Imaging, visual screens, and optical surgery are frequently applied to the nematode Caenorhabditis elegans at subcellular resolution for in vivo biological research. However, these approaches remain low-throughput and require significant manual effort. To improve throughput and enable automation in these techniques, we implement a novel cooling method to immobilize C. elegans directly on their cultivation plate. Previous studies cooled animals in microfluidics or flooded wells to 1-4 C. Counterintuitively, we find that cooling to 5-7 C immobilizes animals more effectively than lower temperatures. At 6 C, animal movement consists of bouts of submicron nose tip movement occurring at a sufficiently low magnitude and frequency to permit clear imaging. We demonstrate the ability to perform subcellular-resolution fluorescence imaging, including 64x magnification 3D image stacks and 2-min long timelapse recordings of the ASJ neuron without blurring from animal motion. We also observe no long-term side effects from cooling immobilization on animal lifespan or fecundity. We believe our cooling method enables high-throughput and high-resolution microscopy with no chemical or mechanical interventions.

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A conditioning lesion of the peripheral sensory axon triggers robust central axon regeneration in mammals. We visualize the ASJ neuron of C. elegans with a cell-specific green fluorescent protein reporter driven by a thioredoxin trx-1 promoter and trigger conditioned regeneration by laser surgery or genetic disruption of sensory pathways. Utilizing calibrated fluorescent beads, we demonstrate that these neurons brighten when conditioned, suggesting that trx-1 expression indicates regenerative capacity. We show that trx-1 functionally enhances conditioned regeneration but inhibits non-conditioned regeneration. Finally, six strains isolated in a forward genetic screen for reduced fluorescence also show reduced axon outgrowth. We demonstrate a link between trx-1 expression and the conditioned state that we leverage to rapidly assess regenerative capacity.