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We have found that several types of cultured mammalian cells, including both normal and transformed human, rat, and mouse cell lines, have an active transport system for a diverse group of structurally related compounds possessing an... more
We have found that several types of cultured mammalian cells, including both normal and transformed human, rat, and mouse cell lines, have an active transport system for a diverse group of structurally related compounds possessing an amine group and various types of aromatic ring structures. Ligands such as the beta-adrenergic antagonists (-)-[3H] dihydroalprenolol (DHA), (-)-[3H]propranolol, and (-)-[125I] iodocyanopindolol, and the tricyclic antidepressant [3H]imipramine, which are used to assess cell surface receptors for catecholamines and serotonin, appear to be actively transported into cells rather than simply bound to cell surface sites or accumulated by passive diffusion. DHA transport was competed by many structurally related amines including imipramine and certain alpha-and beta-adrenergic ligands, but not by catecholamines or serotonin. Ligand transport in HeLa cells was saturable at micromolar levels, selective, nonstereospecific, temperature- and pH-dependent, and sensitive to the ionophore monensin and the amine transport inhibitor reserpine, thus indicating dependence on a carrier system driven by a transmembrane proton gradient. In C6 glioma cells, amine transport was clearly distinguishable from beta-adrenergic receptor binding which could be measured with the recently developed hydrophilic beta-blocker (+/-)-[3H] 4-(3-tertiarybutylamino-2-hydroxy-propoxy)-benzimidazole-2-on hydrochloride (CGP-12177); binding of this ligand met rigorous pharmacological criteria, was not influenced by monensin or reserpine, and, therefore, did not appear to be transported. Membrane vesicles from HeLa and C6 cells transported DHA but not CGP-12177 via a MgATP-dependent mechanism which was inhibited by N,N'-dicyclohexylcarbodiimide, monensin, and reserpine, indicating a carrier system driven by a proton gradient maintained by a proton-pumping ATPase.
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Platelet-activating factor (PAF) is an unusually potent lipid autacoid with a variety of biological activities. The growing body of evidence suggests that PAF might play an important role in modulation of central nervous system function,... more
Platelet-activating factor (PAF) is an unusually potent lipid autacoid with a variety of biological activities. The growing body of evidence suggests that PAF might play an important role in modulation of central nervous system function, particularly during ischemia- and trauma-induced neuronal damage. However, the mechanisms involved in PAF actions on neuronal or other brain cells is virtually unknown. Therefore, this study was designed to characterize PAF receptor-mediated cellular signal transduction in neurohybrid NG108-15 cells with the aid of a new potent PAF antagonist, BN 50739. PAF induced an immediate and concentration-dependent increase in [Ca++]i with an EC50 of 6.8 nM. PAF-induced [Ca++]i mobilization was inhibited by several structurally unrelated PAF antagonists such as BN 50739, WEB 2086, SRI 63-441 and BN 52021, in a dose-dependent manner with IC50 values of 4.8, 6.9, 809 and 98500 nM, respectively. The calcium channel blockers nifedipine (5 microM) and diltiazem (10 microM) had no effect on the PAF-induced increase in [Ca++]i, but omission of CA++ from the incubation buffer caused an 82% reduction of PAF-induced [Ca++]i elevation; the remainder contributed from intracellular sources was completely inhibited by 10 microM TMB-8, an intracellular Ca++ blocker. NG108-15 cells exhibited homologous desensitization to sequential addition of PAF, but no heterologous desensitization between PAF and other agonists such as bradykinin, endothelin, angiotensin II and ATP was observed. PAF stimulated phosphoinositide metabolism in a dose-dependent manner with an EC50 of 5.1 nM for IP3 formation, which was also inhibited by the PAF antagonist BN 50739 in a dose-dependent manner (IC50 = 3.6 nM).(ABSTRACT TRUNCATED AT 250 WORDS)
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Platelet-activating factor (1-O-hexadecyl-2-acetyl-sn-glycero-3-phosphorylcholine) is a potent lipid autacoid produced by many cell types. Platelet-activating factor is produced by cerebellar granule cells in culture and has been... more
Platelet-activating factor (1-O-hexadecyl-2-acetyl-sn-glycero-3-phosphorylcholine) is a potent lipid autacoid produced by many cell types. Platelet-activating factor is produced by cerebellar granule cells in culture and has been extracted from brain tissue. Multiple platelet-activating factor receptors have been demonstrated in brain tissue. Activation of platelet-activating factor receptors in transformed neuronal cell lines involved increases in intracellular calcium. Platelet-activating factor has potent actions on cerebral vessels and cerebral metabolism when administered in vivo, but may not have direct effects on brain microvessels. Excessive platelet-activating factor production in pathological states of the nervous system such as neurotrauma and stroke has been shown in only a few models (e.g., spinal cord ischemia and reperfusion or focal repercussion brain injury). In multiple studies using highly specific and potent platelet-activating factor antagonists, reversal or prevention of key consequences of brain injury such as hypoperfusion following ischemia, reperfusion and edema, inflammatory cell accumulation, neurologic/motor deficits, and neuronal salvage were demonstrated. This review provides and analyzes evidence in support of the role that platelet-activating factor might have in modulation of brain function and pathophysiological processes in brain ischemia and trauma.
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Research Interests: Cognitive Science, Medicine, Cell Culture, Neuroprotection, Hippocampus, and 15 moreModels, Cerebellum, Ischemia, Animals, Brain Ischemia, Neurons, Lipid peroxidation, Clinical Sciences, Excitotoxicity, Gerbil, Glutamate Receptor, carvedilol, Gerbillinae, Antihypertensive agents, and Cardiovascular medicine and haematology
Publisher Summary The endothelin family of 21-amino acid peptides represents the most potent series of known vasoconstrictors and contributes to the local regulation of vascular homeostasis. Four distinct isoforms—ET-1, ET-2, ET-3, and... more
Publisher Summary The endothelin family of 21-amino acid peptides represents the most potent series of known vasoconstrictors and contributes to the local regulation of vascular homeostasis. Four distinct isoforms—ET-1, ET-2, ET-3, and vasoactive intestinal contractor or ET-β)—which differ in amino acid substitutions, have been identified to date and show widespread distribution in mammalian tissue. Vascular endothelial cells produce only ET-1, but ET-1, ET-2, and ET-3 are present and/or active in brain. Neuropeptides function as neuromodulators or local hormones, which oversee the central nervous system regulation of cardiovascular function. In addition, endothelin is found both in posterior and anterior pituitary, regulating anterior pituitary function by eliciting inositol trisphosphate and diacylglycerol production, calcium mobilization and influx, and gonadotropin secretion. Endothelin distributes in a nonvascular pattern in brain, and high densities of endothelin-binding sites are found in the granule cell layer of the cerebellum. This chapter discusses some systems of both primary cultured cells and established neural cell lines for endothelin receptors and their biochemical significance.
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Research Interests: Pharmacology, Calcium, Medicine, Neuroprotection, Hippocampus, and 15 moreBrain, Cerebellum, Animals, Brain Ischemia, Neurons, Clinical Sciences, Neurosciences, Gerbil, Glutamate Receptor, Hippocampal formation, Kainic acid, NMDA receptor, Cell count, Gerbillinae, and Cardiovascular medicine and haematology
The effects of endothelin(ET)-1, ET-2, ET-3 and Big ET on intracellular level of Ca2+ ([Ca2+]i) were studied in neuroblastoma NG108-15 and NCB-20 cells. All ETs, except Big ET, induced an increase in [CA2+]i in NG108-15 cells in a... more
The effects of endothelin(ET)-1, ET-2, ET-3 and Big ET on intracellular level of Ca2+ ([Ca2+]i) were studied in neuroblastoma NG108-15 and NCB-20 cells. All ETs, except Big ET, induced an increase in [CA2+]i in NG108-15 cells in a dose-dependent manner, with EC50: 6.7, 11.2 and 71 nM, respectively. However, none of the ET increased [Ca2+]i in NCB-20 cells. Calcium channel blockers diltiazem or nicardipine had no effect on ET-induced increase in [CA2+]i, but extracellular Ca2(+)-depletion significantly reduced the response of NG108-15 cells to ETs. NG108-15 cells exhibited a homologous desensitization to sequential addition of ETs, but no heterologous desensitization among ET, bradykinin and PAF was observed. These data suggest that ET-induced receptor activation results in increased intracellular Ca2+ via a non voltage calcium channel mechanism and intracellular Ca2+ release.
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Publisher Summary This chapter explores that gonorrhoea is one of the most commonly reported communicable diseases. With the appearance of antibiotic-resistant strains of Neisseria gonorrhoeae (N. gonorrhoeae), which are thought to be the... more
Publisher Summary This chapter explores that gonorrhoea is one of the most commonly reported communicable diseases. With the appearance of antibiotic-resistant strains of Neisseria gonorrhoeae (N. gonorrhoeae), which are thought to be the result of an intergeneric conjugal transfer, the possible loss of efficacy of drug treatment in control of this disease has been forewarned. The development of a vaccine follows basic understanding of human immunity to gonococcal infection because gonorrhoea has no virulence factor against which the person could be easily immunized. The multifactorial nature of virulence and the mechanisms of pathogenesis are examined by an understanding of the basic biology of the gonococcus. This chapter provides a summary of the important aspects of gonococcal physiology that relate directly or indirectly to the disease process. Certain aspects of the physiology and metabolism may be as yet unrelated to pathogenesis, but they provide a base from which future research can be planned. It discusses the differences and similarities between the physiology of N. gonorrhoeae and other pathogenic and non-pathogenic Neisseria spp., where this information is available and pertinent. Knowledge of the physiology of related species may help to delineate subject areas of gonococcal physiology that require additional effort.
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2-Methoxyestradiol (2-ME) is an endogenous metabolite of estradiol-17beta and the oral contraceptive agent 17-ethylestradiol. 2-ME was recently reported to inhibit endothelial cell proliferation. The current study was undertaken to... more
2-Methoxyestradiol (2-ME) is an endogenous metabolite of estradiol-17beta and the oral contraceptive agent 17-ethylestradiol. 2-ME was recently reported to inhibit endothelial cell proliferation. The current study was undertaken to explore the mechanism of 2-ME effects on endothelial cells, especially whether 2-ME induces apoptosis, a prime mechanism in tissue remodeling and angiogenesis. Cultured bovine pulmonary artery endothelial cells (BPAEC) exposed to 2-ME showed morphological (including ultrastructural) features characteristic of apoptosis: cell shrinkage, cytoplasmic and nuclear condensation, and cell blebbing. 2-ME-induced apoptosis in BPAEC was a time- and concentration-dependent process (EC50 = 0.45 +/- 0.09 microM, n = 8). Nucleosomal DNA fragmentation in BPAEC treated with 2-ME was identified by agarose gel electrophoresis (DNA ladder) as well as in situ nick end labeling. Under the same experimental conditions, estradiol-17beta and two of its other metabolites, estriol and 2-methoxyestriol (< or =10 microM), did not have an apoptotic effect on BPAEC. 2-ME activated stress-activated protein kinase (SAPK)/c-Jun amino-terminal protein kinase in BPAEC in a concentration-dependent manner. The activity of SAPK was increased by 170 +/- 27% and 314 +/- 22% over the basal level in the presence of 0.4 and 2 microM 2-ME (n = 3-6), respectively. The activation of SAPK was detected at 10 min, peaked at 20 min, and returned to basal levels at 60 min after exposure to 2-ME. Inhibition of SAPK/c-Jun amino-terminal protein kinase activation by basic fibroblast growth factor, insulin-like growth factor, or forskolin reduced 2-ME-induced apoptosis. Immunohistochemical analysis of BPAEC indicated that 2-ME up-regulated expression of both Fas and Bcl-2. In addition, 2-ME inhibited BPAEC migration (IC50 = 0.71 +/- 0.11 microM, n = 4) and basic fibroblast growth factor-induced angiogenesis in the chick chorioallantoic membrane model. Taken together, these results suggest that promotion of endothelial cell apoptosis, thereby inhibiting endothelial cell proliferation and migration, may be a major mechanism by which 2-ME inhibits angiogenesis.
Research Interests: Biology, Drug interactions, Molecular Pharmacology, Apoptosis, Medicine, and 13 moreAngiogenesis, Signal Transduction, DNA, Animals, Vascular endothelium, Fibroblast Growth Factor, Nucleosomes, Cattle, Neurosciences, Estradiol, Mitogen- Activated Protein Kinases, Chick embryo, and Pharmacology and pharmaceutical sciences
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Research Interests: Cognitive Science, Chemistry, Biology, Calcium, Medicine, and 15 moreNeuroprotection, Hippocampus, Cerebellum, Animals, Brain Ischemia, Neurons, Clinical Sciences, Glycine, Excitotoxicity, Glutamic Acid, Glutamate Receptor, carvedilol, Gerbillinae, Antihypertensive agents, and Cardiovascular medicine and haematology
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Class A scavenger receptor (SR-A) antagonists may prevent the initiation of atherosclerosis, because a recent report found that SR-A/apolipoprotein E (apoE) double-knockout mice had 60% smaller lesions than apoE single-knockout... more
Class A scavenger receptor (SR-A) antagonists may prevent the initiation of atherosclerosis, because a recent report found that SR-A/apolipoprotein E (apoE) double-knockout mice had 60% smaller lesions than apoE single-knockout littermates. We transfected human embryonic kidney (HEK) 293 cells with SR-A type I or II receptors to find small-molecule antagonists. Uptake of 1,1'-dioctadecyl-3,3,3', 3'-tetramethylindocarbocyanine perchlorate-labeled acetylated low-density lipoprotein (DiI-AcLDL) showed that among common polyanionic ligands, polyinosine was the most potent, with an IC50 of 0.74 microgram/ml, whereas the novel compound (E)-methyl 4-chloro-alpha-[4-(4-chlorophenyl)-1, 5-dihydro-3-hydroxy-5-oxo-1-(2-thiazolyl)-2H-pyrrol-2-ylidene]benzene acetate gave an IC50 of 6.1 microgram/ml (13 microM). The novel antagonist also inhibited DiI-AcLDL uptake in cultured human peripheral and rat peritoneal macrophages with IC50 values of 21 microM and 17 microM, respectively. With [125I]AcLDL as ligand for transfected HEK 293 cells, binding/uptake and degradation at 37 degrees C for 5 h was saturable and selective. In a comparison of both types of receptor, we found no difference between the capacity of SR-AI or SR-AII for either binding or degradation. Polyinosine competed both [125I]AcLDL binding and degradation with a Ki of 1 microgram/ml, whereas the novel antagonist competed with a Ki of 19 microgram/ml (40 microM) and 8.6 microgram/ml (18 microM), respectively, for binding and degradation. Saturation binding in the presence of the ionophore monensin indicated that the novel compound behaved as a noncompetitive antagonist and perhaps as an allosteric effector. This is the first report to describe a small-molecule macrophage scavenger receptor antagonist. Utilization of this permanently transfected HEK 293 cell line will allow the identification of more potent macrophage scavenger receptor antagonists, so that their utility as therapeutics for atherosclerosis can be determined.
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The antioxidant effect of carvedilol, a new vasodilating, beta adrenoceptor blocker was studied and compared with five other beta blockers. Carvedilol rapidly inhibited Fe(++)-initiated lipid peroxidation, measured as thiobarbituric acid... more
The antioxidant effect of carvedilol, a new vasodilating, beta adrenoceptor blocker was studied and compared with five other beta blockers. Carvedilol rapidly inhibited Fe(++)-initiated lipid peroxidation, measured as thiobarbituric acid reactive substance (TBARS), in rat brain homogenate with an IC50 of 8.1 microM. Under the same conditions, the IC50 values of atenolol, pindolol propranolol, celiprolol and labetalol were over 1.0 mM. Carvedilol protected against Fe(++)-induced alpha-tocopherol depletion in rat brain homogenate with an IC50 of 17.6 microM; propranolol, celiprolol and labetalol, up to 200 microM, did not show any effect. Using dihydroxyfumarate/Fe(++)-ADP as a OH.radical generating system and 5,5-dimethyl pyrroline-N-oxide (DMPO) as a trapping agent, the characteristic DMPO-OH signals were monitored by electron paramagnetic resonance. Carvedilol dose-dependently decreased the intensity of the DMPO-OH signal, with an IC50 of 25 microM, whereas propranolol, at 500 microM, and U74500A, a 21-aminosteroid, at 100 microM, had no effect. The antioxidant effect of carvedilol mainly resides in the carbazole moiety, and the substitution of a hydroxyl group at certain positions on the phenyl ring of either carbazole or the ortho-substituted phenoxylethylamine part of carvedilol resulted in an increase in antioxidant activity. Furthermore, the protective effect of carvedilol analogs against OH.-mediated neuronal death positively correlated to their antioxidant effect. We conclude that carvedilol is a far more potent antioxidant than other commonly used beta blockers. The apparent mechanism of carvedilol's inhibition of lipid peroxidation is mainly via scavenging free radicals. This novel property of carvedilol may contribute to the known cardioprotective activity of this compound.
Research Interests: Pharmacology, Chemistry, Medicine, Antioxidants, Brain, and 14 morePubmed, Animals, Antioxidant, Plant tissue Culture Techniques, Lipid peroxidation, TBARS, Rats, Propranolol, Structure activity Relationship, Atenolol, carvedilol, Pindolol, Antihypertensive agents, and Pharmacology and pharmaceutical sciences
We have found that several types of cultured mammalian cells, including both normal and transformed human, rat, and mouse cell lines, have an active transport system for a diverse group of structurally related compounds possessing an... more
We have found that several types of cultured mammalian cells, including both normal and transformed human, rat, and mouse cell lines, have an active transport system for a diverse group of structurally related compounds possessing an amine group and various types of aromatic ring structures. Ligands such as the f3-adrenergic antagonists (-)-[3H] dihydroalprenolol (DHA), (-)-[3H]propranolol, and (-)-[‘25I]iodocyanopindolol, and the tricyclic antidepressant [3H]imipramine, which are used to assess cell surface receptors for catecholamines and serotonin, appear to be actively transported into cells rather than simply bound to cell surface sites or accumulated by passive diffusion. DHA transport was competed by many structurally related amines including imipramine and certain a-and fl-adrenergic ligands, but not by catecholamines or serotonin. Ligand transport in HeLa cells was saturable at micromolar levels, selective, nonstereospecific, temperature-and pH-dependent, and sensitive to t...
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Class A scavenger receptor (SR-A) antagonists may prevent the initiation of atherosclerosis, because a recent report found that SR-A/apolipoprotein E (apoE) double-knockout mice had 60% smaller lesions than apoE single-knockout... more
Class A scavenger receptor (SR-A) antagonists may prevent the initiation of atherosclerosis, because a recent report found that SR-A/apolipoprotein E (apoE) double-knockout mice had 60% smaller lesions than apoE single-knockout littermates. We transfected human embryonic kidney (HEK) 293 cells with SR-A type I or II receptors to find small-molecule antagonists. Uptake of 1,1'-dioctadecyl-3,3,3', 3'-tetramethylindocarbocyanine perchlorate-labeled acetylated low-density lipoprotein (DiI-AcLDL) showed that among common polyanionic ligands, polyinosine was the most potent, with an IC50 of 0.74 microgram/ml, whereas the novel compound (E)-methyl 4-chloro-alpha-[4-(4-chlorophenyl)-1, 5-dihydro-3-hydroxy-5-oxo-1-(2-thiazolyl)-2H-pyrrol-2-ylidene]benzene acetate gave an IC50 of 6.1 microgram/ml (13 microM). The novel antagonist also inhibited DiI-AcLDL uptake in cultured human peripheral and rat peritoneal macrophages with IC50 values of 21 microM and 17 microM, respectively. Wi...
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Specific, reversible and saturable binding sites for [3H]-PAF were observed in gerbil brain. Scatchard analysis revealed the existence of two apparent classes of binding sites with Kd1=3.7 nM and Kd2=20.4 nM. The maximal density of... more
Specific, reversible and saturable binding sites for [3H]-PAF were observed in gerbil brain. Scatchard analysis revealed the existence of two apparent classes of binding sites with Kd1=3.7 nM and Kd2=20.4 nM. The maximal density of binding sites was found in midbrain and hippocampus (Domingo et al., 1988). The existence of specific binding sites for PAF was also shown in rat hypothalamus (Junier et al., 1988) and synaptosomes and microsomal fractions of rat cerebral cortex (Marcheselli et al., 1990).
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Background and Purpose We tested the hypothesis that intravenous administration of the potent σ-receptor ligand 4-phenyl-1-(4-phenylbutyl) piperidine (PPBP) during transient focal ischemia would decrease postischemic brain infarction... more
Background and Purpose We tested the hypothesis that intravenous administration of the potent σ-receptor ligand 4-phenyl-1-(4-phenylbutyl) piperidine (PPBP) during transient focal ischemia would decrease postischemic brain infarction volume in rats. Methods Rats underwent intravascular focal ischemia for 2 hours followed by 22 hours of reperfusion. Halothane anesthesia was used only during initiation and cessation of ischemia. Rats received saline (n=10) or 1 μmol/kg per hour PPBP (n=10) by continuous intravenous infusion starting 1 hour after the initiation of ischemia and continuing through 22 hours of reperfusion. Results There was no difference between groups in blood pressure, arterial blood gas values, and body temperature. Triphenyltetrazolium-determined infarction volume of ipsilateral cerebral cortex (saline, 39±6%; PPBP, 21±7% of ipsilateral hemisphere; mean±SEM) and striatum (saline, 68±6%; PPBP, 33±8% of ipsilateral striatum) was smaller in rats treated with PPBP than in...
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Bone morphogenetic proteins (BMPs) and their serine/threonine kinase receptors have been identified in atherosclerotic arteries and vascular smooth muscle cells, respectively. Thus, BMPs (the largest subfamily of the TGF-β superfamily)... more
Bone morphogenetic proteins (BMPs) and their serine/threonine kinase receptors have been identified in atherosclerotic arteries and vascular smooth muscle cells, respectively. Thus, BMPs (the largest subfamily of the TGF-β superfamily) have been implicated in the pathogenesis of atherosclerosis. However, the origins of BMP biosynthesis and the functional roles of BMP in blood vessels are unclear. The present study explored BMP-2 gene expression in various human blood vessels and vascular cell types. Functional in vitro studies were also performed to determine the effects of recombinant human BMP-2 on migration (transwell assay) and proliferation ([3H]-thymidine incorporation) of human aortic vascular smooth muscle cells (HASMC). RT-PCR experiments revealed BMP-2 gene expression in normal and atherosclerotic human arteries as well as cultured human aortic and coronary vascular smooth muscle cells, human umbilical vein endothelial cells (HUVECs) and human macrophages. In cellular migr...
Research Interests: Biology, Cell Migration, Medicine, Gene expression, Bone Morphogenetic Proteins, and 15 moreCell Division, Humans, Chemotaxis, Vascular, Time Dependent, Transforming Growth Factor Beta, In Vitro Studies, Recombinant Proteins, Aorta, Blood Vessel, Synergistic effect, Smooth muscle cell, reverse transcriptase polymerase chain reaction, Drug synergism, and Medical and Health Sciences
Carvedilol is a vasodilating beta-blocker and antioxidant approved for treatment of mild to moderate hypertension. angina, and congestive heart failure. Metoprolol is a beta1-selective adrenoceptor antagonist. When carvedilol and... more
Carvedilol is a vasodilating beta-blocker and antioxidant approved for treatment of mild to moderate hypertension. angina, and congestive heart failure. Metoprolol is a beta1-selective adrenoceptor antagonist. When carvedilol and metoprolol were recently compared in clinical trials for heart failure, each showed beneficial beta-blocker effects such as improved symptoms, quality of life, exercise tolerance, and ejection fraction, with no between-group differences. When thiobarbituric acid reactive substance (TBARS) levels were measured in serum as an indirect marker of free radical activity, there were also no between-group differences. However, we had noted superior cardioprotection by carvedilol in comparison to metoprolol in ischemia and reperfusion models. We therefore examined antioxidant activity directly in cells and tissues. Here we show that in cultured rat cerebellar neurons, and in brain and heart membranes, carvedilol has far greater antioxidant activity than metoprolol, which is essentially inactive as an antioxidant in these model systems. The antioxidant activity of carvedilol could be explained by a greater degree of lipophilicity, as measured by its ClogP value of 3.841 as contrasted to a ClogP value of 1.346 for metoprolol. Alternatively, the molecular structure of carvedilol favors redox recycling, which the structure of metoprolol does not. Therefore, carvedilol could have additional pharmacologic effects that are favorable for long-term therapy.
Research Interests: Pharmacology, Free Radicals, Oxidative Stress, Medicine, Heart Failure, and 15 moreAntioxidants, Cardiovascular Pharmacology, Animals, Antioxidant, Cardioprotection, Heart, Neurons, Lipid peroxidation, Antioxidant Activity, Rats, Cell Membrane, Metoprolol, carvedilol, Cardiovascular medicine and haematology, and Pharmacology and pharmaceutical sciences
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The N-methyl-D-aspartate (NMDA) subtype of glutamate receptor appears to play a pivotal role in enabling glutamate to express its neurotoxic potential in a variety of neurological disorders. Our results show that the transition of... more
The N-methyl-D-aspartate (NMDA) subtype of glutamate receptor appears to play a pivotal role in enabling glutamate to express its neurotoxic potential in a variety of neurological disorders. Our results show that the transition of glutamate from neurotransmitter to neurotoxin is facilitated when cellular energy is limited in cultured cerebellar neurons. Omission of glucose, exclusion of oxygen, or inclusion of inhibitors of oxidative phosphorylation or of the sodium/potassium pump, enables the excitatory amino acids glutamate or NMDA to express their neurotoxic potential. We interpret these results as demonstrating that glucose metabolism, ATP production, and functioning Na+,K+-ATPases are necessary to generate a resting potential sufficient to maintain the voltage-dependent Mg2+ block of the NMDA receptor channel; relief of the Mg2+ block enables the excitatory amino acids to act persistently at the NMDA receptor, resulting in the opening of ion channels and subsequent neuronal damage. These findings are discussed in the context of perturbations or abnormalities which lead to decreased availability or utilization of glucose and oxygen in the brain which may trigger endogenous excitatory amino acids to become neurotoxic by this mechanism.
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MK-801 and (+)SK&F 10047 produced a dose-related inhibition of the EEG suppression and cortical hyperemia associated with cortical spreading depression (CSD) and reduced the CSD propagation rate; ED50 = 1 mg/kg, i.v. and 15 mg/kg,... more
MK-801 and (+)SK&F 10047 produced a dose-related inhibition of the EEG suppression and cortical hyperemia associated with cortical spreading depression (CSD) and reduced the CSD propagation rate; ED50 = 1 mg/kg, i.v. and 15 mg/kg, i.v., respectively. MK-801 had a delayed onset of action (inversely related to dose) and a prolonged duration of action at all doses (> 2 h). In contrast, (+)SK&F 10047 had a rapid onset of action (< 30 min) and a predictable dose-related duration of action. These results suggests that an efficacious compound acting with moderate affinity as a non-competitive antagonist at the NMDA-receptor channel may possess a preferable time-course and toxicity profile when compared to agents acting similarly, but with high affinity.
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tetrodotoxin-sensitive ion channels in hypoxic-ischemic neu-ronal damage have concluded that sodium influx is an impor-tant initiating event. We examined the neuroprotectant effect of tetrodotoxin on both cultured cerebellar neurons and... more
tetrodotoxin-sensitive ion channels in hypoxic-ischemic neu-ronal damage have concluded that sodium influx is an impor-tant initiating event. We examined the neuroprotectant effect of tetrodotoxin on both cultured cerebellar neurons and on CA1 hippocampal neurons of gerbils exposed to brain ischemia. Methods We studied neuroprotective mechanisms using cultured rat cerebellar granule cells exposed to veratridine, which induced cytotoxicity, neurotransmitter release, and cal-cium influx. Survival of gerbil CA1 neurons was examined by direct neuron counts 7 days after 6 minutes of global ischemia with reperfusion. Results Tetrodotoxin protected cultured neurons in a dose-dependent manner from veratridine-induced toxicity (protec-tive concentration [PC50]=22 nmol/L). Veratridine induced
Loci designated penA, penB, and mtr contribute additively to penicillin G resistance in Neisseria gonorrhoeae; the mtr locus also confers resistance to hydrophobic dyes, detergents, and antibiotics, env mutations suppress the phenotypic... more
Loci designated penA, penB, and mtr contribute additively to penicillin G resistance in Neisseria gonorrhoeae; the mtr locus also confers resistance to hydrophobic dyes, detergents, and antibiotics, env mutations suppress the phenotypic expression of mtr and penB and are responsible for increased sensitivity to various hydrophobic molecules. We postulated that the host environment is important in the selection of gonococcal strains with these particular outer membrane phenotypes. Thus, mtr strains should predominate in environments that are high in hydrophobic molecules. To test this hypothesis we determined the outer membrane phenotype of 152 strains of N. gonorrhoeae. Rectal and urethral isolates from 58 homosexual men, urethral isolates from 55 heterosexual men, and cervical and rectal isolates from 39 heterosexual women were used in this study. Strains from 43 of the homosexual men were matched with those from heterosexual men with respect to auxotype and year and season of isol...
Research Interests: Microbiology, Biology, Membrane Proteins, Medicine, Homosexuality, and 15 moreBiological Sciences, Lipids, Humans, Female, Male, Feces, Microbial Sensitivity Tests, Erythromycin, Molecular weight, Cell Wall, Neisseria gonorrhoeae, Penicillin G, Cell membrane permeability, Medical and Health Sciences, and gonorrhea
Chemoattraction of monocytes by the CXC chemokine stromal cell-derived factor-1α (SDF-1α) and its receptor CXCR4 may be involved in vascular diseases like atherosclerosis. We studied the regulation of CXCR4 transcription and SDF-1-induced... more
Chemoattraction of monocytes by the CXC chemokine stromal cell-derived factor-1α (SDF-1α) and its receptor CXCR4 may be involved in vascular diseases like atherosclerosis. We studied the regulation of CXCR4 transcription and SDF-1-induced functional responses in human monocytes during their differentiation in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF), oxidized low-density lipoprotein (Ox-LDL), and unmodified LDL. Our results reveal that the rapid decline of SDF-1-mediated [Ca2+]i influx after monocyte isolation is followed by a gradual functional restoration and a concomitant re-expression of CXCR4 mRNA over time. A further three- to fourfold induction of CXCR4 mRNA occurred in macrophage-derived foam cells on treatment with Ox-LDL. HL-60 cells induced with phorbol myristate acetate (PMA) showed a rapid fourfold stimulation of CXCR4 mRNA within 1 h, declining to barely detectable levels at 3 h, with eventual restoration over time, mirroring the expres...
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Using rat cerebellar granule cells in primary culture as our model system, we have shown that excitatory amino acids (EAAs) become neurotoxic via the NMDA (N-methyl-D-aspartate) receptor when neuronal energy levels are compromised.... more
Using rat cerebellar granule cells in primary culture as our model system, we have shown that excitatory amino acids (EAAs) become neurotoxic via the NMDA (N-methyl-D-aspartate) receptor when neuronal energy levels are compromised. Omission of glucose, exclusion of oxygen, or inclusion of inhibitors of oxidative phosphorylation or of Na+/K+-ATPases enables NMDA receptor agonists to express their neurotoxic potential. Both competitive and noncompetitive NMDA receptor antagonists are potent blockers of EAA neurotoxicity, with MK-801 fully effective at 20 nM. We interpret these results as indicating that glucose metabolism, ATP production, and functioning ion pumps are necessary to generate a resting potential sufficient to maintain the voltage-dependent Mg++ block of the NMDA receptor channel; relief of the block enables EAAs to act persistently at the NMDA receptor causing an excessive ion influx which leads to neuronal death by a mechanism not yet understood. These findings are disc...
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Calcitonin gene-related peptide (CGRP) stimulates the adhesiveness of human umbilical vein endothelial cells for U937 cells and human neutrophils in a dose- and time-dependent manner. The onset of CGRP-induced adhesives of HUVEC was rapid... more
Calcitonin gene-related peptide (CGRP) stimulates the adhesiveness of human umbilical vein endothelial cells for U937 cells and human neutrophils in a dose- and time-dependent manner. The onset of CGRP-induced adhesives of HUVEC was rapid (30 min), independent of protein synthesis, and lasted over 24 h in the continuous presence of the peptide. The stimulatory effect of CGRP was completely blocked by the CGRP antagonist, CGRP(8-37). The present study provides evidence in support of the potential role of sensory nerve-derived neuropeptides in the modulation of leukocyte adhesion to vascular endothelial cells.
Research Interests: Endocrinology, Physiology, Biology, Cell Adhesion, Medicine, and 15 moreAdhesion, Neuroimmunology, Thrombin, CGRP, Humans, Internal Medicine, cyclic AMP, Peptides, Neutrophils, Time Factors, Calcitonin, Adhesion molecules, Neuropeptide, Pharmacology and pharmaceutical sciences, and Calcitonin Gene Related Peptide
Calcitonin gene-related peptide (CGRP) is a neuropeptide with diverse biological properties including potent vasodilating activity. Recently, we reported the cloning of complementary DNAs (cDNAs) encoding the human and porcine CGRP... more
Calcitonin gene-related peptide (CGRP) is a neuropeptide with diverse biological properties including potent vasodilating activity. Recently, we reported the cloning of complementary DNAs (cDNAs) encoding the human and porcine CGRP receptors which share significant amino acid sequence homology with the human calcitonin receptor, a member of the recently described novel subfamily of G-protein-coupled 7TM receptors. Activation of this family of receptors has been shown to result in an increase in intracellular cAMP accumulation and calcium release. In this study, we demonstrate that HEK-293 cells expressing recombinant CGRP receptors (HEK-293HR or PR) respond to CGRP with increased intracellular calcium release (EC50 = 1.6 nM) in addition to the activation of adenylyl cyclase (EC50 = 1.4 nM). The effect of CGRP on adenylyl cyclase activation and calcium release was inhibited by CGRP (8-37), a CGRP receptor antagonist. Both effects were mediated by cholera toxin-sensitive G-proteins, b...
Research Interests: Biology, Medicine, Signal Transduction, Cell line, Molecular and cellular biology, and 11 moreSignal Transduction Pathway Models, Receptor, G protein, Amino Acid Sequence, Molecular and Cellular Biochemistry, Biochemistry and cell biology, Intracellular Calcium, Adenylyl Cyclase, Signaling pathway, Calcitonin Gene Related Peptide, and Cholera toxin
Studies examining the role of tetrodotoxin-sensitive ion channels in hypoxic-ischemic neuronal damage have concluded that sodium influx is an important initiating event. We examined the neuroprotectant effect of tetrodotoxin on both... more
Studies examining the role of tetrodotoxin-sensitive ion channels in hypoxic-ischemic neuronal damage have concluded that sodium influx is an important initiating event. We examined the neuroprotectant effect of tetrodotoxin on both cultured cerebellar neurons and on CA1 hippocampal neurons of gerbils exposed to brain ischemia. We studied neuroprotective mechanisms using cultured rat cerebellar granule cells exposed to veratridine, which induced cytotoxicity, neurotransmitter release, and calcium influx. Survival of gerbil CA1 neurons was examined by direct neuron counts 7 days after 6 minutes of global ischemia with reperfusion. Tetrodotoxin protected cultured neurons in a dose-dependent manner from veratridine-induced toxicity (protective concentration [PC50] = 22 nmol/L). Veratridine induced [3H]aspartate efflux that was sodium dependent, only 25% calcium dependent, and was inhibited by tetrodotoxin (inhibitory concentration [IC50] = 60 nmol/L). Veratridine initiated increases in...
Research Interests:
We reported previously that stroke risk factors prepared the brain stem for the development of ischemia and hemorrhage and induced the production of tumor necrosis factor following an intrathecal injection of lipopolysaccharide, a... more
We reported previously that stroke risk factors prepared the brain stem for the development of ischemia and hemorrhage and induced the production of tumor necrosis factor following an intrathecal injection of lipopolysaccharide, a prototypic monocyte-activating stimulus. This study evaluates whether blood or brain cells of hypertensive rats produce more proinflammatory and prothrombotic mediators than do blood or brain cells of normotensive rats. Levels of tumor necrosis factor, platelet-activating factor, 6-ketoprostaglandin F1 alpha, and thromboxane B2 in the cerebrospinal fluid and blood of spontaneously hypertensive and normotensive Wistar-Kyoto rats were monitored before and after a challenge with lipopolysaccharide. Little or no activity from these mediators was found in the cerebrospinal fluid or blood of saline-injected control animals. Intravenous administration of lipopolysaccharide (0.001, 0.1, and 1.8 mg/kg) produced dose-dependent increases in blood levels of all mediat...
Research Interests: Endocrinology, Medicine, Lipopolysaccharide, Stroke, Brain, and 14 moreInternal Medicine, Hypertension, Animals, Risk factors, Clinical Sciences, Lipopolysaccharides, Rats, Tumor necrosis factor-alpha, Risk Factors, Reference Values, Cerebrovascular Disorders, Neurosciences, Cardiovascular medicine and haematology, and leukocyte Count
Excitatory amino acids and their receptors are involved in mediating ischemic neuronal damage. The sigma-agonists are believed to interact with the N-methyl-D-aspartate receptor. Therefore, we studied the neuroprotective, hypothermic, and... more
Excitatory amino acids and their receptors are involved in mediating ischemic neuronal damage. The sigma-agonists are believed to interact with the N-methyl-D-aspartate receptor. Therefore, we studied the neuroprotective, hypothermic, and motor deficit effects of the sigma-agonist SKF 10,047 and the N-methyl-D-aspartate antagonist MK-801. Neuroprotective effects were compared using an in vitro ischemia model of cultured rat cerebellar granule cells and the gerbil model of global brain ischemia induced by 5 minutes of bilateral carotid artery occlusion followed by 7 days of reperfusion. In vitro, (+)MK-801 protected against 100 microM glutamate with a 50% protective concentration of 30 nM, followed by (-)MK-801 (150 nM), cyclazocine (0.5 microM), (+)SKF 10,047 (3.3 microM), pentazocine (5 microM), and (-)SKF 10,047 (10 microM). In vivo, (+)SKF 10,047 pretreatment (60 mg/kg) or multiple postischemic treatments provided neuroprotection comparable with MK-801 pretreatment (10 mg/kg). Wh...