Signaling Pathways in Reactive Astrocytes,a Genetic Perspective

Reactive astrocytes are associated with a vast array of central nervous system (CNS) pathologies. The activation of astrocytes is characterized by changes in their molecular and morphological features, and depending on the type of damage can also be accompanied by inflammatory responses, neuronal damage, and in severe cases, scar formation. Although reactive astrogliosis is the normal physiological response essential for containing damage, it can also have detrimental effects on neuronal survival and axon regeneration, particularly in neurodegenerative diseases. It is believed that progressive changes in astrocytes as they become reactive are finely regulated by complex intercellular and intracellular signaling mechanisms. However, these have yet to be sorted out. Much has been learned from gain-of-function approaches in vivo and culture paradigms, but in most cases, loss-of-function genetic studies, which are a critical complementary approach, have been lacking. Understanding which signaling pathways are required to control different aspects of astrogliosis will be necessary for designing therapeutic strategies to improve their beneficial effects and limit their detrimental ones in CNS pathologies. In this article, we review recent advances in the mechanisms underlying the regulation of aspects of astrogliosis, with the main focus on the signaling pathways that have been studied using loss-of-function genetic mouse models.     

Extracellular Glutamate Is Increased in Thalamus During Thiamine Deficiency-Induced Lesions and Is Blocked by MK-801

The current study measured extracellular fluid (ECF) levels of excitatory amino acids before and during the onset of thiamine deficiency-induced pathologic lesions. Male Sprague-Dawley rats were treated with daily pyrithiamine (0.25 mg/kg i.p.) and a thiamine-deficient diet (PTD). Microdialysates were simultaneously collected from probes inserted acutely via guide cannulae into right paracentral and ventrolateral nuclei of thalamus and left hippocampus of PTD and pair-fed controls. Hourly samples were collected from unanesthetized and freely moving animals. Basal levels obtained at a prelesion stage (day 12 of PTD treatment) were unchanged from levels in pairfed controls. In samples collected 4–5 h after onset of seizures (day 14 of PTD), the levels of glutamate were elevated an average 640% of basal levels in medial thalamus and 200% in hippocampus. Glutamine levels declined, taurine and glycine were elevated, and aspartate, GABA, and alanine were unchanged during this period. Within 7 h after seizure onset glutamine was undetectable in both areas, whereas glutamate had declined to ～200% in thalamus and 70% in hippocampus. No significant change in glutamate, aspartate, or other amino acids was observed in dialysates collected from probes located in undamaged dorsal-lateral regions of thalamus. Number of neurons within ventrolateral nucleus of thalamus was significantly greater in PTD animals in which the probe was dialyzed compared with nondialyzed, suggesting that removal of excitatory amino acids was protective. No significant pathologic damage was evident in hippocampus. Pretreatment with MK-801 completely blocked the rise of ECF glutamate and significantly reduced the pathologic damage within thalamus of PTD rats and produced a significant decrease in ECF glutamate in control rats.     

PDAPP转基因小鼠脑组织内反应性星形胶质细胞的活化程度明显升高

目的　研究PDAPP转基因小鼠脑组织内反应性星形胶质细胞的活化程度。方法　通过免疫组织化学染色方法检测小鼠脑组织内反应性星形胶质细胞表达胶质纤维酸性蛋白 (GFAP)的情况 ,比较PDAPP转基因小鼠和C5 7 BL非转基因小鼠脑组织反应性星形胶质细胞的活化程度。结果　PDAPP转基因小鼠脑组织内反应性星形胶质细胞表达GFAP的水平明显高于C5 7 BL非转基因小鼠。结论　PDAPP转基因小鼠脑组织内存在明显的神经炎症反应     

Dopamine Induces Ca2+ Signaling in Astrocytes through Reactive Oxygen Species Generated by Monoamine Oxidase

Dopamine is a neurotransmitter that plays a major role in a variety of brain functions, as well as in disorders such as Parkinson disease and schizophrenia. In cultured astrocytes, we have found that dopamine induces sporadic cytoplasmic calcium ([Ca²⁺]_c) signals. Importantly, we show that the dopamine-induced calcium signaling is receptor-independent in midbrain, cortical, and hippocampal astrocytes. We demonstrate that the calcium signal is initiated by the metabolism of dopamine by monoamine oxidase, which produces reactive oxygen species and induces lipid peroxidation. This stimulates the activation of phospholipase C and subsequent release of calcium from the endoplasmic reticulum via the inositol 1,4,5-trisphosphate receptor mechanism. These findings have major implications on the function of astrocytes that are exposed to dopamine and may contribute to understanding the physiological role of dopamine.     

Blockade of "reverse tolerance" to cocaine and amphetamine by MK-801

"Reverse tolerance" was produced in rats and mice by repeated exposure to either cocaine or amphetamine. The locomotorstimulant effect was studied in mice; stereotypy and convulsions in rats. MK-801, the NMDA antagonist, blocked the development of "reverse tolerance" to all three effects. In contrast, haloperidol selectively blocked "reverse tolerance" to cocaine-induced stereotypy but not to convulsions. The data suggest that the glutamate system participates in the mechanism of "reverse tolerance" to the dopaminergic effects of cocaine and amphetamine, as well as to the convulsant effect of cocaine.     

N-methyl-D-aspartate prevented memory deficits induced by MK-801 in mice

An interaction between N-methyl-D-aspartate (NMDA) and MK-801 was examined in mice using a modified elevated plus-maze paradigm that allows assessment of the adaptive form of spatial memory. NMDA administered (s.c.) immediately after the acquisition session protected the animals against the amnesia induced by MK-801 given shortly before the retention session. Behavioral performance, expressed as the transfer latency, and therefore spatial memory potency of NMDA plus MK-801 treated animals was comparable with that of both NMDA-treated animals and the controls.     

Disruption of sensitization to methylphenidate by a single administration of MK-801

Blockade of sensitization to methylphenidate by a single injection of MK-801 was investigated using a computerized activity monitoring system. Male Sprague-Dawley rats were housed in test cages and motor activity was recorded continuously for 16 days. After 2 days of baseline recording and a saline injection on day 3, the rats were randomly divided into four experimental groups. All received 2.5 mg/kg of methylphenidate (s.c.) once a day from days 4 to 9, then after five days of no treatment, they were re-challenged with 2.5 mg/kg of methylphenidate on day 15. One group received only methylphenidate, while the other three groups also received a single i.p. injection of MK-801 (0.30 mg/kg) either 24 h (day 3) or 1 h prior to the first of the six methylphenidate injections (day 4), or 1 h prior to the second methylphenidate injection (day 5). A single injection of MK-801 on day 4 (1 h prior to methylphenidate) blocked the development of sensitization to methylphenidate, since a sensitized response could not be elicited six days after cessation of repeated methylphenidate administration (day 15). However, sensitization to methylphenidate still occurred in the groups receiving MK-801 (0.30 mg/kg) on day 5, indicating that the mechanism by which a single injection of MK-801 disrupts sensitization to methylphenidate is sensitive to timing and is not a direct long-term effect. In conclusion, a single injection of MK-801 persistently blocks the development of sensitization to methylphenidate only if it is given with methylphenidate on the first day of the repetitive treatment phase.     

Eps8 Regulates Axonal Filopodia in Hippocampal Neurons in Response to Brain-Derived Neurotrophic Factor (BDNF)

The regulation of filopodia plays a crucial role during neuronal development and synaptogenesis. Axonal filopodia, which are known to originate presynaptic specializations, are regulated in response to neurotrophic factors. The structural components of filopodia are actin filaments, whose dynamics and organization are controlled by ensembles of actin-binding proteins. How neurotrophic factors regulate these latter proteins remains, however, poorly defined. Here, using a combination of mouse genetic, biochemical, and cell biological assays, we show that genetic removal of Eps8, an actin-binding and regulatory protein enriched in the growth cones and developing processes of neurons, significantly augments the number and density of vasodilator-stimulated phosphoprotein (VASP)-dependent axonal filopodia. The reintroduction of Eps8 wild type (WT), but not an Eps8 capping-defective mutant, into primary hippocampal neurons restored axonal filopodia to WT levels. We further show that the actin barbed-end capping activity of Eps8 is inhibited by brain-derived neurotrophic factor (BDNF) treatment through MAPK-dependent phosphorylation of Eps8 residues S624 and T628. Additionally, an Eps8 mutant, impaired in the MAPK target sites (S624A/T628A), displays increased association to actin-rich structures, is resistant to BDNF-mediated release from microfilaments, and inhibits BDNF-induced filopodia. The opposite is observed for a phosphomimetic Eps8 (S624E/T628E) mutant. Thus, collectively, our data identify Eps8 as a critical capping protein in the regulation of axonal filopodia and delineate a molecular pathway by which BDNF, through MAPK-dependent phosphorylation of Eps8, stimulates axonal filopodia formation, a process with crucial impacts on neuronal development and synapse formation.     

The Response to Stimulation in Neurons and Astrocytes

Neurochemical Research - The brain uses mainly glucose as fuel with an index of glucose to oxygen utilization close to 6, the maximal index if all glucose was completely oxidized. However, this...     

Kynurenic acid prevented social recognition deficits induced by MK-801 in rats

MK-801 impaired social recognition potency of adult male rats when given immediately after the initial interaction with a juvenile rat. Administration of kynurenic acid prior to the initial interaction protected the adults against recognition deficits induced by MK-801. When re-exposed at a delay of 30 min to the familiar juvenile, social investigation in the adults was significantly reduced. Thus, the adults are able to remember olfactory stimuli emitted by juvenile con-specifics.     

Behavioral effects of MK-801 in morphine-dependent and non-dependent mice

The behavioral effects of MK-801 were compared in morphine-dependent and non-dependent mice. The dose of MK-801 selected for these studies was previously demonstrated to attenuate some of the morphine withdrawal signs. Subjects were repeatedly exposed to morphine (8 days, b.i.d., 10–100 mg/kg, s.c.). Twenty-four hours after last morphine injection mice received naloxone (0.1 mg/kg, s.c.) and the observation was commenced. Animals were pretreated with either MK-801 (0.1 mg/kg, i.p.) or saline 30 min prior to testing. It was found that the behavioral effects of MK-801 (decreased sociability and increased rate of transitions between behavioral elements, locomotion, grooming) were less pronounced in morphine-dependent compared to non-dependent subjects. However, the intensified almost stereotypic eating possibly reflected increased psychotomimetic potency of MK-801 in morphine-withdrawn animals.     

BMP Signaling in Astrocytes Downregulates EGFR to Modulate Survival and Maturation

Astrocytes constitute a major cell population in the brain with a myriad of essential functions, yet we know remarkably little about the signaling pathways and mechanisms that direct astrocyte maturation. To explore the signals regulating astrocyte development, we prospectively purified and cultured immature postnatal rodent astrocytes. We identified fibroblast growth factors (FGFs) and bone morphogenetic proteins (BMPs) as robust trophic factors for immature astrocytes. We showed that astrocytes respond directly to BMPs via phosphorylation of the smad1/5/8 pathway. In vitro, BMP signaling promoted immature astrocytes to adopt multiple characteristics of mature astrocytes, including a more process-bearing morphology, aquaporin-4 (AQP4) and S100β immunoreactivity, limited proliferation, and strong downregulation of epidermal growth factor receptor (EGFR). In vivo, activation of the smad1/5/8 pathway in astrocytes was seen during early postnatal development, but inhibition of astrocyte proliferation was not observed. These insights can aid in the further dissection of the mechanisms and pathways controlling astrocyte biology and development.     

Chronic Administration of Thymoquinone Enhances Adult Hippocampal Neurogenesis and Improves Memory in Rats Via Regulating the BDNF Signaling Pathway

Neurochemical Research - Thymoquinone is a pharmacologically active component of Nigella sativa Linn. seeds. Despite the diverse neuropharmacological attributes of TQ, limited reports related to...     

Signaling from Nucleotide Receptors to Protein Kinase Cascades in Astrocytes

In the CNS, extracellular ATP can function as an excitatory neurotransmitter as well as a trophic factor. These short-term and long-term actions are mediated by nucleotide receptors. Extracellular ATP can also act as a co-mitogen in conjunction with polypeptide growth factors such as basic fibroblast growth factor (FGF2). Cellular proliferation, differentiation and survival are regulated by signaling cascades composed of protein kinases, including extracellular signal regulated protein kinase (ERK) and protein kinase B (also called Akt). Here we summarize recent studies on nucleotide receptor signaling to ERK and Akt in astrocytes and the role of protein kinase cascades in mediating the trophic actions of extracellular ATP, alone or together with FGF2. Because extracellular ATP and FGF2 contribute to the hyperplastic and hypertrophic response of astrocytes to CNS injuries, an understanding of their protein kinase signaling mechanisms may lead to novel therapeutic approaches for neurological conditions that involve gliosis and the generation of reactive astrocytes, such as trauma, stroke, seizure and neurodegenerative and demyelinating disorders.Special issue dedicated to Lawrence F. Eng.     

Enhanced BDNF Signaling is Associated with an Antidepressant-like Behavioral Response and Changes in Brain Monoamines

1. Neurotrophins and serotonin have both been implicated in the pathophysiology of depression and in the mechanisms of antidepressant treatments. 2. Brain-derived neurotrophic factor (BDNF) influences the growth and plasticity of serotonergic (5-HT) neurons via the activation of trkB receptor. 3. Transgenic mice overexpressing the full-length trkB receptor (TrkB.TK+) and showing increased trkB activity in brain, and their wild type (WT) littermates, were injected with the antidepressant fluoxetine or saline, and analyzed behaviorally in the forced swimming test paradigm and biochemically for the concentrations of brain monoamines and their metabolites. 4. The TrkB.TK+ mice displayed increased latency to immobility in the forced swim test, suggesting resistance to behavioral despair. 5. Fluoxetine increased the latency to immobility in wild-type mice to a similar level as seen in the trkB.TK+ mice after saline treatment, but had no further behavioral effect in the swimming behavior of the trkB.TK+ mice. 6. Only minor differences in the levels of brain monoamines and their metabolites were observed between the transgenic and wild-type mice. 7. These data, together with other recent observations, suggest that trkB activation may play a critical role in the behavioral responses to antidepressant drugs in mice.     

Glutamate Uptake Is Stimulated by Extracellular S100B in Hippocampal Astrocytes

Summary  

Modulation of MK-801-elicited mouse popping behavior by galantamine is complex and dose-dependent

The ability of phencyclidine (PCP), a noncompetitive antagonist of NMDA receptor-mediated neurotransmission, to precipitate a schizophreniform psychosis in susceptible individuals is consistent with the hypothesized pathologic occurrence of NMDA receptor hypofunction in this disorder. Because the psychosis caused by PCP resembles schizophrenia in all of the relevant domains of psychopathology, investigators have sought to characterize animal models of NMDA receptor hypofunction. MK-801 (dizocilpine) binds to the same hydrophobic channel domain in the NMDA receptor-associated ionophore as PCP, and has been shown to elicit intense irregular episodes of jumping behavior in mice, termed "popping." MK-801-elicited mouse popping is an animal model of NMDA receptor hypofunction that has been used to screen novel candidate compounds for the treatment of schizophrenia. Recently, a selective abnormality in the transduction of the acetylcholine signal at the level of the alpha 7 nicotinic receptor has been described in schizophrenia. The existence of a nicotinic cholinergic abnormality in schizophrenia has stimulated interest in a potential therapeutic role for positive allosteric modulation of nicotinic receptors. Galantamine is a compound that possesses two interesting properties: inhibition of acetylcholinesterase and positive allosteric modulation of nicotinic neurotransmission. Theoretically, galantamine would be expected to increase the efficiency or likelihood that acetylcholine will promote channel opening and ionic conductance at nicotinic receptors. As expected, in the current investigation statistically significant popping behavior was elicited by MK-801 in mice (T(22) = 2.16, P < 0.05). This MK-801-elicited popping was significantly attenuated by 100 mg/kg of galantamine (T(22) = 2.24, P < 0.05). The data show that nicotinic interventions can influence NMDA receptor-mediated neurotransmission in the intact mouse.