(1/1620) Studies on the mechanism of action of amantadine.

1 The effect of amantadine hydrochloride on various aspects of catecholamine metabolism in the rat brain has been investigated. 2 Amantadine failed to have any significant effect on brain concentrations of dopamine or noradrenaline even when administered daily for 9 days. 3 Amantadine had no effect on the rate of decline of noradrenaline and dopamine concentrations after alpha-methyl-p-tyrosine. 4 In vitro amantadine inhibited dopamine uptake into synaptosomes only at high concentrations, and caused little release of dopamine from synaptosomes. 5 There is no evidence from these results to suggest that the anti-Parkinsonian effect of amantadine is related to an action on dopaminergic mechanisms.  (+info)

(2/1620) Changes in protein tyrosine phosphorylation in the rat brain after cerebral ischemia in a model of ischemic tolerance.

A brief period of sublethal cerebral ischemia, followed by several days of recovery, renders the brain resistant to a subsequent lethal ischemic insult, a phenomenon termed ischemic preconditioning or tolerance. Ischemic tolerance was established in the rat two-vessel occlusion model of ischemia, induced by occlusion of both carotid arteries in combination with hypotension. Ischemic preconditioning (3 minutes) provided maximal neuroprotection when induced 2 days prior to a lethal ischemic insult of 9-minute duration. Neuroprotection persisted for at least 8 weeks. Since neurotransmission has been implicated in ischemic cell death, the effect of ischemic preconditioning on tyrosine phosphorylation of proteins and on the levels of glutamate receptor subunits in hippocampus and neocortex was studied. Regional levels of tyrosine phosphorylation of proteins in general and the N-methyl-D-aspartate receptor subunit NR2 in particular are markedly enhanced after ischemia in nonconditioned brains, in both the synaptosomal fraction and the whole-tissue homogenate of rat neocortex and hippocampus, but recover to control levels only in the preconditioned brain. Ischemic preconditioning selectively induces a decrease in the levels of the NR2A and NR2B subunits and a modest decrease in the levels of NR1 subunit proteins in the synaptosomal fraction of the neocortex but not hippocampus after the second lethal ischemia. It was concluded that ischemic preconditioning prevents a persistent change in cell signaling as evidenced by the tyrosine phosphorylation of proteins after the second lethal ischemic insult, which may abrogate the activation of detrimental cellular processes leading to cell death.  (+info)

(3/1620) (S)-(-)-Cotinine, the major brain metabolite of nicotine, stimulates nicotinic receptors to evoke [3H]dopamine release from rat striatal slices in a calcium-dependent manner.

Cotinine, a major peripheral metabolite of nicotine, has recently been shown to be the most abundant metabolite in rat brain after peripheral nicotine administration. However, little attention has been focused on the contribution of cotinine to the pharmacological effects of nicotine exposure in either animals or humans. The present study determined the concentration-response relationship for (S)-(-)-cotinine-evoked 3H overflow from superfused rat striatal slices preloaded with [3H]dopamine ([3H]DA) and whether this response was mediated by nicotinic receptor stimulation. (S)-(-)-Cotinine (1 microM to 3 mM) evoked 3H overflow from [3H]DA-preloaded rat striatal slices in a concentration-dependent manner with an EC50 value of 30 microM, indicating a lower potency than either (S)-(-)-nicotine or the active nicotine metabolite, (S)-(-)-nornicotine. As reported for (S)-(-)-nicotine and (S)-(-)-nornicotine, desensitization to the effect of (S)-(-)-cotinine was observed. The classic nicotinic receptor antagonists mecamylamine and dihydro-beta-erythroidine inhibited the response to (S)-(-)-cotinine (1-100 microM). Additionally, 3H overflow evoked by (S)-(-)-cotinine (10-1000 microM) was inhibited by superfusion with a low calcium buffer. Interestingly, over the same concentration range, (S)-(-)-cotinine did not inhibit [3H]DA uptake into striatal synaptosomes. These results demonstrate that (S)-(-)-cotinine, a constituent of tobacco products and the major metabolite of nicotine, stimulates nicotinic receptors to evoke the release of DA in a calcium-dependent manner from superfused rat striatal slices. Thus, (S)-(-)-cotinine likely contributes to the neuropharmacological effects of nicotine and tobacco use.  (+info)

(4/1620) Functional integrity of mitochondrial genomes in human platelets and autopsied brain tissues from elderly patients with Alzheimer's disease.

To determine whether pathogenic mutations in mtDNA are involved in phenotypic expression of Alzheimer's disease (AD), the transfer of mtDNA from elderly patients with AD into mtDNA-less (rho0) HeLa cells was carried out by fusion of platelets or synaptosomal fractions of autopsied brain tissues with rho0 HeLa cells. The results showed that mtDNA in postmortem brain tissue survives for a long time without degradation and could be rescued in rho0 HeLa cells. Next, the cybrid clones repopulated with exogenously imported mtDNA from patients with AD were used for examination of respiratory enzyme activity and transfer of mtDNA with the pathogenic mutations that induce mitochondrial dysfunction. The presence of the mutated mtDNA was restricted to brain tissues and their cybrid clones that formed with synaptosomes as mtDNA donors, whereas no cybrid clones that isolated with platelets as mtDNA donors had detectable mutated mtDNA. However, biochemical analyses showed that all cybrid clones with mtDNA imported from platelets or brain tissues of patients with AD restored mitochondrial respiration activity to almost the same levels as those of cybrid clones with mtDNA from age-matched normal controls, suggesting functional integrity of mtDNA in both platelets and brain tissues of elderly patients with AD. These observations warrant the reassessment of the conventional concept that the accumulation of pathogenic mutations in mtDNA throughout the aging process is responsible for the decrease of mitochondrial respiration capacity with age and with the development of age-associated neurodegenerative diseases.  (+info)

(5/1620) Short-time effects of neuroactive steroids on rat cortical Ca2+-ATPase activity.

Recent experimental evidence indicates that some steroid hormones, apart from their well-documented genomic actions, could produce non-genomic rapid effects, and are potent modulators of the plasma membrane proteins, including voltage- and ligand-operated ion channels or G protein-coupled receptors. Neuroactive steroids, 17beta-estradiol, testosterone, pregnenolone sulfate and dehydroepiandrosterone sulfate, after a short-time incubation directly modulated the activity of plasma membrane Ca2+-ATPase purified from synaptosomal membranes of rat cortex. The sulfate derivatives of dehydroepiandrosterone and pregnenolone applied at concentrations of 10-11-10-6 M, showed an inverted U-shape potency in the regulation of Ca2+-ATPase activity. At physiologically relevant concentrations (10-8-10-9 M) a maximal enhancement of the basal activity reached 200%. Testosterone (10-11-10-6 M) and 17beta-estradiol (10-12-10-9 M) caused a dose-dependent increase in the hydrolytic ability of Ca2+-ATPase, and the activity with the highest concentration of steroids reached 470% and 200%, respectively. All examined steroids decreased the stimulatory effect of a naturally existing activator of the calcium pump, calmodulin. The present study strongly suggests that the plasma membrane calcium pump could be one of the possible membrane targets for a non-genomic neuroactive steroid action.  (+info)

(6/1620) (-)-Stepholidine enhances K+ depolarization-induced activation of synaptosomal tyrosine 3-monooxygenase from rat striatum.

AIM: To study the mechanism of K+ depolarization-induced activation of synaptosomal tyrosine 3-monooxygenase (TM) in rat striatum and the effect of (-)-stepholidine (SPD) on this activation. METHODS: The TM was assayed for DOPA by HPLC-ECD; the activities of Ca2+/calmodulin (CaM)-dependent protein kinase (PK II) and Ca2+/phosphoinositide-dependent protein kinase (PKC) were assayed using histidine as substrate. RESULTS: The incubation of striatal synaptosomes in K(+)-riched (60 mmol.L-1) medium resulted in a marked activation of TM. PKC inhibitor polymyxin B (PMB) completely blocked the activation of K+ 60 mmol.L-1 on TM. Selective D2 receptor agonist quinpirole (QP), Ca2+ removal from incubation medium and CaM antagonist W7 failed to affect the activation. However, SPD enhanced the activation of K+ 60 mmol.L-1 on TM. Meanwhile, the incubation in K+ 60 mmol.L-1 also activated PKC. Neither QP nor SPD affected K+ depolarization-induced activation of PKC. CONCLUSION: The activation of K+ depolarization on synaptosomal TM is enhanced by SPD and this activation is mediated by PKC rather than by PK II.  (+info)

(7/1620) Effect of saponins of Panax notoginseng on synaptosomal 45Ca uptake.

AIM: To explore the calcium uptake antagonism of saponins of Panax notoginseng (PNS). METHODS: Synaptosomes were prepared from rat cerebral cortex by using differential Ficoll gradients. The effects of PNS on synaptosomal 45Ca uptake were measured in vitro or after acute treatment. RESULTS: PNS 50-800 mg.L-1 produced a concentration-rated inhibition of Ca2+ uptake [IC50 = 111 (46-176) mg.L-1]. Both initial and maximal uptake were inhibited. Similar effect was obtained after acute PNS treatment with 200 mg.kg-1 i.p. The blocking effect of PNS was reversed by calcium in media. CONCLUSION: PNS is a calcium channel blocker in neurons.  (+info)

(8/1620) Characterization of [125I]RTI-121 binding to dopamine transporter in vitro.

AIM: To characterize the binding of [125I]3 beta-(4-iodophenyl) tropane-2 beta-carboxylic acid isopropyl ester (RTI-121) to the dopamine transporter (DAT) under physiologically relevant conditions. METHODS: [125I]RTI-121 was used to label the DAT on fresh rat striatum synaptosomal membranes in artificial cerebrospinal fluid (ACSF) at 37 degrees C. RESULTS: [125I]RTI-121 binding reached equilibrium within 3 min and remained at its plateau value for at least 9 min. The data from kinetic, saturation, and competition studies supported a one-site model for the binding of [125I]RTI-121 to the DAT. Various DAT blockers (oocaine, GBR12935, and BTCP) and substrates (dopamine and d-amphetamine) competitively inhibited the binding of [125I]RTI-121. Compared with NaPhos-KCl-NaCl assay buffer, ACSF containing Ca2+ and Mg2+ markedly increased the IC50 of DAT blockers for inhibiting [125I]RTI-121 binding with less effect on that of substrates. Various D2 receptor ligands (pergolide, quinirole, sulpiride, and l-stepholidine) had no direct effect on the binding of [125I]RTI-121. CONCLUSION: [125I]RTI-121 binding under physiologically relevant conditions fulfills the basic criteria for DAT binding assay.  (+info)