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BACKGROUND: While many studies have assayed behavioral responses of animals to chemical, temperature and light gradients, fewer studies have assayed how animals respond to humidity gradients. Our novel humidity chamber has allowed us to study the neuromolecular basis of humidity sensation in the nematode Caenorhabditis elegans (Russell et al., 2014). NEW METHOD: We describe an easy-to-construct, low-cost humidity chamber to assay the behavior of small animals, including soft-bodied invertebrates, in controlled humidity gradients. RESULTS: We show that our humidity-chamber design is amenable to soft-bodied invertebrates and can produce reliable gradients ranging 0.3-8% RH/cm across a 9-cm long × 7.5-cm wide gel-covered arena. COMPARISON WITH EXISTING METHOD(S): Previous humidity chambers relied on circulating dry and moist air to produce a steep humidity gradient in a small arena (e.g. Sayeed and Benzer, 1996). To remove the confound of moving air that may elicit mechanical responses independent of humidity responses, our chamber controlled the humidity gradient using reservoirs of hygroscopic materials. Additionally, to better observe the behavioral mechanisms for humidity responses, our chamber provided a larger arena. Although similar chambers have been described previously, these approaches were not suitable for soft-bodied invertebrates or for easy imaging of behavior because they required that animals move across wire or fabric mesh. CONCLUSION: The general applicability of our humidity chamber overcomes limitations of previous designs and opens the door to observe the behavioral responses of soft-bodied invertebrates, including genetically powerful C. elegans and Drosophila larvae.
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Cámaras de Exposición Atmosférica , Humedad , Movimiento/fisiología , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Larva , Mutación/genética , SefarosaRESUMEN
Alcohol directly modulates the BK potassium channel to alter behaviors in species ranging from invertebrates to humans. In the nematode Caenorhabditis elegans, mutations that eliminate the BK channel, SLO-1, convey dramatic resistance to intoxication by ethanol. We hypothesized that certain conserved amino acids are critical for ethanol modulation, but not for basal channel function. To identify such residues, we screened C. elegans strains with different missense mutations in the SLO-1 channel. A strain with the SLO-1 missense mutation T381I in the RCK1 domain was highly resistant to intoxication. This mutation did not interfere with other BK channel-dependent behaviors, suggesting that the mutant channel retained normal in vivo function. Knock-in of wild-type versions of the worm or human BK channel rescued intoxication and other BK channel-dependent behaviors in a slo-1-null mutant background. In contrast, knock-in of the worm T381I or equivalent human T352I mutant BK channel selectively rescued BK channel-dependent behaviors while conveying resistance to intoxication. Single-channel patch-clamp recordings confirmed that the human BK channel engineered with the T352I missense mutation was insensitive to activation by ethanol, but otherwise had normal conductance, potassium selectivity, and only subtle differences in voltage dependence. Together, our behavioral and electrophysiological results demonstrate that the T352I mutation selectively disrupts ethanol modulation of the BK channel. The T352I mutation may alter a binding site for ethanol and/or interfere with ethanol-induced conformational changes that are critical for behavioral responses to ethanol.
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Proteínas de Caenorhabditis elegans/genética , Depresores del Sistema Nervioso Central/farmacología , Etanol/farmacología , Canales de Potasio de Gran Conductancia Activados por el Calcio/genética , Locomoción/efectos de los fármacos , Mutación Missense/genética , Aldicarb/farmacología , Animales , Animales Modificados Genéticamente , Células del Asta Anterior/fisiología , Caenorhabditis elegans , Moléculas de Adhesión Celular Neuronal/genética , Inhibidores de la Colinesterasa/farmacología , Células HEK293 , Humanos , Inmunoglobulinas/genética , Locomoción/genética , Proteínas Luminiscentes/genética , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Mutagénesis Sitio-Dirigida , Estructura Terciaria de Proteína/genéticaRESUMEN
All terrestrial animals must find a proper level of moisture to ensure their health and survival. The cellular-molecular basis for sensing humidity is unknown in most animals, however. We used the model nematode Caenorhabditis elegans to uncover a mechanism for sensing humidity. We found that whereas C. elegans showed no obvious preference for humidity levels under standard culture conditions, worms displayed a strong preference after pairing starvation with different humidity levels, orienting to gradients as shallow as 0.03% relative humidity per millimeter. Cell-specific ablation and rescue experiments demonstrate that orientation to humidity in C. elegans requires the obligatory combination of distinct mechanosensitive and thermosensitive pathways. The mechanosensitive pathway requires a conserved DEG/ENaC/ASIC mechanoreceptor complex in the FLP neuron pair. Because humidity levels influence the hydration of the worm's cuticle, our results suggest that FLP may convey humidity information by reporting the degree that subcuticular dendritic sensory branches of FLP neurons are stretched by hydration. The thermosensitive pathway requires cGMP-gated channels in the AFD neuron pair. Because humidity levels affect evaporative cooling, AFD may convey humidity information by reporting thermal flux. Thus, humidity sensation arises as a metamodality in C. elegans that requires the integration of parallel mechanosensory and thermosensory pathways. This hygrosensation strategy, first proposed by Thunberg more than 100 y ago, may be conserved because the underlying pathways have cellular and molecular equivalents across a wide range of species, including insects and humans.
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Caenorhabditis elegans/fisiología , Humedad , Mecanorreceptores/fisiología , Células Receptoras Sensoriales/fisiología , Sensación Térmica/fisiología , Canales Iónicos Sensibles al Ácido/fisiología , Animales , Conducta Animal/fisiología , Caenorhabditis elegans/citología , Proteínas de Caenorhabditis elegans/fisiología , GMP Cíclico/fisiología , Canales Epiteliales de Sodio/fisiología , Humanos , Activación del Canal Iónico/fisiología , Canales Iónicos/fisiología , Proteínas de la Membrana/fisiología , Actividad Motora/fisiología , Complejos Multiproteicos/fisiología , Inanición/fisiopatologíaRESUMEN
Alcohol has a wide variety of effects on physiology and behavior. One of the most well-recognized behavioral effects is disinhibition, where behaviors that are normally suppressed are displayed following intoxication. A large body of evidence has shown that alcohol-induced disinhibition in humans affects attention, verbal, sexual, and locomotor behaviors. Similar behavioral disinhibition is also seen in many animal models of ethanol response, from invertebrates to mammals and primates. Here we describe several examples of disinhibition in the nematode C. elegans. The nematode displays distinct behavioral states associated with locomotion (crawling on land and swimming in water) that are mediated by dopamine. On land, animals crawl and feed freely, but these behaviors are inhibited in water. We found that additional behaviors, including a variety of escape responses are also inhibited in water. Whereas alcohol non-specifically impaired locomotion, feeding, and escape responses in worms on land, alcohol specifically disinhibited these behaviors in worms immersed in water. Loss of dopamine signaling relieved disinhibition of feeding behavior, while loss of the D1-like dopamine receptor DOP-4 impaired the ethanol-induced disinhibition of crawling. The powerful genetics and simple nervous system of C. elegans may help uncover conserved molecular mechanisms that underlie alcohol-induced disinhibition of behaviors in higher animals.
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Conducta Animal/efectos de los fármacos , Caenorhabditis elegans/efectos de los fármacos , Etanol/farmacología , Animales , Dopamina/farmacología , Locomoción/efectos de los fármacos , Sistema Nervioso/efectos de los fármacos , Fenómenos Fisiológicos del Sistema Nervioso/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Natación/fisiologíaRESUMEN
Dopamine is an ancient signaling molecule. It is responsible for maintaining the adaptability of behavioral outputs and is found across taxa. The following is a summary of the role of dopamine and the mechanisms of its function and dysfunction. We discuss our recent findings on dopaminergic control of behaviors in C. elegans and discuss its potential implications for work in the fields of C. elegans and Parkinson research.
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BACKGROUND: Ethanol (EtOH) is metabolized by a 2-step process in which alcohol dehydrogenase (ADH) oxidizes EtOH to acetaldehyde, which is further oxidized to acetate by aldehyde dehydrogenase (ALDH). Although variation in EtOH metabolism in humans strongly influences the propensity to chronically abuse alcohol, few data exist on the behavioral effects of altered EtOH metabolism. Here, we used the nematode Caenorhabditis elegans to directly examine how changes in EtOH metabolism alter behavioral responses to alcohol during an acute exposure. Additionally, we investigated EtOH solution osmolarity as a potential explanation for contrasting published data on C. elegans EtOH sensitivity. METHODS: We developed a gas chromatography assay and validated a spectrophotometric method to measure internal EtOH in EtOH-exposed worms. Further, we tested the effects of mutations in ADH and ALDH genes on EtOH tissue accumulation and behavioral sensitivity to the drug. Finally, we tested the effects of EtOH solution osmolarity on behavioral responses and tissue EtOH accumulation. RESULTS: Only a small amount of exogenously applied EtOH accumulated in the tissues of C. elegans and consequently their tissue concentrations were similar to those that intoxicate humans. Independent inactivation of an ADH-encoding gene (sodh-1) or an ALDH-encoding gene (alh-6 or alh-13) increased the EtOH concentration in worms and caused hypersensitivity to the acute sedative effects of EtOH on locomotion. We also found that the sensitivity to the depressive effects of EtOH on locomotion is strongly influenced by the osmolarity of the exogenous EtOH solution. CONCLUSIONS: Our results indicate that EtOH metabolism via ADH and ALDH has a statistically discernable but surprisingly minor influence on EtOH sedation and internal EtOH accumulation in worms. In contrast, the osmolarity of the medium in which EtOH is delivered to the animals has a more substantial effect on the observed sensitivity to EtOH.
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Caenorhabditis elegans/efectos de los fármacos , Caenorhabditis elegans/metabolismo , Etanol/administración & dosificación , Etanol/metabolismo , Locomoción/efectos de los fármacos , Alcohol Deshidrogenasa/metabolismo , Aldehído Deshidrogenasa/metabolismo , Animales , Conducta Animal/efectos de los fármacos , Conducta Animal/fisiología , Cromatografía de Gases/métodos , Locomoción/fisiología , Concentración OsmolarRESUMEN
For animals inhabiting multiple environments, the ability to select appropriate behaviors is crucial as their adaptability is often context dependent. Caenorhabditis elegans uses distinct gaits to move on land and in water. Gait transitions can potentially coordinate behaviors associated with distinct environments. We investigated whether land and water differentially affect the behavioral repertoire of C. elegans. Swimming worms interrupted foraging, feeding, egg-laying and defecation. Exogenous dopamine induced bouts of these land-associated behaviors in water. Our finding that worms do not drink fluid while immersed may explain why higher drug doses are required in water than on land to elicit the same effects. C. elegans is a valid model to study behavioral hierarchies and how environmental pressures alter their balance.
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Many animals, including humans, select alternate forms of motion (gaits) to move efficiently in different environments. However, it is unclear whether primitive animals, such as nematodes, also use this strategy. We used a multifaceted approach to study how the nematode Caenorhabditis elegans freely moves into and out of water. We demonstrate that C. elegans uses biogenic amines to switch between distinct crawling and swimming gaits. Dopamine is necessary and sufficient to initiate and maintain crawling after swimming. Serotonin is necessary and sufficient to transition from crawling to swimming and to inhibit a set of crawl-specific behaviors. Further study of locomotory switching in C. elegans and its dependence on biogenic amines may provide insight into how gait transitions are performed in other animals.
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Caenorhabditis elegans/fisiología , Dopamina/fisiología , Locomoción/fisiología , Serotonina/fisiología , Animales , Animales Modificados Genéticamente , Conducta Animal/fisiología , Fenómenos Biomecánicos , Neuronas Dopaminérgicas/fisiología , Marcha/fisiología , Neuronas Serotoninérgicas/fisiología , Transducción de Señal/fisiología , Natación/fisiología , Grabación en Video , Viscosidad , AguaRESUMEN
The mechanisms by which ethanol and inhaled anesthetics influence the nervous system are poorly understood. Here we describe the positional cloning and characterization of a new mouse mutation isolated in an N-ethyl-N-nitrosourea (ENU) forward mutagenesis screen for animals with enhanced locomotor activity. This allele, Lightweight (Lwt), disrupts the homolog of the Caenorhabditis elegans (C. elegans) unc-79 gene. While Lwt/Lwt homozygotes are perinatal lethal, Lightweight heterozygotes are dramatically hypersensitive to acute ethanol exposure. Experiments in C. elegans demonstrate a conserved hypersensitivity to ethanol in unc-79 mutants and extend this observation to the related unc-80 mutant and nca-1;nca-2 double mutants. Lightweight heterozygotes also exhibit an altered response to the anesthetic isoflurane, reminiscent of unc-79 invertebrate mutant phenotypes. Consistent with our initial mapping results, Lightweight heterozygotes are mildly hyperactive when exposed to a novel environment and are smaller than wild-type animals. In addition, Lightweight heterozygotes exhibit increased food consumption yet have a leaner body composition. Interestingly, Lightweight heterozygotes voluntarily consume more ethanol than wild-type littermates. The acute hypersensitivity to and increased voluntary consumption of ethanol observed in Lightweight heterozygous mice in combination with the observed hypersensitivity to ethanol in C. elegans unc-79, unc-80, and nca-1;nca-2 double mutants suggests a novel conserved pathway that might influence alcohol-related behaviors in humans.
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Peso Corporal , Etanol/metabolismo , Ratones/metabolismo , Mutación , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/crecimiento & desarrollo , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiología , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Femenino , Canales Iónicos/genética , Canales Iónicos/metabolismo , Masculino , Proteínas de la Membrana , Ratones/genética , Ratones/crecimiento & desarrollo , Ratones/fisiología , Ratones Endogámicos C57BL , Actividad MotoraRESUMEN
Animals frequently switch from one behavior to another, often to meet the demands of their changing environment or internal state. What factors control these behavioral switches and the selection of what to do or what not to do? To address these issues, we will focus on the locomotor behaviors of two distantly related "worms," the medicinal leech Hirudo verbana (clade Lophotrochozoa) and the nematode Caenorhabditis elegans (clade Ecdysozoa). Although the neural architecture and body morphology of these organisms are quite distinct, they appear to switch between different forms of locomotion by using similar strategies of decision-making. For example, information that distinguishes between liquid and more solid environments dictates whether an animal swims or crawls. In the leech, dopamine biases locomotor neural networks so that crawling is turned on and swimming is turned off. In C. elegans, dopamine may also promote crawling, a form of locomotion that has gained new attention.
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Normal aging leads to an inexorable decline in motor performance, contributing to medical morbidity and decreased quality of life. While much has been discovered about genetic determinants of lifespan, less is known about modifiers of age-related behavioral decline and whether new gene targets may be found which extend vigorous activity, with or without extending lifespan. Using Caenorhabditis elegans, we have developed a model of declining neuromuscular function and conducted a screen for increased behavioral activity in aged animals. In this model, behavioral function suffers from profound reductions in locomotory frequency, but coordination is strikingly preserved until very old age. By screening for enhancers of locomotion at advanced ages we identified the ras-related Rag GTPase raga-1 as a novel modifier of behavioral aging. raga-1 loss of function mutants showed vigorous swimming late in life. Genetic manipulations revealed that a gain of function raga-1 curtailed behavioral vitality and shortened lifespan, while a dominant negative raga-1 lengthened lifespan. Dietary restriction results indicated that a raga-1 mutant is relatively protected from the life-shortening effects of highly concentrated food, while RNAi experiments suggested that raga-1 acts in the highly conserved target of rapamycin (TOR) pathway in C. elegans. Rag GTPases were recently shown to mediate nutrient-dependent activation of TOR. This is the first demonstration of their dramatic effects on behavior and aging. This work indicates that novel modulators of behavioral function can be identified in screens, with implications for future study of the clinical amelioration of age-related decline.
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Conducta Animal , Caenorhabditis elegans/fisiología , GTP Fosfohidrolasas/metabolismo , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , GTP Fosfohidrolasas/genética , Esperanza de Vida , NataciónRESUMEN
Genetic defects in the dystrophin-associated protein complex (DAPC) are responsible for a variety of pathological conditions including muscular dystrophy, cardiomyopathy, and vasospasm. Conserved DAPC components from humans to Caenorhabditis elegans suggest a similar molecular function. C. elegans DAPC mutants exhibit a unique locomotory deficit resulting from prolonged muscle excitation and contraction. Here we show that the C. elegans DAPC is essential for proper localization of SLO-1, the large conductance, voltage-, and calcium-dependent potassium (BK) channel, which conducts a major outward rectifying current in muscle under the normal physiological condition. Through analysis of mutants with the same phenotype as the DAPC mutants, we identified the novel islo-1 gene that encodes a protein with two predicted transmembrane domains. We demonstrate that ISLO-1 acts as a novel adapter molecule that links the DAPC to SLO-1 in muscle. We show that a defect in either the DAPC or ISLO-1 disrupts normal SLO-1 localization in muscle. Consistent with observations that SLO-1 requires a high calcium concentration for full activation, we find that SLO-1 is localized near L-type calcium channels in muscle, thereby providing a mechanism coupling calcium influx with the outward rectifying current. Our results indicate that the DAPC modulates muscle excitability by localizing the SLO-1 channel to calcium-rich regions of C. elegans muscle.
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Proteínas de Caenorhabditis elegans/metabolismo , Complejo de Proteínas Asociado a la Distrofina/fisiología , Canales de Potasio de Gran Conductancia Activados por el Calcio/metabolismo , Músculos/fisiología , Animales , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Calcio , Distrofina , Electrofisiología , Canales de Potasio de Gran Conductancia Activados por el Calcio/genética , Proteínas MutantesRESUMEN
Alternative patterns of neural activity drive different rhythmic locomotory patterns in both invertebrates and mammals. The neuro-molecular mechanisms responsible for the expression of rhythmic behavioral patterns are poorly understood. Here we show that Caenorhabditis elegans switches between distinct forms of locomotion, or crawling versus swimming, when transitioning between solid and liquid environments. These forms of locomotion are distinguished by distinct kinematics and different underlying patterns of neuromuscular activity, as determined by in vivo calcium imaging. The expression of swimming versus crawling rhythms is regulated by sensory input. In a screen for mutants that are defective in transitioning between crawl and swim behavior, we identified unc-79 and unc-80, two mutants known to be defective in NCA ion channel stabilization. Genetic and behavioral analyses suggest that the NCA channels enable the transition to rapid rhythmic behaviors in C. elegans. unc-79, unc-80, and the NCA channels represent a conserved set of genes critical for behavioral pattern generation.
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Conducta Animal/fisiología , Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Canales Iónicos/genética , Natación/fisiología , Animales , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Canales Iónicos/metabolismoRESUMEN
C. elegans advances up a chemical gradient by modulating the probability of occasional large, course-correcting turns called pirouettes. However, it remains uncertain whether C. elegans also uses other behavioral strategies for chemotaxis. Previous observations of the unusual, spiral-shaped chemotaxis tracks made by the bent-head mutant unc-23 point to a different strategy in which the animal continuously makes more subtle course corrections. In the present study we have combined automated tracking of individual animals with computer modeling to test the hypothesis that the pirouette strategy is sufficient on its own to account for the spiral tracks. Tracking experiments showed that the bent-head phenotype causes a strong turning bias and disrupts pirouette execution but does not disrupt pirouette initiation. A computer simulation of disrupted pirouette behavior and turning bias reproduced the spiral tracks of unc-23 chemotaxis behavior, showing that the pirouette strategy is sufficient to account for the mutant phenotype. In addition, the simulation reproduced higher order features of the behavior such as the relationship between the handedness of the spiral and the side to which the head was bent. Our results suggest that the pirouette mechanism is sufficient to account for a diverse range of chemotaxis trajectories.
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Caenorhabditis elegans/fisiología , Quimiotaxis/fisiología , Locomoción/fisiología , Modelos Biológicos , Orientación/fisiología , Animales , Simulación por ComputadorRESUMEN
The anatomical connectivity of the nervous system of the nematode Caenorhabditis elegans has been almost completely described, but determination of the neurophysiological basis of behavior in this system is just beginning. Here we used an optimization algorithm to search for patterns of connectivity sufficient to compute the sensorimotor transformation underlying C. elegans chemotaxis, a simple form of spatial orientation behavior in which turning probability is modulated by the rate of change of chemical concentration. Optimization produced differentiator networks capable of simulating chemotaxis. A surprising feature of these networks was inhibitory feedback connections on all neurons. Further analysis showed that feedback regulates the latency between sensory input and behavior. Common patterns of connectivity between the model and biological networks suggest new functions for previously identified connections in the C. elegans nervous system.
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Quimiotaxis/fisiología , Redes Neurales de la Computación , Sinapsis/fisiología , Transmisión Sináptica/fisiología , Potenciales de Acción/fisiología , Algoritmos , Animales , Conducta Animal , Caenorhabditis elegans , Simulación por Computador , Generalización del Estimulo , Modelos Neurológicos , Neuronas/fisiología , Orientación/fisiología , Conducta Espacial/fisiología , Factores de TiempoRESUMEN
The activities of many neuronal proteins are modulated by ethanol, but the fundamental mechanisms underlying behavioral effects of ethanol remain unclear. To identify mechanisms responsible for intoxication, we screened for Caenorhabditis elegans mutants with altered behavioral responses to ethanol. We found that slo-1 mutants, which were previously recognized as having slightly uncoordinated movement, are highly resistant to ethanol in two behavioral assays. Numerous loss-of-function slo-1 alleles emerged from our screens, indicating that slo-1 has a central role in ethanol responses. slo-1 encodes the BK potassium channel. Electrophysiological analysis shows that ethanol activates the channel in vivo, which would inhibit neuronal activity. Moreover, behaviors of slo-1 gain-of-function mutants resemble those of ethanol-intoxicated animals. These results demonstrate that selective activation of BK channels is responsible for acute intoxicating effects of ethanol in C. elegans. BK channel activation may explain a variety of behavioral responses to ethanol in invertebrate and vertebrate systems.