essential COX enzymes, yet will significantly reduce neuroinflammation by decreasing the enhanced production of PGE 2 by extracellular ATP.

PLoS ONE, 2011

Background: PTEN-induced kinase 1 (PINK1) is linked to recessive Parkinsonism (EOPD). Pink1 delet... more Background: PTEN-induced kinase 1 (PINK1) is linked to recessive Parkinsonism (EOPD). Pink1 deletion results in impaired dopamine (DA) release and decreased mitochondrial respiration in the striatum of mice. To reveal additional mechanisms of Pink1-related dopaminergic dysfunction, we studied Ca 2+ vulnerability of purified brain mitochondria, DA levels and metabolism and whether signaling pathways implicated in Parkinson's disease (PD) display altered activity in the nigrostriatal system of Pink1 2/2 mice.

Scandinavian Journal of Rheumatology, 2004

There is evidence from both human and animal research that 5-hydroxytryptamine 3 (5-HT 3 ) recept... more There is evidence from both human and animal research that 5-hydroxytryptamine 3 (5-HT 3 ) receptor antagonists, particularly tropisetron, exert analgesic and antiinflammatory effects. However, the underlying mechanisms of these effects including the expression of 5-HT 3 receptors in cells of the immune system have not yet been investigated in detail. Therefore, we investigated the expression of the 5-HT 3A receptor in primary human monocytes, chondrocytes, T cells, dendritic cells, and synovial tissue. We found that 5-HT 3A receptors are expressed in monocytes, chondrocytes, T-cells, and synovial tissue but not in dendritic cells. Our data show that 5-HT 3A receptors are widely expressed in cells of the immune system and that they might play an important role in inflammatory events and in the observed antiphlogistic effects of 5-HT 3 receptor antagonists.

PLoS ONE, 2009

Periventricular white matter injury (PWMI) is a common form of brain injury sustained by preterm ... more Periventricular white matter injury (PWMI) is a common form of brain injury sustained by preterm infants. A major factor that predisposes to PWMI is hypoxia. Because oligodendrocytes (OLs) are responsible for myelination of axons, abnormal OL development or function may affect brain myelination. At present our understanding of the influences of hypoxia on OL development is limited. To examine isolated effects of hypoxia on OLs, we examined the influences of hypoxia on OL development in vitro. Cultures of oligodendrocyte precursor cells (OPCs) were prepared from mixed glial cultures and were 99% pure. OPCs were maintained at 21% O(2) or hypoxia (1% or 4% O(2)) for up to 7 days. We observed that 1% O(2) lead to an increase in the proportion of myelin basic protein (MBP)-positive OLs after 1 week in culture, and a decrease in the proportion of platelet-derived growth factor receptor alpha (PDGFRalpha)-positive cells suggesting premature OL maturation. Increased expression of the cell cycle regulatory proteins p27(Kip1) and phospho-cdc2, which play a role in OL differentiation, was seen as well. These results show that hypoxia interferes with the normal process of OL differentiation by inducing premature OPC maturation.

Pharmacological Research, 2006

The fixed combination of aspirin, paracetamol (acetaminophen) and caffeine has been used successf... more The fixed combination of aspirin, paracetamol (acetaminophen) and caffeine has been used successfully to treat different kinds of pain including migraine attacks. Even when this formulation has been marketed for a long time, the exact molecular mechanisms underlying its therapeutic effectiveness have not been completely elucidated. In the present investigation, we have studied the effects of the fixed combination of aspirin, paracetamol and caffeine (APC) on the release of dopamine and noradrenaline from rat striatal slices in an attempt to find potential new mechanisms of action of this widely used analgesic combination. We found that APC produced a significant reduction in extracellular dopamine and a dramatic increase in norepinephrine release from the slices incubated with different concentrations of APC (dose relationship 1:1:0.2, corresponding to the dose-relationship of Thomapyrin ® ). These findings suggest that the modulation of catecholaminergic neurotransmission is a new pharmacological effect of APC which could explain the mechanism of action of this formulation, considering that the independent effect of either compound alone does not explain the potent antinociceptive properties when observed in combination.

Parkinsonism & Related Disorders, 2009

Neurodegenerative Diseases, 2012

possibly increasing the sensitivity to oxidative stress. Mitochondria accumulate in large foci in... more possibly increasing the sensitivity to oxidative stress. Mitochondria accumulate in large foci in PINK1 -/-MEF, indicative of abnormal mitochondrial dynamics and/or transport. Like in PINK1 -/-Drosophila , enlarged/swollen mitochondria accumulate in three different cell types from PINK1 -/mice (MEF, primary cortical neurons and embryonic stem cells). However, mitochondrial enlargement is greatest and most prominent in primary cortical neurons that also develop cristae fragmentation and disintegration. Conclusion: Our results reveal mechanisms of PINK1-related parkinsonism, show that the function of PINK1 is conserved between Drosophila and mammals when studied in primary cells, and demonstrate that the same PINK1 mutation can affect mitochondrial morphology/degeneration in a cell type-specific manner, suggesting that tissue-/cell-specific metabolic capacity and adaptations determine phenotypes and cellular vulnerability in PINK1 -/mice and cells.

Neurochemistry International, 2006

Prostaglandins (PGs) are generated by the enzymatic activity of cyclooxygenase-1 and -2 (COX-1/2)... more Prostaglandins (PGs) are generated by the enzymatic activity of cyclooxygenase-1 and -2 (COX-1/2) and modulate several functions in the CNS such as the generation of fever, the sleep/wake cycle, and the perception of pain. Moreover, the induction of COX-2 and the generation of PGs has been linked to neuroinflammatory aspects of Alzheimer's disease (AD). Non-steroidal anti-inflammatory drugs (NSAIDs) that block COX enzymatic activity have been shown to reduce the incidence of AD in various epidemiological studies. While several reports investigated the expression of COX-2 in neurons and microglia, expression of COX-2 in astroglial cells has not been investigated in detail. Here we show that amyloid beta peptide 25-35 (Abeta(25-35)) induces COX-2 mRNA and protein synthesis and a subsequent release of prostaglandin E(2) (PGE(2)) in primary midbrain astrocytes. We further demonstrate that protein kinase C (PKC) is involved in Abeta(25-35)-induced COX-2/PGE(2) synthesis. PKC-inhibitors prevent Abeta(25-35)-induced COX-2 and PGE(2) synthesis. Furthermore Abeta(25-35) rapidly induces the phosphorylation and enzymatic activation of PKC in primary rat midbrain glial cells and in primary human astrocytes from post mortem tissue. Our data suggest that the PKC isoforms alpha and/or beta are most probably involved in Abeta(25-35)-induced expression of COX-2 in midbrain astrocytes. The potential role of astroglial cells in the phagocytosis of amyloid and the involvement of PGs in this process suggests that a modulation of PGs synthesis may be a putative target in the prevention of amyloid deposition.

Neurobiology of Disease, 2012

Mutations in the PARK6 gene coding for PTEN-induced kinase 1 (PINK1) cause recessive earlyonset p... more Mutations in the PARK6 gene coding for PTEN-induced kinase 1 (PINK1) cause recessive earlyonset parkinsonism. Although PINK1 and Parkin promote the degradation of depolarized mitochondria in cultured cells, little is known about changes in signaling pathways that may additionally contribute to dopamine neuron loss in recessive parkinsonism. Accumulating evidence implicates impaired Akt cell survival signaling in sporadic and familial PD (PD). IGF-1/ Akt signaling inhibits dopamine neuron loss in several animal models of PD and both IGF-1 and insulin are neuroprotective in various settings. Here, we tested whether PINK1 is required for insulin-like growth factor 1 (IGF-1) and insulin dependent phosphorylation of Akt and the regulation of downstream Akt target proteins. Our results show that embryonic fibroblasts from PINK1-deficient mice display significantly reduced Akt phosphorylation in response to both IGF-1 and insulin. Moreover, phosphorylation of glycogen synthase kinase-3β (GSK-3β) and nuclear exclusion of FoxO1 are decreased in IGF-1 treated PINK1-deficient cells. In addition, phosphorylation of ribosomal protein S6 is reduced indicating decreased activity of mitochondrial target of rapamycin (mTOR) in IGF-1 treated PINK1 −/− cells. Importantly, the protection afforded by IGF-1 against staurosporine-induced metabolic dysfunction and apoptosis is abrogated in PINK1-deficient cells. Moreover, IGF-1-induced Akt phosphorylation is impaired in primary cortical neurons from PINK1-deficient mice. Inhibition of cellular Ser/Thr phosphatases did not increase the amount of phosphorylated Akt in PINK1 −/− cells, suggesting that components upstream of Akt phosphorylation are compromised in PINK1-deficient cells. Our studies show that PINK1 is required for optimal IGF-1 and insulin dependent Akt signal transduction, and raise the possibility that impaired IGF-1/Akt signaling is involved in PINK1-related parkinsonism by increasing the vulnerability of dopaminergic neurons to stress-induced cell death.

Journal of Neuroimmunology, 2006

Inhibition of neuronal cyclooxygenase-2 (COX-2) and hence prostaglandin E2 (PGE2) synthesis by no... more Inhibition of neuronal cyclooxygenase-2 (COX-2) and hence prostaglandin E2 (PGE2) synthesis by non-steroidal anti-inflammatory drugs has been suggested to protect neuronal cells in a variety of pathophysiological situations including Alzheimer's disease and ischemic stroke. Ascorbic acid (vitamin C) has also been shown to protect cerebral tissue in a variety of experimental conditions, which has been attributed to its antioxidant capacity. In the present study, we show that ascorbic acid dose-dependently inhibited interleukin-1beta (IL-1beta)-mediated PGE2 synthesis in the human neuronal cell line, SK-N-SH. Furthermore, in combination with aspirin, ascorbic acid augmented the inhibitory effect of aspirin on PGE2 synthesis. However, ascorbic acid had no synergistic effect along with other COX inhibitors (SC-58125 and indomethacin). The inhibition of IL-1beta-mediated PGE2 synthesis by ascorbic acid was not due to the inhibition of the expression of COX-2 or microsomal prostaglandin E synthase (mPGES-1). Rather, ascorbic acid dose-dependently (0.1-100 microM) produced a significant reduction in IL-1beta-mediated production of 8-iso-prostaglandin F2alpha (8-iso-PGF2alpha), a reliable indicator of free radical formation, suggesting that the effects of ascorbic acid on COX-2-mediated PGE2 biosynthesis may be the result of the maintenance of the neuronal redox status since COX activity is known to be enhanced by oxidative stress. Our results provide in vitro evidence that the neuroprotective effects of ascorbic acid may depend, at least in part, on its ability to reduce neuronal COX-2 activity and PGE2 synthesis, owing to its antioxidant properties. Further, these experiments suggest that a combination of aspirin with ascorbic acid constitutes a novel approach to render COX-2 more sensitive to inhibition by aspirin, allowing an anti-inflammatory therapy with lower doses of aspirin, thereby avoiding the side effects of the usually high dose aspirin treatment.

Journal of Neurochemistry, 2004

The neuropeptide substance P (SP) has been hypothesized to be involved in the etiopathology of af... more The neuropeptide substance P (SP) has been hypothesized to be involved in the etiopathology of affective disorders. This hypothesis is based on the findings that neurokinin-1-receptor antagonists have antidepressant effects in depressed patients and that SP may worsen mood. In this study, we investigated the effect of the mood-stabilizing agents valproic acid (VPA), carbamazepine, and lithium on SP-induced gene expression. As a model system, we used primary rat astrocytes and human astrocytoma cells, which both express functional SP-receptors and, upon stimulation with SP, synthesize interleukin-6 (IL-6), a cytokine which has been shown to be elevated during the acute depressive state. We found that VPA dose-dependently inhibited SP-induced IL-6 synthesis which was seen with pre-incubation periods of 30 min, 3, 7 and 14 days, whereas carbamazepine and lithium showed no inhibitory effect. The inhibitory effect of VPA was not mediated by inhibition of the stress-regulated kinases p38 and p42/44 (Erk1/2) but by inhibition of protein kinase C epsilon activation. Furthermore, VPA down-regulated the expression of the substance P receptor (neurokinin(NK)-1-receptor) as assessed by real-time PCR. Whether both mechanisms contribute to the mood-stabilizing properties of VPA has to be evaluated in further studies.

Journal of Neurochemistry, 2006

Both interleukin-1b (IL-1b) and prostaglandins (PGs) are important mediators of physiological and... more Both interleukin-1b (IL-1b) and prostaglandins (PGs) are important mediators of physiological and pathophysiological processes in the brain. PGE 2 exerts its effects by binding to four different types of PGE 2 receptors named EP1-EP4. EP3 has found to be expressed in neurons, whereas expression of EP3 in glial cells has not been reported in the brain yet. Here we describe IL-1b-induced EP3 receptor expression in human astrocytoma cells, primary astrocytes of rat and human origin and in rat brain. Using western blot, we found a marked up-regulation of EP3 receptor synthesis in human and rat primary glial cells. Intracerebroventricular administration of IL-1b stimulated EP3 receptor synthesis in rat hippocampus.

Journal of Neurochemistry, 2005

] is a widely distributed neurotransmitter which is involved in neuroimmunomodulatory processes. ... more ] is a widely distributed neurotransmitter which is involved in neuroimmunomodulatory processes. Previously, it has been demonstrated that 5-HT may induce interleukin (IL)-6 expression in primary rat hippocampal astrocytes. The present study was undertaken to investigate the molecular pathways underlying this induction of IL-6 synthesis. As a model system, we used the human astrocytoma cell line U373 MG, which synthesizes IL-6 upon stimulation with various inducers. 5-HT dose-and timedependently induced IL-6 protein synthesis. We identified several 5-HT receptors to be expressed on U373 MG cells, including the 5-HT 1D , 5-HT 2A , 5-HT 3 and 5-HT 7 receptors. In this report, we show that the 5-HT-induced IL-6 release is mediated by the 5-HT 7 receptor based on several agonist/ antagonists that were used. 5-HT-induced IL-6 synthesis is inhibited by the partially selective 5-HT 7 receptor antagonist, pimozide, and the selective antagonist SB269970. Furthermore, IL-6 synthesis was induced by the 5-HT 7 receptor agonist carboxamidotryptamin. In addition, we found p38 MAPKs and protein kinase C (PKC) e to be involved in 5-HTinduced IL-6 synthesis as specific inhibitors of these enzymes (SB202190 and RO-31-8425, respectively) blocked 5-HTinduced IL-6 synthesis. Furthermore, 5-HT mediated the phosphorylation of both p38 MAPK as well as the PKC e isoform. The p42/44 MAPKs, however, were not involved in 5-HT-induced IL-6 synthesis. This study shows, for the first time, a central role of 5-HT 7 receptor linked to p38 MAPK and PKC e for the induction of cytokine synthesis in astrocytic cells.

Journal of Neurochemistry, 2004

Trichloroethylene, a common industrial solvent and a metabolic precursor of chloral hydrate, occu... more Trichloroethylene, a common industrial solvent and a metabolic precursor of chloral hydrate, occurs widely in the environment. Chloral hydrate, which is also used as a hypnotic, has been found to condense spontaneously with tryptamine, in vivo, to give rise to a highly unpolar 1-trichloromethyl-1,2,3,4-tetrahydro-b-carboline (TaClo) that has a structural analogy to the dopaminergic neurotoxin N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Earlier studies have revealed the relative permeability of the molecule through the blood-brain barrier and its ability to induce Parkinson-like symptoms in rats. In this study, we report that TaClo induces an apoptotic pathway in the human neuroblastoma cell line, SK-N-SH, involving the translocation of mitochondrial cytochrome c to the cytosol and activation of caspase 3. TaClo-induced apoptosis shows considerable differences from that mediated by other Parkinson-inducing agents such as MPTP, rotenone and manganese. Although it is not clear if the clinically administered dosage of chloral hydrate or the relatively high environmental levels of trichloroethylene could lead to an onset of Parkinson's disease, the spontaneous in vivo formation of TaClo and its pro-apoptotic properties, as shown in this report, should be considered.

Journal of Molecular Neuroscience, 2005

Prostaglandin E 2 (PGE 2 ), is a major prostanoid produced by the activity of cyclooxygenases (CO... more Prostaglandin E 2 (PGE 2 ), is a major prostanoid produced by the activity of cyclooxygenases (COX) in response to various physiological and pathological stimuli. PGE 2 exerts its effects by activating four specific E-type prostanoid receptors (EP1, EP2, EP3, and EP4). In the present study, we analyzed the expression of the PGE 2 receptor EP1 (mRNA and protein) in different regions of the adult rat brain (hippocampus, hypothalamus, striatum, prefrontal cerebral cortex, parietal cortex, brain stem, and cerebellum) using reverse transcriptionpolymerase chain reaction, Western blotting, and immunohistochemical methods. On a regional basis, levels of EP1 mRNA were the highest in parietal cortex and cerebellum. At the protein level, we found very strong expression of EP1 in cerebellum, as revealed by Western blotting experiments. Furthermore, the present study provides for the first time evidence that the EP1 receptor is highly expressed in the cerebellum, where the Purkinje cells displayed very high immunolabeling of their perikaryon and dendrites, as observed in the immunohistochemical analysis. Results from the present study indicate that the EP1 prostanoid receptor is expressed in specific neuronal populations, which possibly determine the region-specific response to PGE 2 .

European Journal of Immunology, 2013

Mutations in PTEN-induced kinase 1 (PINK1), a serine/threonine kinase linked to familial early-on... more Mutations in PTEN-induced kinase 1 (PINK1), a serine/threonine kinase linked to familial early-onset Parkinsonism, compromise mitochondrial integrity and metabolism and impair AKT signaling. As the activation of a naïve T cell requires an AKT-dependent reorganization of a cell's metabolic machinery, we sought to determine if PINK1-deficient T cells lack the ability to undergo activation and differentiation. We show that CD4 + T cells from PINK1 knockout mice fail to properly phosphorylate AKT upon activation, resulting in reduced expression of the IL-2 receptor subunit CD25. Following, deficient

European Journal of Clinical Investigation, 2009

Background There is a pressing need for research that will lead to the development of new therape... more Background There is a pressing need for research that will lead to the development of new therapeutic approaches for treating inflammatory bowel disease (IBD). The aim of this study was to investigate the effects of tropisetron, a 5-Hydroxytryptamine (5-HT)-3 receptor antagonist with anti-inflammatory properties in a model of experimental colitis in rat.

Annals of the New York Academy of Sciences, 2003

The former popular hypnotic chloral hydrate has been known to spontaneously condense with tryptam... more The former popular hypnotic chloral hydrate has been known to spontaneously condense with tryptamine in the body to give rise to a highly unpolar 1-trichloromethyl-1,2,3,4-tetrahydro-beta-carboline (TaClo) derivative. Earlier studies have revealed the relative permeability of the molecule through the body, and its ability to induce Parkinson-like symptoms in rats. In this study, we report that TaClo is a highly cytotoxic agent that leads to apoptosis in human neuroblastoma cells involving caspase-3 activation.

Neurochemistry International, 2005

Adenosine binds to a class of G-protein coupled receptors, which are further distinguished as A(1... more Adenosine binds to a class of G-protein coupled receptors, which are further distinguished as A(1), A(2a), A(2b) and A(3) adenosine receptors. As we have shown earlier, the stable adenosine analogue NECA (N6-(R)-phenylisopropyladenosine) stimulates IL-6 expression in the human astrocytoma cell line U373 MG via the A(2b) receptor. The mechanism by which NECA promotes astrocytic IL-6 expression has not been identified. By using various inhibitors of signal transduction, we found that p38 mitogen-activated protein kinases (MAPK) activation (inhibitor SB202190), but not extracellular signal-regulated kinase (ERK) (PD98059) and c-jun N-terminal kinase (JNK)(SP600125), is essential in the NECA-induced signalling cascade that leads to the increase in IL-6 synthesis in U373 MG cells. Results obtained with protein kinase C (PKC) inhibitors that have different substrate specificities, indicated that the PKC delta and epsilon isoforms are also involved in adenosine receptor A(2b) dependent upregulation of IL-6 expression. This is supported by the fact that NECA induced the activation of PKC delta and epsilon in U373 MG cells.

Frontiers in Cellular Neuroscience, 2014

Brain inflammation is a common occurrence following responses to varied insults such as bacterial... more Brain inflammation is a common occurrence following responses to varied insults such as bacterial infections, stroke, traumatic brain injury and neurodegenerative disorders. A common mediator for these varied inflammatory responses is prostaglandin E 2 (PGE 2 ), produced by the enzymatic activity of cyclooxygenases (COX) 1 and 2. Previous attempts to reduce neuronal inflammation through COX inhibition, by use of nonsteroidal anti-inflammatory drugs (NSAIDs), have met with limited success. We are proposing the two-hit model for neuronal injury-an initial localized inflammation mediated by PGE 2 (first hit) and the simultaneous release of adenosine triphosphate (ATP) by injured cells (second hit), which significantly enhances the inflammatory response through increased synthesis of PGE 2 . Several evidences on the role of exogenous ATP in inflammation have been reported, including contrary instances where extracellular ATP reduces inflammatory events. In this review, we will examine the current literature on the role of P2 receptors, to which ATP binds, in modulating inflammatory reactions during neurodegeneration. Targeting the P2 receptors, therefore, provides a therapeutic alternative to reduce inflammation in the brain. P2 receptor-based anti-inflammatory drugs (PBAIDs) will retain the activities of essential COX enzymes, yet will significantly reduce neuroinflammation by decreasing the enhanced production of PGE 2 by extracellular ATP.

PLoS ONE, 2011

Background: PTEN-induced kinase 1 (PINK1) is linked to recessive Parkinsonism (EOPD). Pink1 delet... more Background: PTEN-induced kinase 1 (PINK1) is linked to recessive Parkinsonism (EOPD). Pink1 deletion results in impaired dopamine (DA) release and decreased mitochondrial respiration in the striatum of mice. To reveal additional mechanisms of Pink1-related dopaminergic dysfunction, we studied Ca 2+ vulnerability of purified brain mitochondria, DA levels and metabolism and whether signaling pathways implicated in Parkinson's disease (PD) display altered activity in the nigrostriatal system of Pink1 2/2 mice.

Scandinavian Journal of Rheumatology, 2004

There is evidence from both human and animal research that 5-hydroxytryptamine 3 (5-HT 3 ) recept... more There is evidence from both human and animal research that 5-hydroxytryptamine 3 (5-HT 3 ) receptor antagonists, particularly tropisetron, exert analgesic and antiinflammatory effects. However, the underlying mechanisms of these effects including the expression of 5-HT 3 receptors in cells of the immune system have not yet been investigated in detail. Therefore, we investigated the expression of the 5-HT 3A receptor in primary human monocytes, chondrocytes, T cells, dendritic cells, and synovial tissue. We found that 5-HT 3A receptors are expressed in monocytes, chondrocytes, T-cells, and synovial tissue but not in dendritic cells. Our data show that 5-HT 3A receptors are widely expressed in cells of the immune system and that they might play an important role in inflammatory events and in the observed antiphlogistic effects of 5-HT 3 receptor antagonists.

PLoS ONE, 2009

Periventricular white matter injury (PWMI) is a common form of brain injury sustained by preterm ... more Periventricular white matter injury (PWMI) is a common form of brain injury sustained by preterm infants. A major factor that predisposes to PWMI is hypoxia. Because oligodendrocytes (OLs) are responsible for myelination of axons, abnormal OL development or function may affect brain myelination. At present our understanding of the influences of hypoxia on OL development is limited. To examine isolated effects of hypoxia on OLs, we examined the influences of hypoxia on OL development in vitro. Cultures of oligodendrocyte precursor cells (OPCs) were prepared from mixed glial cultures and were 99% pure. OPCs were maintained at 21% O(2) or hypoxia (1% or 4% O(2)) for up to 7 days. We observed that 1% O(2) lead to an increase in the proportion of myelin basic protein (MBP)-positive OLs after 1 week in culture, and a decrease in the proportion of platelet-derived growth factor receptor alpha (PDGFRalpha)-positive cells suggesting premature OL maturation. Increased expression of the cell cycle regulatory proteins p27(Kip1) and phospho-cdc2, which play a role in OL differentiation, was seen as well. These results show that hypoxia interferes with the normal process of OL differentiation by inducing premature OPC maturation.

Pharmacological Research, 2006

The fixed combination of aspirin, paracetamol (acetaminophen) and caffeine has been used successf... more The fixed combination of aspirin, paracetamol (acetaminophen) and caffeine has been used successfully to treat different kinds of pain including migraine attacks. Even when this formulation has been marketed for a long time, the exact molecular mechanisms underlying its therapeutic effectiveness have not been completely elucidated. In the present investigation, we have studied the effects of the fixed combination of aspirin, paracetamol and caffeine (APC) on the release of dopamine and noradrenaline from rat striatal slices in an attempt to find potential new mechanisms of action of this widely used analgesic combination. We found that APC produced a significant reduction in extracellular dopamine and a dramatic increase in norepinephrine release from the slices incubated with different concentrations of APC (dose relationship 1:1:0.2, corresponding to the dose-relationship of Thomapyrin ® ). These findings suggest that the modulation of catecholaminergic neurotransmission is a new pharmacological effect of APC which could explain the mechanism of action of this formulation, considering that the independent effect of either compound alone does not explain the potent antinociceptive properties when observed in combination.

Parkinsonism & Related Disorders, 2009

Neurodegenerative Diseases, 2012

possibly increasing the sensitivity to oxidative stress. Mitochondria accumulate in large foci in... more possibly increasing the sensitivity to oxidative stress. Mitochondria accumulate in large foci in PINK1 -/-MEF, indicative of abnormal mitochondrial dynamics and/or transport. Like in PINK1 -/-Drosophila , enlarged/swollen mitochondria accumulate in three different cell types from PINK1 -/mice (MEF, primary cortical neurons and embryonic stem cells). However, mitochondrial enlargement is greatest and most prominent in primary cortical neurons that also develop cristae fragmentation and disintegration. Conclusion: Our results reveal mechanisms of PINK1-related parkinsonism, show that the function of PINK1 is conserved between Drosophila and mammals when studied in primary cells, and demonstrate that the same PINK1 mutation can affect mitochondrial morphology/degeneration in a cell type-specific manner, suggesting that tissue-/cell-specific metabolic capacity and adaptations determine phenotypes and cellular vulnerability in PINK1 -/mice and cells.

Neurochemistry International, 2006

Prostaglandins (PGs) are generated by the enzymatic activity of cyclooxygenase-1 and -2 (COX-1/2)... more Prostaglandins (PGs) are generated by the enzymatic activity of cyclooxygenase-1 and -2 (COX-1/2) and modulate several functions in the CNS such as the generation of fever, the sleep/wake cycle, and the perception of pain. Moreover, the induction of COX-2 and the generation of PGs has been linked to neuroinflammatory aspects of Alzheimer's disease (AD). Non-steroidal anti-inflammatory drugs (NSAIDs) that block COX enzymatic activity have been shown to reduce the incidence of AD in various epidemiological studies. While several reports investigated the expression of COX-2 in neurons and microglia, expression of COX-2 in astroglial cells has not been investigated in detail. Here we show that amyloid beta peptide 25-35 (Abeta(25-35)) induces COX-2 mRNA and protein synthesis and a subsequent release of prostaglandin E(2) (PGE(2)) in primary midbrain astrocytes. We further demonstrate that protein kinase C (PKC) is involved in Abeta(25-35)-induced COX-2/PGE(2) synthesis. PKC-inhibitors prevent Abeta(25-35)-induced COX-2 and PGE(2) synthesis. Furthermore Abeta(25-35) rapidly induces the phosphorylation and enzymatic activation of PKC in primary rat midbrain glial cells and in primary human astrocytes from post mortem tissue. Our data suggest that the PKC isoforms alpha and/or beta are most probably involved in Abeta(25-35)-induced expression of COX-2 in midbrain astrocytes. The potential role of astroglial cells in the phagocytosis of amyloid and the involvement of PGs in this process suggests that a modulation of PGs synthesis may be a putative target in the prevention of amyloid deposition.

Neurobiology of Disease, 2012

Mutations in the PARK6 gene coding for PTEN-induced kinase 1 (PINK1) cause recessive earlyonset p... more Mutations in the PARK6 gene coding for PTEN-induced kinase 1 (PINK1) cause recessive earlyonset parkinsonism. Although PINK1 and Parkin promote the degradation of depolarized mitochondria in cultured cells, little is known about changes in signaling pathways that may additionally contribute to dopamine neuron loss in recessive parkinsonism. Accumulating evidence implicates impaired Akt cell survival signaling in sporadic and familial PD (PD). IGF-1/ Akt signaling inhibits dopamine neuron loss in several animal models of PD and both IGF-1 and insulin are neuroprotective in various settings. Here, we tested whether PINK1 is required for insulin-like growth factor 1 (IGF-1) and insulin dependent phosphorylation of Akt and the regulation of downstream Akt target proteins. Our results show that embryonic fibroblasts from PINK1-deficient mice display significantly reduced Akt phosphorylation in response to both IGF-1 and insulin. Moreover, phosphorylation of glycogen synthase kinase-3β (GSK-3β) and nuclear exclusion of FoxO1 are decreased in IGF-1 treated PINK1-deficient cells. In addition, phosphorylation of ribosomal protein S6 is reduced indicating decreased activity of mitochondrial target of rapamycin (mTOR) in IGF-1 treated PINK1 −/− cells. Importantly, the protection afforded by IGF-1 against staurosporine-induced metabolic dysfunction and apoptosis is abrogated in PINK1-deficient cells. Moreover, IGF-1-induced Akt phosphorylation is impaired in primary cortical neurons from PINK1-deficient mice. Inhibition of cellular Ser/Thr phosphatases did not increase the amount of phosphorylated Akt in PINK1 −/− cells, suggesting that components upstream of Akt phosphorylation are compromised in PINK1-deficient cells. Our studies show that PINK1 is required for optimal IGF-1 and insulin dependent Akt signal transduction, and raise the possibility that impaired IGF-1/Akt signaling is involved in PINK1-related parkinsonism by increasing the vulnerability of dopaminergic neurons to stress-induced cell death.

Journal of Neuroimmunology, 2006

Inhibition of neuronal cyclooxygenase-2 (COX-2) and hence prostaglandin E2 (PGE2) synthesis by no... more Inhibition of neuronal cyclooxygenase-2 (COX-2) and hence prostaglandin E2 (PGE2) synthesis by non-steroidal anti-inflammatory drugs has been suggested to protect neuronal cells in a variety of pathophysiological situations including Alzheimer's disease and ischemic stroke. Ascorbic acid (vitamin C) has also been shown to protect cerebral tissue in a variety of experimental conditions, which has been attributed to its antioxidant capacity. In the present study, we show that ascorbic acid dose-dependently inhibited interleukin-1beta (IL-1beta)-mediated PGE2 synthesis in the human neuronal cell line, SK-N-SH. Furthermore, in combination with aspirin, ascorbic acid augmented the inhibitory effect of aspirin on PGE2 synthesis. However, ascorbic acid had no synergistic effect along with other COX inhibitors (SC-58125 and indomethacin). The inhibition of IL-1beta-mediated PGE2 synthesis by ascorbic acid was not due to the inhibition of the expression of COX-2 or microsomal prostaglandin E synthase (mPGES-1). Rather, ascorbic acid dose-dependently (0.1-100 microM) produced a significant reduction in IL-1beta-mediated production of 8-iso-prostaglandin F2alpha (8-iso-PGF2alpha), a reliable indicator of free radical formation, suggesting that the effects of ascorbic acid on COX-2-mediated PGE2 biosynthesis may be the result of the maintenance of the neuronal redox status since COX activity is known to be enhanced by oxidative stress. Our results provide in vitro evidence that the neuroprotective effects of ascorbic acid may depend, at least in part, on its ability to reduce neuronal COX-2 activity and PGE2 synthesis, owing to its antioxidant properties. Further, these experiments suggest that a combination of aspirin with ascorbic acid constitutes a novel approach to render COX-2 more sensitive to inhibition by aspirin, allowing an anti-inflammatory therapy with lower doses of aspirin, thereby avoiding the side effects of the usually high dose aspirin treatment.

Journal of Neurochemistry, 2004

The neuropeptide substance P (SP) has been hypothesized to be involved in the etiopathology of af... more The neuropeptide substance P (SP) has been hypothesized to be involved in the etiopathology of affective disorders. This hypothesis is based on the findings that neurokinin-1-receptor antagonists have antidepressant effects in depressed patients and that SP may worsen mood. In this study, we investigated the effect of the mood-stabilizing agents valproic acid (VPA), carbamazepine, and lithium on SP-induced gene expression. As a model system, we used primary rat astrocytes and human astrocytoma cells, which both express functional SP-receptors and, upon stimulation with SP, synthesize interleukin-6 (IL-6), a cytokine which has been shown to be elevated during the acute depressive state. We found that VPA dose-dependently inhibited SP-induced IL-6 synthesis which was seen with pre-incubation periods of 30 min, 3, 7 and 14 days, whereas carbamazepine and lithium showed no inhibitory effect. The inhibitory effect of VPA was not mediated by inhibition of the stress-regulated kinases p38 and p42/44 (Erk1/2) but by inhibition of protein kinase C epsilon activation. Furthermore, VPA down-regulated the expression of the substance P receptor (neurokinin(NK)-1-receptor) as assessed by real-time PCR. Whether both mechanisms contribute to the mood-stabilizing properties of VPA has to be evaluated in further studies.

Journal of Neurochemistry, 2006

Both interleukin-1b (IL-1b) and prostaglandins (PGs) are important mediators of physiological and... more Both interleukin-1b (IL-1b) and prostaglandins (PGs) are important mediators of physiological and pathophysiological processes in the brain. PGE 2 exerts its effects by binding to four different types of PGE 2 receptors named EP1-EP4. EP3 has found to be expressed in neurons, whereas expression of EP3 in glial cells has not been reported in the brain yet. Here we describe IL-1b-induced EP3 receptor expression in human astrocytoma cells, primary astrocytes of rat and human origin and in rat brain. Using western blot, we found a marked up-regulation of EP3 receptor synthesis in human and rat primary glial cells. Intracerebroventricular administration of IL-1b stimulated EP3 receptor synthesis in rat hippocampus.

Journal of Neurochemistry, 2005

] is a widely distributed neurotransmitter which is involved in neuroimmunomodulatory processes. ... more ] is a widely distributed neurotransmitter which is involved in neuroimmunomodulatory processes. Previously, it has been demonstrated that 5-HT may induce interleukin (IL)-6 expression in primary rat hippocampal astrocytes. The present study was undertaken to investigate the molecular pathways underlying this induction of IL-6 synthesis. As a model system, we used the human astrocytoma cell line U373 MG, which synthesizes IL-6 upon stimulation with various inducers. 5-HT dose-and timedependently induced IL-6 protein synthesis. We identified several 5-HT receptors to be expressed on U373 MG cells, including the 5-HT 1D , 5-HT 2A , 5-HT 3 and 5-HT 7 receptors. In this report, we show that the 5-HT-induced IL-6 release is mediated by the 5-HT 7 receptor based on several agonist/ antagonists that were used. 5-HT-induced IL-6 synthesis is inhibited by the partially selective 5-HT 7 receptor antagonist, pimozide, and the selective antagonist SB269970. Furthermore, IL-6 synthesis was induced by the 5-HT 7 receptor agonist carboxamidotryptamin. In addition, we found p38 MAPKs and protein kinase C (PKC) e to be involved in 5-HTinduced IL-6 synthesis as specific inhibitors of these enzymes (SB202190 and RO-31-8425, respectively) blocked 5-HTinduced IL-6 synthesis. Furthermore, 5-HT mediated the phosphorylation of both p38 MAPK as well as the PKC e isoform. The p42/44 MAPKs, however, were not involved in 5-HT-induced IL-6 synthesis. This study shows, for the first time, a central role of 5-HT 7 receptor linked to p38 MAPK and PKC e for the induction of cytokine synthesis in astrocytic cells.

Journal of Neurochemistry, 2004

Trichloroethylene, a common industrial solvent and a metabolic precursor of chloral hydrate, occu... more Trichloroethylene, a common industrial solvent and a metabolic precursor of chloral hydrate, occurs widely in the environment. Chloral hydrate, which is also used as a hypnotic, has been found to condense spontaneously with tryptamine, in vivo, to give rise to a highly unpolar 1-trichloromethyl-1,2,3,4-tetrahydro-b-carboline (TaClo) that has a structural analogy to the dopaminergic neurotoxin N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Earlier studies have revealed the relative permeability of the molecule through the blood-brain barrier and its ability to induce Parkinson-like symptoms in rats. In this study, we report that TaClo induces an apoptotic pathway in the human neuroblastoma cell line, SK-N-SH, involving the translocation of mitochondrial cytochrome c to the cytosol and activation of caspase 3. TaClo-induced apoptosis shows considerable differences from that mediated by other Parkinson-inducing agents such as MPTP, rotenone and manganese. Although it is not clear if the clinically administered dosage of chloral hydrate or the relatively high environmental levels of trichloroethylene could lead to an onset of Parkinson's disease, the spontaneous in vivo formation of TaClo and its pro-apoptotic properties, as shown in this report, should be considered.

Journal of Molecular Neuroscience, 2005

Prostaglandin E 2 (PGE 2 ), is a major prostanoid produced by the activity of cyclooxygenases (CO... more Prostaglandin E 2 (PGE 2 ), is a major prostanoid produced by the activity of cyclooxygenases (COX) in response to various physiological and pathological stimuli. PGE 2 exerts its effects by activating four specific E-type prostanoid receptors (EP1, EP2, EP3, and EP4). In the present study, we analyzed the expression of the PGE 2 receptor EP1 (mRNA and protein) in different regions of the adult rat brain (hippocampus, hypothalamus, striatum, prefrontal cerebral cortex, parietal cortex, brain stem, and cerebellum) using reverse transcriptionpolymerase chain reaction, Western blotting, and immunohistochemical methods. On a regional basis, levels of EP1 mRNA were the highest in parietal cortex and cerebellum. At the protein level, we found very strong expression of EP1 in cerebellum, as revealed by Western blotting experiments. Furthermore, the present study provides for the first time evidence that the EP1 receptor is highly expressed in the cerebellum, where the Purkinje cells displayed very high immunolabeling of their perikaryon and dendrites, as observed in the immunohistochemical analysis. Results from the present study indicate that the EP1 prostanoid receptor is expressed in specific neuronal populations, which possibly determine the region-specific response to PGE 2 .

European Journal of Immunology, 2013

Mutations in PTEN-induced kinase 1 (PINK1), a serine/threonine kinase linked to familial early-on... more Mutations in PTEN-induced kinase 1 (PINK1), a serine/threonine kinase linked to familial early-onset Parkinsonism, compromise mitochondrial integrity and metabolism and impair AKT signaling. As the activation of a naïve T cell requires an AKT-dependent reorganization of a cell's metabolic machinery, we sought to determine if PINK1-deficient T cells lack the ability to undergo activation and differentiation. We show that CD4 + T cells from PINK1 knockout mice fail to properly phosphorylate AKT upon activation, resulting in reduced expression of the IL-2 receptor subunit CD25. Following, deficient

European Journal of Clinical Investigation, 2009

Background There is a pressing need for research that will lead to the development of new therape... more Background There is a pressing need for research that will lead to the development of new therapeutic approaches for treating inflammatory bowel disease (IBD). The aim of this study was to investigate the effects of tropisetron, a 5-Hydroxytryptamine (5-HT)-3 receptor antagonist with anti-inflammatory properties in a model of experimental colitis in rat.