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Genetic Analysis of Olfactory Behavior and Neural Development
Summary: Cornelia Bargmann uses genetic approaches to ask how neural circuits develop and how they function to generate behavior.
An animal uses stable neural circuits to generate flexible behavioral responses to its environment. We are studying the relationships between genes, the nervous system, and behaviors in the nematode Caenorhabditis elegans. We can define the circuits for specific behaviors in C. elegans because its nervous system consists of just 302 neurons with reproducible functions, morphologies, and synaptic connections. Using this information, we study genes that affect neuronal development and behavior and determine how these genes regulate the assembly and function of neural circuits.
Olfaction and Olfactory Learning
In C. elegans and other animals, odors are detected by large families of G protein–coupled receptors. Each C. elegans olfactory neuron expresses many receptor genes, allowing a few cells to detect many odors. The odors that activate one sensory neuron regulate a common behavioral output, such as attraction or avoidance.
An important function of the olfactory system is to allow animals to evaluate potential food odors. C. elegans requires bacterial food for survival but is susceptible to infection by pathogenic bacteria following ingestion of a tainted food source. One of the most robust forms of learning in any animal is conditioned avoidance of tastes associated with visceral distress. We found that pathogenic bacteria elicit C. elegans learning behaviors similar to mammalian conditioned taste aversion. After exposure to pathogenic bacteria, C. elegans modifies its olfactory preferences to avoid odors from the pathogen. This modification is specific, suggesting that the animal associates the experience of infection with the odor of the noxious bacterium.
Learning to avoid pathogenic bacteria requires the neurotransmitter serotonin. Exposure to pathogenic bacteria elevates serotonin levels in one class of neurons called ADF. Serotonin from ADF acts through a serotonin receptor in sensory interneurons to promote aversive learning, and exogenous serotonin causes accelerated learning. Increased serotonin may represent the negative reinforcing stimulus during pathogenic infection.
Oxygen Sensation and Oxygen-Dependent Behavior
Levels of oxygen vary widely in natural soil and water environments. C. elegans has rapid behavioral responses to oxygen that can be monitored when animals move through a gas-phase oxygen gradient. Wild-type C. elegans accumulates at about 5–10 percent oxygen, avoiding both high and low oxygen levels. Normal aerotaxis behavior requires at least three different soluble guanylate cyclases (sGCs) that can directly bind oxygen through their heme domains (work of our collaborators, David Karow and Michael Marletta [University of California, Berkeley]). These results suggest that sGCs are a new class of molecular receptor for environmental oxygen.
Oxygen levels can modify behavioral responses to other stimuli. One behavior that is regulated by oxygen is aggregation, or social feeding. Aggregation is rare in the standard laboratory strain of C. elegans but prominent in wild C. elegans isolates that bear the neuropeptide receptor allele npr-1(215F). Aggregation in npr-1(215F) strains occurs only at high oxygen levels and is controlled by sensory neurons that detect oxygen. Oxygen levels are low in aggregates, so aggregation appears to be a strategy for avoiding hyperoxia.
The standard laboratory C. elegans strain does not avoid high oxygen in the presence of food. Aerotaxis modulation by food is accomplished by npr-1 neuropeptide signaling, a TGFβ (transforming growth factor-β) peptide that regulates transcription, and the neurotransmitter serotonin. These interacting systems regulate a distributed network of oxygen-sensing neurons to mediate the switch from weak aerotaxis on food to strong aerotaxis in its absence. The complete neuroanatomical "wiring diagram" of the C. elegans nervous system does not include these regulatory pathways, indicating that functional experiments are needed to uncover circuits even in anatomically well-described nervous systems.
The Development of Neural Circuits
To determine the rules for the development of neuronal circuits, we have been studying axon formation, axon guidance, and synapse formation.
UNC-6/netrin and its receptor UNC-40/DCC are conserved regulators of axon guidance. We developed reagents to observe UNC-6–dependent neurons as they develop in vivo and found that UNC-6 and UNC-40 act not only during axon guidance but also earlier in axon formation to initiate, maintain, and orient asymmetric neuronal growth. Early in development, the immature HSN neuron breaks spherical symmetry to extend a leading edge in the direction taken by the future axon. In unc-6 and unc-40 mutants, leading-edge formation fails, the cell remains symmetrical until late in development, and the axon that eventually forms is misguided. Thus netrin has two activities: one that breaks neuronal symmetry, and one that guides the future axon. As the axon forms, the lipid modulators AGE-1/phosphoinositide 3-kinase (PI3K) and DAF-18/PTEN phosphatase drive the actin-regulatory lipid-binding protein MIG-10/lamellipodin ventrally in HSN to promote asymmetric growth. The coupling of a directional cue to sustained asymmetric growth via PI3K signaling is reminiscent of polarization in chemotaxing cells.
A second view of polarized growth and axon morphology was provided by studies of secreted proteins of the Wnt family. We found that C. elegans Wnts acting through Frizzled receptors shape axon and dendrite trajectories by specifying the polarity of neurons. In lin-44/Wnt and lin-17/Frizzled mutants, the entire polarity of the PLM neuron is reversed along the anterior-posterior body axis. The Frizzled receptor LIN-17 is asymmetrically localized to the posterior process of PLM in response to Wnt/LIN-44, indicating that Wnt signaling redistributes LIN-17 to polarize the cell. In this context, Wnts appear to function not as growth cone attractants but as organizers of neuronal polarity.
One of the most interesting periods of neuronal development is the period of circuit assembly, when neurons form synapses, take on their final differentiated properties, and coordinate their activity in networks. We obtained unexpected insight into circuit assembly through studies of a left-right asymmetry in the C. elegans olfactory system. A bilateral pair of olfactory neurons called AWCs communicates during embryogenesis to establish stochastic, asymmetric patterns of gene expression in the left and right neurons. We found that a transient network of cells linked by a gap-junction protein, NSY-5, initiates and coordinates AWC left-right asymmetry. The AWCs and several other classes of neurons are interconnected in an NSY-5–dependent gap-junction network that exists just at the time of synapse formation in the embryo. This network allows the AWCs to communicate and modifies both gene expression and synaptic connections between neurons; after it has established the proper gene expression patterns and synaptic patterns, the gap-junction network melts away. Gap junctions are widespread in immature neuronal circuits, but their functional significance is poorly understood. These results provide new insight into gap-junction activity in developing circuits.
Work on olfaction was funded by a grant from the National Institutes of Health.
Last updated: November 12, 2007
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