Key Words behavior, sensory signal transduction, nervous system, neural plasticity, learning and memory
* Abstract Molecular genetic analysis of chemotaxis and thermotaxis in Caenorhabditis elegans has revealed the molecular bases of olfaction, taste, and thermosensation, which, in turn, has demonstrated that sensory signaling in C. elegans is very similar to that in vertebrates. A cyclic nucleotide-gated channel (TAX-2/TAX-4) that is highly homologous to the olfactory and photoreceptor channels in vertebrates is required for taste and thermosensation, in addition to olfaction. A cation channel (OSM-9) that is closely related to a capsaicin receptor channel is required for olfactory adaptation in one olfactory neuron and olfactory sensation in the other olfactory neuron. A novel G[alpha] protein (ODR-3) is essential for olfactory responses in all olfactory neurons and aversive responses in a polymodal sensory neuron. A G protein--coupled seven-transmembrane receptor (ODR-10) is the first olfactory receptor whose ligand was elucidated. Using chemotaxis and thermotaxis as behavioral paradigms, neural plasticity i ncluding learning and memory can be studied genetically in C. elegans.
Animals respond to a variety of environmental signals, and a rich repertoire of behavioral responses to those signals is directed by the nervous system. How does the nervous system function to receive, process, integrate, and interpret sensory signals to generate appropriate behaviors? Recent genetic approaches to chemotaxis and thermotaxis of the nematode Caenorhabditis elegans have provided molecular and cellular clues to these questions. C. elegans is an ideal model to conduct genetic, cellular, and molecular analyses of behavior. The small body size, short life cycle, hermaphroditic reproduction, and accessibility to classical genetics all make it possible to efficiently analyze mutants exhibiting abnormal behaviors . Our comprehensive knowledge of cell lineage and the wiring diagram of its nervous system facilitate cellular analysis of the mutations that cause defects in neuronal development and function [81,82, 89]. The various molecular methodologies for gene cloning, gene knockout, gene disruption , construction of transgenic animals, etc, can also be used to analyze genes involved in the nervous system [30,45,58,69]. Furthermore, the recent publication of the entire genome sequence of C. elegans should accelerate detailed molecular dissection of its nervous system .
BEHAVIORAL GENETICS IN CAENORHABDITIS ELEGANS
The Nervous System and Behaviors
There are 959 somatic cells in total in the adult hermaphrodite of C. elegans, with the nervous system composed of 302 neurons . This is an extraordinarily small number compared to those of other model animals used in neurobiological studies. These neurons are of 118 types based on their morphologies, positions, and connections with other cells, but most are classified as one of the three neuron types: sensory neuron, interneuron, or motorneuron . There are approximately 5000 chemical synapses and 600 gap junctions that, potentially, allow direct or indirect communication between neurons . In the C. elegans genome, there are many genes encoding components important for synaptic connection and gap junction formation. These include presynaptic components required for neurotransmitter release, such as synaptobrevin, syntaxin, and synaptotagmin; many postsynaptic components, such as various glutamate receptors and acetylcholine receptors; and innexins implicated in the formation of gap junctions . Chemical synapses are formed between nearby axons that run in parallel in C. elegans . For any given axon, synapse formations in principle occur along the whole of its length, although there seem to be clusters of regions rich in synapses. The biological significance of these synapse-clusters is not known. The presynaptic site is found next to the postsynaptic site. Hence, for most axons, there is no straightforward distinction between regions that contain exclusively presynaptic sites or postsynaptic sites. Numerous synapses are found in the largest neuropile, called the nerve ring, which is a huge bundle of many axons encircling the center part of the pharynx. The nerve ring is regarded as the CNS or brain of C. elegans.
C. elegans uses a simple, condensed nervous system to respond to diverse environmental stimuli, such as touch, taste, smell, and temperature. One advantage of using C. elegans as a model is that a behavior can be analyzed in the context of connected neurons that direct it. For example, a gentle touch to the body is detected by a set of six mechanosensory touch receptor neurons; these have special microtubule structures and are situated just below the cuticle, thereby allowing sensitivity to the touch stimuli to be maximized . Further, the exquisite positioning of the touch receptor neurons allows the fine touch stimuli to be detected throughout the length of the animal. A light touch to the anterior half or posterior half of the body induces backward or forward movement, respectively; these opposite responses are consistent with antagonistically regulated neural networks downstream of anteriorly and posteriorly positioned touch receptor neurons . This neural model for the touch response is consistent with the results of laser ablation studies, in which individual neurons were killed by a laser microbeam and the resultant behavioral phenotypes were later examined . Many mec mutations that disrupt the touch response have been isolated, and analysis of mec genes revealed molecular components required for mechanosensory transduction. This proposed molecular model, based on a predicted mechanosensory channel (a member of the Degenerin/ENaC gene family) and the associated proteins [40,43,44,53], provided valuable insight into vertebrate mechanotransductions such as hearing by hair cells in the ear and blood pressure regulation by baroreceptor nerve terminals [22,31].
Chemotaxis C. elegans possesses a remarkable ability to detect and discriminate a wide range of chemical compounds . These include water-soluble chemicals such as cyclic nucleotides (cAMP and cGMP); biotin, anions, and cations ([Cl.sup.-], [Na.sup.+], and [K.sup.+]); amino acids (lysine and histidine); and basic pH, and volatile chemicals such as alcohols, ketones, esters, pyrazines, thiazoles, and aromatic compounds [5,6,87]. In the natural environment, the ability to locate a food source and escape from an unfavorable habitat is dependent on this chemosensation. In the laboratory, the responses to various chemicals are observed in chemotaxis assays. Briefly, the animals are exposed to a concentration gradient consisting of a water-soluble or volatile substance, and the movement of animals is evaluated to determine whether the substance tested acts as an attractant or repellent. In addition, C. elegans senses a pheromone that can induce a dauer larva, a developmentally arrested larval form [7,71]. Dauer l arvae are formed when the animals detect conditions detrimental to survival and reproduction such as high population density (high concentration of constitutively secreted pheromone), a shortage of food, or high temperature . When food is provided, the dauer larvae resume normal development and become reproductive adults. Thus, chemosensation not only directs the animal's behavioral responses, it also influences its developmental decisions.
Thermotaxis Temperature affects almost all aspects of the developmental and physiological processes in any animal. The thermosensory system has evolved not only to protect against exposure to temperatures that trigger undesirable physiological reactions, but also to seek to temperatures conducive to survival and reproduction. That C. elegans can detect a range of temperatures very accurately is reflected in its thermotactic behavior [8, 59]. For free-living soil nematodes, thermotaxis is an essential behavioral response to ensure their propagation. Temperature gradients are created in the laboratory to observe thermotactic responses [36, 60]. The animals that were cultivated normally with food at temperatures ranging from 15[degrees]C to 25[degrees]C migrate to the cultivation temperature on a temperature gradient and move isothermally at that temperature. By contrast, the animals migrate away from the temperature at which they were previously starved. Interestingly, resetting acclimation to a new temperature or inducing avoidance response from an unfavorable temperature takes 2-4 h each (I Mori, unpublished data). How are these behavioral changes accomplished? Thermotaxis of C. elegans provides a behavioral model system for genetic analysis of neural plasticity, and possibly memory and learning also (see below).
Structural and Functional Differentiation in Chemosensory and Thermosensory Neurons
The chemosensory neurons required for chemotaxis were identified through a series of laser killing experiments. There are 32 chemosensory neurons that fall into 14 types; of these, 11 types comprising 22 chemosensory neurons are components of the amphid sensilla, the most complicated sensory organs, situated as a bilaterally symmetric pair at the tip of the head  (Figure 1). Laser ablation studies demonstrated that most of the individual amphid chemosensory neurons detect either water-soluble or volatile chemicals, and direct either attraction or aversion, although some compounds act as an attractant at a low concentration and as a repellent at a high concentration [5, 6, 83, 84] (Figure 1). The ADF, ASE, ASG, ASI, and ASK neurons mediate chemotaxis to water-soluble attractants, whereas the AWA and AWC neurons mediate chemotaxis to volatile attractants, and the AWB neurons mediate volatile avoidance. The ASH and ADL neurons only direct avoidance responses, but they are unusual in that both neurons can det ect distinct classes of stimuli; the ASH neurons sense mechanical and …