Chris Gabel et al. (2004) East Coast Worm Meeting "Quantification of electrotaxis"

Dmitri Pavlichin, Chris Gabel, Aravi Samuel, Albert Kao

[East Coast Worm Meeting, 2004]

In an electric field, the worm has preferred trajectories for forward crawling movement that depend on the direction and magnitude of the electric field. In weak fields (<5 V/cm), worms tend to crawl towards either the positive or negative pole. In moderate fields, ~5 V/cm, worms crawl straight towards the negative pole. In stronger fields, >5 V/cm, worms crawl towards the negative pole in trajectories at an angle to the direction of the field. The angle of approach towards the negative pole increases with field strength, rising from 0 degrees (parallel to the field lines) towards 90 degrees (perpendicular to the field lines). We have quantified these electrotactic movements by tracking wild-type and mutant worms crawling over agar surfaces containing different types and concentrations of ions while responding to electric fields varied in amplitude, frequency, and direction.

Mendoza, S. et al. (2017) International Worm Meeting "W-TEM: A microscope to study C. elegans behavior under external stimulations."

Lam, B., Mendoza, S., D'Orazio, E., Sherry, T., Madruga, B., Mai, P., Arisaka, K., Jiang, K.

[International Worm Meeting, 2017]

The ability to monitor neural activity in freely behaving animals is important in determining neural mechanisms responsible for behavior. Real time worm tracking with neuronal observation has been used to study worm behavior in freely moving C. elegans. However, few setups have attempted to use worm tracking in conjunction with behavioral stimulations. The difficulty being that most behavioral stimulations used to study C. elegans such as electrotaxis and thermotaxis are difficult to incorporate into existing worm tracking platforms. We present a novel microscope platform, W-TEM ( Worm Tracking Epifluorescence Microscope), to bridge this gap in our understanding of worm behavior. The microscope is a standard wide field epi-fluorescence microscope placed on top of a stable platform to move the microscope as it tracks along the sample plane, accommodating most C. elegans behavioral stimulations. This microscope can incorporate a stimulation system that covers 20 cm x 20 cm of worm area movement, 50 cm x 50 cm of total size, and a height of up to 30 cm, which is larger than most other stimulation systems. It can take data at video rate 30 fps, for cameleon ratiometic imaging, as well as monitor whole worm behavior. In addition, our platform can be moved easily between different experiments, using a system of rails that is attached to the microscope platform. Using this system, we have been able to study both Electrotaxis and Thermotactic behaviors using the same system. In the case of Electrotaxis, we have found that high voltages corresponding to 8V/cm create a dampening effect of worm motion, possibly due to paralyzing the lower half of the worm body. We also conduct investigations into the AFD and AIY neurons during isothermal behavior and find a phase lag of 0.8 seconds between neural activity and head location during isothermal tracking.

Ramachandran, S. et al. (2017) International Worm Meeting "Dissection of molecular mechanisms that govern functional connectivity across multiple postsynaptic partners."

Ramachandran, S., Philbrook, A., Francis, MM., Lambert, CM.

[International Worm Meeting, 2017]

Neurons often make synaptic outputs onto multiple partners. The strength of these connections can vary greatly across partners, but we have limited understanding of how synaptic weights with specific partners are established and regulated. We have been addressing these questions using the motor circuit of Caenorhabditis elegans as a model. Cholinergic motor neurons form excitatory dyadic synapses onto body wall muscle cells and GABAergic motor neurons. We have developed a system for investigating functional connectivity onto each of these cell types. Specifically, we have generated transgenic strains that express Chrimson in the cholinergic motor neurons together with the genetically encoded calcium sensor GCaMP6s in either muscle cells or GABAergic motor neurons. Photostimulation of cholinergic neurons in wild type animals produces robust Ca2+ transients in both cell types. Compared to motor neurons, calcium responses in muscles are prolonged and larger in amplitude, likely reflecting differences in both synaptic and cellular physiology across these cell types. In order to dissect synaptic mechanisms required for transmission onto each partner, we have been investigating strains carrying mutations in genes required for neurotransmitter release, specific neurotransmitter receptor subunits, and synaptic organizers. By investigating Ca2+ responses in these mutants, we aim to elucidate mechanisms underlying functional connectivity between cholinergic motor neurons and their dyadic synaptic partners.

Ramachandran, S. et al. (2019) International Worm Meeting "NLP-12 signaling through the cholecystokinin receptor homologue CKR-1 regulates local food search behavior."

Banerjee, N., Ramachandran, S., Bhattacharya, R., Francis, MM., Lambert, CM.

[International Worm Meeting, 2019]

Animals have a remarkable capacity to modify their behavior in response to changing environmental conditions. These behavioral adaptations are often key for survival and, in many cases, are encoded by neuromodulatory signals that alter the functional connectivity of neural circuits in a context-dependent manner. We have been investigating neuromodulatory control of behavior using C. elegans food searching as a model. Immediately following removal from food, C. elegans restrict their movement to a limited region of their environment by increasing their rate of turning, a behavior known as area-restricted search. After several minutes without finding food, C. elegans decrease their rate of turning and disperse over a larger region. We have previously shown that NLP-12/Cholecystokinin neuropeptides released from the interneuron DVA regulate area-restricted search behavior1. Specifically, we found that deletion of nlp-12 strikingly decreases the frequency of turning immediately following removal from food, but does not dramatically affect either exploratory movement or dispersal behavior. Here, we show that NLP-12 regulation of local food searching is mediated primarily through the G-protein coupled receptor CKR-1. Deletion of ckr-1 decreases reorientations during local food searching, similar to deletion of nlp-12. In contrast, deletion of ckr-2, encoding a previously characterized NLP-12 receptor, does not impact local food searching. We have previously shown that overexpression of nlp-12 produces a chronic local search-like behavior1. Overexpression of ckr-1 produces a similar behavioral phenotype in an nlp-12 dependent manner. Our genetic analysis points towards a novel GPCR-mediated NLP-12 signaling mechanism, and importantly, implicates the previously uncharacterized CKR-1 GPCR as a key component in NLP-12 modulation of circuit activity. We are now investigating the cellular requirements for NLP-12/CKR-1 modulation of local searching with the ultimate goal of understanding how NLP-12 signaling alters circuit performance and behavior. 1. Bhattacharya et. al. 2014 (PLoS Genetics)

Mai, P. et al. (2017) International Worm Meeting "Damped sine wave oscillation of Caenorhabditis elegans under uniform electric field."

Mai, P., D'Orazio, E.G., Raghute, R., Carmona, J., Jin, S., Mendoza, S., Arisaka, K., Shrestha, A.

[International Worm Meeting, 2017]

Research has shown that Caenorhabditis elegans demonstrates a highly deterministic behavioral response under an electric field; Under such conditions, worms migrate to the negative pole of the field, coupled with an angular offset proportional to the absolute strength of the field. Under relatively large applied voltages (12 V/cm ) worms migrate to the field's negative pole with a larger opening angle, as compared to a more modest 5 V/cm. While this relatively broad observation is interesting in itself, a more comprehensive motional attractor analysis is yet to be conducted on C. elegans demonstrating electrotactic responses. To address this, we experimentally observed the motion of ten C. elegans on 2% agarose under electric fields varying from 5 - 12 V/cm. A custom built worm-tracking microscope was implemented for these purposes in order to accurately record the worm's body configuration and absolute position during trials. MatLab fitting software was written and used to analytically describe the worm's motion under experimental and controlled conditions. In the absence of an electric field, the motion of C. elegans may be well-modeled as a propagating sinusoid, fit along the entirety of the worm's body. However, under the application of an electric field, the sinusoidal fit requires a damping term to fully represent an apparent partial paralyzation of the worm's posterior half. This result suggests the possibility that the presence of an electric field likely inhibits specific motor neurons responsible for the muscular control of the posterior regions of the worm's body.

Baird, Scott E. et al. (2011) International Worm Meeting "Ray Pattern Variation in C. elegans: Mapping a Major-Effect QTL on LGV."

Baird, Scott E., Bailey, Daniel

[International Worm Meeting, 2011]

Cryptic variation of ray pattern has been observed in recombinant-inbred lines (RIL) derived from crosses of N2 and CB4856 strains of C. elegans (Guess et al., 2007). The variant pattern consists of the anterior displacement of ray 3 into a position immediately adjacent to ray 2. QTL analyses of these RIL identified a major-effect QTL on the left arm of chromosome V (QTL-V). One RIL, QX34, possessed a recombinant chromosome V that contained CB4856 alleles from -12.72 to -7.93 cM, a region that precisely corresponded to QTL-V. When this chromosome (QX34-V) was crossed into an otherwise N2 background (strain PB2034), the variant ray pattern was observed at a frequency of 0.50. Based on genotypes of other RIL with recombination breakpoints in this region, it appeared that QTL-V resulted from allelic variation at two or more loci. To confirm this result, we are mapping the gene(s) in this region responsible for the observed variation in ray pattern. Initial experiments demonstrated that the CB4856 alleles responsible for QTL-V were recessive to the corresponding N2 alleles. From deletion heterozygotes, it was determined that at least one gene responsible for QTL-V lies to the right of -9.00 cM. However, deletion of a region from -12.06 to -8.15 did not uncover the ray pattern variant. Therefore, either allelic variation at two or more genes is required for QTL-V or the gene responsible for QTL-V resides between -8.15 and -7.93 cM. Attempts at recombination mapping of QTL-V are ongoing but have been hindered by an apparent suppression of recombination in PB2034/N2 heterozygotes.

Sternberg PW et al. (2000) West Coast Worm Meeting "Evidence of a Mate-finding Cue in the Free-Living Soil Nematode C. elegans"

Simon JM, Sternberg PW

[West Coast Worm Meeting, 2000]

Use of mate-finding cues is well documented throughout the nematode literature, but no assays for C. elegans have come into common practice. Here we report two assays that suggest young adult males can detect young adult hermaphrodites through some unidentified cue. Accumulation of 14 males (per trial) to agar conditioned with 5 hermaphrodites (the muscle mutant unc-52 was used to construct a point source), which were sequentially removed, suggests a cue given off by hermaphrodites and detected by males [mean number of males observed in 1-cm diameter scoring circles shifted from 3.2+2.2 in control trials (n=29) to 6.5+2.5 in conditioned-agar trials (n=28); p<0.0001 (2a)]. Similar assays testing the effects of such cues on solitary males were also constructed. Single males were followed for 10 minutes and total elapsed time and number of reversals were noted [mean time and reversals in scoring regions (again 1-cm diameter circles) shifted from 4 seconds and 0.2 reversals in control trials (n=30) to 90 seconds and 8.7 reversals in conditioned-agar trials (n=31); p<0.002 and p<0.001, respectively (2a)]. Pilot studies suggest some form of kinesis, but the possibility of an additional short-range taxis, especially in a gas-liquid soil environment, should not be eliminated.

Bronwyn L MacInnis et al. (2005) International Worm Meeting "Behavioral analysis of wild type and mutant worms using a novel thermal response assay"

Bronwyn L MacInnis, Miriam B Goodman, Hau-chen Lee, Daniel Ramot

[International Worm Meeting, 2005]

Classical assays of worm thermotaxis have assessed the thermal tracking abilities of individual worms or the accumulation of a population of worms following exposure to spatial thermal gradients for one hour. Here, we report a novel, rapid thermotaxis assay, in which the distribution of a population of worms on a linear thermal gradient is measured after ~10 min. This assay allows us to assess the initial response of the worm population to a thermal gradient and, thus, we have termed it a thermal response assay. In a typical assay, a stable thermal gradient of approximately 1 C/cm was established across NGM agar in a rectangular culture dish (7 x 5 cm). Worms raised under the desired experimental conditions were distributed along the center of the agar perpendicular to the direction of the thermal gradient at a particular starting temperature. Worms dispersed from this starting position within 1-2 min and were killed by exposure to chloroform vapor after 10 min. Their positions were scored by counting animals distributed above and below the starting temperature; these data were used to compute a thermal response index in which -1 indicates that all animals moved to colder temperatures and +1 indicates that all animals moved to warmer temperatures. Wild type (N2) worms exhibited a robust cryophilic drive when their starting position was 3 C above their cultivation temperature (Tc; Tc

Peliti M et al. (2010) Neuronal Development, Synaptic Function and Behavior, Madison, WI "Biased motion in unbiased environments: are the worms navigating?"

Shaham S, Leibler S, Chuang J S, Peliti M

[Neuronal Development, Synaptic Function and Behavior, Madison, WI, 2010]

Like any mobile organism, C. elegans relies on sensory cues to find food. In the absence of such cues, defined search patterns or other stereotypical behaviors may be observed. We are characterizing the movement pattern of C. elegans in the absence of chemotactic stimuli, over time scales comparable to that of starvation. To this end, we devised a flatbed scanner-based imaging setup that enables us to collect individual animals' trajectories over large (24 cm x 24 cm) Petri dishes. Surprisingly, the majority (~60%) of wild-type trajectories display persistence in the direction of motion over length scales that are 50-100 times the animal's body length. A preliminary dataset of trajectories acquired with an independent camera-based imaging setup qualitatively confirmed this result. Synthetic trajectories generated from the same angle and step distributions of individual trajectories show that persistence of motion cannot be accounted for by a simple random walk model of locomotion. To determine whether sensory perception is required for the animals' directional behavior, we analyzed the trajectories of animals with impaired sensory function. We found that animals mutant for either tax-2 or tax-4, which encode subunits of a cGMP-gated channel required for several sensory modalities, do not show directional behavior. However, daf-19 mutants, which lack all chemosensory and mechanosensory cilia, display wild-type directional behavior, albeit at a lower percentage (~20%). Targeting thermosensation specifically with a gcy-8; gcy-18; gcy-23 triple mutation fails to suppress directional behavior. To gain further insight into the role of sensory neurons in directionality we are currently performing cell-specific rescue of tax-4 function.

Peliti, Margherita et al. (2009) International Worm Meeting "The Dispersal Behavior of C. elegans."

Shaham, Shai, Peliti, Margherita, Chuang, John, Leibler, Stanislas

[International Worm Meeting, 2009]

A major determinant of an organism''s long-term success is its ability to locate and harvest resources, and to disperse once these are depleted. We seek to investigate the dispersal strategy of C. elegans, and to provide a characterization of its long-range spatial pattern of movement. We have devised an imaging setup employing several flatbed scanners to visualize the movement of C. elegans over a large surface. Images of 1-day-old adult individual animals placed at the center of a 24 cm x 24 cm agar plate are collected for 90 minutes or until animals encounter the edge of the plate, and positional information is extracted with a tracking algorithm. We utilize a geometrical criterion to quantify the directionality of paths at different length scales, and to sort trajectories into distinct behavioral classes. Our data shows that about 60% of wild-type trajectories are directional over length scales that are 50-100 times the animal''s body-length. Furthermore, comparison of our data with simulated trajectories shows that directionality cannot be accounted for by a stochastic, random walk model of locomotion. The overall direction of movement with respect to our imaging apparatus differs from animal to animal, suggesting that the observed directional bias is unlikely to result from a bias in the experimental setup. To gain insight into the basis of this large-scale directed movement we have begun to examine the tracks of animals with impaired sensory function. Concurrently, we are developing a camera imaging apparatus to validate these results in a different setup, as well as to investigate directionality at a finer time resolution.