3,4-Dihydroxyphenylacetic acid (DOPAC) as an index of noradrenaline turnover: Effects of hydergine® and vincamine

Summary Among the drugs commonly used in the treatment of memory disorders of the elderly, vincamine and hydergine^® have been shown to moderately increase the firing rate of noradrenergic locus coeruleus (LC) neurons. Since changes in electrical activity of noradrenergic neurons are generally reflected in corresponding alterations of the turnover of this transmitter, the effects of these drugs on the accumulation of 3,4-dihyroxyphenylacetic acid (DOPAC) and dopamine (DA) in the presence and absence of the dopamine--hydroxylase inhibitor, FLA 63, were studied in the LC as well as in two of its projection areas, the hippocampus and the cerebellum. Characterization of this procedure with the ₂-adrenoceptor antagonist, idazoxan, the corresponding agonist, clonidine, the ₁-adrenoceptor antagonist prazosine, and haloperidol, suggested that- DOPAC changes are more suitable than those of DA or DOPAC/DA ratios in reflecting changes in noradrenaline (NA) turnover,- inhibiting DBH is advantageous if NA turnover is to be measured in projection areas, but not in LC, and- haloperidol and prazosine, in principle, did not affect NA turnover.Vincamine and hydergine^® at 10 mg/kg doses, at which they were reported to increase LC firing by 50%, did not induce a change in NA turnover in any of the areas. This, together with the data obtained with haloperidol, suggests that a minimal increase in the firing rate of LC cells (+140%) is required before it could influence the turnover of NA, as measured by DOPAC changes.Thus, the stimulating effect of nootropics on the central noradrenergic system may be more sensitively detected by electrophysiological techniques than by biochemical ones.     

Role of mouse cerebellar nicotinic acetylcholine receptor (nAChR) α(4)β(2)- and α(7) subtypes in the behavioral cross-tolerance between nicotine and ethanol-induced ataxia

We have demonstrated that nicotine attenuated ethanol-induced ataxia via nicotinic-acetylcholine-receptor (nAChR) subtypes α(4)β(2) and α(7). In the present study, ethanol (2g/kg; i.p.)-induced ataxia was assessed by Rotorod performance following repeated intracerebellar infusion of α(4)β(2)- and α(7)-selective agonists. Localization of α(4)β(2) and α(7) nAChRs was confirmed immunohistochemically. Cerebellar NO(x) (nitrite+nitrate) was determined flurometrically. Repeated intracerebellar microinfusion of the α(4)β(2)-selective agonist, RJR-2403 (for 1, 2, 3, 5 or 7 days) or the α(7)-selective agonist, PNU-282987 (1, 2, 3 or 5 days), dose-dependently attenuated ethanol-induced ataxia. These results suggest the development of cross-tolerance between ethanol-induced ataxia and α(4)β(2) and α(7) nAChR agonists. With RJR-2403, the cross-tolerance was maximal after a 5-day treatment and lasted 48h. Cross-tolerance was maximal after a 1-day treatment with PNU-282987 and lasted 72h. Pretreatment with α(4)β(2)- and α(7)-selective antagonists, dihydro-β-erythroidine and methyllycaconitine, respectively, prevented the development of cross-tolerance confirming α(4)β(2) and α(7) involvement. Repeated agonist infusions elevated cerebellar NO(x) 16h after the last treatment while acute ethanol exposure decreased it. Pretreatment with repeated RJR-2403 or PNU-282987 reversed ethanol-induced decrease in NOx. The NO(x) data suggests the involvement of the nitric oxide (NO)-cGMP signaling pathway in the cross-tolerance that develops between α(4)β(2)- and α(7)-selective agonists and ethanol ataxia. Both α(4)β(2) and α(7) subtypes exhibited high immunoreactivity in Purkinje but sparse expression in molecular and granular cell layers. Our results support a role for α(4)β(2) and α(7) nAChR subtypes in the development of cross-tolerance between nicotine and ethanol with the NO signaling pathway as a potential mechanism.     

At low doses ethanol maintains blood–brain barrier (BBB) integrity after hypoxia and reoxygenation: a brain slice study

Abstract

Post-ischemia ethanol (EtOH) treatments have been shown to exhibit neuroprotective effects in stroke. However, the mechanisms underlying these effects and those on blood–brain barrier (BBB) integrity have yet to be elucidated. In the present study, we determined whether administering differing concentrations of EtOH alter the expressions of BBB integral proteins, including aquaporins-4 and -9 (AQP-4, AQP-9), matrix metallopeptidases-2 and -9 (MMP-2, MMP-9), zonula occludens-1 (ZO-1), and basal lamina (laminin).

We employed an organotypic brain slice culture model that utilizes oxygen–glucose deprivation followed by reoxygenation (OGD/R). Brain slices were obtained from 10-day-old Sprague–Dawley rats and divided into the following five groups (n = 8 subjects per group): (1) control, (2) hypoxia (OGD/R), no EtOH, (3) OGD/R and 10 mM EtOH, (4) OGD/R and 30 mM EtOH, and (5) OGD/R and 90 mM EtOH. To assess BBB integrity, levels of AQPs, MMPs, ZO-1, and laminin were determined by Western blot.

Compared to control, OGD/R without EtOH significantly increased AQP-4, AQP-9, MMP-2, and MMP-9 levels, while decreasing ZO-1 and laminin levels. All EtOH concentration treatments (groups 3 through 5) significantly reduced the expressions of AQP-4, AQP-9, MMP-2, and MMP-9, compared to the OGD/R, non-alcohol treated slices. Furthermore, compared to the OGD/R without EtOH group, the 30 mM EtOH treatment significantly increased ZO-1 and laminin levels. In contrast, the 90 mM EtOH level neither enhanced the reduction in AQP and MMP levels nor increased ZO-1 or basal lamina expressions observed in the 30 mM treatment.

In conclusion, at an optimal dose of 30 mM, EtOH improves the expressions of MMP-2, MMP-9, AQP-4, AQP-9, ZO-1, and basal laminin, previously altered by OGD/R. These effects may indicate a beneficial effect of EtOH on BBB integrity after stroke.     

Expression of aromatase and two isozymes of 5α-reductase in the developing green anole forebrain

Neural steroids, as well as the enzymes that produce these hormones, are important for sexual differentiation of the brain during development. Aromatase converts testosterone into oestradiol. 5α‐reductase converts testosterone to 5α‐dihydrotestosterone and occurs in two isozymes: type 1 (5αR1) and type 2 (5αR2). Each of these enzymes is present in the developing brain in many species, although no work has been carried out examining the expression of all three enzymes in non‐avian reptiles with genetic sex determination. In the present study, we evaluated mRNA expression of neural aromatase, 5αR1 and 5αR2, on the day of hatching and at day 50 in one such lizard, the green anole. We describe the distribution of these enzymes throughout the brain and the quantification of mRNA expression in three regions that control adult sexual behaviours: the preoptic area (POA) and ventromedial amygdala (AMY), which are involved in male displays, as well as the ventromedial hypothalamus, which regulates female receptivity. Younger animals had a greater number (POA) and density (AMY) of 5αR1 mRNA expressing cells. We detected no effects of sex or age on aromatase or 5αR2. In comparison with data from adults, the present results support the idea that the green anole forebrain has not completely differentiated by 50 days after hatching and that 5αR1 may play a role in the early development of regions important for masculine function.     

Hyperthyroidism: Specifically increased response to central NA-(α-)receptor stimulation and generally increased monoamine turnover in brain

Summary Repeated treatment with thyroxine (T₄) caused in mice enhanced response,i.e. locomotor stimulation, to central noradrenaline (NA)-receptor activation by clonidine but not to central dopamine (DA)-receptor activation by apomorphine or ET 495 after previous depletion of endogenous catecholamines (CA) by reserpine and inhibition of the CA-synthesis by-methyl-p-tyrosine. Increasing the dosese of the receptor agonists in control animals did not increase the locomotor stimulation but merely prolonged the effect. In chronically T₄-treated rats with a significantly elevated plasma level of T₄ and T₃ the apomorphine-induced stereotypies were not increased and the response to small amounts of DA locally applied in N. accumbens was not significantly affected by the T₄-pretreatment. The synthesis rates of NA and DA, reflected in the amount of L-Dopa accumulated in predominantly NA- and DA-rich brain parts, respectively, during 30 min after administration of the centrally active, aromatic L-amino acid decarboxylase inhibitor NSD 1015, were both increased by T₄-pretreatment. Also the brain serotonin (5-hydroxytryptamine, 5-HT)-synthesis was increased, measured in an analogous way. In contrast, utilization of brain CA or 5-HT seemed not affected by the T₄-pretreatment, as disclosed by unchanged disappearance rates of NA, DA or 5-HT after inhibition of tyrosine hydroxylase and tryptophan hydroxylase, respectively. Release of brain CA, as reflected in the amount of O-methylated metabolites after monoamine oxidase inhibition, was not affected by the hormone pretreatment. We conclude that moderate hyperthyroidism is associated with a specific, increased sensitivity at or beyond central NA-(-)receptors. This effect may be implicated in: 1. The potentiation by thyroid hormone of tricyclic antidepressive drugs; 2. the affective symptoms in hyperthyroidism,e.g. anxiety and increased susceptibility to emotional stress; 3. the cardiovascular symptoms in hyperthyroidism.     

Characterization of the expression of macrophage inflammatory protein-1α (MIP-1α) and C-C chemokine receptor 5 (CCR5) after kainic acid-induced status epilepticus (SE) in juvenile rats

X. B. Zhu, Y. B. Wang, O. Chen, D. Q. Zhang, Z. H. Zhang, A. H. Cao, S. Y. Huang and R. P. Sun (2012) Neuropathology and Applied Neurobiology 38, 602–616 Characterization of the expression of macrophage inflammatory protein‐1α (MIP‐1α) and C‐C chemokine receptor 5 (CCR5) after kainic acid‐induced status epilepticus (SE) in juvenile rats Aims: To identify the potential role of macrophage inflammatory protein‐1α (MIP‐1α) with its C‐C chemokine receptor 5 (CCR5) in epileptogenic brain injury, we examined their expression in juvenile rat hippocampus and explored the potential link between MIP‐1α, CCR5 and neuropathological alterations after status epilepticus (SE) induced by intracerebroventricular (i.c.v.) kainic acid (KA) injection. Methods: Based on the determination of the development of spontaneous seizures initiated by SE in developing rat brain, we firstly examined hippocampal neurone damage through Nissl and Fluoro‐Jade B staining, and evaluated microglial reaction during the early phase following KA‐induced SE in 21‐day‐old rats. MIP‐1α and CCR5 protein were quantified by ELISA and Western blot respectively following mRNA by real‐time PCR. We also mapped MIP‐1α and CCR5 expression in the hippocampus by immunohistochemistry and identified their cellular sources using double‐labelling immunofluorescence. Results: In juvenile rats, KA caused characteristic neurone damage in the hippocampal subfields, with accompanying microglial accumulation. In parallel with mRNA expression, MIP‐1α protein in hippocampus was transiently increased after KA treatment, and peaked from 16 to 72 h. Double‐labelling immunofluorescence revealed that MIP‐1α was localized to microglia. Up‐regulated CCR5 remained prominent at 24 and 72 h and was mainly localized to activated microglia. Further immunohistochemistry revealed that MIP‐1α and CCR5 expression were closely consistent with microglial accumulation in corresponding hippocampal subfields undergoing degenerative changes. Conclusions: Our data indicated that MIP‐1α as a regulator, linking with the CCR5 receptor, may be involved within the early stages of the epileptogenic process following SE by i.c.v. KA injection.     

Effects of rabbit anti-α-melanocyte-stimulating hormone (α-MSH) immunoglobulins on α-MSH signaling related to food intake control

Anti-α-melanocyte-stimulating hormone (α-MSH) polyclonal antibodies have been used for α-MSH neutralization in functional studies, but the results are sometime inconsistent with the antibody expected blocking properties. The present study aimed to determine if rabbit (Rb) anti-α-MSH immunoglobulins (Ig) may inhibit or enhance α-MSH signaling on melanocortin receptor type 4 (MC4R) and α-MSH-induced anorexigenic effect if presented as immune complexes with α-MSH. Polyclonal Rb anti-α-MSH IgG were commercially available and their ability to bind α-MSH has been confirmed by the immunohistochemical detection of α-MSH neurons in the rat hypothalamus. In vitro assay of the cyclic-adenosine mono-phosphate (cAMP) secreted by cells transfected with MC4R was performed to analyze effect of Rb IgG on α-MSH-induced cAMP production. We found that adding Rb IgG to α-MSH resulted in stimulation of cAMP detected at lower peptide concentrations as compared to α-MSH alone. To determine effects of Rb IgG on food intake, rats were injected into the arcuate hypothalamic nucleus with either α-MSH, Rb IgG alone or Rb IgG preincubated with α-MSH. During 2 days after injections, food intake was increased in both groups of rats receiving Rb IgG. However, during following 4 days when food was restricted to 1 h/day, only the Rb IgG group displayed higher food intake. Furthermore, after the refeeding, 24 h food intake was lower in rats receiving Rb IgG - α-MSH immune complexes. This group of rats was also characterized by higher number of immunopositive neurons in the arcuate nucleus expressing α-MSH and agouti-related protein but not tyrosine hydroxylase. Taken together, these results show that Rb anti-α-MSH antisera, although efficient for immunohistochemical detection of α-MSH, does not always display α-MSH blocking properties but, in contrast, may enhance α-MSH binding to MC4R and increase α-MSH anorexigenic effects when presented as immune complexes with the peptide.     

Developmental expression and distribution of GABA(A) receptor α1-, α3- and β2-subunits in pig brain

The principal function of the γ-aminobutyric acid (GABA) system in the adult brain is inhibition; however, in the neonatal brain, GABA provides much of the excitatory drive. As the brain develops, transmembrane chloride gradients change and the inhibitory role of GABA is initiated and continues throughout juvenile and adult life. Previous studies have shown that GABA(A) receptor subunit expression is developmentally regulated, and it is thought that the change in GABA function from excitation to inhibition corresponds to the changeover in expression of 'immature' to 'mature' subunit isoforms. We examined the protein expression pattern and distribution of GABA type A (GABA(A)) receptor α1-, α3- and β2-subunits in the parietal cortex and hippocampus of the developing piglet brain. Four perinatal ages were studied; 14 days preterm (P-14), 10 days preterm (P-10), day of birth (P0) and at postnatal day 7 (P7). Animals were obtained by either caesarean section or spontaneous birth. Protein expression levels and subunit localization were analysed by Western blotting and immunohistochemistry, respectively. In the cortex and hippocampus, GABA(A) receptor α1-subunit showed greatest expression at P7 when compared to all other age groups (p < 0.05). In contrast, α3 expression in the cortex was elevated in preterm brain, peaking at P0, followed by a significant reduction by P7 (p < 0.05); a similar trend was observed in the hippocampus. GABA(A) receptor β2-subunit protein expression appeared relatively constant across all time points studied in both the cortex and hippocampus. Immunolabelling of the α1-, α3- and β2-subunits was observed throughout all cortical layers at every age. GABA(A) receptor α3-subunit appeared to show specific localization to layers V and VI whilst labelling for the β2-subunit was observed in layer IV. In the hippocampus of all animals, the α1- and β2-subunits were shown to immunolabel various cells and processes in the dentate gyrus (DG), CA1 and CA3; the α3-subunit was barely observed except at the stratum moleculare of the DG. We report for the first time the ontogenesis of GABA(A) receptor subunits α1, α3 and β2 in the perinatal pig brain.     

Involvement of 5-HT(2A) receptor and α(2)-adrenoceptor blockade in the asenapine-induced elevation of prefrontal cortical monoamine outflow

The psychotropic drug asenapine is approved for the treatment of schizophrenia and manic or mixed episodes associated with bipolar I disorder. Asenapine exhibits higher affinity for several 5-HT receptors and α(2)-adrenoceptors than for D(2) receptors. Noteworthy, blockage of both the 5-HT(2A) and α(2)-adrenergic receptors has been shown to enhance prefrontal dopamine release induced by D(2) receptor antagonists. Previous results show that asenapine, both systemically and locally, increases dopamine, noradrenaline, and serotonin release in the medial prefrontal cortex (mPFC), and that the increased dopamine release largely depends on an intracortical action. Using reverse microdialysis in freely moving rats, we here assessed the potency of low concentrations of asenapine to cause a pharmacologically significant blockage in vivo of 5-HT(2A) receptors and α(2)-adrenoceptors within the mPFC, and thus its ability to affect cortical monoamine release by these receptors. Intracortical administration of 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI), a 5-HT(2A/2C) receptor agonist, increased cortical monoamine release, effects that were antagonized both by asenapine and the selective 5-HT(2A) antagonist M100907. Application of clonidine, an α(2)-adrenoceptor agonist, significantly reduced monoamine release in the mPFC. The selective α(2)-adrenoceptor antagonist idazoxan blocked, whereas asenapine partially blocked clonidine-induced cortical dopamine and noradrenaline decrease. The effects of asenapine and idazoxan on clonidine-induced serotonin decrease were less pronounced. Our results propose that low concentrations of asenapine in the mPFC exhibit a pharmacologically significant 5-HT(2A) and α(2) receptor antagonistic activity, which may contribute to enhance prefrontal monoamine release in vivo and, secondarily, its clinical effects in schizophrenia and bipolar disorder.     

The causes of the syndromes of Landry (1859) and of Guillain, Barré and Strohl (1916)

Post-infectious and post-vaccinal peripheral neuritis or encephalomyelitis have frequently been considered the human equivalents of experimental allergic neuritis (EAN) or encephalomyelitis (EAE). The major basis for these comparisons between diseases in humans and experimental animals rests on the classical observations of "paralytic accidents" of Pasteur-type vaccination against rabies. These old observations in humans injected with brain tissue indicate a remarkable heterogeneity of periphéral as well as central nervous system syndromes, quite in contrast with the remarkable specificity for either peripheral or central involvement in most experimental animals. The syndromes of Landry (1859) and of Guillain, Barré and Strohl (1916) differ clinically and pathologically, the latter a purely peripheral neuritis and the former a mixture of transverse myelitis and facial neuritis. Each can be caused by many different factors, including 1) direct infection by wild or attenuated rabies virus, 2) direct auto-sensitization by myelin antigens in the vaccine and 3) indirect or cross-reactive sensitization by viral or bacterial antigenic determinants (epitopes) with sufficient chemical homology with aminoacid sequences in central or peripheral myelin antigens to be recognized as immunological homologies.     

Do ACE (rs4646994) and αADDUCIN (rs4961) gene polymorphisms predict the recurrence of hypertensive intracerebral hemorrhage?

This study was undertaken to evaluate the role of ACE and αADDUCIN polymorphisms in patients with recurrent and nonrecurrent hypertensive intracerebral hemorrhage (ICH). A total of 101 nonrecurrent and 33 recurrent hypertensive ICH patients underwent an ACE (rs4646994) and αADDUCIN (rs4961) gene polymorphism study. The risk factors, clinical findings, CT scan abnormalities and functional outcome of recurrent and nonrecurrent ICH were compared. ACE (rs4646994) and αADDUCIN (rs4961) gene polymorphisms were also compared in the two groups and with 198 controls. The patients with recurrent ICH were older compared to those with nonrecurrent ICH and the other stroke risk factors were found in the two groups. Ganglionic-ganglionic pattern of recurrence was the commonest (75.6%) and all had at least one ICH in the location of hypertensive ICH. ACE DD genotype (OR6.18, 95%CI 2.93-13.02) and D allele (OR 2.43, 95%CI 1.70-3.47) were associated with nonrecurrent ICH compared to controls. In patients with recurrent ICH, DD genotype (OR 7.46, 95%CI 2.8-19.4) and D allele (OR 3.16, 95%CI 1.83-5.46) of ACE, and GW (OR 3.49, 95%CI 1.47-8.28), WW (OR 2.9, 95%CI 1.40-4.30) genotypes and W allele (OR 7.46, 95%CI 2.80-19.40) of αADDUCIN were more frequent compared to controls. Recurrent ICH also had higher frequency of WW genotype (OR 9.43, 95%CI 1.49-59.50) and W allele (OR 2.19, 95%CI 1.11-4.03) compared to nonrecurrent ICH. The frequency of DD?+?WW (P?=?0.008) and DD/WW?+?ID/GW (P?=?0.0001) genotypes in the recurrent ICH was higher than in the nonrecurrent ICH and the controls. Variant genotype combinations of ACE and αADDUCIN render the hypertensive patient more vulnerable to recurrent ICH.     

Is brain perfusion a differentiating feature in the comparison of Progressive Supranuclear Palsy Syndrome (PSPS) and Corticobasal Syndrome (CBS)?

The aim of this work is to present whether SPECT ^99mTc-HMPAO can be a method of examination to possibly differentiate the syndromes. 21 patients with PSP syndrome and 14 patients with corticobasal syndrome (CBS) were examined using SPECT ^99mTc-HMPAO. Perfusion single photon emission computed tomography (SPECT) as a method of examination of progressive supranuclear palsy (PSP) and corticobasal syndrome (CBS) has not been extensively analyzed in contemporary literature. Recent criteria of PSP and CBS do not describe this method of examination as primary or additional. Extended analysis was done in the context of regions of interest affected by significant average hypoperfusion (>2 standard deviations). Differences between the syndromes were subsequently evaluated using the U-Mann-Whitney test. Lack of significant differences (p < 0.05) were observed in 92 out of 94 regions of interest. However, certain asymmetries were observed in a minority of regions in both of the syndromes. Additionally, the authors of the study verified possible differences of asymmetry of perfusion of both of the syndromes. The overlapping of clinical manifestations and locations of hypoperfusion leads to a question of whether the syndromes should be interpreted as separate entities or variants of the same disease.     

Changes in rates,methods and characteristics of suicide attempters over a 15-year period: comparison between Stockholm,Sweden, and Würzburg,Germany

Purpose  

To investigate age and sex-specific changes in rates, methods used and characteristics of suicide attempters receiving medical care, over a 15-year period in two European WHO catchment areas (Stockholm, Sweden; and Würzburg, Germany).     

Light and electron microscopic studies of “nude” mice CNS after subcutaneous administration of the E variant of the encephalomyocarditis (EMC) virus

Summary Sixteen 3 month old nude mice, 24 of their litter mates and 30 Swiss mice were injected subcutaneously with 0.1 ml suspension of the E variant of the encephalomyocarditis (EMC) virus. While the mortality rate of the litter mates and Swiss mice during 5–7 days after inoculation was more than 40%, none of the nude mice died during the experiment. The surviving animals were sacrificed at 24 h intervals from day one to seven days after injection. Brain suspensions assayed for the presence of the virus yielded significant titers at 24 h in all groups, which increased during 7 days. The litter mates and Swiss mice showed proliferation of lymphocytes and microglial cells in the perivascular areas of the brain during the fifth to the seventh day. The nude mice, on the other hand, displayed no perivascular lymphocytic infiltration during the same periods. Ultrastructurally, all groups showed aggregates of ribosomes in the cytoplasmic matrix on the third day, which became enlarged in size on the 5th day. At 7 days, both litter mates and Swiss mice showed an increased number of necrotic cells, while these changes were not observed in the nude mice. These findings suggest that the high mortality rate in immunologically normal mice was related to the efforts of T cells to eliminate virus-infected cells and to produce extensive necrosis, while T cell-depleted animals showed good survival rates.