Jonathan T. Pierce-Shimomura

Genetic analysis of crawling and swimming locomotory patterns in C. elegans

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.

The homeobox gene lim-6 is required for distinct chemosensory representations in C. elegans

The ability to discriminate between different chemical stimuli is crucial for food detection, spatial orientation and other adaptive behaviours in animals. In the nematode Caenorhabditis elegans, spatial orientation in gradients of soluble chemoattractants (chemotaxis) is controlled mainly by a single pair of chemosensory neurons. These two neurons, ASEL and ASER, are left–right homologues in terms of the disposition of their somata and processes, morphology of specialized sensory endings, synaptic partners and expression profile of many genes. However, recent gene-expression studies have revealed unexpected asymmetries between ASEL and ASER. ASEL expresses the putative receptor guanylyl cyclase genes gcy-6 and gcy-7, whereas ASER expresses gcy-5 (ref. 4). In addition, only ASEL expresses the homeobox gene lim-6, an orthologue of the human LMX1 subfamily of homeobox genes. Here we show, using laser ablation of neurons and whole-cell patch-clamp electrophysiology, that the asymmetries between ASEL and ASER extend to the functional level. ASEL is primarily sensitive to sodium, whereas ASER is primarily sensitive to chloride and potassium. Furthermore, we find that lim-6 is required for this functional asymmetry and for the ability to distinguish sodium from chloride. Thus, a homeobox gene increases the representational capacity of the nervous system by establishing asymmetric functions in a bilaterally symmetrical neuron pair.

Caenorhabditis elegans selects distinct crawling and swimming gaits via dopamine and serotonin

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.

Magnetosensitive neurons mediate geomagnetic orientation in Caenorhabditis elegans

Many organisms spanning from bacteria to mammals orient to the earths magnetic field. For a few animals, central neurons responsive to earth-strength magnetic fields have been identified; however, magnetosensory neurons have yet to be identified in any animal. We show that the nematode Caenorhabditis elegans orients to the earths magnetic field during vertical burrowing migrations. Well-fed worms migrated up, while starved worms migrated down. Populations isolated from around the world, migrated at angles to the magnetic vector that would optimize vertical translation in their native soil, with northern- and southern-hemisphere worms displaying opposite migratory preferences. Magnetic orientation and vertical migrations required the TAX-4 cyclic nucleotide-gated ion channel in the AFD sensory neuron pair. Calcium imaging showed that these neurons respond to magnetic fields even without synaptic input. C. elegans may have adapted magnetic orientation to simplify their vertical burrowing migration by reducing the orientation task from three dimensions to one. DOI: http://dx.doi.org/10.7554/eLife.07493.001

Humidity sensation requires both mechanosensory and thermosensory pathways in Caenorhabditis elegans

Significance Although the neurons and molecules that mediate the sensing of light, odors, tastants, temperature, and pressure have been elucidated, how humidity is sensed in most animals remains unclear. We used the experimental amenability of the model nematode Caenorhabditis elegans to discover a mechanism for sensing humidity. This worm pairs mechanical and thermal information associated with humidity levels via parallel sensory pathways. This strategy, first proposed more than 100 years ago by Thunberg, is notable for not requiring specialized sensory apparatus (or even hair). Because these neurons and their molecular sensors have equivalents represented in diverse animals, the humidity sensation mechanism that we describe here could be used by many animals, including humans. 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.

Conserved Role of unc-79 in Ethanol Responses in Lightweight Mutant Mice

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.

Erratum: The homeobox gene lim-6 is required for distinct chemosensory representations in C. elegans (Nature (2001) 410 (694-698))

This corrects the article DOI: 35070575

Conserved Single Residue in the BK Potassium Channel Required for Activation by Alcohol and Intoxication in C. elegans

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.

Ethanol Metabolism and Osmolarity Modify Behavioral Responses to Ethanol in C. elegans

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.

Shared Strategies for Behavioral Switching: Understanding How Locomotor Patterns are Turned on and Off

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.