Pubs at Harvard

Caenorhabditis elegans uses olfaction as one of its primary means to sense the quality of its environment throughout its life span. Accordingly, the animal displays experience-dependent plasticity in olfactory sensorimotor responses at different life stages. These various forms of olfactory plasticity include imprinting, adaptation to prolonged odor exposure, conditioning with appetitive or aversive stimuli, and learning to avoid the smells of foods that make it ill. Moreover, a number of these C. elegans olfactory responses are subject to the aging process, as similar responses are in vertebrates. Indeed, the dissection of C. elegans olfactory plasticity has revealed mechanistic underpinnings at molecular, cellular, and circuit levels that show substantial similarities to the mechanisms underlying learning and memory in other animals, including humans.

DOI: 10.1016/B978-0-12-415823-8.00010-1

Dynamic serotonin biosynthesis is important for serotonin function; however, the mechanisms that underlie experience-dependent transcriptional regulation of the rate-limiting serotonin biosynthetic enzyme tryptophan hydroxylase (TPH) are poorly understood. Here, we characterize the molecular and cellular mechanisms that regulate increased transcription of Caenorhabditis elegans tph-1 in a pair of serotonergic neurons ADF during an aversive experience with pathogenic bacteria, a common environmental peril for worms. Training with pathogenic bacteria induces a learned aversion to the smell of the pathogen, a behavioral plasticity that depends on the serotonin signal from ADF neurons. We demonstrate that pathogen training increases ADF neuronal activity. While activating ADF increases tph-1 transcription, inhibiting ADF activity abolishes the training effect on tph-1, demonstrating the dependence of tph-1 transcriptional regulation on ADF neural activity. At the molecular level, the C. elegans homolog of CaMKII, UNC-43, functions cell-autonomously in ADF neurons to generate training-dependent enhancement in neuronal activity and tph-1 transcription, and this cell-autonomous function of UNC-43 is required for learning. Furthermore, selective expression of an activated form of UNC-43 in ADF neurons is sufficient to increase ADF activity and tph-1 transcription, mimicking the training effect. Upstream of ADF, the Gqalpha protein EGL-30 facilitates training-dependent induction of tph-1 by functional regulation of olfactory sensory neurons, which underscores the importance of sensory experience. Together, our work elucidates the molecular and cellular mechanisms whereby experience modulates tph-1 transcription.

The insulin/insulin-like peptides (ILPs) regulate key events in physiology, including neural plasticity. However, the cellular and circuit mechanisms whereby ILPs regulate learning remain largely unknown. Here, we characterize two ILPs that play antagonistic roles in aversive olfactory learning of C. elegans. We show that the ILP ins-6 acts from ASI sensory neurons to enable learning by repressing the transcription of another ILP, ins-7, specifically in URX neurons. A high level of INS-7 from URX disrupts learning by antagonizing the insulin receptor-like homolog DAF-2 in the postsynaptic neurons RIA, which play an essential role in the neural circuit underlying olfactory learning. We also show that increasing URX-generated INS-7 and loss of INS-6, both of which abolish learning, alter RIA neuronal property. Together, our results reveal an "ILP-to-ILP" pathway that links environment-sensing neurons, ASI and URX, to the key neuron, RIA, of a network that underlies olfactory plasticity and modulates its activity.

Reproductive behaviors have manifold consequences on evolutionary processes. Here, we explore mechanisms underlying female reproductive choice in the nematode Caenorhabditis elegans, a species in which females have evolved the ability to produce their own self-fertilizing sperm, thereby allowing these "hermaphrodites" the strategic choice to self-reproduce or outcross with males. We report that hermaphrodites of the wild-type laboratory reference strain N2 favor self-reproduction, whereas a wild isolate CB4856 (HW) favors outcrossing. To characterize underlying neural mechanisms, we show that N2 hermaphrodites deficient in mechanosensation or chemosensation (e.g., mec-3 and osm-6 mutants) exhibit high mating frequency, implicating hermaphrodite perception of males as a requirement for low mating frequency. Within chemosensory networks, we find opposing roles for different sets of neurons that express the cyclic GMP-gated nucleotide channel, suggesting both positive and negative sensory-mediated regulation of hermaphrodite mating frequency. We also show that the ability to self-reproduce negatively regulates hermaphrodite mating. To map genetic variation, we created recombinant inbred lines and identified two QTL that explain a large portion of N2 x HW variation in hermaphrodite mating frequency. Intriguingly, we further show that approximately 40 wild isolates representing C. elegans global diversity exhibit extensive and continuous variation in hermaphrodite reproductive outcome. Together, our findings demonstrate that C. elegans hermaphrodites actively regulate the choice between selfing and crossing, highlight the existence of natural variation in hermaphrodite choice, and lay the groundwork for molecular dissection of this evolutionarily important trait.

The confinement of neuronal activity to specific subcellular regions is a mechanism for expanding the computational properties of neurons. Although the circuit organization underlying compartmentalized activity has been studied in several systems, its cellular basis is still unknown. Here we characterize compartmentalized activity in Caenorhabditis elegans RIA interneurons, which have multiple reciprocal connections to head motor neurons and receive input from sensory pathways. We show that RIA spatially encodes head movement on a subcellular scale through axonal compartmentalization. This subcellular axonal activity is dependent on acetylcholine release from head motor neurons and is simultaneously present and additive with glutamate-dependent globally synchronized activity evoked by sensory inputs. Postsynaptically, the muscarinic acetylcholine receptor GAR-3 acts in RIA to compartmentalize axonal activity through the mobilization of intracellular calcium stores. The compartmentalized activity functions independently of the synchronized activity to modulate locomotory behaviour.

The TGF-beta superfamily is conserved throughout metazoan, and its members play essential roles in development and disease. TGF-beta has also been implicated in adult neural plasticity. However, the underlying mechanisms are not well understood. Here we report that DBL-1, a Caenorhabditis elegans TGF-beta homolog known to control body morphology and immunity, is essential for aversive olfactory learning of potentially harmful bacteria food. We show that DBL-1 generated by the AVA command interneurons, which are critical for sensorimotor responses, regulates aversive olfactory learning, and that the activity of the type I TGF-beta receptor SMA-6 in the hypodermis is needed during adulthood to generate olfactory plasticity. These spatial and temporal mechanisms are critical for the DBL-1 signaling to achieve its diverse functions in development and adult neural plasticity. Interestingly, aversive training decreases AVA calcium response, leading to an increase in the DBL-1 signal secreted from AVA, revealing an experience-dependent change that can underlie the role of TGF-beta signaling in mediating plasticity.

An insulin-like signaling pathway mediates the environmental influence on the switch between the C. elegans developmental programs of reproductive growth versus dauer arrest. However, the specific role of endogenous insulin-like peptide (ILP) ligands in mediating the switch between these programs remains unknown. C. elegans has 40 putative insulin-like genes, many of which are expressed in sensory neurons and interneurons, raising the intriguing possibility that ILPs encode different environmental information to regulate the entry into, and exit from, dauer arrest. These two developmental switches can have different regulatory requirements: here we show that the relative importance of three different ILPs varies between dauer entry and exit. Not only do we find that one ILP, ins-1, ensures dauer arrest under harsh environments and that two other ILPs, daf-28 and ins-6, ensure reproductive growth under good conditions, we also show that daf-28 and ins-6 have non-redundant functions in regulating these developmental switches. Notably, daf-28 plays a more primary role in inhibiting dauer entry, whereas ins-6 has a more significant role in promoting dauer exit. Moreover, the switch into dauer arrest surprisingly shifts ins-6 transcriptional expression from a set of dauer-inhibiting sensory neurons to a different set of neurons, where it promotes dauer exit. Together, our data suggest that specific ILPs generate precise responses to dauer-inducing cues, such as pheromones and low food levels, to control development through stimulus-regulated expression in different neurons.

Many animals use their olfactory systems to learn to avoid dangers, but how neural circuits encode naive and learned olfactory preferences, and switch between those preferences, is poorly understood. Here, we map an olfactory network, from sensory input to motor output, which regulates the learned olfactory aversion of Caenorhabditis elegans for the smell of pathogenic bacteria. Naive animals prefer smells of pathogens but animals trained with pathogens lose this attraction. We find that two different neural circuits subserve these preferences, with one required for the naive preference and the other specifically for the learned preference. Calcium imaging and behavioral analysis reveal that the naive preference reflects the direct transduction of the activity of olfactory sensory neurons into motor response, whereas the learned preference involves modulations to signal transduction to downstream neurons to alter motor response. Thus, two different neural circuits regulate a behavioral switch between naive and learned olfactory preferences.

Caenorhabditis elegans responds to chemical cues using a small number of chemosensory neurons that detect a large variety of molecules in its environment. During chemotaxis, C. elegans biases its migration in spatial chemical gradients by lengthening (/shortening) periods of forward movement when it happens to be moving toward (/away) from preferred locations. In classical assays of chemotactic behavior, a group of crawling worms is placed on an agar plate containing a point source of chemical, the group is allowed to navigate for a period of time, and aggregation of worms near the source is quantified. Here we show that swimming worms exhibit acute motile responses to temporal variations of odor in their surrounding environment, allowing our development of an automated assay of chemotactic behavior with single-animal resolution. By placing individual worms in small microdroplets and quantifying their movements as they respond to the addition and removal of odorized airstreams, we show that the sensorimotor phenotypes of swimming worms (wild-type behavior, the effects of certain mutations, and the effects of laser ablation of specific olfactory neurons) are consistent with aggregation phenotypes previously obtained in crawling assays. The microdroplet swimming assay has certain advantages over crawling assays, including flexibility and precision in defining the stimulus waveform and automated quantification of motor response during stimulus presentation. In this study, we use the microdroplet assay to quantify the temporal dynamics of the olfactory response, the sensitivity to odorant concentration, combinations, and gradients, and the contribution of specific olfactory neurons to overall behavior.

The nematode Caenorhabditis elegans is present in soils and composts, where it can encounter a variety of microorganisms. Some bacteria in these rich environments are innocuous food sources for C. elegans, whereas others are pathogens. Under laboratory conditions, C. elegans will avoid certain pathogens, such as Serratia marcescens, by exiting a bacterial lawn a few hours after entering it. By combining bacterial genetics and nematode genetics, we show that C. elegans specifically avoids certain strains of Serratia based on their production of the cyclic lipodepsipentapeptide serrawettin W2. Lawn-avoidance behavior is chiefly mediated by the two AWB chemosensory neurons, probably through G protein-coupled chemoreceptors, and also involves the nematode Toll-like receptor gene tol-1. Purified serrawettin W2, added to an Escherichia coli lawn, can directly elicit lawn avoidance in an AWB-dependent fashion, as can another chemical detected by AWB. These findings represent an insight into chemical recognition between these two soil organisms and reveal sensory mechanisms for pathogen recognition in C. elegans.