Amantadine mitigates the cytotoxic and genotoxic effects of doxorubicin in SH-SY5Y cells and reduces its mutagenicity.

Amantadine (AMA) is a useful drug in neuronal disorders, but few studies have been performed to access its toxicological profile. Conversely, doxorubicin (Dox) is a well-known antineoplastic drug that has shown neurotoxic effects leading to cognitive impairment. The aims of this study are to evaluate the cytotoxic, genotoxic, and mutagenic effects of AMA, as well as its possible protective actions against deleterious effects of Dox. The Salmonella/microsome assay was performed to assess mutagenicity while cytotoxicity and genotoxicity were evaluated in SH-SY5Y cells using MTT and comet assays. Possible modulating effects of AMA on the cytotoxicity, genotoxicity, and mutagenicity induced by Dox were evaluated through cotreatment procedures. Amantadine did not induce mutations in the Salmonella/microsome assay and decreased Dox-induced mutagenicity in the TA98 strain. AMA reduced cell viability and induced DNA damage in SH-SY5Y cells. In cotreatment with Dox, AMA attenuated the cytotoxicity of Dox and showed an antigenotoxic effect. In conclusion, AMA does not induce gene mutations, although it has shown a genotoxic effect. Furthermore, AMA decreases frameshift mutations induced by Dox as well as the cytotoxic and genotoxic effects of Dox in SH-SY5Y cells, suggesting that AMA can interfere with Dox mutagenic activity and attenuate its neurotoxic effects.

Involvement of dopamine D2 and glutamate NMDA receptors in the antidepressant-like effect of amantadine in mice.

The present study investigated the pharmacological mechanisms of the antidepressant-like effects of amantadine in mice and their influence on hippocampal neurogenesis. To improve the translational validity of preclinical results, reproducibility across laboratories and replication in other animal models and species are crucial. Single amantadine administration at doses of 50 and 75 mg/kg resulted in antidepressant-like effects in mice in the tail suspension test (TST), reflected by an increase in immobility time. The effects of amantadine were seen at doses that did not alter locomotor activity. The tyrosine hydroxylase inhibitor α-methyl-ρ-tyrosine did not influence the anti-immobility effect of amantadine in the TST. Pretreatment with the α1 adrenergic receptor antagonist prazosin, ß adrenergic receptor antagonist propranolol, α2 adrenergic receptor antagonist yohimbine, and α2 adrenergic receptor agonist clonidine did not alter the antidepressant-like effect of amantadine. However, amantadine's effect was blocked by the dopamine D2 receptor antagonist haloperidol and glutamate receptor agonist N-methyl-D-aspartate (NMDA). Repeated amantadine administration (50 mg/kg) also exerted an antidepressant-like effect, paralleled by an increase in hippocampal neurogenesis. The present results demonstrate that the antidepressant-like effects of amantadine may be mediated by its actions on D2 and NMDA receptors and likely involve hippocampal neurogenesis.

Efficacy and safety of amantadine for the treatment of L-DOPA-induced dyskinesia.

L-DOPA induced dyskinesias (LIDs) may affect up to 40% of Parkinson's disease (PD) and impact negatively health-related quality of life. Amantadine has demonstrated significant antidyskinetic effects in animal PD models and in randomized double-blind placebo-controlled trials (RCTs) in patients with PD. These effects are thought to be related to the blockade of NMDA receptors modulating cortico-striatal glutamatergic-dopaminergic interactions involved in the genesis of LIDs. There are three pharmaceutical forms of amantadine currently available in the market: an oral immediate-release (IR) formulation, which is widely available; an extended-release (ER) formulation (ADS-5102) which has been recently developed and approved by the FDA; and an intravenous infusion (IV) solution, which is not commonly used in clinical practice. RCTs with amantadine IR or ER, involving more than 650 patients have shown consistent and long-lasting reductions in LIDs. Interestingly, ADS-5102 not only reduced LIDs, but also reduced significantly at the same time the duration of daily OFF-time, a unique finding compared with other antiparkinsonian medications that usually reduce time spent OFF at the cost of worsening of LIDs. Amantadine IR might also have possible effects on other PD symptoms such as apathy or fatigue. The most common adverse reactions with amantadine are constipation, cardiovascular dysfunction including QT prolongation, orthostatic hypotension and edema, neuropsychiatric symptoms such as hallucinations, confusion and delirium, nausea and livedo reticularis. Corneal degeneration is rare but critical. In summary, amantadine immediate and extended-release are effective and safe for the treatment of LIDs.

Virological surveillance and antiviral resistance of human influenza virus in Argentina, 2005–2008 / Vigilancia virológica y resistencia a los antivíricos del virus de la gripe humana en la Argentina, 2005–2008

Objective. To describe the virological characteristics of the influenza strains circulating in Argentina in 2005–2008 and to assess the prevalence of antiviral resistance. Methods. On the basis of their geographical spread and prevalence, influenza A and B isolates grown in Madin–Darby canine kidney cells were selected after antigenic and genomic characterization to be analyzed for antiviral resistance by enzymatic assay and pyrosequencing. Amantadine susceptibility was evaluated by pyrosequencing for known resistance markers on 45 strains of influenza A. Susceptibility to oseltamivir and zanamivir was evaluated by enzymatic assay of 67 influenza A and 46 influenza B strains, some of which were further analyzed by sequencing the neuraminidase gene. Results. Resistance to amantadine was observed only on A(H3N2) strains (29/33); all of them carried the mutation S31N in their M2 sequence. Oseltamivir resistance was observed in 12 (34.3%) of the 35 A(H1N1) strains from 2008; all of them carried the mutation H275Y in their neuraminidase sequence. All these viruses remained sensitive to zanamivir. Conclusions. This study describes a high incidence of amantadine-resistant influenza A(H3N2) viruses since 2006 and an unprecedented increase in oseltamivir resistance detected only in influenza A(H1N1) viruses isolated in 2008. Influenza A and B viruses were more sensitive to oseltamivir than to zanamivir, and influenza A viruses were more sensitive to both neuraminidase inhibitors than the influenza B viruses. The national data generated and analyzed in this study may help increase knowledge about influenza antiviral drug resistance, which is a problem of global concern.

Objetivo. Describir las características virológicas de las cepas de virus de la gripe que circulaban en la Argentina entre el 2005 y el 2008, y evaluar la prevalencia de la resistencia a los antivíricos. Métodos. Según su diseminación geográfica y su prevalencia, se seleccionaron aislados de gripe A y B cultivados en células renales caninas de Madin-Darby después de su caracterización antigénica y genómica, y se analizó su resistencia a los antivíricos mediante análisis enzimático y pirosecuenciación. La sensibilidad a la amantadina se evaluó por pirosecuenciación para los marcadores conocidos de resistencia en 45 cepas de gripe A. La sensibilidad al oseltamivir y al zanamivir se evaluó mediante análisis enzimático de 67 cepas de gripe A y 46 cepas de gripe B, algunas de las cuales se analizaron en mayor profundidad mediante la secuenciación del gen de la neuraminidasa. Resultados. Se observó resistencia a la amantadina solo en las cepas de gripe A (H3N2) (29/33); todas ellas tenían la mutación S31N en su secuencia de M2. Se observó resistencia al oseltamivir en 12 (34,3%) de las 35 cepas de gripe A (H1N1) aisladas en el 2008; todas ellas tenían la mutación H275Y en su secuencia de neuraminidasa. Todos estos virus conservaron su sensibilidad al zanamivir. Conclusiones. En este estudio se describe una incidencia elevada del virus de la gripe A (H3N2) resistente a la amantadina desde el 2006 y un aumento sin precedentes de la resistencia al oseltamivir detectada solo en los virus de la gripe A (H1N1) aislados en el 2008. Los virus de la gripe A y B fueron más sensibles al oseltamivir que al zanamivir y los virus de la gripe A fueron más sensibles a ambos inhibidores de la neuraminidasa que los virus de la gripe B. Los datos nacionales generados y analizados en este estudio pueden ayudar a aumentar los conocimientos acerca de la resistencia a los fármacos antivíricos dirigidos contra el virus de la gripe, lo que es un motivo de preocupación mundial.

[The 2009 pandemic influenza in Russia. I. Diagnosis and molecular biological characteristics of the virus].

The analysis of 1558 clinical samples revealed influenza virus A(H1N1v) RNA in 339 patients with influenza and 163 fatal cases,which was made in May to December 2009. Data on the antigenic properties of more than 250 of pandemic virus strains isolated at the Research Institute of Influenza and the molecular genetic characteristics of 31 strains are presented. All the test isolates were found to have the S203 substitution in hemagglutinin, which was characteristic of one of 5 minor genome A(H1N1v) virus variants found in the United States and Mexico in 2009. All the test strains contain the S31N substitution in the M2 protein, which determines viral resistance to adamantine, and have no H275Y substitution in neuraminidase, which determines oseltamivir resistance. The substitution of amino acid residue of Asp to Gly at position 222 of HA was found in 8 (73%) of 11 isolates from postmortem lung and trachea samples and in 2 (10%) of 20 isolates from nasopharyngeal swabs. The determination of the pathogenic role of this substitution calls for further investigations.

Virological surveillance and antiviral resistance of human influenza virus in Argentina, 2005-2008.

OBJECTIVE: To describe the virological characteristics of the influenza strains circulating in Argentina in 2005-2008 and to assess the prevalence of antiviral resistance. METHODS: On the basis of their geographical spread and prevalence, influenza A and B isolates grown in Madin-Darby canine kidney cells were selected after antigenic and genomic characterization to be analyzed for antiviral resistance by enzymatic assay and pyrosequencing. Amantadine susceptibility was evaluated by pyrosequencing for known resistance markers on 45 strains of influenza A. Susceptibility to oseltamivir and zanamivir was evaluated by enzymatic assay of 67 influenza A and 46 influenza B strains, some of which were further analyzed by sequencing the neuraminidase gene. RESULTS: Resistance to amantadine was observed only on A(H3N2) strains (29/33); all of them carried the mutation S31N in their M2 sequence. Oseltamivir resistance was observed in 12 (34.3%) of the 35 A(H1N1) strains from 2008; all of them carried the mutation H275Y in their neuraminidase sequence. All these viruses remained sensitive to zanamivir. CONCLUSIONS: This study describes a high incidence of amantadine-resistant influenza A(H3N2) viruses since 2006 and an unprecedented increase in oseltamivir resistance detected only in influenza A(H1N1) viruses isolated in 2008. Influenza A and B viruses were more sensitive to oseltamivir than to zanamivir, and influenza A viruses were more sensitive to both neuraminidase inhibitors than the influenza B viruses. The national data generated and analyzed in this study may help increase knowledge about influenza antiviral drug resistance, which is a problem of global concern.

DNA damage in brain cells and behavioral deficits in mice after treatment with high doses of amantadine.

Amantadine (AMA) is an uncompetitive antagonist of the N-methyl-d-aspartate receptor, with clinical application, acting on treatment of influenza A virus and Parkinson's disease. It has been proposed that AMA can indirectly modulate dopaminergic transmission. In high doses, the central nervous system is its primary site of toxicity. To examine deleterious effects on CNS induced by AMA, this study evaluated possible neurobehavioral alterations induced by AMA such as stereotyped behavior, the effects on locomotion and memory and its possible genotoxic/mutagenic activities. Adult male CF-1 mice were treated with a systemic injection of AMA (15, 30 or 60 mg kg(-1) ) 20 min before behavioral tasks on open field and inhibitory avoidance. Higher AMA doses increased the latency to step-down inhibitory avoidance test in the training session in the inhibitory avoidance task. At 60 mg kg(-1) AMA induced impairing effects on locomotion and exploration and hence impaired habituation to a novel environment. Stereotyped behavior after each administration in a 3-day trial was observed, suggesting effects on dopaminergic system. Amantadine was not able to induce chromosomal mutagenesis or toxicity on bone marrow, as evaluated by the micronucleus assay. At the lowest dose tested, AMA did not induce DNA damage and it was unable to impair memory, locomotion, exploration or motivation in mice. However, higher AMA doses increased DNA damage in brain tissue, produced locomotor disturbances severe enough to preclude testing for learning and memory effects, and induced stereotypy, suggesting neurotoxicity.

Effects of amantadine on circulating neurotransmitters in healthy subjects.

Considering that glutamatergic axons innervate the C1(Ad) medullary nuclei, which are responsible for the excitation of the peripheral adrenal glands, we decided to investigate catecholamines (noradrenaline, adrenaline and dopamine) plus indolamines (plasma serotonin and platelet serotonin) at the blood level, before and after a small oral dose of amantadine, a selective NMDA antagonist. We found that the drug provoked a selective enhancement of noradrenaline plus a minimization of adrenaline, dopamine, plasma serotonin and platelet serotonin circulating levels. Significant enhancement of diastolic blood pressure plus reduction of systolic blood pressure and heart rate paralleled the circulating parameter changes. The above findings allow us to postulate that the drug was able to enhance the peripheral neural sympathetic activity. Minimization of both adrenal sympathetic and parasympathetic activities was also registered after the amantadine challenge. The above findings supported the postulation that this drug should be a powerful therapeutic tool for treating diseases affected by adrenal sympathetic hyperactivity.

Búsqueda de resistencia a amantadina en cepas de virus influenza A aisladas en Santiago de Chile, entre los años 2001 y 2002 / Search of amantadine-resistance in influenza A strains isolated in Santiago, Chile, 2001-2002

Amantadina es un fármaco eficaz para el tratamiento y prevención de influenza A. Su mecanismo de acción es inhibir la proteína M2. Su uso por períodos prolongados puede generar resistencia, la cual ocurre por mutaciones en el gen que codifica para la proteína M2. La mutación más frecuentemente encontrada se ubica en la posición 31. El uso de técnicas de biología molecular permite detectar estas mutaciones. Los objetivos fueron determinar la existencia de resistencia a amantadina en cepas de virus influenza A aisladas entre los años 2001 y 2002 en un laboratorio de virología en Santiago de Chile, y validar un nuevo método de biología molecular para reconocer cepas resistentes. Para ello se utilizó metodología de RPC y análisis de tamaño de fragmentos de restricción. En 31 cepas procesadas no se observó la presencia de cambios en la posición 31. Estos hallazgos sugieren que la resistencia a amantadina es muy baja o está ausente en nuestro medio. Esto podría explicarse por un limitado uso de este fármaco en esta población. El método descrito puede servir de base para un monitoreo prospectivo de resistencia, que pueda ser de utilidad al médico clínico.

[Search of amantadine-resistance in influenza A strains isolated in Santiago, Chile, 2001-2002]. / Búsqueda de resistencia a amantadina en cepas de virus influenza A aisladas en Santiago de Chile, entre los años 2001 y 2002.

Amantadine has been used for prevention and treatment of influenza A infection. It blocks the proton through the M2 ion channel. Drug-resistant viruses appear quickly when this therapy is used. Single amino acids changes in the H2 protein can confer resistance, being the most frequent one in position 31. Different methods to detect resistant strains have been described. The objectives were to determine the existence of amantadine resistance of influenza A strains isolated in a virologic laboratory in Santiago, Chile, between 2001-2002, and to validate a new molecular method to detect resistant strains. A PCR restriction fragment length polymorphism analysis was employed for the detection of resistant viruses. In 31 processed strains no mutation in the position 31 was found. This result supports that amantadine resistance is very low or absent in Chile. This could be explained by a limited use of this drug in the study population. This method could be used as a monitoring system to survey resistant viruses.

Dopamine and NMDA systems modulate long-term nociception in the rat anterior cingulate cortex.

The anterior cingulate cortex (ACC) plays a key role in pain processing. It has been reported that increased activity of glutamatergic projections into the ACC intensifies nociception; whereas dopaminergic projections inhibit it. The aim of this study was to evaluate the role of dopaminergic and NMDA systems of the ACC in the modulation of long-term nociception elicited by sciatic denervation in the rat. Score, onset and incidence of long-term nociception were measured by the autotomy behavior. The effects of a single microinjection into the ACC of different doses of dopamine (100 nM, 100 microM and 100 mM), a NMDA receptor antagonist (MK801 200 nM and 9.34 mM) and amantadine, a dopamine agonist and NMDA receptor antagonist (10, 100 and 1000 microM) were tested on long-term nociception. Dopamine diminished autotomy behavior in an inverse dose-dependent manner, with dopamine 100 nM as most effective concentration. MK801 and amantadine elicited a significant reduction on autotomy score. Prior injections of D1 and D2 receptor antagonists blocked the antinociceptive effects of amantadine on long-term nociceptive behavior. The present study suggests an interaction between dopaminergic and glutamatergic systems within the ACC in the genesis and maintenance of long-term nociception.

Fármacos antiparkinsonianos / Anti-Parkinson drugs

La enfermedad de Parkinson requiere la administración de diversos fármacos. En el siguiente artículo se describen las diferentes drogas utilizadas, -agentes dopaminérgicos, anticolinérgicos y neuroprotectores- sus modos de acción, efectos adversos y precauciones y advertencias (AU)

Study of the biodistribution of the amantadine labelled with technetium-99m in Wistar female rats.

Amantadine (AMA) has been described as dopamine stimulant and norepineprhine release, capable to block the N-methyl-D aspartate (NMDA) glutamatergic and nicotinic receptors, enhancing the sexual behavior of the male rats and inducing hypersexuality in humans. The use of technetium-99m (99mTc) can be justified for its physical and chemical properties. The aim of this study was to label and evaluate the bioavailability of the AMA labelled with 99mTc (99mTc-AMA) in Wistar female rats. The solution of 99mTc-AMA was administered by intraperitoneal way and the animals were sacrificed in CO2 chamber 10 min after the administration of the radiotracer. Various organs were removed, weighted, their radioactivity was determined using an auto-gamma counter and the results were expressed as the percentage of the injected activity per gram of tissue (%ATI/g). In the control group only Na99mTcO4 was administered. The analysis of results shows that the highest uptakes 99mTc-AMA treated group were: ovary (7.11 +/- 1.43), spleen (3.54 +/- 1.05), thyroid (2.67 +/- 0.15), stomach (1.56 +/- 1.10), duodenum (0.87 +/- 0.52), muscular tissue (0.57 +/- 0.06), liver (0.52 +/- 0.25), and at control group: thyroid (16.45 +/- 2.57), ovary (1.28 +/- 0.12), liver (1.10 +/- 0.04), spleen (0.57 +/- 0.07) and muscular tissue (0.26 +/- 0.03). The results obtained suggest that 99mTc-AMA may be used to study the bioavailability of amantadine and evaluate its effect in sexual behavior in female rats.

How REM sleep deprivation and amantadine affects male rat sexual behavior.

There are conflicting findings about the sexual effects of REM sleep deprivation (REMd). Otherwise, several studies show a dopaminergic hypersensitivity after REMd. The effect of REMd and amantadine (AMA) was studied for standard measures and temporal patterning in the first experiment, in four groups: normal with vehicle, normal with AMA (5.0 and 10 mg/kg), REMd with vehicle and REMd with AMA (5.0 and 10.0 mg/kg). REMd reduced mount latency (ML), intromission latency (IL) and mount number (MN) and increased copulatory efficiency (CE) and hit rate factor. REMd also reduced the mount bout number (MBN) and increased the sexual interaction (mount bout time, MBT) among male and female during copula. AMA stimulates initiation and hit rate factors and accelerates the temporal patterning of sexual behavior, evoking fewer and quicker mount bouts. In the experiments with combination of REMd and AMA administration, AMA did not increase behavior effects evoked by REM deprivation, probably due to a top or a bottom effect, depending on the measures considered. A second experiment studied the effects of AMA (1.25 to 5.0 mg/kg) and REMd on the sexual reflexes of nonimmobilized male rats. REMd enhanced the AMA effects upon the seminal emission reflex, but inhibited the penile erection reflex elicited by 1.25 mg/kg of AMA. Curiously, our results showed that REMd, like AMA, a dopaminergic agonist, causes similar effects of sexual behavior in the male rat, particularly those related to arousal mechanism and hit rate factor. The results are discussed and the effects of REMd probably involve dopaminergic hypersensitivity and increased sexual motivational response.

Psicobiologia do comportamento sexual: amantadina como ferramenta farmacológica para o estudo dos diferentes componentes da resposta sexual de ratos / Sexual behavior psicobiology: amantadine as pharmacological tool for studying the different components of rats sexual response

O comportamento sexual humano apresenta três componentes distintos e obviamente integrados, respectivamente denominados de desejo, exitação e orgasmo sexuais, regulados por diferentes circuitos neurais. Em ratos, a resposta sexual também apresenta diferentes componentes, tendo sido descrita a existência de quatro fatores: fator de iniciação, relacionado à motivação sexual, fator de contagem de intromissões, da taxa copulatória e da taxa de sucesso copulatório, relacionadas co componente consumatório da resposta sexual. O comportamento sexual de ratos, e provavelmente de outros mamíferos, apresenta ainda uma organização temporal, organizada em "mount bouts". Neste trabalho, procuramos estudar os diferentes componentes de resposta sexual de ratos, utilizando a amantadina, droga que facilitou todos os componentes da resposta sexual e evocou os reflexos genitais de ratos, como ferramenta farmacológica; desenvolvemos um modelo experimental de disfunção erétil através do tratamento crônico com L-NAME, investigando os efeitos da amantadina nesses animais; estudamos os efeitos da privação de sono REM na resposta sexual de ratos, bem como os efeitos da amantadina nos animais privados de sono REM; e investigamos a influência do bloqueio +1 noradrenérgico e da ativação 5-HT1B serotoninérgica nos efeitos da amantadina sobre o comportamento sexual. Nossos resultados ratificam a ação estimulante da amantadina na resposta sexual, sobretudo nos componente motivacional, no sucesso copulatório e na organização temporal do comportamento sexual; demonstram que a privação de sono REM per se facilita ambos os componentes motivacional e da taxa de sucesso copulatório e potencia as ações da amantadina sobre a organização temporal do comportamento sexual; e que esses efeitos da privação de sono REM não decorrem do estresse associado à técnica de privação utilizada. O tratamento crônico com L-NAME forneceu um importante modelo experimental para o estudo de disfunções eréteis, sugerindo que os efeitos dessa droga envolvem principalmente mecanismos periféricos, uma vez que foi revertido pelo co-tratamento com três potentes vasodilatadores. unção ejaculatória, provavelmente por mecanismos centrais, sem alterar os demais componentes, enquanto a ativação 5-HT1B inibiu o componente motivacional, a resposta ejaculatória, reduziu a taxa de sucesso copulatório, mas não modificou significamente a resposta erétil, nem a organização temporal do comportamento sexual dos animais...

Amantadine stimulates sexual behavior in male rats.

The effects of amantadine on sexual behavior, penile erection, and seminal emission of male rats was studied. Amantadine significantly decreased latency of mounts in all doses (1.25 to 50 mg/kg), and decreased the number of mounts and intromission latency at the highest doses used. The lowest dose of amantadine significantly increased ejaculation latency and intromission frequency, while higher doses significantly reduced it, which indicates a biphasic response of the drug. Additionally, seminal emission, erections, and genital grooming were significantly induced by amantadine. Amantadine-induced seminal emissions were impaired by spinal cord transection, which suggests the involvement of supraspinal structures in the drug action. Haloperidol and atropine sulphate significantly reduced seminal emissions and penile erections induced by amantadine. These results demonstrate that amantadine stimulates sexual behavior and genital reflexes in male rats and suggest a facilitatory effect of the drug that probably involves different mechanisms of action.

Parkinsonismo: aspéctos clínicos, biológicos e terapêuticos / Parkinsonism: clinical, biological and therapeutical aspects

O parkinsonismo idiopático (doença de Parkinson) e secundário, ou seja, que apresenta etiologia conhecida (pós-encefalítico, por toxinas, por drogas ou por algumas doenças degenerativas) såo processos neurodegenerativos que afetam progressivamente as funçÆes motoras normais do indivíduo. Neste trabalho, os aspéctos clínicos, etiológicos, fisiopatológicos e principalmente a descriçåo do arsenal farmacológico e as medidas utilizadas no combate ou retardo medicamentoso do parkinsonismo såo abordados e discutidos

The entry of Junin virus into Vero cells.

The entry mechanism of Junin virus (JV) into Vero cells was studied analyzing the effect of lysosomotropic compounds and acid pH on JV infection. Ammonium chloride, amantadine, chlorpheniramine and procaine inhibited JV production. The action of ammonium chloride was exerted at early times of infection. Virus internalization was inhibited and viral protein expression was not detected. When the extracellular medium was buffered at low pH, the ammonium chloride induced block on JV infection was overcome. Furthermore, JV was able to induce fusion of infected cells at pH 5.5 leading to polykaryocyte formation. Taken together, these results demonstrate that JV entry occurs through an endocytic mechanism requiring a low pH dependent membrane fusion.