Neonatal treatment with 192 IgG-saporin produces long-term forebrain cholinergic deficits and reduces dendritic branching and spine density of neocortical pyramidal neurons

The role of basal forebrain-derived cholinergic afferents in the development of neocortex was studied in postnatal rats. Newborn rat pups received intraventricular injections of 192 IgG-saporin. Following survival periods ranging from 2 days to 6 months, the brains were processed to document the cholinergic lesion and to examine morphological consequences. Immunocytochemistry for choline acetyltransferase (ChAT) and in situ hybridization for ChAT mRNA demonstrate a loss of approximately 75% of the cholinergic neurons in the medial septum and nucleus of the diagonal band of Broca in the basal forebrain. In situ hybridization for glutamic acid decarboxylase mRNA reveals no loss of basal forebrain GABAergic neurons. Acetylcholinesterase histochemistry demonstrates a marked reduction of the cholinergic axons in neocortex. Cholinergic axons are reduced throughout the cortical layers; this reduction is more marked in medial than in lateral cortical areas. The thickness of neocortex is reduced by approximately 10%. Retrograde labeling of layer V cortico-collicular pyramidal cells reveals a reduction in cell body size and also a reduction in numbers of branches of apical dendrites. Spine densities on apical dendrites are reduced by approximately 20-25% in 192 IgG- saporin-treated cases; no change was detected in number of spines on basal dendrites. These results indicate a developmental or maintenance role for cholinergic afferents to cerebral cortical neurons.      

Association study of Glutathione S-transferase P1 (GSTP1) with asthma and bronchial hyper-responsiveness in two German pediatric populations

Glutathione S-Transferase P1 ( GSTP1 ) is an important enzyme in the detoxification of products of oxidative stress. Several studies have shown an association of the amino acid variant Ile105Val with bronchial asthma and the reaction of the lung to inhalant pollutants. The aim of this study was to test the two known amino acid variants in GSTP1 for association with bronchial asthma and airway hyper-responsiveness in two German pediatric populations. We genotyped Ile105Val and Ala114Val in the Multicenter Allergy Study cohort (85 children with asthma, 123 controls) and asthmatic children from Freiburg (n = 178). We did not find association of either polymorphisms with bronchial asthma or airway hyper-responsiveness. We conclude from our data that polymorphisms within GSTP1 do not play a major role in the development of bronchial asthma in German children.     

No association of histamine- N-methyltransferase polymorphism with asthma or bronchial hyperresponsiveness in two German pediatric populations

Histamine plays an important role in the allergic inflammation. Histamin N-Methyltransferase (HNMT) catalyses the major pathway of histamine metabolism in the human lung. A common functional single nucleotide polymorphism (SNP) within the HNMT gene (C314T) was recently related to asthma. We tested this SNP for associations with asthma and asthma associated traits in two German pediatric populations (1. MAS-cohort, n=888, 85 children with asthma; 2. asthmatic children from Freiburg, n=176). Non-asthmatic (n=515) and non-atopic (n=211) children from the MAS-cohort were used as controls. For genotyping melting curve analyses (Light Cycler System) were applied. In contrast to a previous study, no association of the HNMT 314T allele with asthma, bronchial hyperresponsiveness (BHR) or other asthma related phenotypes could be observed in either study population. We conclude that this SNP might not play a major role in the pathogenesis of asthma or BHR in German children.     

ACUTE BUT NOT CHRONIC ANGIOTENSIN-CONVERTING ENZYME INHIBITION INDUCES ENZYME SYNTHESIS IN THE GLOMERULUS OF THE SPONTANEOUSLY HYPERTENSIVE RAT

1. Treatment with angiotensin-converting enzyme (ACE) inhibitors slows the rate of progression of nephropathy in the spontaneously hypertensive rat (SHR) with streptozotocin-induced diabetes. Paradoxically, however, chronic ACE inhibitor therapy has been reported to be associated with induction of ACE in the plasma. We sought to determine whether induction also occurred in the glomerulus. 2. Seven days after induction of diabetes rats were randomized to receive perindopril (4mg/kg per day) in the drinking water or water alone. Blood glucoses were maintained 6–10 mmol/L by daily ultralente insulin. Rats were killed after 1 and 12 weeks of ACE inhibitor therapy and the kidneys were harvested. Angiotensin-converting enzyme activity was determined in isolated glomeruli before and after removal of perindopril and reconstitution with zinc sulphate. 3. After 1 week of ACE inhibitor therapy, glomerular ACE was significantly greater after removal of perindopril than either before its removal (P < 0.025) or in the untreated controls (P < 0.025). After 12 weeks of therapy, ACE activity was significantly lower in the perindopril-treated group than in the untreated controls (P < 0.025). There was no increase in ACE activity following removal of perindopril. 4. These studies suggest that short-term ACE inhibition is associated with induction of ACE in the glomerulus. However, there was no increase in ACE activity after removal of perindopril, suggesting that induction of synthesis of this enzyme in the glomerulus does not occur during chronic ACE inhibition.     

EVIDENCE FOR DIRECT INTERACTION OF KETAMINE WITH α- AND β2-ADRENOCEPTORS

1. Ketamine has a number of effects that suggest that it may interact with α- and β-adrenoceptors. To date, the experimental evidence for this has been indirect and has been based on physiological studies using competitive blocking agents. In the present study we sought to determine from receptor binding studies whether ketamine binds directly to α- and β-adrenoceptors. 2. Membrane preparations o. α₁- and β₂-adrenergic binding sites were obtained from urinary bladder and urethrae of sheep. These binding sites were characterized by saturation analyses using [³H]-prazosin for α₁-adrenoceptor binding sites and [¹²⁵I]-cyanopindolol (CYP) for the β₂-adrenoceptor binding sites. The receptors were further characterized by displacement studies using selective and non-selective antagonists. 3. Studies in which ketamine was used to displac. [³H]-prazosin revealed a K_d of 3.40±1.23× 10^?3 mol/L for ketamine binding to ai-adrenoceptors. Displacement studies of [¹²⁵I]-CYP by ketamine showed a K_d of 0.35±0.03× 10^?3 mol/L for ketamine binding to β₂-adrenoceptors. 4. We conclude that ketamine interacts directly with both ai- an. β₂-adrenoceptors and that such interactions probably explain the reported effects of this agent on the vasculature and the bronchial tree.     

Transfected leukocyte integrin CD11b/CD18 (Mac-1) mediates phorbol ester-activated, homotypic cell:cell adherence in the K562 cell line

The CD11b/CD18 leukocyte integrin molecule mediates diverse neutrophil adherence-related functions, including cell:cell and cell:extracellular matrix attachments. To study the individual role of this leukocyte integrin in cell adherence in hematopoietic cells, we expressed the CD11b/CD18 complex on the surface of K562 cells, a cell line derived from an individual with chronic myelogenous leukemia in blast crisis. We used an amphotrophic retroviral vector designated LCD18SN, harboring the complete coding sequence for the CD18 subunit, to transfer the CD18 cDNA into K562 cells and select stable cell lines. The CD11b subunit in the expression plasmid pREP4 was transfected into these K562/CD18 cells by electroporation and stable cell clones were selected. These K562 cells possessed RNA and intracellular protein for each subunit, and they expressed the CD11b/CD18 heterodimer on the cell surface. When CD11b/CD18 expressing K562 cells were stimulated with phorbol myristate acetate (50 ng/mL) for 24 to 48 hours, these K562 cells formed dense cell:cell aggregates. This homotypic aggregation required both activation of the CD11b/CD18 complex and the induction of the counter- receptor for CD11b/CD18 on the conjugate cell. This cell line will (1) enable the structure-function relationships between cell activation and homotypic adherence to be assessed, (2) provide the opportunity to identify accessory molecules required for activation of the CD11b/CD18 complex, and (3) facilitate the identification of novel ligands for the CD11b/CD18 complex.     

Calculation of signal intensities in hybrid sequences for fast NMR imaging

A number of techniques that recently have been used for fast NMR-imaging are based on a hybrid sequence of echo planar imaging (EPI) and FLASH imaging: after each NMR excitation several k-space lines are measured. The complete k-space is covered by performance of several excitations. It has been observed that there is usually an optimal hybrid sequence that maximizes the signal-to-noise ratio. In this work, a method is presented that allows a determination of the optimal sequence as a function of the relaxation times T₁ and T₂^*.