Salt TA et al. (1986) Steroids "24-Methyl-23-dehydrocholesterol: a new sterol intermediate in C-24 demethylation from the nematodes Panagrellus redivivus and Caenorhabditis elegans?"

Lusby WR, Thompson MJ, Chitwood DJ, Lozano R, Salt TA

[Steroids, 1986]

Panagrellus redivivus produced 24-methyl-23-dehydrocholesterol as 4.0% of the 4-desmethylsterols when propagated in a medium containing campesterol as the dietary sterol. The re-examination of previous data revealed that Caenorhabditis elegans produced 1.8% 24-methyl-23-dehydrocholesterol when propagated in medium containing campesterol. 24-Methyl-23-dehydrocholesterol was not detected when the nematodes were propagated in medium containing 22-dihydrobrassicasterol or 24-methylenecholesterol. This may be a result of the greater efficiency of dealkylation of the latter two sterols. This is the first report of the natural occurrence of this sterol in a non-photosynthetic organism, and the first report in organisms that dealkylate 24-alkylsterols.

Rachel T. Wragg et al. (2007) International Worm Meeting "Tyramine and octopamine inhibit serotonin-stimulated aversive behaviors in Caenorhabditis elegans through distinct receptors and signaling pathways."

Vera M. Hapiak, Rachel T. Wragg, Gareth P. Harris, Richard W. Komuniecki, Sarah B. Miller

[International Worm Meeting, 2007]

Nematodes lack an autonomic nervous system and epinephrine/norepinephrine and instead appear to use the biogenic amines, such as serotonin (5-HT), tyramine (TA) and octopamine (OA) to modulate many rhythmic behaviors. In C. elegans, distinct TA/OA specific neurons, receptors and behaviors have been identified. In general, TA and OA oppose the action of 5-HT causing hyperactivity and inhibiting 5-HT dependent egg-laying and pumping. In the present study, we have demonstrated that both TA and OA abolish 5-HT dependent increases in responses to dilute octanol. Therefore, we have analyzed the roles of previously characterized and putative TA and OA receptors through the use of TA/OA receptor null mutants. Interestingly, TA and OA inhibit 5-HT mediated increases in octanol responsiveness through different receptors; TYRA-3 is essential for inhibition by TA and F14D12.6 is essential for inhibition by OA. In addition, responses to TA and OA desensitize after pre-exposure to the amines and this adaptation appears to be amine-specific. Inhibition binding studies using [3H]LSD confirm that membranes from mammalian cells transiently expressing either TYRA-3 or F14D12.6 exhibit highest affinity for TA or OA, respectively. f14d12.6::gfp is expressed in a number of neurons, including the ASH sensory neuron that is primarily responsible for sensitvity to dilute octanol, while tyra-3::gfp is robustly expressed in the dopaminergic ADE/CEPs. However, the individual neuron(s) required for TA or OA inhibition remain to be identified. Taken together, these results demonstrate that, although tyraminergic and octopaminergic signaling yield identical phenotypes in our behavioral assays, these amines act through separate receptors located in distinct neurons.

Luo, L et al. (2011) International Worm Meeting "How worms move up and down salt gradients."

Samuel, A, Hendricks, M, Zhang, Y, Luo, L

[International Worm Meeting, 2011]

The behavioral and neural mechanisms by which C. elegans moves up NaCl gradients have been well characterized. The ASEL and ASER neurons are capable of sensing temporal upsteps and downsteps in salt concentration to bias a random walk towards regions with higher salt. By studying the movements of worms on linear salt gradients with defined slope and offset, we find that worms can both move up and down salt gradients in search of regions with salt concentrations that corresponds to their previous cultivation conditions. Using calcium imaging and quantitative behavioral analysis, we find that the ASEL and ASER neurons each have distinct roles in mediating movement up or down salt gradients when the worm navigates below or above the set-point of salt memory. Our results show how the salt-sensing circuit is modulated by experience to enable the worm to either ascend or descend salt gradients.

Vera Hapiak et al. (2006) Neuronal Development, Synaptic Function, and Behavior Meeting "Tyramine abolishes 5-HT dependent increases in octanol sensitivity in Caenorhabditis elegans"

Robert Hobson, Vera Hapiak, Rachel Wragg, Richard Komuniecki, Gareth Harris

[Neuronal Development, Synaptic Function, and Behavior Meeting, 2006]

Tyramine (TA) regulates a number of key processes in nematodes, including pharyngeal pumping, egg-laying, and locomotion. Previously, we have identified two C. elegans TA receptors, SER-2 and TYRA-2. In the present study, we have identified a third C. elegans gene, mo3f4.3, that also appears to encode for a TA receptor. mo3f4.3 is conserved in the parasitic nematode, Brugia malayi and is expressed as multiple isoforms with different N-termini. Membranes from COS-7 cells expressing M03F4.3a bind [3H]-LSD in the low nM range (Kd, 30 +/- 4 nM) with higher affinity for TA than other classical biogenic amines. For example, IC50s for TA, octopamine, dopamine and 5-HT are 0.5 +/- 0.07, 59.5 +/- 16.5, 83 +/- 23, and 540 +/- 127 uM, respectively. M03F4.3 does not appear to play any obvious role in TA-dependent hyperactivity or decreases in 5-HT stimulated egg-laying or pharyngeal pumping, based on an analysis of these phenotypes in putative mo3f4.3 null mutants. Interestingly, TA abolished 5-HT increases in sensitivity to 30% octanol and this TA-dependent inhibition was absent in animals containing null mutations in mo3f4.3, but not ser-2 or tyra-2. These studies are continuing to examine the G-protein coupling and downstream signaling of MO3F4.3 and the neuron-specific rescue of the TA dependent phenotypes observed in mo3f4.3 null mutants.

Adachi T et al. (2010) Genetics "Reversal of salt preference is directed by the insulin/PI3K and Gq/PKC signaling in Caenorhabditis elegans."

Tomioka M, Mori I, Adachi T, Iino Y, Kunitomo H, Ohno H, Okochi Y

[Genetics, 2010]

Animals search for foods and decide their behaviors according to previous experience. Caenorhabditis elegans detects chemicals with a limited number of sensory neurons, allowing us to dissect roles of each neuron for innate and learned behaviors. C. elegans is attracted to salt after exposure to the salt (NaCl) with food. In contrast, it learns to avoid the salt after exposure to the salt without food. In salt-attraction behavior, it is known that the ASE taste sensory neurons (ASEL and ASER) play a major role. However, little is known about mechanisms for learned salt avoidance. Here, through dissecting contributions of ASE neurons for salt chemotaxis, we show that both ASEL and ASER generate salt chemotaxis plasticity. In ASER, we have previously shown that the insulin/PI 3-kinase signaling acts for starvation-induced salt chemotaxis plasticity. This study shows that the PI 3-kinase signaling promotes aversive drive of ASER but not of ASEL. Furthermore, the Gq signaling pathway composed of Gq EGL-30, diacylglycerol, and nPKC (novel protein kinase C) TTX-4 promotes attractive drive of ASER but not of ASEL. A putative salt receptor GCY-22 guanylyl cyclase is required in ASER for both salt attraction and avoidance. Our results suggest that ASEL and ASER use distinct molecular mechanisms to regulate salt chemotaxis plasticity.

Kunitomo, Hirofumi et al. (2015) International Worm Meeting "Dissecting the roles of primary interneurons that regulate memory-dependent salt concentration chemotaxis."

Kunitomo, Hirofumi, Satoh, Yohsuke, Iino, Yuichi, Sato, Hirofumi

[International Worm Meeting, 2015]

Mapping the neuronal components of perception, memory, and motor control is critical in understanding how memory-dependent behavior is encoded by the nervous system. Salt chemotaxis of Caenorhabditis elegans is a memory-dependent navigation behavior: animals are attracted to the salt concentration at which they have been fed, whereas they avoid it if they have been starved (salt concentration chemotaxis). The animals employ a biased random walk strategy (also called the pirouette strategy) to migrate towards preferred directions. In this strategy, the frequency of reorientation depends on the temporal change of salt concentration and this bias is modulated by the differences in salt concentration between the current environment and the memorized one. Interestingly, input from a single sensory neuron, ASER, is required and sufficient for salt concentration chemotaxis under well fed conditions, and ASER is activated by salt concentration decrease irrespective of cultivation salt concentrations. To understand how sensory inputs are translated into behavior, we examined the roles of postsynaptic neurons of ASER by ablating them individually or in combination. ASER densely synapses on three pairs of interneurons, AIA, AIB and AIY. Loss of AIY or mutations in ttx-3 resulted in impaired chemotaxis to low salt but not to high salt. On the other hand, AIA-ablated animals showed a weak but significant defect in chemotaxis to high salt. Ablating both AIY and AIA disrupted chemotaxis to both directions. Quantitative analyses of locomotion revealed that these chemotaxis defects were concomitant with altered properties of the pirouette strategy: upon salt concentration decreases, AIY-abated animals showed a defect in suppression of turning after cultivation at low salt, whereas AIA-ablated animals showed a defect in activation of turning after cultivation at high salt. Ablation of AIB interneurons did not cause discernible effect on salt concentration chemotaxis, although these neurons were involved in the regulation of ASER-evoked reorientation. These results indicate that although both AIY and AIA are involved in the regulation of turning frequency upon salt concentration decreases, they differently mediate transmission of sensory information after cultivation at distinct salt concentrations.

Kunitomo, Hirofumi et al. (2009) International Worm Meeting "A combination of salt and food conditions in the habitat affects preference for salt in C. elegans."

Kunitomo, Hirofumi, Iino, Yuichi

[International Worm Meeting, 2009]

Caenorhabditis elegans senses a wide variety of chemicals associated with food, danger, or other animals. Sodium chloride is generally considered as an attractive cue for the animal. We have previously reported, however, that the worms avoid NaCl when animals are pretreated with NaCl under starvation. This behavioral plasticity, salt chemotaxis learning, is regulated by the insulin/PI3-K signaling pathway that functions in ASER, the right member of the bilateral gustatory neurons, ASE (Tomioka et al. 2006). Here we show that cultivation of worms under trace amount of NaCl in the presence of food for several hours or longer also induces avoidance of NaCl. This type of plasticity of salt chemotaxis, tentatively called low salt conditioning, is stimulus-specific and reversible. Salt, but not osmolarity, in the growth media is required for maintaining preference of salt. To reveal the mechanisms of low salt conditioning, we assessed the behavior of the mutants that show defects in salt chemotaxis learning or in the development of specific neurons. Of the insulin signaling pathway components, age-1 and akt-1 were required for low salt conditioning, whereas ins-1 and pdk-1 were not. casy-1, an ortholog of calsyntenin essential for salt chemotaxis learning, was not involved in low salt conditioning. Interestingly, the mutants of odr-7, which is required for proper differentiation of the AWA olfactory neurons, showed severe defects in low salt conditioning but not in salt chemotaxis learning. These results indicate that salt chemotaxis is modulated by a combination of salt and food conditions in the habitat and the two plasticity paradigms, both resulting in avoidance of salt, are regulated at least in part by distinct genetic and neural mechanisms.

Jang, M.S. et al. (2017) International Worm Meeting "Identification of neurons and analysis of the neural circuit involved in the learned salt-avoidance behavior in C. elegans."

Kunitomo, H., Jang, M.S., Toyoshima, Y., Iino, Y.

[International Worm Meeting, 2017]

Animals flexibly adapt to the environment by modifying their behavior, which depends on the plasticity of neural network. Here, we focused on the functional plasticity of neural circuits of Caenorhabditis elegans that sense salts and generate taxis behaviors. C. elegans shows a form of associated learning in which availability of food and salt concentration are associatively memorized. Worms are attracted to the salt concentration at which they have been cultured under well-fed conditions, whereas they avoid it if they have been cultivated under starvation, which hereafter we call salt avoidance learning. The ASER gustatory neuron is known to sense salt and has a major function for salt chemotaxis plasticity. Inputs to the ASER gustatory neuron are sufficient for the salt chemotaxis plasticity under well-fed conditions. However, other sensory neuron(s) in addition to ASER neuron were suggested to be required for salt avoidance learning. To identify the sensory neurons required for salt avoidance learning, we conducted cell-specific rescue experiments using the dysesthesia mutant dyf-11, whose sensory endings are dysfunctional so that these animals are unable to sense water-soluble compounds. As a result, we found that dyf-11 mutants with functionally recovered ASG neurons in addition to ASER showed the salt avoidance learning, but recovery in either ASER or ASG alone did not. Behavioral analyses revealed that the frequency of the turning events upon salt concentration change was recovered in animals whose ASER and ASG were rescued. Moreover, silencing ASG reduced salt chemotaxis after starvation. Unexpectedly, the ASG neurons did not respond to salt stimuli under any conditions we have tested. These results suggest that the ASG neurons have functions to regulate salt chemotaxis behaviors under starved conditions without responding to the salt stimuli themselves.

Sakurai, Yuki et al. (2011) International Worm Meeting "Genetic analysis of food-associated salt concentration memory."

Sakurai, Yuki, Iino, Yuichi, Kunitomo, Hirofumi

[International Worm Meeting, 2011]

Caenorhabditis elegans is usually attracted to NaCl if grown on standard culture conditions. On the other hand, it avoids the salt after exposure to salt under starvation (salt chemotaxis learning). We have reported that the insulin/PI3-kinase signaling regulates salt chemotaxis learning in ASER, one of the major taste neurons of the organism (Tomioka et al. 2006, Adachi et al, 2010). To examine how preference for salt is modified by experiences, we exploited a novel behavioral test, salt preference assay. Briefly, adult animals grown under standard culture conditions (50 mM NaCl with E. coli as food) are transferred to a conditioning plate that contains a defined concentration, 0-100 mM, of NaCl with food. After 6 hours of conditioning, worms are washed out from the plates and tested for chemotaxis to 35-95 mM of NaCl. Through this analysis, we found that wild-type C. elegans is attracted to the NaCl concentrations at which it has been fed (food-associated salt preference), indicating that the behavior on salt gradient is based on salt concentration memory. Mutants of PI3-kinase pathway didnt show significant defects in this assay. This result suggests that the mechanisms for salt preference is distinct, at least in part, from that of salt chemotaxis learning. To elucidate the mechanisms of food-associated salt preference, we screened for mutants of food-associated NaCl concentration preference. 30,000 EMS-mutagenized F2 animals were divided into 30 independent groups and screened by the salt preference assay after 0 or 100 mM NaCl conditioning with food. Animals that were attracted to high or low concentrations of NaCl after 0 or 100 mM conditioning, respectively, were collected and bred for next generation. In this screen we isolated several independent mutant strains JN572-JN577. JN574 showed no food-associated salt preference, whereas JN572 and JN577 showed defects only after conditioning at low concentrations of salt. These mutants are not likely chemosensory mutants because they showed normal salt chemotaxis after starvation. Characterization of these mutants will clarify the molecular mechanisms as to how the animals adapt to ambient salt conditions.

Saul N et al. (2010) J Gerontol A Biol Sci Med Sci "The longevity effect of tannic acid in Caenorhabditis elegans: Disposable Soma meets hormesis."

Sturzenbaum SR, Pietsch K, Menzel R, Steinberg CE, Saul N

[J Gerontol A Biol Sci Med Sci, 2010]

This study shows that exposure to low concentrations of the polyphenol tannic acid (TA) induces potent life-prolonging properties in Caenorhabditis elegans. In addition, enhanced thermal stress resistance, reduced growth, and slightly increased oxidative stress resistance were observed, although reproductive capacities and pharyngeal pumping rate were not modulated. Exploiting a suite of 14 mutant strains revealed that the mitogen-activated protein kinase kinase SEK-1 (SAPK/ERK kinase) is a key player involved in TA-mediated longevity. It is conceivable that TA mimics pathogen action and therefore activates the SEK-1-mediated pathogen resistance pathway. This pathway is thought to inhibit potential detrimental effects of TA and may also be involved in the longevity process. The observed dose response suggests the presence of a hormesis effect, and the growth impairment is in agreement with the "Disposable Soma Theory." This report underlines the uniqueness of TA-mediated longevity and facilitates a first glimpse into its complex mode of action.