Abstract
BACKGROUND: Animal studies demonstrate that anesthetic exposure during neurodevelopment can lead to persistent behavioral impairment. The changes in neuronal function underlying these effects are incompletely understood. Caenorhabditis elegans is well suited for functional imaging of postanesthetic effects on neuronal activity. This study aimed to examine such effects within the neurocircuitry underlying C. elegans locomotion. METHODS: C. elegans were exposed to 8% isoflurane for 3 h during the neurodevelopmentally critical L1 larval stage. Locomotion was assessed during early and late adulthood. Spontaneous activity was measured within the locomotion command interneuron circuitry using confocal and light-sheet microscopy of the calcium-sensitive fluorophore GCaMP6s. RESULTS: C. elegans exposed to isoflurane demonstrated attenuation in spontaneous reversal behavior, persisting throughout the animal's lifespan (reversals/min: untreated early adulthood, 1.14 ± 0.42, vs. isoflurane-exposed early adulthood, 0.83 ± 0.55; untreated late adulthood, 1.75 ± 0.64, vs. isoflurane-exposed late adulthood, 1.14 ± 0.68; P = 0.001 and 0.006, respectively; n > 50 animal tracks/condition). Likewise, isoflurane exposure altered activity dynamics in the command interneuron AVA, which mediates crawling reversals. The rate at which AVA transitions between activity states was found to be increased. These anesthetic-induced effects were more pronounced with age (off-to-on activity state transition time (s): untreated early adulthood, 2.5 ± 1.2, vs. isoflurane-exposed early adulthood, 1.9 ± 1.3; untreated late adulthood, 4.6 ± 3.0, vs. isoflurane-exposed late adulthood, 3.0 ± 2.4; P = 0.028 and 0.008, respectively; n > 35 traces acquired from more than 15 animals/condition). Comparable effects were observed throughout the command interneuron circuitry, indicating that isoflurane exposure alters transition rates between behavioral crawling states of the system overall. These effects were modulated by loss-of-function mutations within the FoxO transcription factor daf-16 and by rapamycin-mediated mechanistic Target of Rapamycin (mTOR) inhibition. CONCLUSIONS: Altered locomotive behavior and activity dynamics indicate a persistent effect on interneuron dynamics and circuit function in C. elegansafter developmental exposure to isoflurane. These effects are modulated by a loss of daf-16 or mTOR activity, consistent with a pathologic activation of stress-response pathways.