Figure 1.
A. Schematic diagram of the recording system. Three subsystems are indicated: Ratiometric calcium imaging (blue), behavior imaging (red), and image recentering (green). Gray objects show the output of each subsystem. B. Performance of the image recentering subsystem, as illustrated by the probability density of radial eccentricities for a single neuron in a freely moving animal. Dashed lines show the region containing 95% of the data. C. Performance of the image recentering subsystem, as illustrated by the cumulative probability of cell displacement between contiguous frames in tracking mode (green line) and for a stationary stage (black line). Data for a stationary stage were predicted based on the animal's speed at the time the frame was taken. Dotted lines indicate the cumulative probabilities for a displacement of one neuron radius in the tracking and stationary modes. Measurements were obtained from the same data set as in B.
Figure 2.
Osmotic avoidance responses in a virtual environment.
Channelrhodopsin-2 was specifically expressed in the osmo-sensitive neuron class ASH. Animals were tracked via a small cluster of neurons expressing a red fluorescent protein located in the head. ASH was photoactivated whenever the animal's head entered an annular region with outside and inside diameters of 1.4 and 1.0 mm, respectively. A. Representative tracks of animals raised in presence (A1) or absence (A2) of the co-factor retinal; blue segments indicate photoactivation. B. Probability density of the distance from the center of the annulus for 25 retinal (+) and retinal (−) animals. The blue bars indicate the photoactivation zone. C. Probability density of spatial error, defined as the distance between the tracking target and the nearest annulus border at the onset of photoactivation.
Figure 3.
The dedicated circuit model of locomotion control in C. elegans.
The forward circuit is composed of the command neuron classes AVB and PVC together with dorsal (D) and ventral (V) classes of B-type motor neurons. The reverse circuit is composed of the command neuron classes AVA and AVD together with dorsal and ventral classes of A-type motor neurons. Each class of command neuron consists of a left-right pair of monopolar neurons whose process runs the length of the animal, forming presynaptic connections en passant with the motor neurons. The number of motor neurons in each class is shown in parentheses. Motor neurons are distributed evenly along the length of the animal.
Figure 4.
Simultaneous recordings of the activity of the command neuron AVA and the locomotory behavior of freely moving animals.
A. Fluorescence image from the calcium imaging camera showing the position of AVA; the position of the interneuron RIM, which lies in a different focal plane, is indicated by the circle. B1, C1. Representative time courses of AVA emission ratio (upper trace) and velocity (lower trace) in two different animals. The colors of the emission ratio trace and background indicate the direction of locomotion (blue, forward; red, reverse). B2, C2. The x-y trajectory of the neurons recorded in B1 and C1; colors as above. D1, D2.Control for movement artifacts in AVA neurons expressing a calcium insensitive fluorescent protein (GFP). Emission ratio, velocity, and x-y trajectory are plotted as in B and C. E. Ensemble averages of emission ratio and velocity during changes in the direction of locomotion (E1, forward to reverse; E2 reverse to forward). Grey shading represents ±SEM.
Figure 5.
Simultaneous recordings of the activity of the command neuron AVB, the interneuron AIA, and the locomotory behavior of freely moving animals.
A. Fluorescence image from the calcium imaging camera showing the positions of the two neurons. B1–B3. Representative time courses of AVB (B1–3) and AIA (B1–2) emission ratios (upper traces) and velocity (lower trace) in three different animals. The colors of the emission ratio trace and background indicate the direction of locomotion (blue, forward; red, reverse). The AIA recordings serve as internal controls for movement artifacts. C1, C2. Ensemble averages of AVB emission ratio and velocity during changes in the direction of locomotion (C1, reverse to forward; C2 forward to reverse). Note that the time axis in C2 was modified to reflect the slow time course of the emission ratio. Grey shading represents ±SEM.
Figure 6.
Simultaneous recordings of the activity of the motor neurons and the locomotory behavior of freely moving animals.
A1. Fluorescence image showing the relative positions of neurons (green) and coelomocytes (red) used as proxy tracking targets. The latter were labeled with the fluorescent protein dsRed. A2. An image from the spotting camera illustrating the analysis of dorsoventral undulations. Undulations were quantified by the angle θ formed by the line segments between three points located on the outline of the animal, as shown. Positive and negative angles correspond to dorsal and ventral bends, respectively. B1, C1. Representative time courses of bending angle (upper trace) and emission ratio in the indicated motor neurons (lower traces) in two different animals. The colors of the emission ratio trace indicate the direction of locomotion (blue, forward; red, reverse); the color of the background indicates the direction of the body bend (dark gray, dorsal; light gray, ventral). B2, C2. Cross-correlations between bending angle and emission ratio in the recordings shown in B1 and c1. Trace color corresponds to motor neuron identity as indicated. D. Control for movement artifacts in motor neurons expressing a calcium insensitive fluorescent protein (GFP). Bending angle, emission ratio, and cross-correlations plotted as above.