Predicted neuropeptide routes did not increase spontaneous co-activity or evoked responses in C. elegans after wiring and neuron controls; a small negative co-activity link was not matching-specific

Question: does the predicted neuropeptide connectome of C. elegans (Ripoll-Sanchez et al. 2023; 92 peptide-receptor pairs, EC50 <= 500 nM) predict neuronal activity beyond the wired connectome? We wrote the plans before each test. Analysis A used spontaneous whole-brain calcium activity of 40 freely moving animals (Atanas, Kim et al. 2023): absolute mean co-activity of 7,872 neuron pairs, with per-neuron fixed effects, gap junctions, chemical synapses, shared input and output, neurite contact, soma distance, ganglion and homology as controls. Analysis B used the optogenetic signal-propagation atlas (Randi et al. 2023): functional connections (q < 0.05) in 23,383 directed wild-type pairs and in unc-31 mutants, which cannot release dense-core vesicles. Inference used neuron-label permutation (QAP) and a new ligand-receptor relabel null that keeps the expression sets but gives them to random receptor genes. Results: in the full model, pairs with more peptide routes were less co-active (partial r = -0.074, QAP p = 0.0005, 39 of 40 animals negative). The pre-specified positive hypothesis therefore failed. The negative link was not specific to the real ligand-receptor matching (receptor-relabel two-sided p = 0.15). The predicted larger effect for slow than for fast activity was not found. In the evoked atlas, peptide routes did not predict functional connections (partial r = 0.003, QAP p = 0.69), while gap junctions did in the same model (r = 0.050, z = 6.8). The unc-31 mutants did not lose a peptide effect (WT 0.009, unc-31 0.021; one-sided p = 0.76). The monoamine network gave a small negative co-activity link (r = -0.031). Limitations: connectome models, expression data and activity come from different animals; expression is binary; short-, mid- and long-range models are identical in the nerve-ring subset; one behavioural condition; slow or global peptide effects can be removed by the per-neuron controls.

C. elegans · neuropeptide connectome · extrasynaptic signalling · functional connectivity · whole-brain imaging · signal propagation atlas · unc-31 · structure-function · null result · QAP permutation · ligand-receptor specificity · gap junctions