Showing papers by "Valina L. Dawson published in 2000"

Parkin functions as an E2-dependent ubiquitin– protein ligase and promotes the degradation of the synaptic vesicle-associated protein, CDCrel-1

Abstract: Parkinson's disease is a common neurodegenerative disorder in which familial-linked genes have provided novel insights into the pathogenesis of this disorder Mutations in Parkin, a ring-finger-containing protein of unknown function, are implicated in the pathogenesis of autosomal recessive familial Parkinson's disease Here, we show that Parkin binds to the E2 ubiquitin-conjugating human enzyme 8 (UbcH8) through its C-terminal ring-finger Parkin has ubiquitin–protein ligase activity in the presence of UbcH8 Parkin also ubiquitinates itself and promotes its own degradation We also identify and show that the synaptic vesicle-associated protein, CDCrel-1, interacts with Parkin through its ring-finger domains Furthermore, Parkin ubiquitinates and promotes the degradation of CDCrel-1 Familial-linked mutations disrupt the ubiquitin–protein ligase function of Parkin and impair Parkin and CDCrel-1 degradation These results suggest that Parkin functions as an E3 ubiquitin–protein ligase through its ring domains and that it may control protein levels via ubiquitination The loss of Parkin's ubiquitin–protein ligase function in familial-linked mutations suggests that this may be the cause of familial autosomal recessive Parkinson's disease

Requirement for nitric oxide activation of p21ras/extracellular regulated kinase in neuronal ischemic preconditioning

Abstract: The mechanisms underlying neuronal ischemic preconditioning, a phenomenon in which brief episodes of ischemia protect against the lethal effects of subsequent periods of prolonged ischemia, are poorly understood. Ischemia can be modeled in vitro by oxygen-glucose deprivation (OGD). We report here that OGD preconditioning induces p21ras (Ras) activation in an N-methyl-d-aspartate receptor- and NO-dependent, but cGMP-independent, manner. We demonstrate that Ras activity is necessary and sufficient for OGD tolerance in neurons. Pharmacological inhibition of Ras, as well as a dominant negative mutant Ras, block OGD preconditioning whereas a constitutively active form of Ras promotes neuroprotection against lethal OGD insults. In contrast, the activity of phosphatidyl inositol 3-kinase is not required for OGD preconditioning because inhibition of phosphatidyl inositol 3-kinase with a chemical inhibitor or with a dominant negative mutant does not have any effect on the development of OGD tolerance. Furthermore, using recombinant adenoviruses and pharmacological inhibitors, we show that downstream of Ras the extracellular regulated kinase cascade is required for OGD preconditioning. Our observations indicate that activation of the Ras/extracellular regulated kinase cascade by NO is a critical mechanism for the development of OGD tolerance in cortical neurons, which may also play an important role in ischemic preconditioning in vivo.

NMDA But Not Non-NMDA Excitotoxicity is Mediated by Poly(ADP-Ribose) Polymerase

Abstract: Poly(ADP-ribose) polymerase (PARP-1), a nuclear enzyme that facilitates DNA repair, may be instrumental in acute neuronal cell death in a variety of insults including, cerebral ischemia, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced parkinsonism, and CNS trauma. Excitotoxicity is thought to underlie these and other toxic models of neuronal death. Different glutamate agonists may trigger different downstream pathways toward neurotoxicity. We examine the role of PARP-1 in NMDA- and non-NMDA-mediated excitotoxicity. NMDA and non-NMDA agonists were stereotactically delivered into the striatum of mice lacking PARP-1 and control mice in acute (48 hr) and chronic (3 week) toxicity paradigms. Mice lacking PARP-1 are highly resistant to the excitoxicity induced by NMDA but are as equally susceptible to AMPA excitotoxicity as wild-type mice. Restoring PARP-1 protein in mice lacking PARP-1 by viral transfection restored susceptibility to NMDA, supporting the requirement of PARP-1 in NMDA neurotoxicity. Furthermore, Western blot analyses demonstrate that PARP-1 is activated after NMDA delivery but not after AMPA administration. Consistent with the theory that nitric oxide (NO) and peroxynitrite are prominent in NMDA-induced neurotoxicity, PARP-1 was not activated in mice lacking the gene for neuronal NO synthase after NMDA administration. These results suggest a selective role of PARP-1 in glutamate excitoxicity, and strategies of inhibiting PARP-1 in NMDA-mediated neurotoxicity may offer substantial acute and chronic neuroprotection.

Dynamic regulation of neuronal NO synthase transcription by calcium influx through a CREB family transcription factor-dependent mechanism

Abstract: Neuronal nitric oxide (NO) synthase (nNOS) is dynamically regulated in response to a variety of physiologic and pathologic stimuli. Although the dynamic regulation of nNOS is well established, the molecular mechanisms by which such diverse stimuli regulate nNOS expression have not yet been identified. We describe experiments demonstrating that Ca(2+) entry through voltage-sensitive Ca(2+) channels regulates nNOS expression through alternate promoter usage in cortical neurons and that nNOS exon 2 contains the regulatory sequences that respond to Ca(2+). Deletion and mutational analysis of the nNOS exon 2 promoter reveals two critical cAMP/Ca(2+) response elements (CREs) that are immediately upstream of the transcription start site. CREB binds to the CREs within the nNOS gene. Mutation of the nNOS CREs as well as blockade of CREB function results in a dramatic loss of nNOS transcription. These findings suggest that nNOS is a Ca(2+)-regulated gene through the interactions of CREB on the CREs within the nNOS exon 2 promoter and that these interactions are likely to be centrally involved in the regulation of nNOS in response to neuronal injury and activity-dependent plasticity.

Stroke outcome in double-mutant antioxidant transgenic mice.

Abstract: Background and Purpose—Both NO and superoxide cytotoxicity are important in experimental stroke; however, it is unclear whether these molecules act within parallel pathological pathways or as coreagents in a common reaction. We examined these alternatives by comparing outcomes after middle cerebral artery occlusion in male and female neuronal NO synthase (nNOS)-deficient (nNOS−/−) or human CuZn superoxide dismutase–overexpressing (hSOD1+/−) mice and a novel strain with both mutations. Methods—Permanent middle cerebral artery occlusion was performed by use of the intraluminal filament technique (18 hours). Neurological status was scored, and tissue infarction volume was determined by 2,3,5-triphenyltetrazolium staining and image analysis. Results—Hemispheric infarction volume was reduced in each transgenic strain relative to the genetically matched, wild-type, control cohorts (WT mice): nNOS−/− (80±6 mm3) and double-mutant (49±6 mm3) mice versus WT mice (114±7 mm3) and hSOD1+/− mice (52±7 mm3) versus WT mi...

Method of using selective parp inhibitors to prevent or treat neurotoxicity

Abstract: Neutral tissue damage resulting for ischemia and reperfusion injury or neurodegenerative diseases can be prevented by administering therapeutically effective amounts of certain selective inhibitors of poly(ADP-ribose)polymerase. The inhibitors can be administered intravenously, intraperitoneally, intramuscularly, intraventricularly, or orally. They can be administered as a capsule or tablet containing single or divided dose. Alternatively, the inhibitors can be administered as a sterile solution, suspension or emulsion.

Of Flies and Mice

Abstract: Parkinson's disease (PD) is one of the commonest neurodegenerative diseases of old age but its causes remain unclear. It is known that a cardinal feature of the disease is loss of dopamine neurons in the substantia nigra of the brain with concomitant loss of motor control. Several genes that are mutated in the rare inherited form of the disease have been discovered. Now, as [Dawson][1] outlines, two recent papers report the first animal models of PD in flies and mice, fueling optimism that there will be rapid further progress in working out the pathogenesis of this disease and in designing more effective treatments. [1]: http://www.sciencemag.org/cgi/content/full/288/5466/631

Requirement for nitric oxide activation of p21 ras yextracellular regulated kinase in

Abstract: receptor- and NO-dependent, but cGMP-independent, manner. We demonstrate that Ras activity is necessary and sufficient for OGD tolerance in neurons. Pharmacological inhibition of Ras, as well as a dominant negative mutant Ras, block OGD preconditioning whereas a constitutively active form of Ras promotes neuropro- tection against lethal OGD insults. In contrast, the activity of phosphatidyl inositol 3-kinase is not required for OGD precondi- tioning because inhibition of phosphatidyl inositol 3-kinase with a chemical inhibitor or with a dominant negative mutant does not have any effect on the development of OGD tolerance. Further- more, using recombinant adenoviruses and pharmacological inhib- itors, we show that downstream of Ras the extracellular regulated kinase cascade is required for OGD preconditioning. Our observa- tions indicate that activation of the Rasyextracellular regulated kinase cascade by NO is a critical mechanism for the development of OGD tolerance in cortical neurons, which may also play an important role in ischemic preconditioning in vivo.

Influence of Nitric Oxide on Neuroendocrine Function and Behavior

Abstract: Publisher Summary Nitric oxide (NO) functions as a neurotransmitter and neuromodulator in the central and peripheral nervous systems. This chapter describes how the use of pharmacological agents that block synthesis of NO, the use of knockout mice that lack one of the various isoforms of nitric oxide synthase, and neuroanatomical studies have revealed that NO appears to play an important role in the control of several neuroendocrine pathways and behaviors. The effects of NO on blood vessel tone and neuronal function form the basis for an important role of NO on neuroendocrine function and behavior. NO mediates hypothalamic-portal blood flow, and thus, affects neuropeptide secretion, as well as mediates neuroendocrine function in the hypothalamuspituitary-adrenal and the hypothalamuspituitary-gonadal axes. NO influences several motivated behaviors including sexual, aggressive, and ingestive. Learning and memory functions are also influenced by NO. Taken together, NO is emerging as an important chemical mediator of neuroendocrine function and behavior.

Neurotoxic Actions and Mechanisms of Nitric Oxide

Abstract: Publisher Summary Nitric oxide (NO) has revolutionized the perception of neurotransmission and neuronal signaling. It is emerging as a key regulator of numerous physiological responses. However, excessive generation of NO can mediate neuronal damage in a variety of neurologic diseases. Understanding the pathways through which NO causes neuronal cell death is key in the development of more effective therapies. This chapter reviews the current knowledge on the mechanisms of NO-mediated neurotoxicity. NO damages neurons in the reaction with superoxide anion to generate peroxynitrite. Peroxynitrite is a potent cellular toxin and is capable of altering proteins, lipids, and DNA. NO may interact directly with a large number of proteins and cause subsequent alteration of their function. NO-induced cell death is characterized by typical features of both necrosis and/or apoptosis. NO plays a role as a neuronal-cell-death mediator in a variety of disorders of the nervous system. This chapter discusses the involvement of NO in excitoxicity, stroke, ischemic reconditioning, and Parkinson's disease.

Nitric Oxide in Brain Ischemia/Reperfusion Injury

Abstract: The brain is a highly energetic organ, which requires a continuous supply of nutrients and oxygen from the circulatory system in order to function properly. Cessation of the blood supply to the brain for even a few minutes results in neuronal injury following initiation of a cascade of secondary mechanisms. Ischemia results in the reduction of resting membrane potential of glia and neurons in the brain. The leakage of potassium out of cells results in the depolarization of neurons, leading to a massive release of glutamate. Glutamate elicits its actions by acting on series of post-synaptic receptors, including the N-methyl-D-aspartate receptor (NMDA), non-NMDA receptors, and metabotropic glutamate receptors (SAMDANI et al. 1997). For nearly two decades, the actions of glutamate have been linked as important mediators of ischemic brain injury. The observation that activation of NMDA receptors generates nitric oxide (NO) in a calcium-dependent manner (GARTHWAITE et al. 1988; BREDT and SNYDER 1989; GARTHWAITE et al. 1989) raised the possibility that NO might be an important mediator in regulating glutamate neurotoxicity. The observation that non-selective NO synthase (NOS) inhibitors could reduce glutamate neurotoxicity in vitro (DAWSON et al. 1991b) and reduce infarct volume following transient focal ischemia in mice (NOWICKI et al. 1991) suggested a role for NO as a neurotoxin. Immediately, there was controversy over the role of NO in neurotoxicity and ischemic damage (DAWSON et al. 1994a; DAWSON and DAWSON 1996). Since then, a large body of scientific literature has evolved describing the role of NO in glutamate toxicity and ischemia-reperfusion injury. The early controversies were due to the important but opposing effects of NO generated from different NOS isoforms in the central nervous system (CNS), the use of nonselective NOS inhibitors, and the lack of understanding of the complex chemistry of NO in a biologic setting. Advances in pharmacology and chemistry, in addition to the generation of genetically engineered mice, have greatly expanded our understanding of NO biology in ischemic injury.

The NO Signaling Pathway in the Brain

Abstract: Nitric oxide (NO) is a widespread and multifunctional biological messenger molecule. It mediates vasodilation of blood vessels, host defense against infectious agents and tumors, and neurotransmission of the central and peripheral nervous systems (1–3). The discovery of NO as a messenger molecule in the nervous system also revised conventional concepts of neurotransmitters. Compared with the traditional neuronal messenger molecules, NO has a variety of distinguished features. NO is probably the smallest and most versatile bioactive molecule identified, it diffuses freely across membranes, it is not stored in synaptic vesicles, and it is not released by exocytosis upon membrane depolarization. NO seems to be terminated primarily by reactions with its targets. In the nervous system, NO may play a role not only in physiologic neuronal functions, such as neurotransmitter release, neural development, regeneration, synaptic plasticity, and regulation of gene expression, but also in pathological conditions in which deregulated excessive production of NO leads to neural injury. Furthermore, rapid progress is now being made in understanding the regulation of NOS activity and the cellular and molecular targets of NO under physiologic and pathologic conditions. Some of the newly revealed roles for NO in the nervous system include regulation or control of neuronal morphogenesis, short-term or long-term synaptic plasticity, regulation of gene expression, and modification of sexual and aggressive behavior. Excess formation of NO plays a role in neural injury in several kinds of neurologic insults, which has promoted the development of selective NOS inhibitors for the treatment of neurologic disorders.