Altered neuropeptide Y Y1 responses in mesenteric arteries in rats with congestive heart failure

The aim of the present study was to elucidate if the potentiating effect of neuropeptide Y on various vasoactive agents in vitro is (1) altered in mesenteric arteries from rats with congestive heart failure and (2) mediated by the neuropeptide Y Y₁ receptor. The direct vascular effects of neuropeptide Y and its modulating effects on the contractions induced by endothelin-1-, noradrenaline-, 5-hydroxytryptamine (5-HT)-, U46619-(9, 11-dideoxy-11, 9-epoxymethano-prostaglandin F₂) and ATP, and acetylcholine-induced dilatations were studied in the presence and absence of the neuropeptide Y Y₁ antagonist, BIBP3226 (BIBP3226{(R)-N2-(diphenylacetyl)-N-[(4-hydroxyphenyl)methyl]- -arginine-amide}). Neuropeptide Y, per se, had no vasoactive effect in the arteries. The potency of endothelin-1 was significantly decreased in congestive heart failure rats. Neuropeptide Y and neuropeptide Y-(13–36) potentiated the endothelin-1-induced contraction in congestive heart failure mesenteric arteries. In 20% of the congestive heart failure rats, sarafotoxin 6c induced a contraction of 31±4%. Neuropeptide Y also potentiated U46619- and noradrenaline-induced contractions but not 5-HT-induced contractions in congestive heart failure arteries. In sham-operated animals neuropeptide Y potentiated noradrenaline- and 5-HT-induced contractions. These potentiations were inhibited by BIBP3226. Acetylcholine induced an equipotent relaxation in both groups which was unaffected by neuropeptide Y. In conclusion, neuropeptide Y responses are altered in congestive heart failure rats. The potentiating effect differs between vasoactive substances. Neuropeptide Y Y₁ and non-neuropeptide Y₁ receptors are involved.     

Correlation between soluble transferrin receptor and serum ferritin levels following bone marrow transplantation for thalassemia

Abstract: This study analyzes the serum transferrin receptor (sTfR) levels in a series of 230 ex-thalassemics with a follow-up of 1 to 9 years after bone marrow transplantation (BMT) for homozygous β thalassemia. Ex-thalassemics are individuals, cured of homozygous β thalassemia by BMT, who maintain different degrees of iron overload acquired during the pretransplant period. Both in experimental and clinical conditions, sTfR concentrations have been shown to be a quantitative measure of body iron status. This study was carried out to assess whether the level of sTfR may be of help in determining the extent of iron overload in ex-thalassemics. Patients who received the marrow from their HLA-identical sibling donor heterozygous for β thalassemia, namely heterozygous ex-thalassemics, displayed significantly higher levels of sTfR than patients transplanted from their normal sibling donors (normal ex-thalassemics). This finding suggests that increased erythropoiesis, albeit in part ineffective in heterozygous ex-thalassemics, is responsible for the sTfR increment. Both heterozygous and normal ex-thalassemics had significant lower sTfR levels than their heterozygous (p < 0.003) or normal (p < 0.0001) donors, respectively. These differences may be ascribed to the presence of iron overload in ex-thalassemics in comparison to their normal or heterozygous donors who did not present excess of iron in the body. A significant inverse correlation between sTfR and serum ferritin levels (r = –0.54, p < 0.0001) was found when normal ex-thalassemics were considered. In heterozygous ex-thalassemics, the lack of correlation between these two parameters may be explained by the enhanced erythropoietic activity of individuals with thalassemic trait. These results suggest that the level of sTfR may be a useful indicator of iron overload in normal ex-thalassemics.     

Inhibition of ligand binding to g protein-coupled receptors by arachidonic acid

Arachidonic acid (AA), released in response to muscarinic acetylcholine receptor (mAChR) stimulation, previously has been reported to function as a reversible feedback inhibitor of the mAChR. To determine if the effects of AA on binding to the mAChR are subtype specific and whether AA inhibits ligand binding to other G protein-coupled receptors (GPCRs), the effects of AA on ligand binding to the mAChR subtypes (M₁, M₂, M₃, M₄, and M₅) and to the μ-opioid receptor, β₂-adrenergic receptor (β₂-AR), 5-hydroxytryptamine receptor (5-HTR), and nicotinic receptors were examined. AA was found to inhibit ligand binding to all mAChR subtypes, to the β₂-AR, the 5-HTR, and to the μ-opioid receptor. However, AA does not inhibit ligand binding to the nicotinic receptor, even at high concentrations of AA. Thus, AA inhibits several types of GPCRs, with 50% inhibition occurring at 3–25 μM, whereas the nicotinic receptor, a non-GPCR, remains unaffected. Further research is needed to determine the mechanism by which AA inhibits GPCR function.     

Diabetes alters μ and κ opioid binding in rat brain regions: comparison with effects of food restriction

Diabetic rats display changes in opioid pharmacology and brain regional levels of opioid peptides and prodynorphin mRNA. Previous investigations of opioid receptor binding, carried out in whole-brain homogenates, have, however, failed to detect changes. In the present study, quantitative autoradiography was used to measure μ and κ opioid receptor binding in discrete brain regions of streptozotocin-treated diabetic rats. Measurement was limited to regions that previously displayed opioid binding changes in chronically food-restricted rats, since our primary aim is to identify brain mechanisms that mediate adaptive responses to persistent metabolic need and adipose depletion. Diabetics displayed strong trends or statistically significant changes which matched seven of the thirteen binding changes observed in food-restricted rats. In no case did diabetics display changes in the opposite direction. The two statistically significant changes common to food-restricted and diabetic rats are increased κ binding in the medial preoptic area and decreased μ binding in the lateral habenula. The possible functional significance of these changes is discussed.     

Inhibition and superactivation of the calcium-stimulated isoforms of adenylyl cyclase

It was shown previously that chronic exposure to opiate agonists increases adenylyl cyclase (AC) activity, a phenomenon termed AC superactivation (or supersensitization). More recently, we showed that acute G_i/o-coupled receptor activation inhibits the activity of several AC isozymes, including Ca²⁺/calmodulin-stimulated AC-I and -VIII, whereas chronic receptor activation induces their superactivation. Here, we report that both acute μ-opioid receptor-induced inhibition and chronic induced superactivation of AC-I and -VIII are pertussis toxin sensitive. In addition, we show that proteins that interfere with the activity of {ie195-2} subunits ({ie195-3} scavengers) strongly attenuate the acute inhibition of AC-I and -VIII and the superactivation of AC-I, and abolish the superactivation of AC-VIII. Based on these results, we suggest that {ie195-4} is involved in the acute inhibition and chronic agonist-induced superactivation of AC types I and VIII.     

Block of the endplate acetylcholine receptor channel by the sympathomimetic agents ephedrine, pseudoephedrine, and albuterol

Recent observations suggest that some patients with congenital myasthenic syndromes respond favorably to ephedrine, pseudoephedrine, or albuterol. Conventional microelectrode studies, however, provide no clear explanation for a beneficial effect of ephedrine in endplate diseases. To gain further insight into how these drugs affect neuromuscular transmission, we investigated their effects on the kinetic properties of the acetylcholine (ACh) receptor. Single channel currents were recorded from rat lumbrical muscles endplates using low concentrations of ACh and 2.5–100 μM of drugs. Between 10–100 μM, each drug progressively increased the rate of channel closure in a concentration dependent manner, consistent with an open-channel block. Albuterol acted as a sequential fast-acting channel blocker, increasing the mean burst duration in a concentration dependent manner without altering the total open time per burst or the duration of intraburst blockages. Increasing concentrations of ephedrine and pseudoephedrine also increased the number of intraburst closures but decreased the total open time per burst. None of the drugs altered single channel conductance. The channel blocking effects of ephedrine and pseudoephedrine might reduce the synaptic overactivity that occurs in the slow-channel myasthenic syndromes or in endplate ACh esterase deficiency, but these effects occur at concentrations not attainable in clinical practice.     

Hippocampal NMDA receptor mRNA undergoes subunit specific changes during developmental lead exposure

The N-methyl- -aspartate (NMDA) receptor has shown to play an important role in the cognitive deficits associated with developmental lead (Pb) exposure. In this study, we examined the effects of low-level Pb exposure on NMDA receptor subunit gene expression in the developing rat brain. The pattern of NR1, NR2A, NR2B, and NR2C subunit mRNA in situ hybridization was consistent with previous studies. Brain levels of NR1 and NR2A mRNAs were lowest shortly after birth, increasing to reach peak levels by 14 or 21 days of age and subsequently decreasing at 28 days of age. NR2B mRNA levels were highest during early development and decreased as the animals aged. NR2C subunit mRNA was restricted to the cerebellum and a signal was not detectable until the second week of life. Lead exposure resulted in significant and opposite effects in NR1 and NR2A subunit mRNA expression with no changes in NR2B or NR2C subunit expression. The Pb-induced changes in NR1 and NR2A subunit mRNA were mainly present in the hippocampus. Hippocampal NR1 mRNA levels were significantly increased in the CA1 (15.3%) and CA4 (26.8%) pyramidal cells from 14-day-old Pb-exposed rats. At 21 days of age, only the NR1 mRNA at the CA4 subfield remained significantly elevated (10.3%). Lead exposure caused reductions of NR2A mRNA levels (11.9–19.3%) in the pyramidal and granule cell layers of the hippocampus at 14 and 21 days of age. NR1 mRNA levels were also significantly increased (14.0%) in the cerebellum of 28-day-old rats with no change in NR2A mRNA at any age. No significant changes in subunit mRNA levels were present in cortical or subcortical regions at any age. The Pb-induced changes in hippocampal NMDA receptor subunit mRNA expression measured in the present study may lead to modifications in receptor levels or subtypes and alter the development of defined neuronal connections which require NMDA receptor activation.     

Differential changes of nicotinic receptors in the ratbrain following ibotenic acid and 192-IgG saporin lesionsof the nucleus basalis magnocellularis

The basal forebrain cholinergic neurons are implicated in the pathogenesis ofneurodegenerative diseases including Alzheimer^fn2s disease (AD). The nicotinic acetylcholine receptors (nAChRs) have been found to besignificantly afflicted in AD. To study the underlying mechanisms for dysfunction of the basalforebrain cholinergic neurons development of suitable animal models is warranted. In this studywe investigated the effects of bilateral lesions of the nucleus basalis magnocellularis on nAChRs inthe rat brain using the cholinergic system selective immunotoxin 192-IgG saporin andnon-selective excitotoxin ibotenic acid. Changes in nAChRs were measured by ³H-cytisineand ³H-epibatidine, two ligands with different selectivity for nAChRs subtypes. Inthe parietal cortex of ibotenic acid lesioned rates, the choline acetyltransferase activity (ChAT)was decreased by 24% while no changes were detected in the frontal cortex or hippocampus.Similarly, a 40% decrease was observed in the number of nAChRs labelled by ³H-cytisine,but not by ³H-epibatidine, in the parietal cortex, while no changes were found in thefrontal cortex or hippocampus. Although the 192-IgG saporin induced lesions reduced the ChATactivity in the frontal cortex, parietal cortex and hippocampus by 77, 50 and 21%, respectively, nochanges were observed in the number of nAChRs as studied by ³H-cytisine or ³H-epibatidine. The results indicate a difference in vulnerability of the cortical nAChRsubtypes to experimental lesions of the nucleus basalis magnocellularis. The findings in this studysuggest that a major portion of the nAChRs might be located on non-cholinergic neurons in thebrain.     

Blockade of K+ channels induced by AMPA/kainate receptor activation in mouse oligodendrocyte precursor cells is mediated by NA+ entry

AMPA/kainate receptor activation in cultured oligodendrocyte precursor cells from embryonic mouse cortex leads to a blockade of delayed rectifying K⁺ currents. In the present study, we provide evidence using the patch-clamp technique in the whole-cell configuration that the mechanism linking kainate receptor activation and K⁺ conductance blockade is due to the receptor-mediated Na⁺ entry: (1) The blockade was not observed in Na⁺ -free bathing solution nor when intracellular [Na⁺] was elevated by dialzying the cell with a pipette solution containing high [Na⁺]. (2) Elevation of intracellular [Na⁺] alone led to a blockade of outward currents in contrast to cells dialyzed by sucrose. High [Li⁺]_i also reduced the outward currents, and in Li⁺-containing bathing solution the kainate-induced blockade of K⁺ channels was more pronounced. Probably, Li⁺ accumulates intracellularly after permeation through the receptor pore due to slower extrusion mechanisms. Experiments with GTPγS or GDPβS and pertussis toxin indicated that GTP-binding protein-mediated mechanisms were not of importance for the kainate-induced K⁺ conductance blockade. Our data suggest that in glial precursor cells AMPA/kainate receptor activation leads to an intracellular [Na⁺] increase which blocks delayed rectifying K⁺ channels. © 1995 Wiley-Liss, Inc.     

Chlorocarbamate-mustard-linked 1,1,2-triphenylbut-1-enes with a selective antitumor activity on mammary tumors containing estrogen receptors

Summary A 1,1,2-triphenylbut-1-ene with a 4-OH group at one C-1 phenyl ring and a chlorocarbamate mustard moiety at the second C-1 ring (compound 3) was synthesized in order to obtain a cytotoxic estrogen with a specific antitumor effect on estrogen-receptor-containing tumors. This compound was tested in comparison to the carrier (compound 1) and a compound (2) having a carbamate mustard group on both C-1 phenyl rings. The estrogen receptor affinity of compound 3 was only about one-quarter lower than that of compound 1, but much higher than that of compound 2. Compounds 2 and 3 showed only partially irreversible binding to the receptor owing to their relatively low alkylating properties. The growth inhibition of the receptor-positive MCF-7 breast cancer cell line by compound 3, but not by compound 1 or 2, was more pronounced than the inhibition of the receptor-negative line MDA. In vivo the hormone-dependent, transplantable mammary tumor MXT M3.2 of the mouse was much better inhibited by compound 3 than its hormone-resistent line MXT OVEX. Compounds 1–3 had no antiestrogenic properties in the mouse, but estrogenic activity was in the order 1>3>2. From these results and because the antitumor activity of compound 3 was superior to that of compounds 1 and 2 in the hormone-dependent tumor models, a selective, receptor-mediated cytotoxic effect of compound 3 on estrogen-receptor-positive tumors in obvious.Supported by the Deutsche Forschungsgemeinschaft, SFB 234 and the Matthias Lackas StiftungDedicated to Prof. Dr. M. F. El Etreby on the occasion of his 50th birthday