Compartmentalized calcium dynamics in a C. elegans interneuron encode head movement

Abstract

The confinement of neuronal activity to specific subcellular regions is a mechanism for expanding the computational properties of neurons. Although the circuit organization underlying compartmentalized activity has been studied in several systems, its cellular basis is still unknown. Here we characterize compartmentalized activity in Caenorhabditis elegans RIA interneurons, which have multiple reciprocal connections to head motor neurons and receive input from sensory pathways. We show that RIA spatially encodes head movement on a subcellular scale through axonal compartmentalization. This subcellular axonal activity is dependent on acetylcholine release from head motor neurons and is simultaneously present and additive with glutamate-dependent globally synchronized activity evoked by sensory inputs. Postsynaptically, the muscarinic acetylcholine receptor GAR-3 acts in RIA to compartmentalize axonal activity through the mobilization of intracellular calcium stores. The compartmentalized activity functions independently of the synchronized activity to modulate locomotory behaviour.

Figure 1. Compartmentalized calcium signals in RIA encode head movement

a, Highly simplified RIA and SMD synaptic connections. b, RIA anatomy showing nrD, nrV and loop subdomains. c, Single frames of GCaMP3 fluorescence recording in RIA. Dashed ovals denote subdomains in b. Anterior, left; D, dorsal; V, ventral. d, Calcium dynamics in RIA axonal domains and corresponding head bending. Open arrowheads, time points in c; filled arrowhead, a synchronous calcium event. e, Cross-correlations between nrV or nrD Ca²⁺ responses and head movement. Gray lines, individual animals (n = 53); solid lines, mean values.

Figure 2. RIA compartmentalized activity requires transmission from SMD motor neurons

a, Topography of SMD synapses onto RIA and muscles. b, Overlay of representative SMD calcium recording and movement. The SMDD trace is inverted to show correlation with dorsal head bending. c, Cross-correlations of SMDV/D with head movement (n = 8). d, Representative calcium recordings of RIA subdomains or SMDs in levamisole-treated animals. e, RIA correlations with head movement in glr-1::TeTx or lad-2::TeTx transgenic strains (glr-1 n = 18, lad-2 n = 17, control n = 28). f, Internal cross-correlations in SMD::TeTx strains. g, Comparison of peak correlations between strains. Cross-correlation plots and bar charts are mean +/− s.e.m.. ***p < 0.001, **p < 0.01, ANOVA corrected for multiple comparisons.

Figure 3. A muscarinic acetylcholine pathway establishes motor-correlated RIA dynamics

a, Sample calcium dynamics of nrV, nrD, and SMDs. b, Cross-correlations between RIA dynamics and head movement and internal cross-correlations in cha-1(p1152) (n = 18, control n = 25) and scopolamine-treated wild type animals (100 μM, n = 10). c, Cross-correlation plots as in b for mutants: gar-1(ok755) (n = 5), gar-3(gk305) (n = 16), gar-3(vu78) (n = 6), egl-8(md1971) (n = 7). d, Cross-correlations of transgenic (n = 13) and nontransgenic (n = 9) siblings in a strain expressing a gar-3 cDNA in RIA in gar-3(gk305) mutants. e, f, Peak correlation comparisons for data in b-d. g, Images of wild type and gar-3(gk305) animals every 5 seconds over 50 seconds, beneath each image series is the projection of each animal’s midline every 0.5 seconds over the same time period. h, Comparison of mean postural aspect ratios during forward locomotion for wild type (n = 17), gar-3(gk305) (n = 10) and transgenic (n = 10) and non-transgenic (n = 15) siblings in the RIA::gar-3 rescue line. Cross-correlation plots and bar charts are mean +/− s.e.m. ***p < 0.001, **p < 0.01, *p < 0.05, ANOVA corrected for multiple comparisons.

Figure 4. Sensory-evoked synchronized RIA activity is dependent on glutamatergic transmission and is additive with compartmentalized dynamics

a, , Raster plot and histogram of synchronous calcium influx (blue) or efflux (red). Dots, synchronous events; histogram, mean rate of Ca²⁺ flux (n = 53). b, c, Mean responses across two trials in wild type (n = 19) and eat-4(ky5) mutants (n = 18). ***p < 0.001, significant interaction between stimulus response and genotype, MANOVA with repeated measures. Error bars, s.e.m. d. Example of nrV and nrD responses to switching between IAA and buffer every 2 seconds. e, Spectral density of nrV and nrD activity and cross-correlations (mean +/− s.e.m.) between nrV or nrD and head movement before and after synchronous signal subtraction (see text).