bluuebunny/crossref_metadata_2025_split huggingface dataset:
Differential monoaminergic, neuroendocrine and behavioural responses after central administration of corticotropin‐releasing factor receptor type 1 and type 2 agonists
Authors: De Groote Lotte , Peñalva Rosana G. , Flachskamm Cornelia , Reul Johannes M. H. M. , Linthorst Astrid C. E. 
Published: September 2005

AbstractCorticotropin‐releasing factor (CRF) mediates various aspects of the stress response. To differentiate between the roles of CRF1 and CRF2 receptor subtypes in monoaminergic neurotransmission, hypothalamic–pituitary–adrenocortical axis activity and behaviour we compared the effects of CRF and urocortin 1 with those of the selective CRF2 receptor ligands urocortin 2 and urocortin 3. In vivo microdialysis in the rat hippocampus was used to assess free corticosterone, extracellular levels of serotonin (5‐HT) and noradrenaline (NA), and their metabolites 5‐hydroxyindoleacetic acid (5‐HIAA) and 3‐methoxy‐4‐hydroxyphenylglycol (MHPG), respectively. Intracerebroventricular (i.c.v.) injection of CRF and urocortin 1, 2 and 3 (1.0 µg) increased hippocampal levels of 5‐HT and 5‐HIAA. CRF and urocortin 1 increased NA and MHPG, whereas urocortin 2 and urocortin 3 elevated MHPG, but not NA levels. CRF and the urocortins induced an immediate increase in behavioural activity. CRF and urocortin 1 mainly caused grooming and exploratory behaviour. In contrast, urocortin 2 and urocortin 3 both induced exploratory behaviour, but not grooming, and increased time spent eating food pellets. All urocortins, but not CRF, suppressed food intake 4–6 h after injection. Hippocampal free corticosterone levels were elevated by CRF, urocortin 1 and 3, but not by urocortin 2. The time courses of the CRF‐ and urocortin 1‐induced responses were significantly prolonged as compared to those of the CRF2 receptor ligands. The stimulatory changes evoked by CRF and urocortin 1 were present up to 4–6 h after injection, whereas the effects of urocortin 2 and urocortin 3 returned to baseline within 2.5 h after injection. Pre‐treatment with the selective antagonist antisauvagine‐30 (5.0 µg, i.c.v.) confirmed that the effects of urocortin 3 were CRF2 receptor‐mediated. The differential time course of the monoaminergic, neuroendocrine and behavioural effects of CRF and urocortin 1, as compared to urocortin 2 and urocortin 3, and the specific behavioural pattern induced by the CRF2 receptor ligands, suggest a distinct role for CRF2 receptors in the stress response.
Length and overall sequence of the PEN‐2 C‐terminal domain determines its function in the stabilization of presenilin fragments
Authors: Prokop Stefan , Haass Christian , Steiner Harald 
Published: September 2005

Abstractγ‐Secretase is an aspartyl protease complex that catalyzes the intramembrane cleavage of a subset of type I transmembrane proteins including the β‐amyloid precursor protein (APP) implicated in Alzheimer's disease. Presenilin (PS), nicastrin (NCT), anterior pharynx defective (APH‐1) and presenilin enhancer‐2 (PEN‐2) constitute the active γ‐secretase complex. PEN‐2, the smallest subunit, is required for triggering PS endoproteolysis. Stabilization of the resultant N‐ and C‐terminal fragments, which carry the catalytically active site aspartates, but not endoproteolysis itself, requires the C‐terminal domain of PEN‐2. To functionally dissect the C‐terminal domain we created C‐terminal deletion mutants and mutagenized several evolutionary highly conserved residues. The PEN‐2 mutants were then probed for functional complementation of a PEN‐2 knockdown, which displays deficient PS1 endoproteolysis and impaired NCT maturation. Progressive truncation of the C‐terminus caused increasing loss of function. This was also observed for an internal deletion mutant as well as for C‐terminally tagged PEN‐2 with a twofold elongated C‐terminal domain. Interestingly, only simultaneous, but not individual substitution of the highly conserved D90, F94, P97 and G99 residues with alanine interfered with PEN‐2 function. All loss of function mutants identified allowed PS1 endoproteolysis, but failed to stably associate with the resultant PS1 fragments, which like the PEN‐2 loss of function mutants underwent proteasomal degradation. However, complex formation of the PEN‐2 mutants with PS1 fragments could be recovered when proteasomal degradation was blocked. Taken together, our data suggest that the PS‐subunit stabilizing function of PEN‐2 depends on length and overall sequence of its C‐terminal domain.
Constitutive high‐affinity choline transporter endocytosis is determined by a carboxyl‐terminal tail dileucine motif
Authors: Ribeiro Fabiola M. , Black Stefanie A. G. , Cregan Sean P. , Prado Vania F. , Prado Marco A. M. , Rylett R. Jane , Ferguson Stephen S. G. 
Published: September 2005

AbstractMaintenance of acetylcholine synthesis depends on the effective functioning of a high‐affinity sodium‐dependent choline transporter (CHT1). Recent studies have shown that this transporter is predominantly localized inside the cell, unlike other neurotransmitter transporters, suggesting that the trafficking of CHT1 to and from the plasma membrane may play a crucial role in regulating choline uptake. Here we found that CHT1 is rapidly and constitutively internalized in clathrin‐coated vesicles to Rab5‐positive early endosomes. CHT1 internalization is controlled by an atypical carboxyl‐terminal dileucine‐like motif (L531, V532) which, upon replacement by alanine residues, blocks CHT1 internalization in both human embryonic kidney 293 cells and primary cortical neurons and results in both increased CHT1 cell surface expression and choline transport activity. Perturbation of clathrin‐mediated endocytosis with dynamin‐I K44A increases cell surface expression and transport activity to a similar extent as mutating the dileucine motif, suggesting that we have identified the motif responsible for constitutive CHT1 internalization. Based on the observation that the localization of CHT1 to the plasma membrane is transient, we propose that acetylcholine synthesis may be influenced by processes that lead to the attenuation of constitutive CHT1 endocytosis.
Caspase 7: increased expression and activation after traumatic brain injury in rats
Authors: Larner Stephen F. , McKinsey Deborah M. , Hayes Ronald L. , W. Wang Kevin K. 
Published: September 2005

AbstractCaspases, a cysteine proteinase family, are required for the initiation and execution phases of apoptosis. It has been suggested that caspase 7, an apoptosis executioner implicated in cell death proteolysis, is redundant to the main executioner caspase 3 and it is generally believed that it is not present in the brain or present in only minute amounts with highly restricted activity. Here we report evidence that caspase 7 is up‐regulated and activated after traumatic brain injury (TBI) in rats. TBI disrupts homeostasis resulting in pathological apoptotic activation. After controlled cortical impact TBI of adult male rats we observed, by semiquantitative real‐time PCR, increased mRNA levels within the traumatized cortex and hippocampus peaking in the former about 5 days post‐injury and in the latter within 6–24 h of trauma. The activation of caspase 7 protein after TBI, demonstrated by immunoblot by the increase of the active form of caspase 7 peaking 5 days post‐injury in the cortex and hippocampus, was found to be up‐regulated in both neurons and astrocytes by immunohistochemistry. These findings, the first to document the up‐regulation of caspase 7 in the brain after acute brain injury in rats, suggest that caspase 7 activation could contribute to neuronal cell death on a scale not previously recognized.
Nuclear translocation of the SRF co‐activator MAL in cortical neurons: role of RhoA signalling
Authors: Tabuchi Akiko , Estevez Marcel , Henderson Jennifer A. , Marx Ruth , Shiota Jun , Nakano Hiroyasu , Baraban Jay M. 
Published: September 2005

AbstractAlthough it is well established that RhoA signaling pathways play key roles in regulating neuronal morphology, their involvement in other aspects of neuronal function has received little attention. Recent studies have elucidated a novel intracellular signaling pathway used by RhoA to elicit activation of serum response factor (SRF)‐mediated transcription. In this pathway, activation of RhoA triggers nuclear translocation of the SRF co‐activator, megakaryocytic acute leukemia (MAL). In assessing whether RhoA regulates transcription in neurons via this pathway, we have found that a constitutively active form of Tech (transcript‐enriched in cortex and hippocampus), a RhoA guanine nucleotide exchange factor (GEF) that is expressed in forebrain neurons, stimulates SRF reporter activity in extracts of primary cortical cultures and induces nuclear translocation of MAL in cortical neurons. Both of these responses appear to be mediated by Tech's activation of RhoA as they are not mimicked by a mutant Tech construct lacking RhoA GEF activity and are blocked by C3 transferase, a selective inhibitor of RhoA. Furthermore, Tech‐induced increases in SRF activity are suppressed by a dominant negative MAL construct. These findings demonstrate that RhoA signaling pathways are able to regulate transcription in neurons by triggering translocation of the SRF co‐activator MAL.
Sequestosome 1/p62 shuttles polyubiquitinated tau for proteasomal degradation
Authors: Babu Jeganathan Ramesh , Geetha Thangiah , Wooten Marie W. 
Published: September 2005

AbstractInclusions isolated from several neurodegenerative diseases, including Alzheimer's disease (AD), are characterized by ubiquitin‐positive proteinaceous aggregates. Employing confocal and immunoelectron microscopy, we find that the ubiquitin‐associating protein sequestosome1/p62, co‐localizes to aggregates isolated from AD but not control brain, along with the E3 ubiquitin ligase, TRAF6. This interaction could be recapitulated by co‐transfection in HEK293 cells. Employing both in vitro and in vivo approaches, tau was found to be a substrate of the TRAF6, possessing lysine 63 polyubiquitin chains. Moreover, tau recovered from brain of TRAF6 knockout mice, compared with wild type, was not ubiquitinated. Tau degradation took place through the ubiquitin–proteasome pathway and was dependent upon either the K63‐polyubiquitin chains or upon p62. In brain lysates of p62 knockout mice, tau fails to co‐interact with Rpt1, a proteasomal subunit, thereby indicating a requirement for p62 shuttling of tau to the proteasome. Our results demonstrate that p62 interacts with K63‐polyubiquitinated tau through its UBA domain and serves a novel role in regulating tau proteasomal degradation. We propose a model whereby either a decline in p62 expression or a decrease in proteasome activity may contribute to accumulation of insoluble/aggregated K63‐polyubiquitinated tau.
Excitotoxic and apoptotic neuronal death induce different patterns of glial activation in vitro
Authors: Pérez‐Capote Kamil , Serratosa Joan , Solà Carme 
Published: September 2005

AbstractWe have studied glial activation in rat cerebellar neuronal‐glial cultures after inducing neuronal death using various stimuli. Cultures were exposed to 100 µm glutamate for 20 min, which induces excitotoxic neuronal death, or to potassium/serum deprivation, which induces apoptosis of granule neurons. We evaluated alterations in several parameters related to glial activation: nuclear factor‐κB activation, nitric oxide and tumour necrosis factor‐α production, which are associated with a pro‐inflammatory response, glial proliferation and phagocytic activity. Although the two experimental models of neuronal damage resulted in the death of most neuronal cells within 24 h, differences were observed in the response of the various glial parameters evaluated. While nitric oxide production was not detected in any case, tumour necrosis factor‐α production, nuclear factor‐κB activation and glial proliferation were only induced in the presence of excitotoxic neuronal death. However, phagocytosis was induced in both cases, although earlier in the case of apoptotic neuronal death. These results show that glial cells respond to excitotoxic neuronal death with an inflammatory response associated with proliferation and phagocytosis. In contrast, whilst glial cells do not produce pro‐inflammatory molecules in the presence of apoptotic neuronal death, phagocytic activity is rapidly induced.
ALS‐IgG‐induced selective motor neurone apoptosis in rat mixed primary spinal cord cultures
Authors: Demestre M. , Pullen A. , Orrell R. W. , Orth M. 
Published: September 2005

AbstractThere is evidence that in sporadic amyotrophic lateral sclerosis (ALS) immunological mechanisms may be involved in the pathophysiology of the disease. We tested whether purified IgG from ALS patients induce cell death in rat mixed primary spinal cord cultures and compared this with the effect of IgG purified from patients with Guillain–Barré syndrome (GBS) or from healthy donors. Treatment with ALS‐IgG increases caspase‐3 apoptosis when compared with control IgG or with GBS‐IgG, but does not induce death by necrosis. Because ALS is characterized by the selective loss of motor neurones, we next assessed the differential effect of ALS‐IgG on motor neurones or astrocytes. We showed, semiquantitatively, that motor neurones are more susceptible to apoptosis when cultures were treated with ALS‐IgG compared with control‐IgG. In conclusion, we have demonstrated in primary spinal cord cultures that IgG from patients with ALS induces apoptosis selectively in motor neurones, and that the caspase‐3 pathway is involved. This suggests that immunological mechanisms may contribute to the selective loss of motor neurones in ALS.
Acute and long‐term effects of IL‐6 on cultured dorsal root ganglion neurones from adult rat
Authors: Segond von Banchet Gisela , Kiehl Michaela , Schaible Hans‐Georg 
Published: September 2005

AbstractIL‐6 contributes to pain and hyperalgesia in inflamed tissue. We have investigated short‐ and long‐term effects of IL‐6 on dorsal root ganglion (DRG) neurones. Glycoprotein 130‐like immunoreactivity (the signal transduction receptor subunit) was found in almost all neurones in DRG sections and in cultured DRG neurones from adult rat. In calcium‐imaging studies bath application of IL‐6 caused an increase of intracellular calcium in about one‐third of the DRG neurones suggesting functional IL‐6 receptors in a proportion of neurones. Long‐term but not short‐term exposure of DRG neurones to IL‐6 in vitro significantly enhanced the proportion of DRG neurones expressing neurokinin 1 receptor‐like immunoreactivity from 10% to up to 40%. This up‐regulation was dependent on the activation of mitogen‐activated protein kinase kinase (MEK) in the neurones, suggesting that the mitogen‐activated protein kinase (MAPK) pathway is important for this effects of IL‐6. Calcium‐imaging studies demonstrated that previous exposure of DRG neurones to IL‐6 enhanced the proportion of neurones that exhibit a substance P‐induced rise in intracellular calcium. These data show that IL‐6 has short‐ and long‐term effects on a proportion of DRG neurones. These effects are likely to contribute to pro‐nociceptive effects of IL‐6.
Proline mutations induce negative‐dosage effects on uptake velocity of the dopamine transporter
Authors: Lin Zhicheng , Uhl George R. 
Published: September 2005

AbstractAla and Gly substitutions for Pro 101 (P101) located in transmembrane domain 2 of the dopamine transporter (DAT) abolished transport activity but did not disrupt plasma membrane expression. Due to the high conservation of P101 in all neurotransmitter transporters and the capability of Pro to add flexibility to helices, we hypothesized that P101 contributes to the dynamic feature of substrate translocation. To test this hypothesis, here we analysed transport activity for DAT mutants where this Pro was mutated into different amino acids, including Ser, Val, Leu and Phe. The transmembrane domain 2 helix of P101F, unlike the other mutants, was computationally predicted to have a Van der Waals energy threefold higher than the wild‐type helix. P101F mutant expression was consistently disrupted in COS cells. Among all the other mutants that express normally, P101V, with a side‐chain size close to that of Pro, restores the transport activity of P101A by sevenfold. Most importantly, P101V, P101L and P101S display negative‐dosage effects on dopamine (DA) transport, i.e. the velocity–concentration curve for DA uptake does not show a plateau with increasing [DA] but rather peaks and then goes down. These data support the view that P101 of DAT plays an essential role in DA translocation.
The membrane environment of endogenous cellular prion protein in primary rat cerebellar neurons
Authors: Loberto Nicoletta , Prioni Simona , Bettiga Arianna , Chigorno Vanna , Prinetti Alessandro , Sonnino Sandro 
Published: September 2005

AbstractWe studied the membrane environment of cellular prion protein in primary cultured rat cerebellar neurons differentiated in vitro. In these cells, about 45% of total cellular prion protein (corresponding to a 35‐fold enrichment) is associated with a low‐density, sphingolipid‐ and cholesterol‐enriched membrane fraction, that can be separated by flotation on sucrose gradient. Biotinylation experiments indicated that almost all prion protein recovered in this fraction was exposed at the cell surface. Prion protein was efficiently separated from this fraction by a monoclonal antibody immunoseparation procedure. Under conditions designed to preserve lipid‐mediated membrane organization, several proteins were found in the prion protein‐enriched membrane domains (i.e. the non‐receptor tyrosine kinases Lyn and Fyn and the neuronal glycosylphosphatidylinositol‐anchored protein Thy‐1). The prion protein‐rich membrane domains contained, as well, about 50% of the sphingolipids, cholesterol and phosphatidylcholine present in the sphingolipid‐enriched membrane fraction. All main sphingolipids, including sphingomyelin, neutral glycosphingolipids and gangliosides, were similarly enriched in the prion protein‐rich membrane domains. Thus, prion protein plasma membrane environment in differentiated neurons resulted to be a complex entity, whose integrity requires a network of lipid‐mediated non‐covalent interactions.
Cumulative irritation patch test of sanitary pads on sensitive skin
Authors: Farage Miranda , Stadler Angela 
Published: September 2005

SummaryBackground  Tools are needed to assess the effects of various products on sensitive skin.Aim  To investigate the cutaneous compatibility of various sanitary pads in people with self‐declared sensitive skin.Patients/methods  Eight subjects, who considered their skin to be sensitive and who reported adverse skin responses to everyday products or clothing, were patch tested on the arm with two, low‐irritancy sanitary pads for four consecutive 24‐hour periods. Test products differed only in their surface covering. Sodium lauryl sulfate solution (0.1% w/v) and physiological saline served as standard irritant and nonirritant controls, respectively. Skin irritation (erythema) was graded after each 24‐hour period.Results  No significant difference in skin erythema scores was observed between groups (scores on day 4: 1.06 ± 0.11 vs. 1.25 ± 0.13, on a five‐point scale, P < 0.05). The temporal development of cumulative irritation associated with the pads was comparable to that observed with the nonirritant control (a plateau in maximal group scores) and distinct from that of the irritant control (continuously rising scores).Conclusion  Sanitary pads under investigation elicited negligible cumulative irritation in a four‐day patch test on subjects with self‐declared sensitive skin. The temporal pattern of cumulative response was consistent with the inherently low irritation potential of the products.
An evaluation of topical 3% salicylic acid and 1% hydrocortisone in the maintenance of scalp pruritus
Authors: Draelos Zoe Diana 
Published: September 2005

SummaryBackground  Scalp pruritus is a common condition causing dermatologic distress. The presence of skin scale on clothing is cosmetically undesirable and scalp scratching in public is socially embarrassing. Scratching can also result in removal of the cuticle and premature hair shaft fracture.Objective  To demonstrate the efficacy of 3% salicylic acid in combination with 1% hydrocortisone in the treatment of scalp pruritus.Methods  Sixty subjects with moderate scalp scaling and scalp pruritus were enrolled in a three‐arm double blind 2‐week study. The 20 subjects in arm 1 applied a topical 1% hydrocortisone product twice daily. The 20 subjects in arm 2 applied a topical 3% salicylic acid product twice daily. Lastly, the 20 subjects in arm 3 applied a topical 3% salicylic acid product in the morning and a topical 1% hydrocortisone product in the evening. Evaluations were performed at baseline, after 1 week of treatment, and after 2 weeks of treatment. The study investigator evaluated the subjects for scalp scale, erythema, excoriation, and overall assessment. In addition, scalp scale scrapings were collected and analyzed to gain further insight into scalp scale morphology. Subject assessments and scalp photography was also performed.Results  The investigator assessments revealed less excoriation in the hydrocortisone twice daily arm over the salicylic acid twice daily arm (P = 0.03), which might be expected because of its anti‐inflammatory effect. The morning salicylic acid application and evening hydrocortisone application arm performed better than the salicylic acid twice daily group at week 2 in terms of erythema (P = 0.02), excoriation (P = 0.03), and overall assessment (P = 0.01). Scalp scale scrapings revealed the least amount of retained skin scale in the combination salicylic acid/hydrocortisone group.Conclusion  The combination of a 3% salicylic acid keratolytic combined with a 1% hydrocortisone anti‐inflammatory provides the best relief of scalp pruritus.