RESUMO
Molecular vibrations and quantum tunneling may link ligand binding to the function of pharmacological receptors. The well-established lock-and-key model explains a ligand's binding and recognition by a receptor; however, a general mechanism by which receptors translate binding into activation, inactivation, or modulation remains elusive. The Vibration Theory of Olfaction was proposed in the 1930s to explain this subset of receptor-mediated phenomena by correlating odorant molecular vibrations to smell, but a mechanism was lacking. In the 1990s, inelastic electron tunneling was proposed as a plausible mechanism for translating molecular vibration to odorant physiology. More recently, studies of ligands' vibrational spectra and the use of deuterated ligand analogs have provided helpful information to study this admittedly controversial hypothesis in metabotropic receptors other than olfactory receptors. In the present work, based in part on published experiments from our laboratory using planarians as an experimental organism, I will present a rationale and possible experimental approach for extending this idea to ligand-gated ion channels.
Assuntos
Vibração , Ligantes , Animais , Teoria Quântica , Humanos , Receptores Odorantes/metabolismo , Receptores Odorantes/química , Ligação ProteicaRESUMO
Planarians are rapidly developing into very useful research subjects in pharmacology and neuroscience research. Here we report that curare, a cholinergic nicotinic receptor antagonist, alleviates the nicotine-induced planarian seizure-like movements (pSLM) by up to 50 % at equimolar concentrations of nicotine and curare (1 mM), while curare alone does not induce significant pSLMs. The simplest interpretation of our data is that there are nicotine induced behaviors insensitive to curare in our experimental organism. To the best of our knowledge, this is the first report on curare-insensitive, nicotine-induced effects in any organism.
Assuntos
Comportamento Animal/efeitos dos fármacos , Curare/farmacologia , Movimento/efeitos dos fármacos , Nicotina/farmacologia , Planárias/efeitos dos fármacos , Animais , Planárias/metabolismo , Convulsões/induzido quimicamenteRESUMO
Previous work from our laboratory showed that cotinine, a nicotine metabolite, reverses three nicotine-induced behavioral effects in freshwater planarians: motility decrease, seizure-like movements, and withdrawal-like behaviors. The present work explored whether cotinine, a nicotine metabolite, antagonized the nicotine-induced effects on planarian motility in a concentration-dependent manner. We found that nicotine decreased planarian motility at nicotine concentrations above 60 µM but increased planarian velocity at concentrations equal to or below 50 µM, in agreement with previous data. Cotinine did not affect planarian motility at a concentration range between 250 and 2750 µM. Furthermore, we found that cotinine alleviated the 100 µM nicotine-induced motility decrease in a concentration-dependent manner and reversed the low nicotine concentration motility increase, albeit in a concentration-independent manner. The apparent concentration-dependent alleviation of >60 µM nicotine-induced motility decrease by cotinine suggests an orthosteric relationship between nicotine and cotinine. On the other hand, the evident concentration-independent cotinine alleviation of the increase in motility induced by 50 µM nicotine suggests an allosteric relationship. Our data is consistent with the existing literature about the relationship between nicotine and cotinine in various models, reinforcing the case for the usefulness of the planarian model in pharmacological studies.
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The tobacco cembranoid (1S,2E,4R,6R,7E,11E)-2,7,11-cembratriene-4,6-diol (4R) interacts with nicotinic acetylcholine receptors, which results in neuroprotection against organophosphate toxicity, brain ischemia, and Parkinson's disease. The present study is a continuation of our previous research in which we applied a single dose of 4R 1 h before or 24 h after exposure to diisopropylfluorophosphate (DFP) (analog of the nerve agent sarin). The 4R dose robustly decreased neuroinflammation and neuronal death at both timepoints. Here, we investigated the toxicity of a single dose of 4R in male and female Sprague Dawley (SD) rats after a subcutaneous (s.c.) injection of 6, 24, or 98 mg/kg. Body weight was not affected by 4R during the 7-day observation period. No histopathologic changes in the organs were attributed to 4R. Minor hematological and blood composition variations were detected on Day 3 in the mid- and the high-dose males, but these were resolved by Day 8. At the area of the s.c. injection site, alopecia and dry skin were detected in both the 4R-treated males and females and in the female controls.
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Combinatorially synthesized nucleotide polymers have been used during the last decade to find ligands that bind to specific sites on biological molecules, including membrane-bound proteins such as the nicotinic acetylcholine receptors (nAChRs). The neurotransmitter receptors belong to a group of four structurally related proteins that regulate signal transmission between ~10(11) neurons of the mammalian nervous system. The nAChRs are inhibited by compounds such as the anticonvulsant MK-801 [(+)-dizocilpine] and abused drugs such as cocaine. Based on predictions arising from the mechanism of receptor inhibition by MK-801 and cocaine, we developed two classes of RNA aptamers: class I members, which inhibit the nAChR, and class II members, which alleviate inhibition of the receptor by MK-801 and cocaine. The systematic evolution of ligands by the exponential enrichment (SELEX) method was used to obtain these compounds. Here, we report that we have truncated RNA aptamers in each class to determine the minimal nucleic acid sequence that retains the characteristic function for which the aptamer was originally selected. We demonstrate that a truncated class I aptamer containing a sequence of seven nucleotides inhibits the nAChR and that a truncated class II aptamer containing a sequence of only four nucleotides can alleviate MK-801 inhibition.
Assuntos
Aptâmeros de Nucleotídeos/farmacologia , Receptores Nicotínicos/efeitos dos fármacos , Animais , Aptâmeros de Nucleotídeos/síntese química , Aptâmeros de Nucleotídeos/química , Sequência de Bases , Linhagem Celular , Cocaína/farmacologia , Maleato de Dizocilpina/farmacologia , Cinética , Camundongos , Modelos Teóricos , Dados de Sequência Molecular , Conformação de Ácido NucleicoRESUMO
Planarians are traditional model invertebrates in regeneration and developmental biology research that also display a variety of quantifiable behaviors useful to screen for pharmacologically active compounds. One such behavior is the expression of seizure-like movements (pSLMs) induced by a variety of substances. Previous work from our laboratory showed that cocaine, but not nicotine, induced pSLMs in intact but not decapitated planarians. Interestingly, as decapitated planarians regenerated their heads, they gradually recovered their sensitivity to cocaine. These results suggested a method to assess planarian brain regeneration and a possible way of identifying compounds that could enhance or hold back brain regeneration. In the present work, we demonstrate that the cholinergic agent cytisine is a suitable reference compound to apply our method. Cytisine induces pSLMs in a concentration-dependent manner in intact (but not decapitated) planarians of the species Girardia tigrina. Based on our data, we developed a behavioral protocol to assess planarian brain regeneration over time. We tested this method to measure the effect of ethanol on G. tigrina's brain regeneration. We found that ethanol slows down the rate of planarian brain regeneration in a concentration-dependent manner, consistently with data from other research groups that tested ethanol effects on planarian brain regeneration using different behavioral protocols. Thus, here we establish a general method using cytisine-induced pSLMs as an indicator of brain regeneration in planarians, a method that shows potential for assessing the effect of pharmacologically active compounds in this process.
Assuntos
Alcaloides/farmacologia , Encéfalo/efeitos dos fármacos , Colinérgicos/farmacologia , Planárias/fisiologia , Regeneração/efeitos dos fármacos , Anestésicos Locais/farmacologia , Animais , Azocinas/farmacologia , Encéfalo/fisiologia , Carbacol/farmacologia , Cocaína/farmacologia , Cotinina/farmacologia , Relação Dose-Resposta a Droga , Etanol/farmacologia , Nicotina/farmacologia , Quinolizinas/farmacologia , Regeneração/fisiologia , Convulsões/fisiopatologiaRESUMO
In this work we describe a series of simple protocols using planaria as an animal model in toxicology and behavioral pharmacology. These procedures have proven useful to provide significant research experience to undergraduate students, including coauthorship in peer-reviewed publications. The methods described in this work have proven useful to allow students to visualize concepts related to concentration-effect curves for toxicity and behavioral experiments, without the need to consider factors that must be taken into account when working with vertebrate animals.
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One of the most important aspects of the scientific endeavour is the definition of specific concepts as precisely as possible. However, it is also important not to lose sight of two facts: (i) we divide the study of nature into manageable parts in order to better understand it owing to our limited cognitive capacities and (ii) definitions are inherently arbitrary and heavily influenced by cultural norms, language, the current political climate, and even personal preferences, among many other factors. As a consequence of these facts, clear-cut definitions, despite their evident importance, are oftentimes quite difficult to formulate. One of the most illustrative examples about the difficulty of articulating precise scientific definitions is trying to define the concept of a brain. Even though the current thinking about the brain is beginning to take into account a variety of organisms, a vertebrocentric bias still tends to dominate the scientific discourse about this concept. Here I will briefly explore the evolution of our 'thoughts about the brain', highlighting the difficulty of constructing a universally (or even a generally) accepted formal definition of it and using planarians as one of the earliest examples of organisms proposed to possess a 'traditional', vertebrate-style brain. I also suggest that the time is right to attempt to expand our view of what a brain is, going beyond exclusively structural and taxa-specific criteria. Thus, I propose a classification that could represent a starting point in an effort to expand our current definitions of the brain, hopefully to help initiate conversations leading to changes of perspective on how we think about this concept. This article is part of the theme issue 'Liquid brains, solid brains: How distributed cognitive architectures process information'.
Assuntos
Encéfalo/fisiologia , Planárias/fisiologia , Animais , Terminologia como AssuntoRESUMO
We recently reported that parthenolide and related sesquiterpene lactones are able to prevent and reverse behavioral responses in planarian worms induced by acute cocaine exposure. Previous reports indicate that when planarians are chronically exposed to microM concentrations of cocaine, they display stereotypical withdrawal-like behaviors when the cocaine is removed. Here we report that parthenolide prevents this cocaine-induced expression of planarian withdrawal-like behaviors.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Comportamento Animal/efeitos dos fármacos , Cocaína/efeitos adversos , Planárias/fisiologia , Sesquiterpenos/farmacologia , Síndrome de Abstinência a Substâncias/tratamento farmacológico , Síndrome de Abstinência a Substâncias/psicologia , Animais , Comportamento Estereotipado/efeitos dos fármacosRESUMO
Here we report the prevention and reversal of cocaine-induced behaviors in planarian worms by parthenolide and two related cyclic sesquiterpene lactones (SL), costunolide and santonin. Using established protocols, we studied two cocaine-induced behavioral effects in planaria; the induction of motility decrease and the induction of C-like hyperkinesia. Cocaine, parthenolide, costunolide, santonin, and a lactone-less cyclic sesquiterpene, beta-eudesmol, decreased planarian motility in a concentration-dependent manner. Only cocaine induced C-like hyperkinesia. At concentrations that did not show any motility decrease, parthenolide, costunolide and santonin, but not beta-eudesmol, significantly reduced the cocaine-induced motility decrease and C-like hyperkinesia, in a concentration-dependent manner. Furthermore, parthenolide, costunolide and santonin were able to rescue planaria from C-like hyperkinesia, after the worms were exposed to cocaine. Conversely, cocaine at a concentration that did not show any measurable effects (10 microM), was able to alleviate the SL-, but not the beta-eudesmol-induced motility decrease. Liquid Chromatography/Mass Spectrometry experiments demonstrated that cocaine does not interact directly with any of the cyclic sesquiterpenoids, which suggests specific biochemical targets for these compounds in planarians. Our data suggests a common binding site for cocaine and the sesquiterpene lactones in planarians.
Assuntos
Comportamento Animal/efeitos dos fármacos , Cocaína/antagonistas & inibidores , Cocaína/farmacologia , Inibidores da Captação de Dopamina/antagonistas & inibidores , Inibidores da Captação de Dopamina/farmacologia , Planárias/fisiologia , Sesquiterpenos/farmacologia , Algoritmos , Animais , Cromatografia Líquida de Alta Pressão , Cocaína/análise , Inibidores da Captação de Dopamina/análise , Relação Dose-Resposta a Droga , Hipercinese/induzido quimicamente , Hipercinese/psicologia , Espectrometria de Massas , Atividade Motora/efeitos dos fármacos , Santonina/farmacologia , Sesquiterpenos/análise , Sesquiterpenos de Eudesmano/farmacologia , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
Although planarians are established model organisms in developmental biology and regeneration studies, in the last forty years or so, they have caught the attention of pharmacologists, especially to study the pharmacology of drugs of abuse. This review covers the following topics: some fundamentals of the history of animal models and planarians in biomedical research; an abbreviated story of systematic pharmacology research using planarians as a model organism; an example of how planarians are contributing to the search for compounds against acute cocaine toxicity; an analysis of the number of papers on planarians and pharmacological topics from 1900-2016; some perspectives on pharmacology in developmental and regeneration studies, arguing in favor of the planarian model as a leading subject for this interdisciplinary area of research, and finally some concluding thoughts.
Assuntos
Comportamento Animal/efeitos dos fármacos , Cocaína/toxicidade , Biologia do Desenvolvimento , Modelos Animais de Doenças , Planárias/citologia , Planárias/fisiologia , Regeneração/fisiologia , Animais , Inibidores da Captação de Dopamina/toxicidade , Planárias/efeitos dos fármacosRESUMO
In this work, we describe aspects of the toxicity and behavioral effects of dimethylsulfoxide (DMSO) in planaria. Planarian worms have traditionally been a favored animal model in developmental biology. More recently, this organism is being recognized as an animal model in neuropharmacology research. DMSO is often used in cell and tissue culture as a cryoprotectant agent and is also commonly used to enhance the solubility of hydrophobic drugs in aqueous solutions. This compound can elicit various physiological effects in both vertebrates and invertebrates. Many drugs and drug candidates are hydrophobic, needing solvents like DMSO to be able to reach their physiological targets. As planaria becomes increasingly popular in neuropharmacology research, a description of the DMSO effects in this organism is essential. We found that DMSO is toxic to planarians at concentrations above 5% (705 mM), with an LD(50) of 10% (1.4M) at exposure times above 5 min. At sub-toxic concentrations, DMSO decreases planarian exploratory behavior in a concentration-dependent manner. This reduction in locomotor behavior is reversible and preincubation-independent. DMSO at a concentration of 0.1% (14.1 mM), which is usually enough to solubilize hydrophobic substances in aqueous solutions, did not display any toxic or behavioral effects in planaria. Therefore, in this animal model, DMSO concentrations above 0.1% should be avoided in order to be able to reliably observe any behavioral or toxic effects of hydrophobic drugs.
Assuntos
Dimetil Sulfóxido/toxicidade , Animais , Comportamento Animal , Crioprotetores/toxicidade , Dose Letal Mediana , Planárias , Solventes/toxicidadeRESUMO
Nicotine is one of the most addictive drugs abused by humans. Our laboratory and others have demonstrated that nicotine decreases motility and induces seizure-like behavior in planarians (pSLM, which are vigorous writhing and bending of the body) in a concentration-dependent manner. Nicotine also induces withdrawal-like behaviors in these worms. Cotinine is the major nicotine metabolite in humans, although it is not the final product of nicotine metabolism. Cotinine is mostly inactive in vertebrate nervous systems and is currently being explored as a molecule which possess most of nicotine's beneficial effects and few of its undesirable ones. It is not known whether cotinine is a product of nicotine metabolism in planarians. We found that cotinine by itself does not seem to elicit any behavioral effects in planarians up to a concentration of 1mM. We also show that cotinine antagonizes the aforementioned nicotine-induced motility decrease and also decreases the expression of nicotine-induced pSLMs in a concentration-dependent manner. Also cotinine prevents the manifestation of some of the withdrawal-like behaviors induced by nicotine in our experimental organism. Thus, we obtained evidence supporting that cotinine antagonizes nicotine in this planarian species. Possible explanations include competitive binding of both compounds at overlapping binding sites, at different nicotinic receptor subtypes, or maybe allosteric interactions.
Assuntos
Comportamento Animal/efeitos dos fármacos , Cotinina/farmacologia , Nicotina/farmacologia , Agonistas Nicotínicos/farmacologia , Antagonistas Nicotínicos/farmacologia , Planárias/efeitos dos fármacos , Animais , Sítios de Ligação , Relação Dose-Resposta a Droga , Receptores Nicotínicos/metabolismoRESUMO
Planarians are traditional animal models in developmental and regeneration biology. Recently, these organisms are arising as vertebrate-relevant animal models in neuropharmacology. Using an adaptation of published behavioral protocols, we have described the alleviation of cocaine-induced planarian seizure-like movements (pSLM) by a naturally-occurring sesquiterpene lactone, parthenolide. Interestingly, parthenolide does not prevent the expression of pSLM induced by amphetamines; in vertebrates, amphetamines interact with the same protein target as cocaine. Parthenolide is also unable to prevent pSLM elicited by the cholinergic com-pounds nicotine and cytisine or by the glutamatergic agents L- or D- glutamic acid or NMDA. Thus, we conclude that parthenolide is a specific anti-cocaine agent in this experimental organism.
Assuntos
Comportamento Animal/efeitos dos fármacos , Cocaína/antagonistas & inibidores , Cocaína/toxicidade , Planárias/efeitos dos fármacos , Convulsões/tratamento farmacológico , Sesquiterpenos/uso terapêutico , Animais , Anti-Inflamatórios não Esteroides/uso terapêutico , Inibidores da Captação de Dopamina/antagonistas & inibidores , Inibidores da Captação de Dopamina/toxicidade , Agonistas de Aminoácidos Excitatórios/toxicidade , Metanfetamina/toxicidade , N-Metilaspartato/toxicidade , Nicotina/toxicidade , Agonistas Nicotínicos/toxicidade , Planárias/citologia , Regeneração/efeitos dos fármacos , Convulsões/induzido quimicamenteRESUMO
Chronic cocaine administration leads to catecholamine reuptake inhibition which enhances reward and motivational behaviors. Ventral Tegmental Area dopaminergic (VTA DA) neuronal firing is associated with changes in reward predictive signals. Acute cocaine injections inhibit putative VTA DA cell firing in vertebrates. Parthenolide, a compound isolated from the feverfew plant (Tanacetum parthenium), has been shown to substantially inhibit cocaine's locomotion effects in a planarian animal model (Pagán et al., 2008). Here we investigated the effects of parthenolide on the spontaneous firing activity of putative VTA DA neurons in anesthetized male rats (250-300g). Single-unit recordings were analyzed after intravenous (i.v.) parthenolide administration followed by 1mg/kg i.v. cocaine injection. Results showed that parthenolide at 0.125 mg/kg and 0.250mg/kg significantly blocked cocaine's inhibitory effect on DA neuronal firing rate and bursting activity (p< 0.05, two way ANOVA). We propose that parthenolide might inhibit cocaine's effects on VTA DA neurons via its interaction with a common binding site at monoamine transporters. It is suggested that parthenolide could have a potential use as an overdose antidote or therapeutic agent to cocaine intoxication.
RESUMO
We recently reported that the natural cyclic lactone, parthenolide, and related analogs prevent the expression of behavioral effects induced by cocaine in planarians and that parthenolide's γ-lactone ring is required for this effect. In the present work, we tested a series of alkyl γ-lactones with varying chain length (1-8 carbons) to determine their ability to antagonize the planarian motility decrease induced by 200 µM cocaine. Alkyl lactones with up to a 4-carbon alkyl chain did not affect planarian motility or antagonized the cocaine-induced motility decrease; only the compound γ-nonalactone (a γ-lactone with a 5-carbon chain) was able to prevent the cocaine-induced behavioral patterns, while alkyl lactones with longer carbon chains failed to prevent the cocaine-induced effects. Thus, we conclude that the optimal structural features of this family of compounds to antagonize cocaine's effect in this experimental system is a γ-lactone ring with at a 5-carbon long functional group.
Assuntos
Cocaína/antagonistas & inibidores , Lactonas/química , Lactonas/farmacologia , Atividade Motora/efeitos dos fármacos , Planárias/efeitos dos fármacos , Animais , Atividade Motora/fisiologia , Planárias/química , Planárias/fisiologia , Relação Estrutura-AtividadeRESUMO
Nicotinic acetylcholine receptors (AChRs) are pentameric proteins that form agonist-gated cation channels through the plasma membrane. AChR agonists and antagonists are potential candidates for the treatment of neurodegenerative diseases. Cembranoids are naturally occurring diterpenoids that contain a 14-carbon ring. These diterpenoids interact with AChRs in complex ways: as irreversible inhibitors at the agonist sites, as noncompetitive inhibitors, or as positive modulators, but no cembranoid was ever shown to have agonistic activity on AChRs. The cembranoid eupalmerin acetate displays positive modulation of agonist-induced currents in the muscle-type AChR and in the related gamma-aminobutyric acid (GABA) type A receptor. Moreover, cembranoids display important biological effects, many of them mediated by nicotinic receptors. Cembranoids from tobacco are neuroprotective through a nicotinic anti-apoptotic mechanism preventing excitotoxic neuronal death which in part could result from anti-inflammatory properties of cembranoids. Moreover, tobacco cembranoids also have anti-inflammatory properties which could enhance their neuroprotective properties. Cembranoids from tobacco affect nicotine-related behavior: they increase the transient initial ataxia caused by first nicotine injection into naive rats and inhibit the expression of locomotor sensitization to repeated injections of nicotine. In addition, cembranoids are known to act as anti-tumor compounds. In conclusion, cembranoids provide a promising source of lead drugs for many clinical areas, including neuroprotection, smoking-cessation, and anti-cancer therapies.
Assuntos
Antozoários/química , Diterpenos/farmacologia , Nicotiana/química , Receptores Nicotínicos/efeitos dos fármacos , Animais , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Diterpenos/metabolismo , Fármacos Neuroprotetores/metabolismo , Fármacos Neuroprotetores/farmacologia , Receptores Nicotínicos/metabolismoRESUMO
Using an adaptation of published behavioral protocols, we determined that acute exposure to the cholinergic compounds nicotine and carbamylcholine decreased planarian motility in a concentration-dependent manner. A tobacco cembranoid (1S,2E,4R,6R,7E,11E)-cembra-2,7,11-triene-4,6-diol (4R-cembranoid), also decreased planarian motility. Experiments in the presence of 1 microM 4R-cembranoid did increase the IC50 for nicotine- but not carbamylcholine-induced decrease in planarian motility. When planarians were exposed for 24 h to either nicotine or carbamylcholine at concentrations near their respective IC50 values and then transferred to plain media, nicotine-exposed, but not carbamylcholine- or cembranoid-exposed worms displayed withdrawal-like distress behaviors. In experiments where planarians were pre-exposed to 100 microM nicotine for 24 h in the presence of 1 microM 4R-cembranoid, the withdrawal-like effects were significantly reduced. These results indicate that the 4R-cembranoid might have valuable applications for tobacco abuse research. This experimental approach using planarians is useful for the initial screening of compounds relevant to drug abuse and dependence.
Assuntos
Comportamento Animal/efeitos dos fármacos , Diterpenos/farmacologia , Nicotiana/química , Nicotina/efeitos adversos , Síndrome de Abstinência a Substâncias/tratamento farmacológico , Síndrome de Abstinência a Substâncias/psicologia , Animais , Carbacol/efeitos adversos , Atividade Motora/efeitos dos fármacos , PlanáriasRESUMO
In spinal anesthesia, the effects of local anesthetics (LAs) are not completely explained by sodium channel inhibition. Other targets include neuronal nicotinic acetylcholine receptors (nAChRs). LA affinities for the Torpedo californica nAChR were measured by inhibition of [(3)H]TCP binding and correlated with molecular volume, surface area, molecular weight, and log of the octanol-water partition coefficients (P and D). To understand the molecular determinants important for interaction with the nAChR, ester and amide LAs were compared separately. Also, correlations with the aromatic/linker half and the hydrophilic half of the LA molecules were considered individually. The IC(50)s of the ester LAs correlated better with the molecular volume, surface area, molecular weight, and log P of the aromatic/linker half of the molecules; whereas the IC(50)s for amide LAs correlated better with the four parameters based on the hydrophilic half. These correlations were used to predict the IC(50) of various LAs (including several not studied here) and to compare these values with the published values. The predicted values of IC(50) correlated well with the published results both for neuronal and for electroplaque-desensitized nAChR, suggesting that the results can be generalized to include neuronal nAChRs.