Niels Ringstad, PhD - US grants

DESCRIPTION (provided by applicant): description Serotonin plays critical roles as a neurotransmitter, neuromodulator and hormone. Therapeutics for many psychiatric disorders, including major depression, anxiety disorders and eating disorders, target serotonin signaling pathways in the brain. Understanding molecular mechanisms that regulate serotonin signaling is therefore essential both for understanding the causes of many types of neurological and psychiatric illness and for developing new therapeutics. The roundworm C. elegans is a powerful model for genetic and molecular studies of nervous system function. A synapse between serotonergic motor neurons and muscles of the reproductive system drives the reproductive behavior of the C. elegans hermaphrodite, egg laying. Through genetic studies of egg-laying behavior, we have discovered that neuropeptides modulate serotonin signaling in the C. elegans reproductive system by directly inhibiting serotonergic motor neurons. Some of these modulatory neuropeptides are provided by a pair of sensory neurons, the BAG neurons, which are stimulated by environmental carbon dioxide. The aim of this proposal is to determine physiological and molecular mechanisms required for the activation of peptidergic sensory neurons by carbon dioxide, and mechanisms by which neuropeptides inhibit the serotonergic neurons that they target.

Serotonin plays critical roles as a neurotransmitter, neuromodulator and hormone. Therapeutics for many psychiatric disorders, including major depression, anxiety disorders and eating disorders, target serotonin signaling pathways in the brain. Understanding molecular mechanisms that regulate serotonin signaling is therefore essential both for understanding the causes of many types of neurological and psychiatric illness and for developing new therapeutics. The roundworm C. elegans is a powerful model for genetic and molecular studies of nervous system function. A synapse between serotonergic motor neurons and muscles of the reproductive system drives the reproductive behavior of the C. elegans hermaphrodite, egg laying. Through genetic studies of egg-laying behavior, we have discovered that neuropeptides modulate serotonin signaling in the C. elegans reproductive system by directly inhibiting serotonergic motor neurons. Some of these modulatory neuropeptides are provided by a pair of sensory neurons, the BAG neurons, which are stimulated by environmental carbon dioxide. The aim of this proposal is to determine physiological and molecular mechanisms required for the activation of peptidergic sensory neurons by carbon dioxide, and mechanisms by which neuropeptides inhibit the serotonergic neurons that they target.

DESCRIPTION (provided by applicant): Chemosensory neurons specialized for the detection of carbon dioxide (CO2), a major product of aerobic metabolism, are present in the nervous systems of diverse animals. In vertebrates these neurons regulate breathing rhythms, and defects in CO2-sensing brain circuits are thought to underlie neurological disorders such as apneas and Sudden Infant Death Syndrome. Despite the critical roles that CO2-sensing neurons play in physiology, the molecular mechanisms required for their development and function remain poorly understood. Through genetic and physiological studies of CO2-sensing neurons of the nematode C. elegans we have established a powerful model for the study of such mechanisms. We have discovered that CO2-sensing neurons of C. elegans mediate a pathogen-avoidance behavior and require a Toll-like receptor (TLR) and its associated signaling pathway for their function. TLRs are evolutionarily conserved receptors that canonically function in embryonic patterning and innate immunity, and also function in the vertebrate nervous system to mediate inflammation. Our data indicate a previously unknown function for TLRs in the differentiation and function of sensory neurons and suggest a new role for TLRs in the vertebrate nervous system. Here we propose experiments to determine the molecular mechanisms by which TLR signaling promotes the differentiation and function of CO2-sensing neurons. These mechanisms will involve molecules required for TLR signaling, which is well known to play important roles in inflammation in the vertebrate brain and which our data suggest might also function in neuronal differentiation, as well as new components of the chemotransduction apparatus used for CO2-sensing.

PROJECT SUMMARY The neuromodulator dopamine is important for many brain functions: loss of dopamine neurons causes movement disorders, such as Parkinson's disease; dopamine signaling is targeted by drugs of abuse and integral to the neurobiology of reward and addiction; and dopamine signaling is a therapeutic target for the treatment of many neuropsychiatric disorders. Despite its importance in the brain, relatively little is known about mechanisms that regulate dopamine release in vivo or are otherwise required for the function of dopamine neurons. To directly address this question, we have used a simple invertebrate model ? the nematode C. elegans - to identify molecules required in vivo for dopamine signaling. We have identified genes whose expression is highly enriched in dopamine neurons, and by testing the corresponding mutants for defects in a behavior that requires endogenous dopamine we have identified five evolutionarily conserved genes that are likely required for the function of dopamine neurons. Using live-cell imaging and studies of dopamine neuron physiology we will determine the function of these genes. We will also use genetic analyses to determine how these genes interact with known components of dopamine signaling pathways. We expect that these studies will identify new therapeutic targets for the treatment of neurological and psychiatric disorders. Importantly, because extant therapeutic targets act downstream of dopamine release (they are receptors, transporters and dopamine-degrading enzymes), we also anticipate that our studies will identify targets with fundamentally new mechanisms of action in dopamine signaling.

PROJECT SUMMARY Neuromodulator and neurotransmitter signaling pathways are critical for brain function and are targeted for the treatment of diverse neurological and psychiatric disorders. The nervous system of the simple invertebrate Caenorhabditis elegans is endowed with many of the same neurotransmitters and neuromodulators that are critical for brain function and whose dysfunction is linked to human disease. We study a C. elegans neural circuit in which sensory neurons use neuropeptides to modulate serotonin neurons that drive reproductive behavior and the neurotransmitter glutamate to mediate avoidance behavior. Genetic analysis of the function of this circuit and its development permits the discovery of molecular mechanisms required in vivo for inhibitory neuropeptide signaling and excitatory glutamate signaling. Sensory neurons that coordinately regulate locomotory and reproductive behaviors are activated by carbon dioxide (CO2) evolved by microbial respiration. This simple circuit, therefore, also offers the opportunity to study mechanisms by which neurons sense respiratory gases - a critical chemosensory modality that in humans controls breathing rhythms and remains poorly understood at the molecular level. Our studies of this circuit have yielded genes that are conserved between invertebrates and humans, and we have discovered functions for these genes in transcriptional control of gene expression during sensory neuron development, CO2-chemosensing, control of glutamate release from neurons, and control of neuronal excitability by neuropeptide receptors. Our studies have yielded more genes that function in these processes and that remain to be characterized, and we expect that this sensory-motor circuit will continue to serve as a powerful platform for the discovery of genes that function in neuromodulator and neurotransmitter signaling and that might be eventually be developed as therapeutic targets.

PROJECT SUMMARY Sensory neurons concentrate and organize molecules used to detect environmental stimuli into cilia, which are specialized microtubule-based structures on the cell surface that function as cellular antennas. The proteins that constitute the machinery of sensory transduction are synthesized elsewhere and must be separated from other cellular proteins and transported to the cilium. The importance of mechanisms that mediate trafficking of proteins to the sensory cilium is illustrated by disease-causing mutations that disrupt this process. Mutations that compromise ciliary trafficking of the photopigment rhodopsin or the enzyme guanylyl cyclase cause retinal dystrophies marked by photoreceptor degeneration and, ultimately, blindness. Despite the importance of protein trafficking to the cilium, its underlying molecular mechanisms remain poorly understood. We propose to use chemosensory BAG neurons of the nematode C. elegans as a model for discovery of mechanisms that select and transport cargo destined for the sensory cilium. Like vertebrate photoreceptor neurons, BAG neurons use cyclic GMP as a second messenger for sensory transduction, and the enzymes and effectors that control cyclic GMP signals and turn them into electrical signals are highly similar to those found in photoreceptor neurons. Trafficking of proteins to BAG cilia can be measured in situ using high-resolution fluorescence microscopy assays, and powerful genetic tools are available to acutely or chronically manipulate specific molecular pathways in BAG neurons and determine their function in trafficking to the cilium. Importantly, C. elegans permits discovery of novel factors that mediate ciliary trafficking through genetic screens and biochemical approaches. We propose to use this powerful experimental system to (1) delineate a molecular pathway that matches cargo destined for the sensory cilium with specific motors that will carry it through the dendrite to its destination, and (2) determine how trafficking mechanisms are regulated by physiological or developmental cues that trigger remodeling of the BAG cilium. These studies will advance understanding of a cellular process that is essential for sensory neuron function and viability and will integrate cellular trafficking mechanisms with physiological and developmental programs that impact sensory cilia in vivo.

The neuromodulator dopamine is important for many brain functions: loss of dopamine neurons causes movement disorders, such as Parkinson?s disease; dopamine signaling is targeted by drugs of abuse and integral to the neurobiology of reward and addiction; and dopamine signaling is a therapeutic target for the treatment of many neuropsychiatric disorders. Despite its importance in the brain, relatively little is known about mechanisms that regulate synaptic dopamine release in vivo. And although the effects of dopamine on individual cells have been extensively studied, how dopamine signals are processed to change the dynamics of post synaptic neurons to execute changes of behavior is not well understood. The microscopic roundworm C. elegans offers the opportunity to study these aspects of dopamine signaling using powerful tools of molecular genetics and in vivo circuit analysis. Using behavioral genetics and newly developed methods for analysis of neural circuits in behaving animals we will (1) determine mechanisms that regulate dopamine release in response to appetitive stimuli and postsynaptic and (2) determine circuit mechanisms that transform dopamine signaling events into lasting changes in behavior. Because of the ancient and conserved functions of dopamine signaling in the animal nervous system, we propose that our studies will also advance understanding of pre- and postsynaptic mechanisms in dopamine systems of the human brain and accelerate discovery of new approaches to understanding and treating diseases linked to dysfunction of dopaminergic systems.