Antiproliferative properties of the lazaroids U-83836E and U-74389G on glioma cells in vitro.
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The 21-aminosteroids (lazaroids) are a new family of steroid compounds that inhibit lipid peroxidation reactions. They are novel antioxidant agents, which have been shown to have antiproliferative properties on cancer cells and also are thought to prevent free radical-mediated blood-brain barrier damage. In order to understand the effect of lazaroids on glioma, we tested U-83836E and U-74389G at doses ranging between 0.1 100 m mM on primary cultures of glioblastoma multiforme from three patients, rat C6 glioma cell line, and 5 th subculture established from one of the patients. The effects of both compounds on cell proliferation were determined using 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) colorimetric assay. U-83836E in the primary cultures was found to have 50% inhibitory concentrations (IC50 ) of 6.30, 6.75 and 6.50 m mM, respectively. The IC50 value of U-74389G was calculated as 91 m mM in only one of the patients. On C6 glioma cells, while the IC50 of U-83836E was 45 m mM, U-74389G showed no cytotoxic effect. On the 5 th subculture, U-83836E had an IC50 of 37.5 m mM, but the cytotoxic effects of U-74389G was less than in that of the primary culture. In conclusion, these compounds were found to be more cytotoxic in primary culture than the cell lines and there were also differences between their members in the inhibition of cell survival. (+info)
The effects of MK-801 and U-83836E on post-ischemic reperfusion injury in rat brain.
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Rats were subjected to incomplete cerebral ischemia induced by occlusion of common carotid arteries for 30 min, and subsequent reperfusion for 15 min. The concentrations of reduced glutathione (GSH), malondialdehyde (MDA) and superoxide dismutase (SOD) activity were determined in the dorsal hippocampus in order to evaluate their changes during ischemia and reperfusion following ischemia. The depletion of GSH was observed during ischemia with a further depletion during post-ischemic reperfusion (P < 0.001), while a significant increase in SOD activity and MDA levels was found only after reperfusion following ischemia (P < 0.001). Animals in which ischemia was followed by reperfusion were treated with a non-competitive NMDA receptor antagonist, MK-801 (1 mg/kg, i.v.), and a radical scavenger, U-83836E (5 mg/kg, i.v.), prior to ischemia. Although a full recovery of GSH levels was not observed following MK-801 and U-83836E pretreatment as compared to control (P < 0.05), MK-801 was more potent than U-83836E in the partial protection of the GSH pool (P < 0.05 and P < 0.01, respectively). The rise in SOD activity and MDA level were brought close to those of control due to the effects of both MK-801 and U-83836E (P > 0.05). In conclusion, the tissue changes in GSH concentrations evoked by ischemia and reperfusion were partially prevented by the effects of both drugs, MK-801 having the greater effect. This suggests that the NMDA receptor activation may play a role in the generation of reactive oxygen species. On the other hand, the inhibition of lipid peroxidation brought about by both MK-801 or U-83836E suggests the therapeutic efficiency of these agents in ischemia/reperfusion injury. (+info)
Recovery of dopamine neuronal transporter but lack of change of its mRNA in substantia nigra after inactivation by a new irreversible inhibitor characterized in vitro and ex vivo in the rat.
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1. In vitro, the ability of DEEP-NCS {1-[2-(diphenylmethoxy)ethyl]-4-[2-(4-isothiocyanatophenyl)ethyl]- piperazine} to inhibit [3H]-dopamine uptake by rat striatal synaptosomes was concentration-dependent and inversely related to the protein concentration. This inhibition was irreversible and resulted from changes in Vmax and KM. DEEP-NCS was less potent on noradrenaline, serotonin and choline transport. 2. One day after intrastriatal injections of DEEP-NCS (100 and 1000 pmol) in 20% dimethylsulphoxide, moderate decreases in the ex vivo dopamine uptake were observed in synaptosomes obtained from striatum injected with DEEP-NCS or solvent, and the contralateral uninjected striatum. 3. In similar conditions, 300 pmol DEEP-NCS in 45% 2 hydroxypropyl-gamma-cyclodextrin - 0.5% dimethylsulphoxide solution sub-totally reduced ex vivo dopamine uptake and mazindol binding, and moderately decreased choline and serotonin transport. These reductions were specific to DEEP-NCS-injected striata. A clomipramine pretreatment (16 mg kg-1 i.p. 1 h before) was performed in following experiments, since it reduced the DEEP-NCS-elicited decrease in serotonin uptake without affecting other indices. 4. One day after intrastriatal injection, DEEP-NCS elicited similar dose-dependent decreases in ex vivo dopamine uptake and mazindol binding (ID50=6.9-8 ng striatum-1). Changes in KM and Vmax for ex vivo dopamine transport produced by DEEP-NCS disappeared according to similar time-courses. 5. The t(1/2) for transporter recovery was 6. 1 days. This value should correspond to its actual turnover rate in vivo, since no change in transporter mRNA level was observed in substantia nigra ipsilateral to 300 pmol DEEP-NCS-injected striatum. 6. The results indicate that DEEP-NCS behaves as a potent, quite selective, irreversible inhibitor of the DAT, in vitro and in vivo. Its use in vivo suggests that the physiological half-life of the rat striatal DAT is close to 6 days. (+info)
Selective glutamate receptor antagonists can induce or prevent axonal sprouting in rat hippocampal slice cultures.
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After the transection of the Schaffer collateral pathway in hippocampal slice cultures, reactive sprouting is induced in the CA3 area, and eventually synaptic transmission between areas CA1 and CA3 is restored. Using this model, we have studied the role of ionotropic glutamate receptors in the initiation of axonal sprouting and the regeneration of functional synapses. We show that neither reactive sprouting nor functional recovery of synaptic transmission occur in the presence of the non-N-methyl-D-aspartate (NMDA) receptor antagonist 6-nitro-7-sulfamoylbenzoquinoxaline-2,3-dione (CNQX). In contrast, the NMDA receptor antagonists methyl-10, 11-dihydro-5-H-dibenzocyclohepten-5,10-imine (MK-801) or 3-(RS)-2-carboxypiperazine-4-yl)-propyl-1-phosphonic acid (CPP) did not interfere with these processes. Moreover, we observed that the application of NMDA receptor antagonists induced massive axonal sprouting and an increase in the frequency of miniature excitatory postsynaptic currents in unlesioned cultures. Our results thus indicate that NMDA and non-NMDA receptors exert a differential effect on reactive sprouting and the recovery of synaptic transmission after injury in the hippocampus. Activation of non-NMDA receptors appears necessary for these processes to occur, whereas activation of NMDA receptors suppresses growth-associated protein -43 expression and axonal outgrowth. (+info)
Newer pharmacologic alternatives for erectile dysfunction.
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With the introduction of effective pharmacologic therapies for erectile dysfunction, more men are seeking treatment. The underlying cause of erectile dysfunction is usually a chronic medical illness or a side effect of certain drugs. Less commonly, the problem is psychogenic. Even after optimal treatment of common medical disorders such as diabetes mellitus and hypertension, erectile dysfunction may persist. Pharmacologic treatments, such as the intracavernosal or transurethral administration of alprostadil or the use of the new oral medication sildenafil, may offer patients substantial benefit. Before any of these drugs are prescribed, consideration should be given to existing medical illnesses and medications, partner satisfaction, comfort with the method of administration and the side effect profile. (+info)
Novel antimigraineur dotarizine releases Ca2+ from caffeine-sensitive Ca2+ stores of chromaffin cells.
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1. The novel antimigraineur, dotarizine (30 microM), increased cytosolic Ca2+ concentration, [Ca2+]c, in fura-2-loaded bovine adrenal chromaffin cells. This increase was transient, reached a peak in about 2 - 5 min (0.53+/-0.07 microM; n=19) and then declined to basal levels over a further 5 min period. 2. This transient rise of [Ca2+]c was mimicked by 1 microM thapsigargin and by 30 microM cyclopiazonic acid (CPA), but not by 30 microM flunarizine. Both thapsigargin and CPA occluded the effects of dotarizine and vice versa. 3. All three compounds suppressed the transient [Ca2+]c rises induced by caffeine (10 mM, 10 s); blockade induced by thapsigargin was irreversible and that induced by CPA and dotarizine was reversible. 4. Of the three compounds, only dotarizine blocked reversibly the [Ca2+]c spikes induced by short pulses of high K+ (70 mM, 5 s), suggesting that dotarizine blocks voltage-dependent Ca2+ channels but CPA and thapsigargin do not. 5. Dotarizine caused a gradual and reversible depletion of endoplasmic reticulum (ER) Ca2+ in chromaffin cells transfected with ER-targeted aequorin. CPA had a similar effect. 6. These data show that dotarizine shares with thapsigargin and CPA the ability to deplete Ca2+ in the ER; this novel action of dotarizine could be relevant to its prophylactic effects in migraine. Unlike thapsigargin and CPA, however, dotarizine additionally and reversibly blocks Ca2+ entry through voltage-dependent Ca2+ channels. (+info)
5-hydroxytryptamine receptors mediating contraction in human small muscular pulmonary arteries: importance of the 5-HT1B receptor.
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1. The 5-hydroxytryptamine (5-HT) receptors mediating vasoconstriction in isolated human small muscular pulmonary arteries (SMPAs) were determined using techniques of wire myography and reverse transcription-polymerase chain reaction (RT - PCR). 2. The agonists 5-HT, 5-carboxamidotryptamine (5-CT, unselective for 5-HT1 receptors) and sumatriptan (selective for 5-HT1B/D receptors) all caused contraction and were equipotent (pEC50s: 7.0+/-0.2, 7.1+/-0.3 and 6.7+/-0.1, respectively) suggesting the presence of a 5-HT1 receptor. 3. Ketanserin (5-HT2A-selective antagonist, 0.1 microM) inhibited 5-HT-induced contractions only at non-physiological/pathological concentrations of 5-HT (>0.1 microM) whilst GR55562 (5-HT1B/1D-selective antagonist, 1 microM) inhibited 5-HT-induced contractions at all concentrations of 5-HT (estimated pKB=7.7+/-0.2). SB-224289 (5-HT1B-selective antagonist, 0.2 microM) inhibited sumatriptan-induced contractions (estimated pKB=8.4+/-0.1) whilst these were unaffected by the 5-HT1D-selective antagonist BRL15572 (0.5 microM) suggesting that the 5-HT1B receptor mediates vasoconstriction in this vessel. 4. RT - PCR confirmed the presence of substantial amounts of mRNA for the 5-HT2A and 5-HT1B receptor subtypes in these arteries whilst only trace amounts of 5-HT1D receptor message were evident. 5. These findings suggest that a heterogeneous population of 5-HT2A and 5-HT1B receptors co-exist in human small muscular pulmonary arteries but that the 5-HT1B receptor mediates 5-HT-induced vasoconstriction at physiological and pathophysiological concentrations of 5-HT. These results have important implications for the treatment of pulmonary hypertension in which the 5-HT1B receptor may provide a novel and potentially important therapeutic target. (+info)
Comparison of the novel antipsychotic ziprasidone with clozapine and olanzapine: inhibition of dorsal raphe cell firing and the role of 5-HT1A receptor activation.
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Ziprasidone is a novel antipsychotic agent which binds with high affinity to 5-HT1A receptors (Ki = 3.4 nM), in addition to 5-HT1D, 5-HT2, and D2 sites. While it is an antagonist at these latter receptors, ziprasidone behaves as a 5-HT1A agonist in vitro in adenylate cyclase measurements. The goal of the present study was to examine the 5-HT1A properties of ziprasidone in vivo using as a marker of central 5-HT1A activity the inhibition of firing of serotonin-containing neurons in the dorsal raphe nucleus. In anesthetized rats, ziprasidone dose-dependently slowed raphe unit activity (ED50 = 300 micrograms/kg i.v.) as did the atypical antipsychotics clozapine (ED50 = 250 micrograms/kg i.v.) and olanzapine (ED50 = 1000 micrograms/kg i.v.). Pretreatment with the 5-HT1A antagonist WAY-100,635 (10 micrograms/kg i.v.) prevented the ziprasidone-induced inhibition; the same dose of WAY-100,635 had little effect on the inhibition produced by clozapine and olanzapine. Because all three agents also bind to alpha 1 receptors, antagonists of which inhibit serotonin neuronal firing, this aspect of their pharmacology was assessed with desipramine (DMI), a NE re-uptake blocker previously shown to reverse the effects of alpha 1 antagonists on raphe unit activity. DMI (5 mg/kg i.v.) failed to reverse the inhibitory effect of ziprasidone but produced nearly complete reversal of that of clozapine and olanzapine. These profiles suggest a mechanism of action for each agent, 5-HT1A agonism for ziprasidone and alpha 1 antagonism for clozapine and olanzapine. The 5-HT1A agonist activity reported here clearly distinguishes ziprasidone from currently available antipsychotic agents and suggests that this property may play a significant role in its pharmacologic actions. (+info)