Navin Pokala

Approaches for inhibiting and activating neurons are essential for understanding how neurons and neuronal circuits produce behavior. Optogenetics is a recently-developed technique which uses light to manipulate neuronal activity with temporal precision in behaving animals and is widely-used by neuroscience researchers. Optogenetics is also an excellent method to incorporate into an undergraduate neuroscience laboratory module because students can learn to conduct and analyze quantitative behavioral assays, reinforce their understanding of synaptic transmission, and investigate the genetic and neuronal basis of behavior. Here, we describe a module in which students use light to activate serotonergic neurons expressing the light-activated ion channel channelrhodopsin in wildtype and mutant Caenorhabditis elegans, and observe and analyze the effects on the movement behavior of the nematode. The methods described here provide a foundation which students can use to design and conduct additional experiments that may have never been done before. Thus, this laboratory module provides an opportunity for students to learn a state-of-the-art neuroscience technique, think about neuroscience on genetic, cellular and behavioral levels, and to develop an independent research project.

SUMMARY: Foraging strategies that enable animals to locate food efficiently are composed of highly conserved behavioral states with characteristic features. Here, we identify parallel multimodal circuit modules that control an innate foraging state -- local search behavior -- after food removal in the nematode Caenorhabditis elegans . Two parallel groups of chemosensory and mechanosensory glutamatergic neurons that detect food-related cues trigger local search by inhibiting separate integrating neurons through a metabotropic glutamate receptor, MGL-1. The chemosensory and mechanosensory modules are separate and redundant, as glutamate release from either can drive the full behavior. Spontaneous activity in the chemosensory module encodes information about the time since the last food encounter and correlates with the foraging behavior. In addition, the ability of the sensory modules to control local search is gated by the internal nutritional state of the animal. This multimodal circuit configuration provides robust control of an innate adaptive behavior.

We describe a chemical-genetic approach for inducible silencing ofCaenorhabditis elegansneurons in intact animals, using the histamine-gated chloride channel HisCl1 fromDrosophilaand exogenous histamine. Administering histamine to freely movingC. elegansthat express HisCl1 transgenes in neurons leads to rapid and potent inhibition of neural activity within minutes.C. elegansdoes not use histamine as an endogenous neurotransmitter, and exogenous histamine has little apparent effect on wild-typeC. elegansbehavior.

Standardized driver and effector lines that use optimized GAL4 from a cryophilic yeast species enable bipartite control of transgene expression in Caenorhabditis elegans. The GAL4–UAS system is a powerful tool for manipulating gene expression, but its application in Caenorhabditis elegans has not been described. Here we systematically optimize the system's three main components to develop a temperature-optimized GAL4–UAS system (cGAL) that robustly controls gene expression in C. elegans from 15 to 25 °C. We demonstrate this system's utility in transcriptional reporter analysis, site-of-action experiments and exogenous transgene expression; and we provide a basic driver and effector toolkit.

Random search is a behavioral strategy used by organisms from bacteria to humans to locate food that is randomly distributed and undetectable at a distance. We investigated this behavior in the nematode Caenorhabditis elegans, an organism with a small, well-described nervous system. Here we formulate a mathematical model of random search abstracted from the C. elegans connectome and fit to a large-scale kinematic analysis of C. elegans behavior at submicron resolution. The model predicts behavioral effects of neuronal ablations and genetic perturbations, as well as unexpected aspects of wild type behavior. The predictive success of the model indicates that random search in C. elegans can be understood in terms of a neuronal flip-flop circuit involving reciprocal inhibition between two populations of stochastic neurons. Our findings establish a unified theoretical framework for understanding C. elegans locomotion and a testable neuronal model of random search that can be applied to other organisms.

Significance: Inducible silencing of neurons is a powerful method for studying the functions of neural circuits. We describe a system for inducible silencing ofCaenorhabditis elegansneurons, based on transgenicC. elegansnematodes engineered to produce the inhibitoryDrosophilahistamine-gated chloride channel (HisCl1) in specific neurons. The activity of HisCl1-expressing neurons is acutely and reversibly silenced by exogenous histamine, whichC. elegansabsorbs from the media. In addition to completely silencing neurons, the histamine-HisCl1 system can be used to titrate neural activity, demonstrating quantitative relationships between neural activity and behavior. The histamine-HisCl1 system is complementary to other methods of monitoring and manipulating neuronal activity, providing opportunities for combinatorial control of circuits.