Wen JYM et al. (1995) International C. elegans Meeting "Olfactory Conditioning in C. elegans."

Negandhi H, Wen JYM, van der Kooy D, Runciman S, Morrison GE

[International C. elegans Meeting, 1995]

The nematode C. elegans as is an ideal system to study the cellular and molecular basis for learning and memory because of its simple genetics and well defined neuroanatomy. We have devised a new associative learning paradigm which uses the ability of C. elegans to track odors. An attractive odor (Diacetyl) is used as the conditioned stimulus (CS) while acetic acid (which is aversive) is used as the unconditioned stimulus (US). Non-food deprived worms are washed onto a fine wire filter and exposed to diacetyl followed by a brief exposure to acetic acid. After six rounds of conditioning, individual worms are tested for their tracking response to a small drop of the CS. A high proportion of naive worms show a tracking response to the CS. However, after conditioning a significant percentage of worms no longer track the CS suggesting that they no longer find it attractive. Several controls have been performed to demonstrate that the failure of conditioned worms to track Diacetyl is due to associative conditioning rather than some non-associative process. CS only and US only controls have no significant effect on the tracking response of worms to the CS when compared to the tracking response of naive worms. In addition, explicitly unpaired and pseudo-random pairings also have no significant effects on their tracking response. Interestingly, the lrn-1 mutant which was previously isolated using a Na+/Cl- + E. coli learning paradigm and has subsequently failed to learn other associative paradigms is also impaired in this task. This suggests that the lrn-1 mutation is necessary for C. elegans to acquire several different associative learning paradigms that cross many CS/US boundaries. Finally, the efficiency of this assay will facilitate the screening of large numbers of mutant worms in order to identify novel genes involved in associative learning.

Wen JYM et al. (1997) Behav Neurosci "Mutations that prevent associative learning in C. elegans."

Rousseau J, van der Kooy D, Runciman S, Kumar N, Rambaldini G, Morrison GE, Wen JYM

[Behav Neurosci, 1997]

The nematode Caenorhabditis elegans offers a promising system for the reductionist study of learning and memory. In this article, classical conditioning in C. elegans is demonstrated with a variety of associative learning assays. These assays allowed for the isolation and behavioral characterization of 2 mutant C. elegans lines impaired in associative learning. Both lines show no short-term or long-term associative conditioning; however, they appear relatively normal in tests of nonassociative learning and sensorimotor function. In combination with the well-described genetics and neuroanatomy of C. elegans, the isolation of mutants selectively, yet completely, blocked in associative learning provides the basis for an effective characterization of the cellular and molecular aspects of associative learning.

Morrison GE et al. (1999) Behav Neurosci "Olfactory associative learning in Caenorhabditis elegans is impaired in lrn-1 and lrn-2 mutants."

Runciman S, van der Kooy D, Wen JYM, Morrison GE

[Behav Neurosci, 1999]

The C. elegans mutants, lrn-1 and lrn-2, are impaired in associative learning using conditioned taste cues. Both mutants are defective in associative learning about appetitive and aversive events, indicating that lrn-1 and lrn-2 exert effects across motivational boundaries. In a new olfactory associative learning paradigm, in which wild type worms learn to avoid a previously attractive diacetyl odor after it has been paired with an aversive acetic acid solution, lrn-1 and lrn-2 are impaired. Although defective in associative learning using a conditioned olfactory cue, nonassociative learning (habituation and dishabituation) using this same olfactory cue is unaffected. The discovery that lrn-1 and lrn-2 are defective in associative learning with both taste and olfactory cues may suggest that associative learning in different sensory modalities converges on a common genetic pathway in C. elegans that is subserved by lrn-1 and lrn-2.

Kumar N et al. (1991) International C. elegans Meeting "CLASSICAL CONDITIONING IN C. ELEGANS;"

van der Kooy D, Wen JYM, Culotti JG, Kumar N

[International C. elegans Meeting, 1991]

The nematode is an excellent model system in which to pursue a neurogenetic analysis of behavior. In particular, we chose to train nematodes in a classical conditioning paradigm in order to focus on the cellular and molecular bases of learning and memory. Initially, 2 chemotactically attractive ions Na+ (NaCH3COO) and Cl- (NH4Cl) were chosen as conditioned stimuli (CSs) and E. coli, a rewarding food source, was used as the unconditioned stimuli (US). The animals were trained by exposing them to a constant concentration of one ion (CS+) paired with the US followed by exposure to the other ion (CS-) in the absence of the US. The experiment was counterbalanced for which CS was paired with the US and for the order of CS presentation. The testing consisted of placing approximately 100 of the trained animals between point gradients of each CS (in the absence of the US) and allowing them to migrate towards one of the gradient centers. CS levels were balanced such that naive animals displayed equal chemotactic preferences for either CS. Immediately after one CS+ and one CS- pairing, 67.1_3.1% of the animals demonstrated a preference for the CS paired with the US. A 5 hour food deprivation prior to training increased the preference for the CS+ ion to 77.3_2.6%. These animals showed evidence of memory for the association at 7 hours post-training but by 27 hours post-training their test performance was no longer significantly different (56.5+9.8 %) from controls. The nematodes learned inhibitory relationships (the CS- predicts the absence of food) as well as excitatory ones (the CS+ predicts the presence of food). We have also been able to demonstrate conditioned aversion learning by replacing E. coli, an appetitive US, with an aversive garlic extract. In this case nematodes can be trained to avoid the CS+ ion. In addition to being sensitive to changes in the US, the nematode can also learn about different CSs. If distinct temperatures are used as CSs instead of ions, food deprived nematodes learn to exhibit a preference for the temperature that was paired with the food reward. Thus, a desirable range of conditioned stimuli and conditioning responses can be utilized in learning paradigms with C. elegans. Presently we are undertaking a mutational screen for learning and memory mutants.

Goulding M (2012) Neuron "Motor neurons that multitask."

Goulding M

[Neuron, 2012]

Animals use a form of sensory feedback termed proprioception to monitor their body position and modify the motor programs that control movement. In this issue of Neuron, Wen etal. (2012) provide evidence that a subset of motor neurons function as proprioceptors in C.elegans, where B-type motor neurons sense body curvature to control the bending movements that drive forward locomotion.

Kumar N et al. (1993) International C. elegans Meeting "Isolation of learning specific mutants in C. elegans."

Wen JYM, Morrison GE, van der Kooy D, Parker JA, Kumar N

[International C. elegans Meeting, 1993]

Morrison GE et al. (1995) International C. elegans Meeting "lrn-1 and lrn-2 Block Associative Learning in C. elegans."

Morrison GE, Wen JYM, Runciman S, Neghandi H, van der Kooy D, Kumar N

[International C. elegans Meeting, 1995]

The nematode C. elegans can be conditioned using a classical conditioning paradigm based on its chemotactic responses to the conditioning stimuli (CS), Na+ (NaCH3CHOO) and Cl- (NH4Cl), which are equally preferred under baseline conditions. After exposure to one of the CS ions (CS+) in the presence of the US (E. coli, a food source), counterbalanced by an equal exposure to the alternate ion (CS-) in the absence of the US, the animals demonstrate a significant preference for the CS+ ion on testing. Using an EMS screen, we isolated two lines of mutant animals (lrn-1 and lrn-2) that after conditioning displayed the equivalent preferences for the CS+ and CS- ions shown by unconditioned wild type worms. Using garlic (an aversive stimulus) as a US, wild type worms show a conditioned avoidance for the ion (CS+) that was paired with the garlic, whereas the two mutant lines displayed equivalent preferences for the CS+ and CS- ions after conditioning. We used a short-term assay to determine the effects of the two mutations on short- and long-term memory. After conditioning for minutes, the initial test headings (after 30 sec) of individual wild type animals towards the CS+ and CS- ions are not significantly different from the learning scores in our original, 90 min test paradigm (~75% approach to the CS+ ion). However, after conditioning, initial test headings of individual mutant animals remain balanced, 50% approaching the CS+ ion and 50% approaching the CS- ion. These two mutants are normal in motor and sensory capabilities since the approaches to point sources of both CS ions and the US (E. coli) are similar to those of wild type animals in both the accumulation rates and overall preferences. As well, the two mutants are capable of normal non-associative learning (habituation) to the ions used as conditioning stimuli in the associative learning paradigm. Using the sequence tagged site strategy for C. elegans and multiplex PCR techniques, we have begun mapping the two mutations through the generation of Bergerac/mutant N2 hybrids.

Bukhari, Syed Irfan Ahmad et al. (2011) International Worm Meeting "Implication of microRNA pathway in C. elegans germline biogenesis."

Vasquez-Rifo, Alejandro, Simard, Martin J, Bukhari, Syed Irfan Ahmad, Masson, Jean-Yves, Zetka, Monique

[International Worm Meeting, 2011]

Small non-coding RNA pathways have been reported to assume important roles in the regulation of nearly all developmental and cellular processes. However, the contribution of the microRNA pathway in the germline remains poorly characterized. We therefore decided to address whether this small non-coding RNA pathway plays an important role in germ cell formation using C. elegans as animal model. We first observed that the loss-of-function of key players in the microRNA pathway lead to severe defects in animal fertility. Animals carrying null alleles of Drosha and Dicer are sterile while animals lacking the Argonaute alg-1 and alg-2 genes display various germline defects. An extensive analysis of the germline of these argonaute mutants reveals defects in the gamete formation resulting in reduced brood size. We observed that ALG-1 and ALG-2 proteins are localized to the distal tip cell or DTC, a specialized cell located at the tip of both gonadal arms that regulates mitosis-meiosis transition. Re-establishing the expression of these proteins in the DTC partially rescue the germline defects observed in mutant animals. Our data suggests the paramount importance of the microRNA pathway in the C. elegans germline biogenesis. Using germline-specific microarrays, genetic and in silico analysis, we are currently attempting to identify the specific regulatory pathway in germ line which is targeted by microRNAs. This work is supported by the Natural Sciences and Engineering Research Council of Canada (MJS and JYM).

Antebi A et al. (1996) East Coast Worm Meeting "daf-12 Coordinates Multiple Developmental Events"

Hedgecock EM, Antebi A

[East Coast Worm Meeting, 1996]

In the nematode, developmental events within or between tissues are precisely coordinated. The DAF-12 nuclear hormone receptor plays multiple roles in synchronizing development. As a heterochronic regulator, it instructs stage specific programs in both gonad and soma. In the dauer pathway, it selects dauer or continuous development in response to growth conditions. Gonadal, somatic and dauer functions are genetically separable, suggesting a complex locus, and alleles can be grouped into six idealized classes. In collaboration with Don Riddle, Pam Larsen and Wen Hui Yeh, we have begun a molecular analysis of alleles to correlate structure with function. So far, we have found a sharp clustering of alleles which selectively disrupt dauer formation within the zinc fingers of the receptor. A unifying hypothesis is that a hormone, acting through DAF-12 and related receptors, coordinates events in gonad and some, as well as dauer development. We suggest a simple model in which the commitments or starts to each developmental stage are synchronized by a hormonal pulse available to all tissues. Moreover, we propose that the starts to each developmental stage lie between the molts, as judged by new cuticle synthesis in the hypodermic and transitions in the migratory behavior of gonadal leader cells. These "parastages" may more accurately reflect the actual functional boundaries of developmental stages.

Kim, Jinmahn et al. (2019) International Worm Meeting "Decoding head neural circuit underlying rhythmic forward movement in C. elegans."

Pyo, Seondong, Kim, Kyuhyung, Yeon, Jihye, Kim, Jongrae, Park, Cheon-Gyu, Kim, Jinmahn, Suh, Byung-Chang

[International Worm Meeting, 2019]

Forward locomotion of C. elegans is a rhythmic behavior which is initiated by contraction and relaxation of head muscles. However, the neuronal and molecular mechanisms in which to generate and maintain forward movement are not fully understood. Previously, cholinergic B-type and GABAergic D-type motor neurons process proprioceptive signals and regulate activity of body muscles during forward movement (Wen et al., 2012). To investigate the neural circuit underlying head movement, we analyzed neural connectome data and found that the two cholinergic SMB and SMD neurons innervate the head muscles and connect to the RME GABAergic neurons. When we genetically ablated SMB or SMD using recCaspase system, the SMD ablated worms displayed poor forward but normal backward movement. While the SMB ablated worms move normally, they exhibit increased wave width of sinusoidal waveform. We then activated SMD or SMB via optogenetic manipulation and found that transgenic animals expressing ReaChR (red-activatable ChR) in SMD or SMB elicit immediate forward movement or increase of wave width, respectively, upon light exposure. Additionally, to identify the molecules that regulate forward movement, we examined expression pattern of 29 TRP and DEG/ENaC channels and found that unc-8 DEG/ENaC channel gene is expressed in SMB. unc-8 mutant animals fail to maintain wave form during forward movement. Furthermore, expression of UNC-8 in HEK293T cells generates outward currents upon mechanical deformation in a heterologous system. Currently, we are investigating function of the SMB, SMD and RME neurons and their circuit mechanisms in regulation of forward movement.