Figure 1.
Optical discovery of cell Tr2 targets. A, A schematic of the preparation used to find cell Tr2 targets. The drawing represents the isolated leech nerve cord, consisting of the head brain (HB), 21 midbody ganglia (circles), and the tail brain (TB). Cell Tr2 was recorded intracellularly in the head brain, and the voltage-sensitive dye components were applied to a midbody ganglion, in this case ganglion 10. Ant, Anterior. B, The raw fluorescence image of ganglion 10, obtained by averaging all frames from a movie of cellular fluorescence over time. The frame includes the posterior medial packet and parts of the right and left posterior lateral packets, all of which are on the ventral side of the animal. Scale bar, 50 μm. C, Ellipses that were drawn by hand and used to average the pixels for each cell in the field. Each cell was given an arbitrary alphanumeric label unless it was impaled and definitively identified in that preparation. Cell p54(L) is likely to be cell 54(L) (p is for putative), but we did not impale it and verify its identity in this preparation. Colored cells were significantly coherent with the cell Tr2(L) electrical recording, at the α = 0.001 level (see Materials and Methods). Cells in black were not significantly coherent. Colors were chosen only to match up cells in C with traces in D and points in E; they are otherwise arbitrary. D, Simultaneous electrical recording of cell Tr2(L) and optical recordings from six of the cells shown in C. An increase in fluorescence reflects a depolarization of the cell. The optical traces are ordered by the magnitude of their coherence with the cell Tr2(L) electrical recording at 1 Hz. The coherence magnitude values are given to the right of each trace. The bars above the cell Tr2(L) voltage trace show when current was being passed into cell Tr2(L). E, Polar plot of the coherence between each optical recording and the cell Tr2 electrical recording, at the 1 Hz drive frequency, for the 43 cells circled in C. The distance from the center represents the coherence magnitude, and the angle represents the coherence phase. The outer circle represents a coherence magnitude of 1, the highest possible value. The dashed line indicates the α = 0.001 threshold for significance. The error bars represent 1 SE. A positive-negative phase angle means the signal leads or lags the cell Tr2 signal by that amount. All coherence estimates for fluorescence signals are shifted by +54°, to correct for the phase shift caused by the time constant of the dye response (see Materials and Methods). The data shown in A-E can also be presented as a movie, which is available at www.jneurosci.org. F, Analogous to C, but for a preparation in which cell 54(L) was impaled to identify it definitively. The circled cells are in the left posterior packet of the ventral side. Cells in the posterior medial packet cannot be seen clearly because they are above the focal plane. Scale bar, 50 μm. G, H, Analogous to D and E, but for the field shown in F.