Figure 2. Recordings of the CPG pattern, the motor pattern, and the beat pattern.

(A1, B1, C1) Recording sites and methods for all levels of the heartbeat control system. (A2, B2, C2) Recordings for all levels of the heartbeat control system. (A3, B3, C3) The base of the triangle represents the intersegmental phase differences (Δϕ) between the front segment and the rear segment in the two coordinations (left side). (A4, B4, C4) Circular phase plots illustrate the cycle-to-cycle variability in peristaltic coordination (left side). The vector length (value next to each vector) was used to calculate the angular variance (s² = 2(1 r)). Longer vectors indicate less variance. (A1) Suction electrodes were placed on the left and right HN(4) and HN(7) interneurons. (A2) Simultaneous extracellular recordings from these four premotor HN interneurons across a switch in coordination state (top: left heart interneurons; HN(L,4) and HN(L,7); below: right heart interneurons (HN(R,4) and HN(R,7)). Phase marker (♦) is the middle spike in each burst. Dashed lines and white triangles aid in assessing the phase differences between the two interneurons in the two coordinations. Initially, the left side is peristaltic (magenta shaded box) and the right side is synchronous (light blue shaded box). State switches midway. (A3) Colored triangles illustrate the average intersegmental Δϕ (black bars) between the ipsilateral pair of HN interneurons in the peristaltic (magenta) and synchronous (light blue) coordinations of the recording of (A2). (A4) The circular phase plot shows the cycle-to-cycle variability in peristaltic coordination for the recording of (A2). Each circle represents the Δϕ between the two ipsilateral HN interneurons of one burst cycle (n = 22 bursts, mean Δϕ = 0.19). (B1) Suction electrodes were placed on the left and right HE(8) and HN(12) motor neurons. (B2) Simultaneous extracellular recordings from these four heart motor neurons across a switch in coordination state (top, left motor neurons HE(L,8) and HE(L,12); below, right motor neurons (HE(R,8) and HE(R,12)). Phase marker (♦) is the middle spike in each burst. Labeling as in (A2). Initially, the left side is peristaltic and the right side is synchronous. State switches midway. (B3) Labels as in (A3). (B4) The circular phase plot shows the cycle-to-cycle variability in peristaltic coordination for the recording of (B2). Each triangle represents the Δϕ between the two ipsilateral heart motor neurons for one burst cycle (n = 55 bursts, mean Δϕ = 0.16). Labeling as in (A4). Phase scale bars as in (A3). (C1) Video imaging of intact animals yielded optical signals to extract the constriction/relaxation cycles for both hearts in multiple segments. (C2) The beat cycles of two bilateral pairs of heart segments are shown across a switch in coordination state (top: Heart (L,8) and Heart (L,12); below: Heart (R,8) and Heart (R,12)). Phase marker (♦) is the maximum rate of rise (MRR) during the constriction (Wenning et al., 2014). Initially, the left side is peristaltic and the right side is synchronous. State switches midway. (C3) Labels as in (A3). (C4) The circular phase plot shows the cycle-to-cycle variability in peristaltic coordination for the video recording of (C2). Each diamond represents the Δϕ between the two ipsilateral heart segments for one beat cycle (n = 18 beats, mean Δϕ = 0.25). Labeling as in (A4) Phase scale bars as in (A3). Data from the animal shown in Figure 1—video 1. Animal Groups: Bilateral Recordings (Figure 2—figure supplement 1C) and Intact Animal Database (Figure 2—figure supplement 1D).

Figure 2—figure supplement 1. Project Database and animals used in this study.

(A) The entire Project Database comprises mostly unilateral recordings collected between 2008 and 2016. Extracellular recordings from the HN(4) and HN(7) interneurons were made in 129 preparations (green circle), 17 of these were bilateral recordings (dotted green circle). Extracellular recordings from the HE(8) and HE(12) motor neurons were made in 83 preparations (blue circle), 24 of these were bilateral recordings (dotted blue circle). In 59 of these recordings (overlap, chocolate), HN interneurons and HE motor neurons were recorded simultaneously. (B) Of the 59 Simultaneous Recordings from HN interneurons and HE motor neurons (chocolate circle), nine were bilateral recordings (i.e., 8-point recordings, from the HE(R/L8), HE(R/L,12), (HN(R/L,4), HN(R/L,7); yellow circle). (C) Bilateral recordings are the focus of this study. In all, there were 26 bilateral recordings from the HN(R/L,4) and HN(R/L,7) interneurons (17 (A) + 9 (B), green circle) and 33 bilateral recordings from the HE(R/L8) and the HE(R/L,12) motor neurons (24 (A) + 9 (B), blue circle). 16 of the 26 bilateral HN interneuron recordings were also used to determine the synaptic strength of the HE(R/L8), HE(R/L,10), and HE(R/L,12) motor neurons (‘Synaptics’, dotted green circle). In 9 of the 33 bilateral HE(R/L8) and HE(R/L,12) motor neuron recordings, the HE(R/L,10) motor neurons were also recorded, in 15 of the 33 bilateral HE(R/L8) and HE(R/L,12) motor neuron recordings, the HE(R/L,14) motor neurons were also recorded (smaller blue circles). (D) Heart segments 7 to 14 were imaged bilaterally in 12 intact animals. For this study, we analyzed the intersegmental phase relations for heart segments 8 and 12 on both sides.