Encephalitogenicity of murine, but not bovine, DM20 in SJL mice is due to a single amino acid difference in the immunodominant encephalitogenic epitope

Myelin proteolipid protein (PLP) contains 2 immunodominant encephalitogenic epitopes in SJL mice, namely PLP residues 139-151 and 178-191. DM20, a minor isoform of PLP, lacks residues 116-150 and consequently contains only the single major encephalitogenic epitope 178-191. However, it has been found previously that bovine DM20 is not encephalitogenic in SJL mice. Since residue 188 within peptide 178-191 is phenylalanine (F) in murine DM20 and alanine (A) in bovine DM20, we tested the effect of this difference on the immune responses and induction of EAE. SJL mice were immunized with either highly purified murine or bovine DM20. Residues 178-191 were found to be immunodominant for each, but only murine and not bovine DM20 was encephalitogenic. A synthetic peptide corresponding to the murine 178-191 sequence (F188) was also encephalitogenic, whereas the peptide corresponding to the bovine sequence (A188) was not. Both F188 and A188 bind with high affinity to I-A(s) and both are recognized by the SJL T cell repertoire. A188-specific T cell lines reacted to both A188 and F188, but F188-specific T cell lines were not stimulated by A188. F188-specific T cell lines produced mRNA for the Th1 cytokines IL2 and IFN gamma and, in passive transfer experiments, were encephalitogenic upon stimulation with F188, but not A188. In contrast, A188-specific T cell lines produced mRNA for IL4, IL5 and IL10, in addition to IL2 and IFN gamma, and were not encephalitogenic after stimulation with either F188 or A188. Cotransfer of A188-specific T cell lines with F188-specific T cell lines resulted in protection from EAE. Thus, A188 induces a functionally different phenotype of T cells from that induced by F188. Taken together these data suggest that the failure of bovine DM20 to induce EAE may be attributable to induction of protective rather than pathogenic T cells by the immunodominant epitope.

Myelin proteolipid protein expressed in COS-1 cells is targeted to actin-associated surfaces

The compact myelin sheath represents one of the largest expanses of membrane-membrane contact in the body and, in the central nervous system, requires the myelin proteolipid protein (PLP) for assembly, To determine whether the molecular properties of PLP promote membrane adhesion and direct its subcellular localization in the absence of oligodendrocyte-specific targeting mechanisms, PLP was expressed in COS-I fibroblasts, Immunofluorescence staining indicated that PUP was translated effectively, transited the rough endoplasmic reticulum and Golgi apparatus, was delivered to the cell surface, and was endocytosed, In the plasma membrane, the PLP distribution was patchy and only sporadically coincided with sites of membrane-membrane contact between PLP-expressing cells, PLP was not randomly distributed, however, but correlated closely with microfilament locations in leading edge membranes and microvilli, as demonstrated by phalloidin double labeling, Our results indicate that even in non-myelinating cells, PLP can be concentrated in membranes associated with movement and growth, and suggest possible roles for the actin cytoskeleton in PLP localization, As PLP, DM20, and the DM20-like M6 protein all associate with actin-enriched membranes, this may be a common feature of PLP/DM20 gene family members. (C) 1997 Wiley-Liss, Inc.

Peripheral neuropathy in patients with primary antiphospholipid (Hughes`) syndrome

The involvement of the peripheral nervous system in diverse autoimmune diseases is well established. However, no appropriately designed studies have been performed in primary antiphospholipid syndrome (PAPS)-related peripheral neuropathy. We aimed to investigate the occurrence of peripheral neuropathy in patients diagnosed with PAPS. Twenty-six consecutive patients with PAPS (Sapporo criteria) and 20 age-and gender-matched healthy controls were enrolled at two referral centers. Exclusion criteria were secondary causes of peripheral neuropathy. A complete clinical neurologic exam followed by nerve conduction studies (NCS) was performed. Paresthesias were reported in eight patients (31%). Objective mild distal weakness and abnormal symmetric deep tendon reflexes were observed in three patients (11.5%). With regard to the electrophysiologic evidence of peripheral neuropathy, nine patients (35.0%) had alterations: four (15.5%) had pure sensory or sensorimotor distal axonal neuropathy (in two of them a carpal tunnel syndrome was also present) and one (4%) had sensorimotor demyelinating and axonal neuropathy involving upper and lower extremities, while four patients (15.5%) showed isolated carpal tunnel syndrome. Clinical and serologic results were similar in all the patients with PAPS, regardless of the presence of electrophysiologic alterations. In conclusion, peripheral neuropathy is a common asymptomatic abnormality in patients with PAPS. The routine performance of NCS may be considered when evaluating such patients. Lupus (2010) 19, 583-590.

Ca2+-activated K+ channels in rat otic ganglion cells: Role of Ca2+ entry via Ca2+ channels and nicotinic receptors

1. Intracellular recordings were made from neurones in the rat otic ganglion in vitro in order to investigate their morphological, physiological and synaptic properties. We took advantage of the simple structure of these cells to test for a possible role of calcium influx via nicotinic acetylcholine receptors during synaptic transmission. 2. Cells filled with biocytin comprised a homogeneous population with ovoid somata and sparse dendritic trees. Neurones had resting membrane potentials of -53 +/- 0.7 mV (n = 69), input resistances of 112 + 7 M Omega, and membrane time constants of 14 +/- 0.9 ms (n = 60). Upon depolarization, all cells fired overshooting action potentials which mere followed by an apamin-sensitive after-hyperpolarization (AHP). In response to a prolonged current injection, all neurones fired tonically. 3. The repolarization phase of action potentials had a calcium component which was mediated by N-type calcium channels. Application of omega-conotoxin abolished both the repolarizing hump and the after-hgrperpolarization suggesting that calcium influx via N-type channels activates SK-type calcium-activated potassium channels which underlie the AHP. 4. The majority (70%) of neurones received innervation from a single preganglionic fibre which generated a suprathreshold excitatory postsynaptic potential mediated by nicotinic acetylcholine receptors. The other 30% of neurones also had one or more subthreshold nicotinic inputs. 5. Calcium influx via synaptic nicotinic receptors contributed to the AHP current, indicating that this calcium has access to the calcium-activated potassium channels and therefore plays a role in regulating cell excitability.

The central pathway of primary olfactory axons is abnormal in mice lacking the N-CAM-180 isoform

Although N-CAM has previously been implicated in the growth and fasciculation of axons, the development of axon tracts in transgenic mice with a targeted deletion of the 180-kD isoform of the neural cell adhesion molecule (N-CAM-180) appears grossly normal in comparison to wild-type mice. We examined the organization of the olfactory nerve projection from the olfactory neuroepithelium to glomeruli in the olfactory bulb of postnatal N-CAM-180 null mutant mice. Immunostaining for olfactory marker protein revealed the normal presence of fully mature primary olfactory neurons within the olfactory neuroepithelium of mutant mice. The axons of these neurons form an olfactory nerve, enter the nerve fiber layer of the olfactory bulb, and terminate in olfactory glomeruli as in wild-type control animals. The olfactory bulb is smaller and the nerve fiber layer is relatively thicker in mutants than in wild-type mice. Previous studies have revealed that the plant lectin Dolichos biflorus agglutinin (DBA) clearly stains the perikarya and axons of a subpopulation of primary olfactory neurons. Thus, DBA staining enabled the morphology of the olfactory nerve pathway to be examined at higher resolution in both control and mutant animals. Despite a normal spatial pattern of DBA-stained neurons within the nasal cavity, there was a distorted axonal projection of these neurons onto the surface of the olfactory bulb in N-CAM-180 null mutants. In particular, DBA-stained axons formed fewer and smaller glomeruli in the olfactory bulbs of mutants in comparison to wild-type mice. Many primary olfactory axons failed to exit the nerve fiber layer and contribute to glomerular formation. These results indicate that N-CAM-180 plays an important role in the growth and fasciculation of primary olfactory axons and is essential for normal development of olfactory glomeruli. (C) 1997 John Wiley & Sons, Inc.

Heart Transplantation in 107 Cases of Chagas` Disease

Introduction. Chagas` disease is endemic in South America. Objective. This research reviewed the experience with cardiac transplantation in Chagas` disease, emphasizing reactivation, immunosuppression, and mortality. Methods. Over 25 years from March 1985 to March 2010, 107/409 (26.2%) patients with Chagas` disease underwent heart transplantation, patients including 74 (71.1%) men and 72 (67.2%), in functional class IV with 33 (30.8%) on vasopressors and 17 (10.7%) on mechanical circulatory support. Results. The diagnosis of disease reactivation was performed by identifying the parasite in the myocardium (n = 23; 71.8%) in the subcutaneous tissue (n = 8; 25.0%), in blood (n = 11; 34.3%), or in central nervous tissue (n = 1; 3.1%). Hospital mortality was 17.7% (n = 19) due to infection (n = 6; 31.5%), graft dysfunction (n = 6; 31.5%), rejection (n 4; 21.1%), or sudden death (n = 2; 10.5%). Late mortality was 27 (25.2%) cases, which were distributed as: rejection (n = 6; 22.2%), infection (n = 6; 22.2%), (n = lymphoma 4; 14.8%), sarcoma (n = 2; 7.4%), for constrictive pericarditis (n = 2; 7.4%) reactivation of Chagas` disease in the central nervous system (n = 1; 7.1%). Conclusions. Transplantation in Chagas` disease has peculiar problems that differ from other etiologies due to the possibility of disease reactivation and the increased possibility of emergence of cancers. However, transplantation is the only treatment able to modify the natural progression of the disease in its terminal phase. Early diagnosis and rapid introduction of benzonidazole reverses the histological patterns. Immunosuppression, especially steroids, predisposes to the development of cancer and disease reactivation.

Effects of high-intensity aerobic interval training vs. moderate exercise on hemodynamic, metabolic and neuro-humoral abnormalities of young normotensive women at high familial risk for hypertension

Exercise training has an important role in the prevention and treatment of hypertension, but its effects on the early metabolic and hemodynamic abnormalities observed in normotensive offspring of hypertensive parents (FH+) have not been studied. We compared high-intensity interval (aerobic interval training, AIT) and moderate-intensity continuous exercise training (CMT) with regard to hemodynamic, metabolic and hormonal variables in FH+ subjects. Forty-four healthy FH+ women (25.0+/-4.4 years) randomized to control (ConFH+) or to a three times per week equal-volume AIT (80-90% of VO(2MAX)) or CMT (50-60% of VO(2MAX)) regimen, and 15 healthy women with normotensive parents (ConFH-; 25.3+/-3.1 years) had their hemodynamic, metabolic and hormonal variables analyzed at baseline and after 16 weeks of follow-up. Ambulatorial blood pressure (ABP), glucose and cholesterol levels were similar among all groups, but the FH+ groups showed higher insulin, insulin sensitivity, carotid-femoral pulse wave velocity (PWV), norepinephrine and endothelin-1 (ET-1) levels and lower nitrite/ nitrate (NOx) levels than ConFH- subjects. AIT and CMT were equally effective in improving ABP (P<0.05), insulin and insulin sensitivity (P<0.001); however, AIT was superior in improving cardiorespiratory fitness (15 vs. 8%; P<0.05), PWV (P<0.01), and BP, norepinephrine, ET-1 and NOx response to exercise (P<0.05). Exercise intensity was an important factor in improving cardiorespiratory fitness and reversing hemodynamic, metabolic and hormonal alterations involved in the pathophysiology of hypertension. These findings may have important implications for the exercise training programs used for the prevention of inherited hypertensive disorder. Hypertension Research (2010) 33, 836-843; doi:10.1038/hr.2010.72; published online 7 May 2010

Biochemical synthesis, purification and preliminary pharmacological evaluation of normorphine-3-glucuronide

Normorphine was synthesised from morphine by thermal decomposition of an N-alpha-chloroethylchloroformate adduct, and purified (> 98% purity) using semipreparative HPLC with ultraviolet detection. Normorphine-3-glucuronide (NM3G) was biochemically synthesised using the substrate normorphine, uridine diphosphoglucuronic acid and Sprague-Dawley rat liver microsomes in a 75% yield (relative to normorphine base). The synthesised NM3G was purified by precipitation and washing with acetonitrile. Determinations of purity using HPLC with electrochemical and ultraviolet detection confirmed that the NM3G produced was of high (> 99%) purity. Mass spectrometry, fourier transform infrared spectrophotometry and nuclear magnetic resonance spectrometry confirmed the structure, especially placement of the glucuronide moiety at the 3-phenolic position and not at the 17-nitrogen. Administration of NM3G by the intracerebroventricular (icy) route to rats in doses of 2.5 and 7.5 mu g resulted in the development of central nervous system (CNS) excitatory behavioural effects including myoclonus, chewing, wet-dog shakes, ataxia and explosive motor behaviour. At an icy dose of 7.5 mu g, NM3G also induced short periods of tonic-clonic convulsive activity. Thus, NM3G elicits CNS excitation following supraspinal administration in a manner analogous to morphine-3-glucuronide (M3G), the major metabolite of morphine (1). Further studies are required to determine whether NM3G attenuates morphine-induced antinociception in se similar manner to M3G.

The alcohol withdrawal syndrome

The alcohol withdrawal syndrome (AWS) is a set of signs and symptoms that typically develops in alcohol-dependent people within 6–24 h of their last drink. It may occur unintentionally if abstinence is enforced by illness or injury, or deliberately if the person voluntarily stops drinking because of an alcohol-related illness, or as a prelude to becoming and remaining abstinent. The signs and symptoms of the syndrome (panel) are largely, but not exclusively, those of autonomic hyperactivity, the reverse of the effects of alcohol intoxication. They represent a homoeostatic readjustment of the central nervous system (CNS) to the neuroadaptation that occurs with prolonged alcohol intoxication.1 RC Turner, PR Lichstein and JG Peden et al., Alcohol withdrawal syndromes: a review of pathophysiology, clinical presentation and treatment, J Gen Intern Med 4 (1989), pp. 432–444. Full Text via CrossRef | View Record in Scopus | Cited By in Scopus (39)1 They vary in severity from mild to severe.1

Repetitive transcranial magnetic stimulation improve tinnitus in normal hearing patients: a double-blind controlled, clinical and neuroimaging outcome study

Background and purpose: Tinnitus is a frequent disorder which is very difficult to treat and there is compelling evidence that tinnitus is associated with functional alterations in the central nervous system. Targeted modulation of tinnitus-related cortical activity has been proposed as a promising new treatment approach. We aimed to investigate both immediate and long-term effects of low frequency (1 Hz) repetitive transcranial magnetic stimulation (rTMS) in patients with tinnitus and normal hearing. Methods: Using a parallel design, 20 patients were randomized to receive either active or placebo stimulation over the left temporoparietal cortex for five consecutive days. Treatment results were assessed by using the Tinnitus Handicap Inventory. Ethyl cysteinate dimmer-single photon emission computed tomography (SPECT) imaging was performed before and 14 days after rTMS. Results: After active rTMS there was significant improvement of the tinnitus score as compared to sham rTMS for up to 6 months after stimulation. SPECT measurements demonstrated a reduction of metabolic activity in the inferior left temporal lobe after active rTMS. Conclusion: These results support the potential of rTMS as a new therapeutic tool for the treatment of chronic tinnitus, by demonstrating a significant reduction of tinnitus complaints over a period of at least 6 months and significant reduction of neural activity in the inferior temporal cortex, despite the stimulation applied on the superior temporal cortex.

Prion protein ablation increases cellular aggregation and embolization contributing to mechanisms of metastasis

Cellular Prion Protein (PrP(C)) is a cell surface protein highly expressed in the nervous system, and to a lesser extent in other tissues. PrP(C) binds to the extracellular matrix laminin and vitronectin, to mediate cell adhesion and differentiation. Herein, we investigate how PrP(C) expression modulates the aggressiveness of transformed cells. Mesenchymal embryonic cells (MEC) from wildtype (Prnp(+/+)) and PrP(C)-null (Prnp(0/0)) mice were immortalized and transformed by co-expression of ras and myc. These cells presented similar growth rates and tumor formation in vivo. When injected in the tail vein, PrnP(0/0)raS/myc cells exhibited increased lung colonization compared with Prnp(+/+)ras/myc cells. Additionally, Prnp(0/0)ras/myc cells form more aggregates with blood components than Prnp(+/+)ras/myc cells, facilitating the arrest of Prnp(0/0)ras/myc cells in the lung vasculature. Integrin alpha(v)beta(3) is more expressed and activated in MEC and in transformed Prnp(0/0) cells than in the respective Prnp(+/+) cells. The blocking of integrin alpha(v)beta(3) by RGD peptide reduces lung colonization in transformed Prnp(0/0) cells to similar levels of those presented by transformed Prnp(+/+) cells. Our data indicate that PrP(C) negatively modulates the expression and activation of integrin alpha(v)beta(3) resulting in a more aggressive phenotype. These results indicate that PrP(C) may have main implications in modulating metastasis formation. (C) 2009 UICC

Genetic basis of the formation and identity of type I and type II neurons in Drosophila embryos

The embryonic peripheral nervous system of Drosophila contains two main types of sensory neurons: type I neurons, which innervate external sense organs and chordotonal organs, and type II multidendritic neurons, Here, we analyse the origin of the difference between type I and type II in the case of the neurons that depend on the proneural genes of the achaete-scute complex (ASC), We show that, in Notch(-) embryos, the type I neurons are missing while type nr neurons are produced in excess, indicating that the type I/type II choice relies on Notch-mediated cell communication, In contrast, both type I and type II neurons are absent in numb(-) embryos and after ubiquitous expression of tramtrack, indicating that the activity of numb and the absence of tramtrack are required to produce both external sense organ and multidendritic neural fates, The analysis of string(-) embryos reveals that when the precursors are unable to divide they differentiate mostly into type II neurons, indicating that the type II is the default neuronal fate, We also report a new mutant phenotype where the ASC-dependent neurons are converted into-type II neurons, providing evidence for the existence of one or more genes required for maintaining the alternative (type I) fate, Our results suggest that the same mechanism of type I/type II specification may operate at a late step of the ASC-dependent lineages, when multidendritic neurons arise as siblings of the external sense organ neurons and, at an early step, when other multidendritic neurons precursors arise as siblings of external sense organ precursors.

Expression of vitamin D receptor mRNA in the hippocampal formation of rats submitted to a model of temporal lobe epilepsy induced by pilocarpine

Vitamin D (VD), is a steroid hormone with multiple functions in the central nervous system (CNS), producing numerous physiological effects mediated by its receptor (VDR). Clinical and experimental studies have shown a link between VD dysfunction and epilepsy. Along these lines, the purpose of our work was to analyze the relative expression of VDR mRNA in the hippocampal formation of rats during the three periods of pilocarpine-induced epilepsy. Male Wistar rats were divided into five groups: (1) control group; rats that received saline 0.9%, i.p. and were killed 7 days after its administration (CTRL, n = 8), (2) SE group; rats that received pilocarpine and were killed 4 h after SE (SE, n = 8), (3) Silent group-7 days; rats that received pilocarpine and were killed 7 days after SE (SIL 7d, n = 8), (4) Silent group-14 days; rats that received pilocarpine and were killed 14 days after SE (SIL 14d, n = 8), (5) Chronic group; rats that received pilocarpine and were killed 60 days after the first spontaneous seizure, (chronic, n = 8). The relative expression of VDR mRNA was determined by real-time PCR. Our results showed an increase of the relative expression of VDR mRNA in the SIL 7 days, SIL 14 days and Chronic groups, respectively (0.060 +/- 0.024; 0.052 +/- 0.035; 0.085 +/- 0.055) when compared with the CTRL and SE groups (0.019 +/- 0.017; 0.019 +/- 0.025). These data suggest the VDR as a possible candidate participating in the epileptogenesis process of the pilocarpine model of epilepsy. (C) 2008 Elsevier Inc. All rights reserved.

Novel gene containing multiple epidermal growth factor-like motifs transiently expressed in the papillae of the ascidian tadpole larvae

We have investigated molecular mechanisms of the embryonic development of an ascidian, a primitive chordate which shares features of both invertebrates and vertebrates, with a view to identifying genes involved in development and metamorphosis, We isolated 12 partial cDNA sequences which were expressed in a stage-specific manner using differential display, We report here the isolation of a full-length cDNA sequence for one of these genes which was specifically expressed during the tailbud and larval stages of ascidian development, This cDNA, 1213 bp in length, is predicted to encode a protein of 337 amino acids containing four epidermal growth factor (EGF)-like repeats and three novel cysteine-rich repeats, Characterization of its spatial expression pattern by in situ hybridisation in late tailbud and larval embryos demonstrated strong expression localised throughout the papillae and anteriormost trunk and weaker expression in the epidermis of the remainder of the embryo, As recent evidence indicates that the signal for metamorphosis originates in the anterior trunk region, these results suggest that this gene may have a role in signalling the initiation of metamorphosis. (C) 1997 Wiley-Liss, Inc.

Differential expression of Egr-1-like DNA-binding activities in the naive rat brain and after excitatory stimulation

Egr-1 and related proteins are inducible transcription factors within the brain recognizing the same consensus DNA sequence. Three Egr DNA-binding activities were observed in regions of the naive rat brain. Egr-1 was present in all brain regions examined. Bands composed, at least in part, of Egr-2 and Egr-3 were present in different relative amounts in the cerebral cortex, striatum, hippocampus, thalamus, and midbrain. All had similar affinity and specificity for the Egr consensus DNA recognition sequence. Administration of the convulsants NMDA, kainate, and pentylenetetrazole differentially induced Egr-1 and Egr-2/3 DNA-binding activities in the cerebral cortex, hippocampus, and cerebellum. All convulsants induced Egr-1 and Egr-2 immunoreactivity in the cerebral cortex and hippocampus. These data indicate that the members of the Egr family are regulated at different levels and may interact at promoters containing the Egr consensus sequence to fine tune a program of gene expression resulting from excitatory stimuli.