Fig. 3. Autonomous BCF propulsion.

(A to G) Mechano-electrical signaling of the muscular bilayer. (A) Spontaneous activation of one-side muscle induces consecutive contraction of the opposite-side muscle through mechano-electrical signaling between muscular bilayer tissues. (B) Representative time lapse images of consecutive antagonistic muscle contraction of 49-day-old biohybrid fish. (C) Midline kinematics (time step: 100 ms). (D) Correlation between spontaneous muscle activation and the moving distance. (E) Curvature of the midline during five consecutive left and right muscle strokes. (F) Empirical probability of antagonistic contraction and (G) moving speed of self-paced biohybrid fish treated with stretch-activated channel blockers, 250 μM streptomycin (n = 4 biohybrid fish) and 100 μM Gd³⁺ (n = 5 biohybrid fish) (box plot: center line, box limits, and whiskers indicates mean, SEM, and the first and third quartiles, respectively). The treatment of stretch-activated channel blockers, streptomycin and Gd³⁺ reduced the chance of antagonistic muscle contraction as well as the swimming speed of the biohybrid fish. (H to L) Geometrically insulated node (G-node). Activation pattern of biohybrid fish (H) without G-node and (I) with G-node. (J) Probability of muscle activation sites. Spontaneous muscle activation from G-node dominates spontaneous activation from the corners (n = 6 biologically independent samples without G-node and 5 samples with G-node). (K) Tail-beat frequency of biohybrid fish equipped with single-layer (n = 9 videos from nine fish), muscular bilayer (n = 20 videos from 14 fish), and muscular bilayer with G-node (n = 18 videos from five fish). Significance was determined by the Tukey-Kramer honestly significant difference test. (L) Positive relationship between pacing frequency and moving speed of autonomously swimming biohybrid fish (n = 30 videos from 19 autonomously swimming biohybrid fish and 54 videos from optogenetically swimming biohybrid fish). Data represent mean ± SEM.