bluuebunny/crossref_metadata_2025 huggingface dataset:
Glucose transport and utilization are altered in the brain of rats deficient in n‐3 polyunsaturated fatty acids
Authors: Ximenes da Silva Adriana , Lavialle Françoise , Gendrot Ghislaine , Guesnet Philippe , Alessandri Jean‐Marc , Lavialle Monique
Published: March 2003

Long‐chain polyunsaturated (n‐3) fatty acids have been reported to influence the efficiency of membrane receptors, transporters and enzymes. Because the brain is particularly rich in docosahexaenoic acid (DHA, 22:6 n‐3), the present study addresses the question of whether the 22:6 n‐3 fatty acid deficiency induces disorder in regulation of energy metabolism in the CNS. Three brain regions that share a high rate of energy metabolism were studied: fronto‐parietal cortex, hippocampus and suprachiasmatic nucleus. The effect of the diet deficient in n‐3 fatty acids resulted in a 30–50% decrease in DHA in membrane phospholipids. Moreover, a 30% decrease in glucose uptake and a 20–40% decrease in cytochrome oxidase activity were observed in the three brain regions. The n‐3 deficient diet also altered the immunoreactivity of glucose transporters, namely GLUT1 in endothelial cells and GLUT3 in neurones. In n‐3 fatty acid deficient rats, GLUT1‐immunoreactivity readily detectable in microvessels became sparse, whereas the number of GLUT3 immunoreactive neurones was increased. However, western blot analysis showed no significant difference in GLUT1 and GLUT3 protein levels between rats deficient in n‐3 fatty acids and control rats. The present results suggest that changes in energy metabolism induced by n‐3 deficiency could result from functional alteration in glucose transporters.
Temporal patterns of cytokine and apoptosis‐related gene expression in spinal cords of the G93A‐SOD1 mouse model of amyotrophic lateral sclerosis
Authors: Hensley Kenneth , Floyd Robert A. , Gordon Brian , Mou Shenyun , Pye Quentin N. , Stewart Charles , West Melinda , Williamson Kelly
Published: March 2003

Familial amyotrophic lateral sclerosis (FALS) is often caused by gain‐of‐function mutations in Cu,Zn‐superoxide dismutase (SOD1). Multiprobe ribonuclease protection assays (RPAs) were used to investigate expression of 36 different cytokines and apoptosis‐related genes in spinal cords of mice that ubiquitously express human SOD1 bearing a glycine (r) alanine substitution at residue 93 (G93A‐SOD1). Mice were studied at late presymptomatic stage (80 days), and at 120 days when the animals experience severe hindlimb paralysis and accumulation of oxidatively modified proteins. Spinal cord tissue from G93A‐SOD1 mice expressed a selective subset of macrophage‐typical cytokines (monokines) including interleukin (IL)1α, IL1β and IL1RA at 80 days increasing by 120 days. Contrastingly, T‐cell derived cytokines (lymphokines) including IL2, IL3 and IL4 were detected at low levels in non‐transgenic mice but these were not elevated in G93A‐SOD1 mice even at 120 days. Apoptosis‐related genes were generally unaffected at 80 days but multiple caspases and death receptor components were up‐regulated at 120 days; the only exceptions being FADD and the tumor necrosis factor (TNF)α receptor p55 which was up‐regulated at 80 days and increased further at 120 days. These data indicate that in the G93A‐SOD1 mouse: (i) cytokine expression changes precede bulk protein oxidation and apoptosis gene expression; (ii) lymphocyte contributions to cytokine expression in FALS are likely minor; and (iii) TNFα and its receptors may link inflammation to apoptosis in ALS.
Oxidative damage increases with age in a canine model of human brain aging
Authors: Head E. , Liu J. , Hagen T. M. , Muggenburg B. A. , Milgram N. W. , Ames B. N. , Cotman C. W.
Published: March 2003

We assayed levels of lipid peroxidation, protein carbonyl formation, glutamine synthetase (GS) activity and both oxidized and reduced glutathione to study the link between oxidative damage, aging and β‐amyloid (Aβ) in the canine brain. The aged canine brain, a model of human brain aging, naturally develops extensive diffuse deposits of human‐type Aβ. Aβ was measured in immunostained prefrontal cortex from 19 beagle dogs (4–15 years). Increased malondialdehyde (MDA), which indicates increased lipid peroxidation, was observed in the prefrontal cortex and serum but not in cerebrospinal fluid (CSF). Oxidative damage to proteins (carbonyl formation) also increased in brain. An age‐dependent decline in GS activity, an enzyme vulnerable to oxidative damage, and in the level of glutathione (GSH) was observed in the prefrontal cortex. MDA level in serum correlated with MDA accumulation in the prefrontal cortex. Although 11/19 animals exhibited Aβ, the extent of deposition did not correlate with any of the oxidative damage measures, suggesting that each form of neuropathology accumulates in parallel with age. This evidence of widespread oxidative damage and Aβ deposition is further justification for using the canine model for studying human brain aging and neurodegenerative diseases.
α‐Synuclein exhibits competitive interaction between calmodulin and synthetic membranes
Authors: Lee Daekyun , Lee Sun‐Young , Lee Eui‐Nam , Chang Chung‐Soon , Paik Seung R.
Published: March 2003

α‐Synuclein, a pathological component of Parkinson's disease by constituting the Lewy bodies, has been suggested to be involved in membrane biogenesis via induction of amphipathic α‐helices. Since the amphipathic α‐helix is also known as a recognition signal of calmodulin for its target proteins, molecular interaction between α‐synuclein and calmodulin has been investigated. By employing a chemical coupling reagent ofN‐(ethoxycarbonyl)‐2‐ethoxy‐1,2‐dihydroquinoline, α‐synuclein has been shown to yield a heterodimeric 1 : 1 complex with calmodulin on sodium dodecyl sulfate–polyacrylamide gel electrophoresis in the presence and even absence of calcium, whereas β‐synuclein was more dependent upon calcium for its calmodulin interaction. The selective calmodulin interaction of α‐synuclein in the absence of calcium was also demonstrated with the aggregation kinetics of the synucleins in which only the α‐synuclein aggregation was affected by calmodulin. A reversible binding assay confirmed that α‐synuclein interacted with the Ca2+‐free as well as the Ca2+‐bound calmodulins with almost identicalKds of 0.35 µmand 0.31 µm, respectively, while β‐synuclein preferentially recognized the Ca2+‐bound form with aKdof 0.68 µm. By using a C‐terminally truncated α‐synuclein of α‐syn97, the calmodulin binding site(s) on α‐synuclein was(were) shown to be located on the N‐terminal region where the amphipathic α‐helices have been suggested to be induced upon membrane interaction. By employing liposome and calmodulin in a state of being either soluble or immobilized on agarose, actual competition of α‐synuclein between membranes and calmodulin was demonstrated with the observation that α‐synuclein previously bound to the liposome was released upon specific interaction with the calmodulins. Taken together, these data may suggest that α‐synuclein could act not only as a negative regulator for calmodulin in the presence and even absence of calcium, but it could also exert its activity at the interface between calmodulin and membranes.
Differential effects of hypoxia–ischemia on subunit expression and tyrosine phosphorylation of the NMDA receptor in 7‐ and 21‐day‐old rats
Authors: Gurd James W. , Bissoon N. , Beesley P. W. , Nakazawa Takanobu , Yamamoto Tadashi , Vannucci S. J.
Published: March 2003

The effect of cerebral hypoxia–ischemia (HI) on levels and tyrosine phosphorylation of the NMDA receptor was examined in 7‐ (P7) and 21 (P21)‐day‐old rats. Unilateral HI was administered by ligation of the right common carotid artery and exposure to an atmosphere of 8% O2/92% N2for 2 (P7) or 1.5 (P21) h. This duration of HI produces significant infarction in nearly all of the survivors with damage being largely restricted to the cortex, striatum, and hippocampus of the hemisphere ipsilateral to the carotid artery ligation. NR2A levels in the right hemisphere of P7 pups were markedly reduced after 24 h of recovery, while NR1 and NR2B remained unchanged. In contrast, NR2B, but not NR2A, was reduced after HI at P21. At both ages, HI resulted in a transient increase in tyrosine phosphorylation of a number of forebrain proteins that peaked between 1 and 6 h of recovery. At both P7 and P21, tyrosine phosphorylation of NR2B was enhanced 1 h after HI and had returned to basal levels by 24 h. HI induced an increase in tyrosine phosphorylation of NR2A in 21 day, but not in 7‐day‐old animals. The differential effects of HI on the NMDA receptor at different post‐natal ages may contribute to changing sensitivity to hypoxia–ischemia.
A new brain‐derived neurotrophic factor transcript and decrease inbrain‐derived neurotrophic factor transcripts 1, 2 and 3 in Alzheimer's disease parietal cortex
Authors: Garzon Diego , Yu Guanhua , Fahnestock Margaret
Published: March 2003

Brain‐derived neurotrophic factor (BDNF) supports hippocampal, cortical and basal forebrain cholinergic neurons, which lose function in Alzheimer's disease. In Alzheimer's tissues such as hippocampus and parietal cortex, brain‐ derived neurotrophic factor mRNA is decreased three‐ to four‐fold compared with controls. However, the molecular mechanism of the down‐regulation of BDNF in Alzheimer's disease is unknown. The human brain‐derived neurotrophic factor gene has multiple promoters governing six non‐coding upstream exons that are spliced to one downstream coding exon, leading to six different transcripts. Here we report an alternate human splice variant within exon 4I for a total of seven transcripts. Previous brain‐derived neurotrophic factor mRNA measurements in Alzheimer's disease tissue were done using the downstream coding exon present in all transcripts. Using RT‐PCR primers specific for each upstream exon, we observe a significant decrease in three human brain‐derived neurotrophic factor mRNA transcripts in Alzheimer's disease samples compared with controls. Transcripts 1 and 3 each exhibit a two‐fold decrease, and transcript 2 shows a five‐fold decrease. There are no significant differences between control and Alzheimer's disease samples for the other transcripts, including the new splice variant. In rat, both transcripts 1 and 3 are regulated through the transcription factor cAMP response element binding protein, whose phosphorylation is decreased in the Alzheimer's disease brain. This could lead to specific down‐regulation of the brain‐derivedneurotrophic factor transcripts shown here.
Restricted, but abundant, expression of the novel rat gene‐3 (R3) relaxin in the dorsal tegmental region of brain
Authors: Burazin Tanya C. D. , Bathgate Ross A. D. , Macris Mary , Layfield Sharon , Gundlach Andrew L. , Tregear Geoffrey W.
Published: March 2003

Relaxin is a peptide hormone with known actions associated with female reproductive physiology, but it has also been identified in the brain. Only one relaxin gene had been characterized in rodents until recently when a novel human relaxin gene, human gene‐3 (H3) and its mouse equivalent (M3) were identified. The current study reports the identification of a rat homologue, rat gene‐3 (R3) relaxin that is highly expressed in a discrete region of the adult brain. The full R3 relaxin cDNA was generated using RT‐PCR and 3′ and 5′ RACE protocols. The derived amino acid sequence of R3 relaxin retains all the characteristic features of a relaxin peptide and has a high degree of homology with H3 and M3 relaxin. The distribution of R3 relaxin mRNA in adult rat brain was determined and highly abundant expression was only detected in neurons of the ventromedial dorsal tegmental nucleus (vmDTg) in the pons, whereas all other brain areas were unlabelled or contained much lower mRNA levels. Relaxin binding sites and relaxin immunoreactivity were also detected in the vmDTg. These together with earlier findings provide strong evidence for a role(s) for multiple relaxin peptides as neurotransmitters and/or modulators in the rat CNS.
Selective expression of Narp, a secreted neuronal pentraxin, in orexin neurons
Authors: Reti Irving M. , Reddy Radhika , Worley Paul F. , Baraban Jay M.
Published: March 2003

Recent studies have provided compelling evidence demonstrating that orexin (also known as hypocretin) neurons play a central role in the pathophysiology of narcolepsy. However, targeted deletion of orexin does not fully mimic the functional deficits induced by selective ablation of these neurons; implying that other secreted signaling molecules expressed in these neurons mediate key aspects of their function. In this study, we demonstrate that orexin neurons display robust expression of neuronal activity‐regulated pentraxin (Narp), a secreted neuronal pentraxin, implicated in regulating clustering of α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate (AMPA) receptors. Furthermore, we have found that hypothalamic melanin‐concentrating hormone (MCH) neurons, which form a peptidergic pathway thought to oppose the effects of the orexin system, express another neuronal pentraxin, NP1. Thus, these findings suggest that these pathways utilize neuronal pentraxins, in addition to neuropeptides, as synaptic signaling molecules.
Regulation of hydrogen peroxide production by brain mitochondria by calcium and Bax
Authors: Starkov Anatoly A. , Polster Brian M. , Fiskum Gary
Published: March 2003

Abnormal accumulation of Ca2+ and exposure to pro‐apoptotic proteins, such as Bax, is believed to stimulate mitochondrial generation of reactive oxygen species (ROS) and contribute to neural cell death during acute ischemic and traumatic brain injury, and in neurodegenerative diseases, e.g. Parkinson's disease. However, the mechanism by which Ca2+ or apoptotic proteins stimulate mitochondrial ROS production is unclear. We used a sensitive fluorescent probe to compare the effects of Ca2+ on H2O2 emission by isolated rat brain mitochondria in the presence of physiological concentrations of ATP and Mg2+ and different respiratory substrates. In the absence of respiratory chain inhibitors, Ca2+ suppressed H2O2 generation and reduced the membrane potential of mitochondria oxidizing succinate, or glutamate plus malate. In the presence of the respiratory chain Complex I inhibitor rotenone, accumulation of Ca2+ stimulated H2O2 production by mitochondria oxidizing succinate, and this stimulation was associated with release of mitochondrial cytochrome c. In the presence of glutamate plus malate, or succinate, cytochrome c release and H2O2 formation were stimulated by human recombinant full‐length Bax in the presence of a BH3 cell death domain peptide. These results indicate that in the presence of ATP and Mg2+, Ca2+ accumulation either inhibits or stimulates mitochondrial H2O2 production, depending on the respiratory substrate and the effect of Ca2+ on the mitochondrial membrane potential. Bax plus a BH3 domain peptide stimulate H2O2 production by brain mitochondria due to release of cytochrome c and this stimulation is insensitive to changes in membrane potential.
Ethanol impairs monosaccharide uptake and glycosylation in cultured rat astrocytes
Authors: Tomás M. , Fornas E. , Megías L. , Durán J.‐M. , Portolés M. , Guerri C. , Egea G. , Renau‐Piqueras J.
Published: March 2003

Astrocyte and glial–neuron interactions have a critical role in brain development, which is partially mediated by glycoproteins, including adhesion molecules and growth factors. Ethanol affects the synthesis, intracellular transport, subcellular distribution and secretion of these glycoproteins, suggesting alterations in glycosylation. We analyzed the effect of long‐term exposure to low doses of ethanol (30 mm) on glycosylation process in growing cultured astrocytes in vitro. Cells were incubated for short (5 min) and long (90 min) periods with several radioactively labeled carbohydrate precursors. The uptake, kinetics and metabolism of these precursors, as well as the radioactivity distribution in protein gels were analyzed. The levels of GLUT1 and mannosidase II were also determined. Ethanol increased the uptake of monosaccharides and the protein levels of GLUT1 but decreased those of mannosidase II. It altered the carbohydrate moiety of proteins and increased cell surface glycoproteins containing terminal non‐reduced mannose. These results indicate that ethanol impairs glycosylation in rat astrocytes, thus disrupting brain development.
Interaction between GABAA receptor subunit intracellular loops: implications for higher order complex formation
Authors: Nymann‐Andersen Jesper , Wang Hongbing , Sawyer Gregory W. , Olsen Richard W.
Published: March 2003

The majority of fast inhibitory neurotransmission in the CNS is mediated by the GABA type‐A (GABAA) receptor, a ligand‐gated chloride channel. Of the approximately 20 different subunits composing the hetero‐pentameric GABAA receptor, the γ2 subunit in particular seems to be important in several aspects of GABAA receptor function, including clustering of the receptor at synapses. In this study, we report that the intracellular loop of the γ2 subunit interacts with itself as well as with γ1, γ3 and β1–3 subunits, but not with the α subunits. We further show that γ2 subunits interact with photolabeled pentameric GABAA receptors composed of α1, β2/3 and γ2 subunits, and calculate the dissociation constant to be in the micromolar range. By using deletion constructs of the γ2 subunit in a yeast two‐hybrid assay, we identified a 23‐amino acid motif that mediates self‐association, residues 389–411. We confirmed this interaction motif by inhibiting the interaction in a glutathione‐S‐transferase pull‐down assay by adding a corresponding γ2‐derived peptide. Using similar approaches, we identified the interaction motif in the γ2 subunit mediating interaction with the β2 subunit as a 47‐amino acid motif that includes the γ2 self‐interacting motif. The identified γ2 self‐association motif is identical to the interaction motif reported between GABAA receptor and GABAA receptor‐associated protein (GABARAP). We propose a model for GABAA receptor clustering based on GABARAP and GABAA receptor subunit–subunit interaction.
Identification of a peptide mimic of the L2/HNK‐1 carbohydrate epitope
Authors: Simon‐Haldi Maryline , Mantei Ned , Franke Jens , Voshol Hans , Schachner Melitta
Published: March 2003

The L2/HNK‐1 carbohydrate is carried by many neural recognition molecules and is involved in neural cell interactions during development, regeneration in the peripheral nervous system, synaptic plasticity, and autoimmune‐based neuropathies. Its key structure consists of a sulfated glucuronic acid linked to lactosaminyl residues. Because of its biological importance but limited availability, the phage display method was used to isolate a collection of peptide mimics that bind specifically to an L2/HNK‐1 antibody. The phages isolated from a 15‐mer peptide library by adsorption to this antibody share a consensus sequence of amino acids. The peptide mimicked several important functions of the L2/HNK‐1 carbohydrate, such as binding to motor neurons in vitro, and preferential promotion of in vitro neurite outgrowth from motor axons compared with sensory neurons. A scrambled version of the peptide had no activity. The combined observations indicate that we have isolated a mimic of the L2/HNK‐1 carbohydrate that is able to act as its functional substitute.
Agonist‐induced internalization of metabotropic glutamate receptor 1A: structural determinants for protein kinase C‐ and G protein‐coupled receptor kinase‐mediated internalization
Authors: Mundell Stuart J. , Pula Giordano , Carswell Katy , Roberts Peter J. , Kelly Eamonn
Published: March 2003

To investigate the role of the intracellular C‐terminal tail of the rat metabotropic glutamate receptor 1a (mGlu1a) in receptor regulation, we constructed three C‐terminal tail deletion mutants (Arg847stop, DM‐I; Arg868stop, DM‐II; Val893stop, DM‐III). Quantification of glutamate‐induced internalization provided by ELISA indicated that DM‐III, like the wild‐type mGlu1a, underwent rapid internalization whilst internalization of DM‐I and DM‐II was impaired. The selective inhibitor of protein kinase C (PKC), GF109203X, which significantly reduced glutamate‐induced mGlu1a internalization, had no effect on the internalization of DM‐I, DM‐II, or DM‐III. In addition activation by carbachol of endogenously expressed M1 muscarinic acetylcholine receptors, which induces PKC‐ and Ca2+‐calmodulin‐dependent protein kinase II‐dependent internalization of mGlu1a, produced negligible internalization of the deletion mutants. Co‐expression of a dominant negative mutant form of G protein‐coupled receptor kinase 2 (DNM‐GRK2; Lys220Arg) significantly attenuated glutamate‐induced internalization of mGlu1a and DM‐III, whilst internalization of DM‐I and DM‐II was not significantly affected. The glutamate‐induced internalization of mGlu1a and DM‐III, but not of DM‐I or DM‐II, was inhibited by expression of DNM‐arrestin [arrestin‐2(319–418)]. In addition glutamate‐induced rapid translocation of arrestin‐2‐Green Fluorescent Protein (arr‐2‐GFP) from cytosol to membrane was only observed in cells expressing mGlu1a or DM‐III. Functionally, in cells expressing mGlu1a, glutamate‐stimulated inositol phosphate accumulation was increased in the presence of PKC inhibition, but so too was that in cells expressing DM‐II and DM‐III. Together these results indicate that different PKC mechanisms regulate the desensitization and internalization of mGlu1a. Furthermore, PKC regulation of mGlu1a internalization requires the distal C terminus of the receptor (Ser894–Leu1199), whilst in contrast glutamate‐stimulated GRK‐ and arrestin‐dependent regulation of this receptor depends on a region of 25 amino acids (Ser869–Val893) in the proximal C‐terminal tail.
Specific interactions of neuronal focal adhesion kinase isoforms with Src kinases and amphiphysin
Authors: Messina Samantha , Onofri Franco , Bongiorno‐Borbone Lucilla , Giovedì Silvia , Valtorta Flavia , Girault Jean‐Antoine , Benfenati Fabio
Published: March 2003

Focal adhesion kinase (FAK) is a non‐receptor tyrosine kinase that activates Src family kinases via SH2‐ and SH3‐mediated interactions. Specific FAK isoforms (FAK+), responsive to depolarization and neurotransmitters, are enriched in neurons. We analyzed the interactions of endogenous FAK+ and recombinant FAK+ isoforms containing amino acid insertions (boxes 6,7,28) with an array of SH3 domains and the c‐Src SH2/SH3 domain tandem. Endogenous FAK+ bound specifically to the SH3 domains of c‐Src (but not n‐Src), Fyn, Yes, phosphtidylinositol‐3 kinase, amphiphysin II, amphiphysin I, phospholipase Cγ and NH2‐terminal Grb2. The inclusion of boxes 6,7 was associated with a significant decrease in the binding of FAK+ to the c‐Src and Fyn SH3 domains, and a significant increase in the binding to the Src SH2 domain, as a consequence of the higher phosphorylation of Tyr‐397. The novel interaction with the amphiphysin SH3 domain, involving the COOH‐terminal proline‐rich region of FAK, was confirmed by coimmunoprecipitation of the two proteins and a closely similar response to stimuli affecting the actin cytoskeleton. Moreover, an impairment of endocytosis was observed in synaptosomes after internalization of a proline‐rich peptide corresponding to the site of interaction. The data account for the different subcellular distribution of FAK and Src kinases and the specific regulation of the transduction pathways linked to FAK activation in the brain and implicate FAK in the regulation of membrane trafficking in nerve terminals.
Stabilization of the cyclin‐dependent kinase 5 activator, p35, by paclitaxel decreases β‐amyloid toxicity in cortical neurons
Authors: Li Guibin , Faibushevich Alexander , Turunen Brandon J. , Yoon Sung Ok , Georg Gunda , Michaelis Mary L. , Dobrowsky Rick T.
Published: March 2003

One hallmark of Alzheimer's disease (AD) is the formation of neurofibrillary tangles, aggregated paired helical filaments composed of hyperphosphorylated tau. Amyloid‐β (Aβ) induces tau hyperphosphorylation, decreases microtubule (MT) stability and induces neuronal death. MT stabilizing agents have been proposed as potential therapeutics that may minimize Aβ toxicity and here we report that paclitaxel (taxol) prevents cell death induced by Aβ peptides, inhibits Aβ‐induced activation of cyclin‐dependent kinase 5 (cdk5) and decreases tau hyperphosphorylation. Taxol did not inhibit cdk5 directly but significantly blocked Aβ‐induced calpain activation and decreased formation of the cdk5 activator, p25, from p35. Taxol specifically inhibited the Aβ‐induced activation of the cytosolic cdk5‐p25 complex, but not the membrane‐associated cdk5‐p35 complex. MT‐stabilization was necessary for neuroprotection and inhibition of cdk5 but was not sufficient to prevent cell death induced by overexpression of p25. As taxol is not permeable to the blood–brain barrier, we assessed the potential of taxanes to attenuate Aβ toxicity in adult animals using a succinylated taxol analog (TX67) permeable to the blood–brain barrier. TX67, but not taxol, attenuated the magnitude of both basal and Aβ‐induced cdk5 activation in acutely dissociated cortical cultures prepared from drug treated adult mice. These results suggest that MT‐stabilizing agents may provide a therapeutic approach to decrease Aβ toxicity and neurofibrillary pathology in AD and other tauopathies.
Effect of S 17092, a novel prolyl endopeptidase inhibitor, on substance P and α‐melanocyte‐stimulating hormone breakdown in the rat brain
Authors: Bellemère Gaëlle , Morain Philippe , Vaudry Hubert , Jégou Sylvie
Published: March 2003

In the present study, we have investigated the effects of a novel prolyl endopeptidase (EC 3.4.21.26, PEP) inhibitor, compound S 17092, on substance P (SP) and α‐melanocyte‐stimulating hormone (α‐MSH) metabolism in the rat brain. In vitro experiments revealed that S 17092 inhibits in a dose‐dependent manner PEP activity in rat cortical extracts (IC50 = 8.3 nm). In addition, S 17092 totally abolished the degradation of SP and α‐MSH induced by bacterial PEP. In vivo, a significant decrease in PEP activity was observed in the medulla oblongata after a single oral administration of S 17092 at doses of 10 and 30 mg/kg (−78% and −82%, respectively) and after chronic oral treatment with S 17092 at doses of 10 and 30 mg/kg per day (−75% and −88%, respectively). Concurrently, a single administration of S 17092 (30 mg/kg) caused a significant increase in SP‐ and α‐MSH‐like immunoreactivity (LI) in the frontal cortex (+41% and +122%, respectively) and hypothalamus (+84% and +49%, respectively). In contrast, chronic treatment with S 17092 did not significantly modify SP‐ and α‐MSH‐LI in the frontal cortex and hypothalamus. Collectively, the present results show that S 17092 elevates SP and α‐MSH concentrations in the rat brain by inhibiting PEP activity. These data suggest that the effect of S 17092 on memory impairment can be accounted for, at least in part, by inhibition of catabolism of promnesic neuropeptides such as SP and α‐MSH.
Protein kinase C‐ε is involved in the adenosine‐activated signal transduction pathway conferring protection against ischemia‐reperfusion injury in primary rat neuronal cultures
Authors: Di‐Capua Noam , Sperling Oded , Zoref‐Shani Esther
Published: March 2003

Adenosine activates a signal transduction pathway (STP) in the heart and the brain, conferring protection against ischemia‐reperfusion insult. Activation of protein kinase C (PKC), probably mainly PKC‐ε, has been demonstrated to be part of the heart STP, but its role in the neuronal pathway is less clear. Here, we provide proof for the participation of PKC‐ε in the neuronal adenosine‐activated STP. Primary rat neuronal cultures were exposed to chemical ischemia by iodoacetate, followed by reperfusion. The cultured neurons were protected against this insult by activation of the adenosine mechanism, by N6‐(R)‐phenylisopropyladenosine [R(–)‐PIA], a specific A1 adenosine receptor agonist. Exposure of the cultures to bisindolylmaleimide I, a highly selective PKC inhibitor, abrogated the protection. The exposure of the cultures to R(–)‐PIA was found to result in phosphorylation (activation) of PKC‐ε. Furthermore, insertion into the cells of a specific peptide inhibitor of PKC‐ε translocation (εV1‐2), also abrogated the protection conferred by R(–)‐PIA. These results demonstrate that activation of PKC‐ε is a vital step in the neuronal adenosine‐activated STP.
Peripheral benzodiazepine receptors: on mice and human brain imaging
Authors: Weissman Ben Avi , Raveh Lily
Published: March 2003

There are numerous methods designed to monitor brain neuropathologies resulting from a wide arsenal of insults. Regardless of the cause of neuronal death, reactive glial cells always appear at and around the site of degeneration. These cells are distinguished by the exceptional abundance of peripheral benzodiazepine receptors, particularly compared with surrounding neurons. Measuring the binding of specific ligands to these peripheral benzodiazepine receptors offers a unique indirect marker for reliable damage assessment in the CNS and a faithful indicator for the accompanying cognitive deficits.
In vivo nicotine treatment regulates mesocorticolimbic CREB and ERK signaling in C57Bl/6J mice
Authors: Brunzell Darlene H. , Russell David S. , Picciotto Marina R.
Published: March 2003

The extracellular regulated kinase (ERK) pathway was studied to determine its role in neuronal plasticity related to the development of nicotine dependence. Levels and phosphorylation state of ERK, cAMP response element binding protein (CREB) and proline‐rich/Ca2+‐activated tyrosine kinase (PYK2), and levels of tyrosine hydroxylase (TH), were determined using western blotting. C57Bl/6J mice received acute or chronic nicotine (200 µg/mL) in their drinking water or were withdrawn from nicotine for 24 h following chronic exposure. CREB phosphorylation was reduced in the nucleus accumbens following chronic nicotine, consistent with previous reports that decreased accumbens CREB activity increases drug reinforcement. In contrast, CREB phosphorylation was increased in the prefrontal cortex following chronic nicotine exposure and in the ventral tegmental area during nicotine withdrawal. In addition, total and phosphorylated ERK decreased in the amygdala following chronic nicotine exposure, but ERK phosphorylation increased in the prefrontal cortex. TH levels increased in both the amygdala and prefrontal cortex, supporting the hypothesis that increased catecholaminergic tone contributes to nicotine reinforcement. Overall, these results support a role for ERK and CREB activity in neural plasticity associated with nicotine dependence.
Relationship between a global rating of speech ability at the age of 3 yrs and a phonological screening 1 yr later
Authors: Westerlund Monica
Published: March 2003

Relationship between a global rating of speech ability at the age of 3 yrs and a phonological screening 1 yr later

Objective: This study, which was carried out under routine conditions, evaluates the predictability of a global rating of children’s speaking manner made by Child Health Centre nurses. The global rating was added to a formalized screening of 3‐yr olds, which intended to identify severely language disabled children. This screening procedure is used in several Swedish counties and the participation rate is nearly 100%. The main questions of the study are: (i) can a global rating of speech ability in children, who have passed the 3‐yr screening, predict the outcome of a new population screening 1 yr later? and if so (ii) what is the most efficient cut‐off point? Methods: The global ratings of the 3‐yr olds were made using a 100 mm visual analogue scale (VAS). At the age of 4 yrs, 2065 globally rated children with no objections from the 3‐yr screening participated in a second, well‐established assessment. The purpose of this second screening was to identify not only severe but also moderate disability. Results: The mean VAS value of the global ratings was 62 mm. One‐third of the ratings fell on the lower half of the scale. With a cut‐off level set at ≤50 mm on the VAS two newly‐identified, severely disabled children and 74% of the moderately disabled 4‐yr olds were identified. Conclusion: The results suggest that the global rating in connection to the 3‐yr screening would serve as a useful tool for selective screening of 4‐yr olds, and thus, reduce the work load.
Managing pain in older persons who receive home‐help for their daily living. Perceptions by older persons and care providers
Authors: Blomqvist Kerstin , Hallberg Ingalill R.
Published: March 2003

Managing pain in older persons who receive home‐help for their daily living. Perceptions by older persons and care providers

Pain is a common problem for older persons who need professional help for their daily living. In this study 94 older persons (75+) in persistent pain were compared with 52 care providers concerning the pain management methods they had used/administered during the previous week and how helpful they perceived these methods to be. Interviews were based on 16 items from the original version of the pain management inventory (PMI). Both groups perceived prescribed medication, rest and distraction as the most frequently utilized methods. Specific methods such as transcutaneous electrical nerve stimulation (TENS) or bracing/splinting or bandaging the affected body part were seldom employed, although most users perceived these methods as helpful. Care providers perceived most methods for managing pain as more effective than older persons did. The results imply that care providers need skills in a variety of pharmacological and nonpharmacological methods to manage pain and a need to evaluate effectiveness of the methods in a systematic way.
Microsatellite markers from a commercially important South‐east Asian cyprinid, the silver barb (Barbodes gonionotus Bleeker)
Authors: McConnell S. K. J. , Skibinski D. O. F. , Mair G. C.
Published: March 2003

Barbodes gonionotus (Puntius gonionotus/javanicus, Bleeker) is a cyprinid fish, endemic to South‐east Asia and is of great importance in both wild fisheries and in aquaculture in the region. It is a common to dominant species in the middle and lower stretches of medium and large rivers. Five highly polymorphic microsatellite loci are described (HO = 0.7–0.9, number of alleles per locus = 10–37), which will be of use both in population genetic studies and for the assessment of true effective breeding population sizes in aquaculture stocks.
Development of species‐specific markers in an organism with endosymbionts: microsatellites in the scleractinian coral Seriatopora hystrix
Authors: Maier Elke , Tollrian Ralph , Nürnberger Beate
Published: March 2003

We report on the development of microsatellites in Seriatopora hystrix, a coral with algal endosymbionts. In order to obtain a genomic library free of algal DNA, we conducted a whole genome preamplification from minute amounts of symbiont‐free tissue. The resulting fragments were cloned into pUC18, and Escherichia coli were transformed with the recombinant plasmids. Twenty‐nine microsatellites were isolated from a library screen with a fluorescently labelled (CA)15 probe. Five of these yielded reliable polymorphic markers.
Development of polymorphic microsatellite markers in a perennial herbaceous plant, Polygonum cuspidatum (Polygonaceae)
Authors: Miwa, M. , Zhou Z. , Shinone M. , Maruta E. , Hogetsu T.
Published: March 2003

Six single‐locus, polymorphic microsatellite markers in a perennial herbaceous plant, Polygonum cuspidatum, were developed. Tests to amplify these six loci in another Polygonum species failed, suggesting that the six markers are specific to P. cuspidatum.
Development of microsatellite loci for population studies of the pink ling, Genypterus blacodes (Teleostei: Ophidiidae)
Authors: Ward R. D. , Reilly A.
Published: March 2003

Fifteen microsatellite loci were developed for pink ling (Genypterus blacodes), a fish of significant commercial importance to Australasia. Nine loci were examined in samples from five regions of Australia’s South East Fishery. All nine were highly polymorphic; numbers of alleles per locus ranged from 11 to 52 in total samples of 270–306 individuals. The average observed heterozygosity per locus per sample (0.823) was a little lower than the average Hardy–Weinberg expected heterozygosity per sample (0.895), perhaps reflecting the possible presence of null alleles at two loci. There was no significant evidence of genetic stock heterogeneity.
Microsatellite loci in the soil‐dwelling collembolan, Orchesella cincta
Authors: Van Der Wurff A. W. G. , Gols R. , Ernsting G. , Van Straalen N. M.
Published: March 2003

Three total genomic libraries of the springtail Orchesella cincta (Insecta; Collembola) were screened for the presence of microsatellites using phosphor and colour detection. PCR primers were successfully constructed for six dinucleotide, one trinucleotide, and one interrupted dinucleotide microsatellite repeat. None of the microsatellite arrays were longer than 11 repeat units. Individuals from three forests were investigated: five out of eight microsatellite markers were polymophic in all forests (2–5 alleles per locus) and the observed heterozygosity was 0.1–0.9 per locus.
Polymorphic microsatellite repeats are not conserved between Leishmania donovani and Leishmania major
Authors: Jamjoom M. B. , Ashford R. W. , Bates P. A. , Kemp S. J. , Noyes H.A.
Published: March 2003

Thirteen sets of polymerase chain reaction (PCR) primers were designed to amplify microsatellite loci identified in the genome sequence of Leishmania major. Polymorphisms were detected in L. major at all loci. In Leishmania donovani only two of these loci were informative for classification purposes with this data set. The PCR products of all loci from one L. donovani strain were sequenced and it was found that the number of repeats in the microsatellite loci were either substantially reduced with respect to L. major or absent altogether. Consequently it is unlikely to be possible to use the genome sequence of L. major to identify polymorphic microsatellite loci in other Leishmania species.
Isolation and characterization of microsatellites in a perennial Apiaceae, Eryngium alpinum L.
Authors: Gaudeul M. , Naciri‐Graven Y. , Gauthier P. , Pompanon F.
Published: March 2003

In order to investigate the genetic structure in an endangered Alpine plant (Eryngium alpinum L.), we developed microsatellites. Two different approaches were used: an enrichment protocol and the classical technique of hybridization on nylon membranes. We identified 25 loci, 13 of which revealed to be polymorphic. The polymorphism was rather low (2–6 alleles; HE = 0.49 ± 0.16), probably due to the short size of microsatellites (6–10 dinucleotide repeats) and to the fine spatial scale investigated. However, these markers are expected to provide a new insight about the genetic processes at work within and among E. alpinum populations.
Polymerase chain reaction primers for polymorphic microsatellite loci from the túngara frog Physalaemus pustulosus
Authors: Pröhl H. , Adams R. M. M. , Mueller U. , Rand S. , Ryan M. J.
Published: March 2003

We developed eight PCR−primer pairs of polymorphic microsatellite loci for the túngara frog Physalaemus pustulosus. Genomic libraries were enriched for one of four microsatellite repeat sequences (CAn, GAn, ATGn and TAGAn). Following characterization of microsatellite loci by sequencing, primers were designed and PCR conditions optimized. Microsatellite PCR‐amplification was tested in 37 frogs from 8 populations in Costa Rica and Panama. Primer sequences, PCR conditions, allelelic diversities and observed as well as expected heterozygosities in the screened populations are described.
Novel universal primers establish identity of an enormous number of animal species for forensic application
Authors: Verma S. K. , Singh L.
Published: March 2003

This study describes a polymerase chain reaction (PCR)‐based approach, which without knowing the history of a forensic sample, is able to reveal whether the source of the sample is human or animal, and, if animal, which of the 221 animal species included in the study, simply by using one set of novel primers to amplify and sequence the PCR amplicons. The primers described in this study universally amplify a specific segment of mitochondrial cytochrome b sequence from a sample of unknown origin and delineate its identity to the level of family, genus and species. Because these primers are universal, this approach can be applied to an enormous number of other species, which are not included in the study, and could be an ultimate solution for the identification of species for forensic application.
Characterization of 12 polymorphic microsatellite loci in the Japanese bush warbler Cettia diphone
Authors: Otsuka R. , Nishiumi I. , Wada M.
Published: March 2003

Japanese bush warblers, Cettia diphone, are a common species in Japan and have a polygynous breeding system. In the breeding ground some males have their own territories with more than one female. Male floaters are also not uncommon in the breeding ground. To understand breeding strategy in this species, exact parentage should be elucidated. In order to obtain a tool for this purpose, we isolated 34 microsatellite loci from a genomic library in this species and developed primers for 12 loci. These primers were tested in the Japanese bush warbler and successfully amplified. In analyses of 49 unrelated individuals, allelic numbers ranged from two to 22, and observed heterozygosity (HO) ranged from 0.27 to 0.854 except for two loci (Cdi29 and Cdi35a) with HO < 0.1.
Isolation and characterization of polymorphic microsatellite loci in nurse shark (Ginglymostoma cirratum)
Authors: Heist Edward J. , Jenkot Jennifer L. , Keeney Devon B. , Lane Ryan L. , Moyer Gregory R. , Reading Benjamin J. , Smith Noelle L.
Published: March 2003

We report on the development of nine polymorphic microsatellite loci in nurse shark (Ginglymostoma cirratum) using a combination of enriched and unenriched subgenomic libraries. Based on the small percentage of positive clones in the unenriched library (0.4%) it appears that microsatellites are very scarce in nurse shark genomes. Numbers of alleles at polymorphic loci ranged from two to 15; observed heterozygosity ranged from 0.17 to 0.90. We expect these loci to be useful for studies of breeding structure and paternity.
Microsatellites in the freshwater snail Bulinus globosus (Gastropoda: Planorbidae) from Zanzibar
Authors: Emery Aidan M. , Loxton Nicola J. , Stothard J. Russell , Jones Catherine S. , Spinks Jenny , Llewellyn‐Hughes Julia , Noble Leslie R. , Rollinson David
Published: March 2003

Six microsatellite loci were isolated and characterized in Bulinus globosus, a freshwater snail with a wide distribution throughout sub‐Saharan Africa. Bulinus globosus is an intermediate host of Schistosoma haematobium, the causative agent of human urinary schistosomiasis. Microsatellites were tested using 32 snails from four populations collected from Pemba and Unguja islands of Zanzibar. The microsatellite loci displayed relatively low levels of variation, with between two and five alleles per locus. FST estimates indicate that gene flow is low, as has previously been suggested for other species of Bulinus.
The physical basis of cosmetic defects of the nail plate
Authors: De Berker David
Published: March 2003

Summary Cosmetic defects of the nail cover a range of changes. Some are variants of normal which are considered unattractive, others are part of the normal ageing process and some changes are manifestations of local or systemic disease. Interpretation of these changes relies on recognition of the specific characteristics in question and an understanding of the anatomy and biology of nails. In many instances, therapies are limited and explanation of the changes represents one of the most useful contributions the professional can make to the affected individual. This article covers some of the most common cosmetic defects of nail involving colour, surface, brittleness and behaviour of surrounding tissues. Understanding of the physical basis of these defects is limited in many cases, but current thinking is recorded.
Selection through predation, snowfall and microclimate on nest‐site preferences in the Water Pipit Anthus spinoletta
Authors: Rauter Claudia M. , Reyer Heinz‐Ulrich , Bollmann Kurt
Published: March 2003

Nest‐site characteristics can have a strong impact on reproductive success in birds. Nest sites should simultaneously protect from predators, offer shelter and provide a favourable microclimate. We studied the relationship between three agents of natural selection (predators [i.e. Adders and birds/mammals], snowfall and microclimate), nest‐site characteristics and reproductive success to determine whether these influenced preference for specific nest‐site characteristics in the Water Pipit Anthus spinoletta. Pooled over all nests, the relative importance as agents of natural selection decreased from mammalian/avian predation (15% of all nests) through Adder predation (12%) to snowfall (7%), but there were clear differences in space and time. Predation by Adders selected for nest sites surrounded by few medium‐sized shrubs. Selection by mammalian and avian predators favoured no specific nest‐site characteristics. Protection from snowfall was best in nests surrounded by relatively few medium‐sized shrubs. Microclimate had a strong influence on nestling survival and duration of nestling period. In nests on ENE‐facing slopes, where maximum temperatures were reached in the morning, nestling survival was higher than on WSW‐facing slopes, where temperature maxima occurred in the afternoon. Our results indicate that weak, but significant, directional selection is acting on preference for certain nest‐site characteristics through effects on survival and development of nestlings. As predation and snowfall are unpredictable, the evolution of an optimal nest placement strategy is unlikely on a small scale. On a larger scale, however, choice of one breeding area over another may be favoured because of predictable differences between locations in terms of survival and nestling development.
Factors affecting divorce in the Great TitParus major
Authors: Saitou Takashi
Published: March 2003

In a population of the Great TitParus majorin Sagamihara, the divorce rate was 17.8% over 10 years from 1984/85 to 1993/94. Most of the divorces occurred in the year following pair formation. There were no differences in breeding success between faithful and divorced pairs. The birds were divided into three groups in relation to their home ranges: residents, emigrants and migrants. The residents established their home ranges within their flock ranges and remained there for life. Non‐residents (of which two sorts were identified) left their flock ranges to breed, but returned to their original flock ranges after breeding. Therefore, their home ranges were separated into breeding and previous flock ranges. Most of the faithful pairs consisted of residents whose home ranges overlapped extensively. The divorce rate was higher in the pairs consisting of residents and non‐residents and in those of non‐residents. Since the non‐breeding home ranges of emigrants and migrants did not overlap with their mates’ non‐breeding home ranges, an important factor affecting divorce seems to be the non‐overlap of non‐breeding home ranges between both mates of pairs after breeding.