Glutamate-induced homocysteic acid release from astrocytes: possible implication in glia-neuron signaling.

Glial cells synthesise neuroactive substances and release them upon neurotransmitter receptor activation. Homocysteic acid (HCA), an endogenous agonist for glutamatergic N-methyl-D-aspartate (NMDA) receptors, is predominantly localised in glial cells. We have previously demonstrated the release of HCA from mouse astrocytes in culture following activation of beta-adrenergic receptors. Moreover, a release of HCA has also been observed in vivo upon physiological stimulation of sensory afferents in the thalamus. Here we report the glutamate-induced release of HCA from astrocytes. The effect of glutamate was mediated by the activation of ionotropic (NMDA and non-NMDA) as well as by metabotropic receptors. In addition, the release of HCA was Ca(2+)- and Na(+)-dependent, and its mechanism involved the activation of the Na+/Ca(2+)-exchanger. Furthermore, we provide evidence for the presence of functional NMDA receptors on astrocytes, which are coupled to an intracellular Ca2+ increase via stimulation of the Na+/Ca(2+)-exchanger. Our data thus favour a participation of glial cells in excitatory neurotransmission and corroborate the role of HCA as a "gliotransmitter."

Release of homocysteic acid from rat thalamus following stimulation of somatosensory afferents in vivo: feasibility of glial participation in synaptic transmission.

The sulphur-containing amino acid homocysteic acid (HCA) is present in and released in vitro from nervous tissue and is a potent neuronal excitant, predominantly activating N-methyl-d-aspartate (NMDA) receptors. However, HCA is localised not in neurones but in glial cells [Eur J Neurosci 3 (1991) 1370], and we have shown that it is released from astrocytes in culture upon glutamate receptor activation [Neuroscience 124 (2004) 377]. We now report the in vivo release of HCA from ventrobasal (VB) thalamus following natural stimulation of somatosensory afferents arising from the facial vibrissae of the rat. Simultaneously with multi-unit recording, [35S]-methionine, a HCA precursor, was perfused through a push-pull cannula in VB thalamus of anaesthetized rats. Perfusates were collected before, during and after 4 min stimulation of the vibrissal afferents with an air jet. A marked release of radiolabeled HCA was observed during and after the stimulation. Furthermore, the beta-adrenoreceptor agonist isoproterenol, which is known to evoke HCA release from glia in vitro, was found to increase the efflux of HCA in the perfusate in vivo. In separate experiments, the excitatory actions of iontophoretically applied HCA on VB neurones were inhibited by the NMDA receptor antagonist CPP, but not by the non-NMDA antagonist CNQX. These results suggest a possible "gliotransmitter" role for HCA in VB thalamus. The release of HCA from glia might exert a direct response or modulate responses to other neurotransmitters in postsynaptic neurons, thus enhancing excitatory processes.

Quality of life in children with traumatic brain injury--basic issues, assessment, and recommendations.

An international Task Force was convened under the guidance of BMBF Conference so as to review the "State of the art" for measuring quality of life (QoL) in children who have suffered traumatic brain injury (TBI). After expert review of instruments and evaluation of two independent literature reviews this work group established "inclusion criteria" for the review of current tools that could contribute to the measurement of QoL in children with TBI. Six instruments were determined to meet all or most of the criteria required to be used in current clinical practice and research for children with TBI.

Glial-neuronal transfer of arginine and S-nitrosothiols in nitric oxide transmission.

The arginine-nitric oxide (Arg-NO) and the S-nitrosothiols systems, two less well-studied aspects of NO transmission in the central nervous system, are reviewed. A growing body of evidence suggested that they play a crucial role in NO synthesis and activity. l-Arginine, the NO precursor, is predominantly localized in glia. Together with in vitro and in vivo results of arginine release, this suggests a transfer of arginine from glia to neurons in order to supply NO synthase with its substrate. NO biosynthesis may thus involve the co-occurrence of the glial-neuronal transfer of arginine and of NOS activation. The arginine availability may shed light on the dual, beneficial and toxic effects of NO. At low arginine concentrations, neuronal NO synthase generates NO and superoxide, favouring the production of the toxin peroxynitrite. NMDA-induced excitotoxicity in neuronal cells is dependent on arginine availability and glia may play a neuroprotective role by supplying arginine. The reversible S-nitros(yl)ation of thiol containing molecules may represent an important cellular signal transduction mechanism, probably comparable to phosphorylation. S-nitrosothiols, in particular through the presence and release of S-nitroso-cysteinylglycine in sensory thalamus, may act as a local buffering system in NO transmission. This may represent a novel specific facilitating mechanism in order to enhance transmission of persistent stimuli.

Long-term follow-up of an epidemiologically defined cohort of patients with Tourette syndrome.

The goal of this study was to collect prospective longitudinal information on the development of an epidemiologically defined cohort of patients with Tourette syndrome. These data may improve prognostic understanding of the condition. This information will also be important for specification of an adult phenotype for genetic marker studies. A prospective longitudinal cohort study was conducted. Fifty-four of 73 patients from our 1986 prevalence study of Tourette syndrome in North Dakota school-aged children were eligible for inclusion. The subjects were diagnosed in 1984 and 1985. We were able to interview 39 of 54 eligible patients for 507 person-years of follow-up. For the cohort, tic severity declined by 59%, global assessment of functioning improved by 50%, and the average number of comorbidities decreased by 42%. Forty-four percent of patients were essentially symptom free at follow-up. Only 22% were on medication as adults. Tourette syndrome is a developmental neuropsychiatric disorder with a long-term course that is favorable for most patients. Males demonstrated substantially more variability in improvement but overall demonstrated more improvement than females.

Archaeological evidence of teosinte domestication from Guilá Naquitz, Oaxaca.

Analysis of the three most ancient Zea mays inflorescence fragments from Guilá Naquitz, Oaxaca, Mexico shows they did not disarticulate naturally, indicating that agricultural selection of domesticated teosinte was underway by 5,400 (14)C years before the present (about 4,200 dendrocalibrated years B.C.). The cooccurrence of two-ranked specimens with two rows and four rows of grain and numerous additional morphological characteristics of these specimens support hypotheses based on molecular and quantitative genetic analyses that maize evolved from teosinte. Domestication of the wild ancestor of maize occurred before the end of the 5th millennium B.C.

Children's spatial behavior is differentially affected after traumatic brain injury.

Spatial behavior in 20 children with severe traumatic brain injury (TBI) and 20 healthy controls was investigated using the Kiel Locomotor Maze. Children had to remember defined locations in an experimental chamber with completely controlled intra- and extra-maze cues until learning criterion was reached. In a second experiment, spatial orientation strategies were assessed. Children with TBI were shown to be impaired in spatial learning and spatial memory. Spatial orientation was found to be deficient even in cases where spatial learning and memory proved to be unimpaired, especially in tasks that demanded the use of relational place strategies. Children who suffered a TBI at an early age proved to be more severely impaired in spatial learning and orientation than older children.

Glial-derived arginine, the nitric oxide precursor, protects neurons from NMDA-induced excitotoxicity.

Excitotoxic neuronal cell death is characterized by an overactivation of glutamate receptors, in particular of the NMDA subtype, and the stimulation of the neuronal nitric oxide synthase (nNOS), which catalyses the formation of nitric oxide (NO) from l-arginine (L-Arg). At low L-Arg concentrations, nNOS generates NO and superoxide (O2(.)(-)), favouring the production of the toxin peroxynitrite (ONOO-). Here we report that NMDA application for five minutes in the absence of added L-Arg induces neuronal cell death, and that the presence of L-Arg during NMDA application prevents cell loss by blocking O2(.)(-) and ONOO- formation and by inhibiting mitochondrial depolarization. Because L-Arg is transferred from glial cells to neurons upon activation of glial glutamate receptors, we hypothesized that glial cells play an important modulator role in excitotoxicity by releasing L-Arg. Indeed, as we further show, glial-derived L-Arg inhibits NMDA-induced toxic radical formation, mitochondrial dysfunction and cell death. Glial cells thus may protect neurons from excitotoxicity by supplying L-Arg. This potential neuroprotective mechanism may lead to an alternative approach for the treatment of neurodegenerative diseases involving excitotoxic processes, such as ischemia.

Population structure and ecology of a tropical rare rhizomatous species of teosinte Zea diploperennis (Gramineae).

One of the objectives of the Sierra de Manantlán Biosphere Reserve (Jalisco, México) is the conservation in situ of the teosinte Zea diploperennis Iltis, Doebley, Guzman & Pazzi. Zea diploperennis is perennial, shade intolerant and its 1-3 m shoots are architecturally similar to maize. Clonal growth is of the phalanx type. Genets are iteroparous (modules semelparous). The demography of seven module and genet populations was studied in seven sites representing three stages of old-field succession. Seven permanent one-meter-square plots were randomly established in each site. All genets initially present and those that became established during our study were mapped and labeled according to year of establishment. The magnitude of demographic fluctuations was greater in module populations. Genet population dynamics followed a seasonal rhythm with a maximum population size obtained at the onset of the rainy season. A relation was documented between percent annual mortality of a cohort and its age: the younger the cohort, the greater the mortality. This was a statistically significant relationship, Y = [sin(-0.288x + 1.657)]2 (r = 0.92, p < 0.01), where is proportion annual mortality of genets and is the age of the cohort. The maximum rates of genet mortality occurred during the rainy season when population densities were greatest. As a consequence, we postulate that competition occurs principally during the rainy season. Linear relationships were observed between rate of population increase of genets versus old-field successional stage and soil type. Those areas with poor soil (Ultisol), degraded soil or soils with similar physical characteristics could be rehabilitated by introducing Z. diploperennis. Such rehabilitation would achieve two distinct objectives, promote propagation of this rare endemic and reclaim areas that are susceptible to erosion and further degradation.

Novel mode of nitric oxide neurotransmission mediated via S-nitroso-cysteinyl-glycine.

S-nitroso-cysteinyl-glycine, a novel nitric oxide-adduct thiol compound, can be detected in the brain (2.3+/-0.6 pmol/mg protein), and released following stimulation of sensory afferents to the rat ventrobasal thalamus in vivo (resting conditions 17 nM; stimulation: 186 nM). Iontophoretic application of CysNOGly (20-80 nA) onto thalamic neurons in vivo resulted in enhancements of excitatory responses to either NMDA or AMPA (182+/-13.6% and 244+/-27.8% of control values, n = 15). CysNOGly enhanced responses to stimulation of vibrissal afferents to 132+/-2.2% (n = 7) of control values. In contrast, the dipeptide CysGly reduced responses of ventrobasal neurons to NMDA and AMPA (54+/-8.4% and 55+/-10.8% of control, n = 5). CysNOGly was also a potent activator of soluble guanylate cyclase in vitro. Moreover, we found that NMDA elevated CysNOGly levels in vitro and this stimulatory effect was reduced by inhibitors of the neuronal NO synthase and of the gamma-glutamyl transpeptidase, suggesting that production of NO and CysGly is a prelude to CysNOGly synthesis. These findings suggest that the nitrosothiol CysNOGly plays a role in synaptic transmission in the ventrobasal thalamus. We propose a novel synaptic buffering mechanism where S-nitroso-cysteinyl-glycine serves to restrict the locus of action of nitric oxide and so increase its local availability for target delivery. This could lead to a change in neuronal responses favouring sensory transmission similar to that seen in wakefulness or arousal in order to locally enhance transmission of persistent sensory stimuli.

Influence of age-related factors on long-term outcome after traumatic brain injury (TBI) in children: A review of recent literature and some preliminary findings.

Cerebral plasticity of the immature brain is often inferred to lead to less serious consequences of early traumatic brain injury (TBI) in the pediatric age group. This notion is seriously challenged by recent research findings. Data from prospective studies point to some children's dif-ficulties in ongoing skill-acquisition and the possibility of late-emerging deficits. Accordingly, preliminary group data of an own ongoing study support the notion of an increased risk for pervasive neuropsychological impairment in subjects with severe TBI and early age at trauma. The pattern of neuropsychological deficits may depend on the developmental level at the time of injury, although effects of hemispheric site of lesion were also found to persist in individual cases. Theoretical considerations and empirical findings stress the importance of a longitudinal developmental perspective for the evaluation of long-term outcome after pediatric TBI. ("Verbund Neurotrauma Kiel / Project 4: Evaluation of neurological rehabilitation and course of cognitive development in children and adoles-cents with secondarily acquired brain damage", funded through the Research Program "Gesundheit 2000" of the German government, FKZ 01 KO 9512.).

Assessment of sensorimotor functions after traumatic brain injury (TBI) in childhood - Methodological aspects.

Various basic qualitative and quantitative methods for the evaluation of sensorimotor functions after Traumatic Brain Injury (TBI) are introduced and discussed. Methodological aspects are illustrated by a single case follow-up study of a child after severe TBI (age 11; 7–12;1 yrs; 6, 8 and 12 month post TBI) in comparison to an age-matched healthy control group (N=16). The evaluation consisted of neurological investigation, Barthel-Index, Terver Numeric Score for Functional Assessment, Rappaport Disability Rating Scale (modified version), a coordination-test for children (KTK), a pilot-tested Motor Function Score, quantitative evaluation of spatiotemporal gait parameters on a walkway and on a treadmill, and the kinematic assessment of hand motor functions. Quantitative movement analyses revealed two general types of motor disorder: Slowing of movements and compensatory motor strategies. Averaged z-scores showed deficits, which were pronounced in fine motor skills (hand movements: 1.86, gait: 1.3). During follow-up, a strong improvement rate during the first (-0.48 z-scores) and nearly no improvement rate (-0.03 z-scores) during the second time interval was seen. Clinical scores and developmental tests were not able to document the whole restitutional course, whereas motor tests with special emphasis on functional aspects and the quantitative movement assessment seemed to be suitable methods. We conclude that a sufficient evaluation of sensorimotor functions after TBI in childhood needs an increase in procedural uniformity on onehand and the combination of various qualitative and quantitative methods on the other hand. To connect both claims, further research is necessary.

beta-Adrenergic stimulation promotes homocysteic acid release from astrocyte cultures: evidence for a role of astrocytes in the modulation of synaptic transmission.

The sulfur-containing amino acid homocysteic acid (HCA) is present in and released from nervous tissue, exerting excitatory effects on neurons by predominantly activating NMDA receptors. It is interesting that HCA appears to be exclusively localized in glial cells, not in neurons. This profile of glial localization and excitatory action on neurons has led to the hypothesis that HCA could participate in intercellular communication in the brain as a "gliotransmitter." To test this hypothesis further, we searched for specific, receptor-mediated stimuli that could induce release of HCA from cultured astrocytes. For this reason we tested the effect of noradrenaline and vasoactive intestinal peptide, two transmitters known to interact with specific receptors on astrocytes, on the release of HCA from these cells. Noradrenaline and the beta-adrenergic agonist isoproterenol induced an efflux of HCA from astrocyte cultures. Further stressing the beta-adrenergic mediation of this effect is the blockade by atenolol of the HCA release evoked by isoproterenol. The stimulation of HCA release from astrocytes was not observed with the alpha-noradrenergic agonist methoxamine and with vasoactive intestinal peptide. These results taken together further strengthen the role of HCA as a gliotransmitter. Its efflux from glia could be controlled by noradrenaline, activating beta-adrenergic receptors on astrocytes. The present study provides the first evidence for an influence of beta-adrenergic receptor activation on the release of an excitatory amino acid from astrocytes and further supports the notion that glial-neuronal interactions play a role in synaptic transmission.

Glutamate-induced release of the nitric oxide precursor, arginine, from glial cells.

Arginine, the nitric oxide precursor, is predominantly localized in glial cells, whereas the constitutive nitric oxide synthase is mainly found in neurons. Therefore, a transfer of arginine from glial cells to neurons is needed to replenish the neuronal precursor pool. This is further supported by the finding that arginine is released upon selective pathway stimulation both in vitro and in vivo. We investigated the mechanism underlying this glial-neuronal interaction by analysing the effect of glutamate receptor agonists on the extracellular [3H]arginine level in cerebellar and cortical slices and in cultures of either cortical astroglial cells or neurons. We present data indicating that arginine is released from cerebellar and cortical slices and astroglial cell cultures upon activation of ionotropic non-NMDA glutamate receptors. Glutamate had no effect on the extracellular [3H]arginine level in neuronal cultures. Moreover, the effect of glutamate in cerebellar slices was tetrodotoxin-insensitive, and the calcium ionophore A23187 evoked the release of [3H]arginine from astroglial cell cultures. Thus, nitric oxide synthesis and nitric oxide transmission may be based on the glial-neuronal transfer of arginine which is induced by activation of excitatory amino acid receptors on glial cells.

Nitric oxide precursor arginine and S-nitrosoglutathione in synaptic and glial function.

In the last few years, there has been an important increase in interest in nitric oxide (NO) as an intercellular messenger, and its putative role in numerous CNS functions is being continually updated. Arginine, the nitric oxide precursor, has been found in our laboratory to be released following stimulation of the white matter in the cerebellum and of sensory afferents in the thalamus. Since arginine is localized in glial cells while the nitric oxide synthesizing enzyme is localized in different cells (predominantly in neurons), these findings may represent a transfer of arginine from glia to neurons in order to supply the nitric oxide synthase with its substrate. The mechanism underlying this glial-neuronal interaction seems to involve the activation of excitatory amino acid receptors present on glial cells. Our results speak for an intense crosstalk between neurons and glia (activation of glial receptors by neurotransmitter released from neurons) and between glia and neurons (supply of the nitric precursor arginine from glia to neurons). The form in which NO is released from cells has been much debated. The chemical identity of the endothelial-derived relaxing factor in particular is still a matter of dispute, the major contender being NO. and a S-nitrosothiol compound. Based on the strong reactivity of NO for thiols and on the presence of cysteine and glutathione at the mM level intracellularly and microM level extracellularly, we have investigated whether S-nitrosothiols, i.e. S-nitrosoglutathione, may be the potential "package" form in which NO could be stored. We demonstrated, with HPLC coupled to mass spectrometry techniques, the presence of endogenous nitrosoglutathione in rat brain tissue. This packaging of NO in the form of nitrosothiols might serve to facilitate its transfer, prolong its life, and target its delivery to specific effectors. That could confer a specificity of action to the widely diffusable messenger NO, may determine the range of effectiveness of NO and mitigate its adverse cytotoxic effects.

DNA replication specificity of TYLCV geminivirus is mediated by the amino-terminal 116 amino acids of the Rep protein.

Geminiviruses are plant DNA viruses replicating by a rolling circle mechanism. We have investigated the specificity of replication origin recognition of two different isolates of tomato yellow leaf curl virus (TYLCV). Here, we show that TYLCV-Sardinian and -Israeli replication proteins display a high degree of specificity for their respective origins. The DNA sequences recognized are located on the left part of the intergenic region whereas the amino-terminal 116 amino acids of the Rep protein determine the specificity of origin recognition.

A mydriatic eye-drop combination without systemic effects for premature infants: a prospective double-blind study.

Eye drops used for diagnostic mydriasis may produce systemic side effects in preterm infants. Studies on the pupil dilating and systemic effect of various mydriatic agents yielded conflicting results. We conducted a prospective randomized double-blind study on the systemic effect of two mydriatic eye-drop combinations. Thirty-nine preterm infants were randomly assigned to two groups. An eye-drop combination of 2.5% phenylephrine and 0.5% tropicamide (group D) was compared with the combination of 0.5% cyclopentolate and 0.5% tropicamide (group F). Either eye-drop combination was followed by 0.5% tropicamide given 20 minutes later. Heart rate (HR) and the systolic, mean, and diastolic blood pressure (BP) were recorded before and after eye-drop instillation and after ophthalmoscopy. A control session with NaCl eye drops was added for each infant. A significant increase of BP and HR peak values was observed within 7 to 10 minutes after the cyclopentolate/tropicamide combination only. On the other hand, the mydriatic effect of the phenylephrine/tropicamide combination was significantly superior to that of the cyclopentolate/tropicamide combination. We recommend the combination of 2.5% phenylephrine and 0.5% tropicamide to achieve a sufficient diagnostic mydriasis without systemic side effects in preterm infants.

Effects of verapamil enantiomers and major metabolites on the cytotoxicity of vincristine and daunomycin in human lymphoma cell lines.

Verapamil, a calcium channel blocker, is used as the racemate. Recently, racemic verapamil has been shown to increase the cytotoxicity of vinca alkaloid and anthracycline derivatives in several resistant tumour cell lines. With respect to its cardiovascular activity S-verapamil is an order of magnitude more potent than R-verapamil. Since it was not known whether the effect on multi-drug resistance was also enantioselective a comparison has been made of the potency of the R and S enantiomers and racemic verapamil in their ability to increase the cytotoxicity of vincristine and daunomycin in sensitive (MOLT 4B) and drug resistant human T-lymphoma cell lines (MOLT/VCR-5 x 9, MOLT/DAU-8 and VCR 1000, a highly resistant subline of CCRF-CEM). Two major metabolites, norverapamil and D617 were tested in the same system. (+)-R, (-)-S-, racemic verapamil, norverapamil and D617 alone had no effect on cell growth in sensitive or resistant cell lines in concentrations up to 20 microM. In combination with vincristine, verapamil and norverapamil but not D617 produced a concentration dependent increase in the sensitivity of the resistant lines. Racemic verapamil, its individual enantiomers and norverapamil were equipotent. The concentration of the modifiers required to elicit 50% of the maximum effect (EC50) was of the order of 0.5 microM. No significant difference in the slopes of the concentration-effect curves were observed. The effect of verapamil and norverapamil was additive. In the sensitive MOLT 4B cell line both enantiomers and norverapamil increased sensitivity towards vincristine. However, the EC50 values were at least an order of magnitude higher (2.5-8 microM) than in the resistant cell lines.(ABSTRACT TRUNCATED AT 250 WORDS)