No abstract

SUMMARY Many animals use their olfactory systems to learn to avoid dangers, but how neural circuits encode naïve 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. Naïve 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 naïve preference and the other specifically for the learned preference. Calcium imaging and behavioral analysis reveal that the naïve 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 naïve and learned olfactory preferences.

One advantage of the nematode Caenorhabditis elegans as a model organism is its suitability for in vivo optical microscopy. Imaging C. elegans often requires animals to be immobilized to avoid movement-related artifacts. Immobilization has been performed by application of anesthetics or by introducing physical constraints using glue or specialized microfluidic devices. Here we present a method for immobilizing C. elegans using polystyrene nanoparticles and agarose pads. Our technique is technically simple, does not expose the worm to toxic substances, and allows recovery of animals. We evaluate the method and show that the polystyrene beads increase friction between the worm and agarose pad. We use our method to quantify calcium transients and long-term regrowth in single neurons following axotomy by a femtosecond laser.

A protonmotive force (pmf) across the cell's inner membrane powers the flagellar rotary motor of Escherichia coli. Speed is known to be proportional to pmf when viscous loads are heavy. Here we show that speed also is proportional to pmf when viscous loads are light. Two motors on the same bacterium were monitored as the cell was slowly deenergized. The first motor rotated the entire cell body (a heavy load), while the second motor rotated a small latex bead (a light load). The first motor rotated slowly and provided a measure of the cell's pmf. The second motor rotated rapidly and was compared with the first, to give the speed-pmf relation for light loads. Experiments were done at 24.0°C and 16.2°C, with initial speeds indicating operation well into the high-speed, low-torque regime. Speed was found to be proportional to pmf over the entire (accessible) dynamic range (0 -270 Hz). If the passage of a fixed number of protons carries the motor through each revolution, i.e., if the motor is tightly coupled, a linear speed-pmf relation is expected close to stall, where the work done against the viscous load matches the energy dissipated in proton flow. A linear relation is expected at high speeds if proton translocation is rate-limiting and involves multiple steps, a model that also applies to simple proton channels. The present work shows that a linear relation is true more generally, providing an additional constraint on possible motor mechanisms.bacteria ͉ energetics ͉ molecular motors ͉ motility T he flagellar rotary motor of an Escherichia coli bacterium couples proton flux across the inner cell membrane to motor rotation. Protons move down an electrochemical gradient through specific MotA͞B protein complexes anchored to the cell wall. As a result, these complexes, known as torque-generating units, step along the rotor, causing it to rotate. At heavy viscous loads and low speeds, motor torque is approximately constant (and efficiencies are high), whereas at light viscous loads and high speeds, motor torque declines rapidly (and efficiencies become small). For a review of motor structure and function, see ref.1. Fung and Berg (2) found that when the motor operates in the low-speed regime near stall (Ͻ10 Hz), its speed is proportional to the voltage applied across the inner cell membrane. The experiments reported here extend these measurements well into the motor's high-speed regime. In this regime, the speedprotonmotive force (pmf) relation will depend explicitly on the mechanochemical cycle of the motor; therefore, its characterization puts new restrictions on possible motor mechanisms. MethodsCells of E. coli strain KAF95, a strain with sticky filaments that rotate exclusively counterclockwise (3), were inoculated from frozen stocks, grown in Luria broth [1% tryptone (Difco)͞0.5% yeast extract (Difco)͞0.5% NaCl] at 30°C until mid-exponential phase, and then sheared by using syringes with 26-gauge needles connected by 0.58-mm i.d. polyethylene tubing, producing cells with short flagellar stubs. These cells were...

The nematode Caenorhabditis elegans deliberately crawls toward the negative pole in an electric field. By quantifying the movements of individual worms navigating electric fields, we show that C. elegans prefers to crawl at specific angles to the direction of the electric field in persistent periods of forward movement and that the preferred angle is proportional to field strength. C. elegans reorients itself in response to time-varying electric fields by using sudden turns and reversals, standard reorientation maneuvers that C. elegans uses during other modes of motile behavior. Mutation or laser ablation that disrupts the structure and function of amphid sensory neurons also disrupts electrosensory behavior. By imaging intracellular calcium dynamics among the amphid sensory neurons of immobilized worms, we show that specific amphid sensory neurons are sensitive to the direction and strength of electric fields. We extend our analysis to the motor level by showing that specific interneurons affect the utilization of sudden turns and reversals during electrosensory steering. Thus, electrosensory behavior may be used as a model system for understanding how sensory inputs are transformed into motor outputs by the C. elegans nervous system.