How do gap junctions of the backward circuit allow and establish a bias for forward movement? In this and the next section, we show that AVA-A coupling reduces VX-809 supplier the activity of the backward circuit through two concurrent effects, both of which are required to permit the higher forward-circuit output that drives forward motion. First, AVA-A coupling reduces AVA activity to prevent hyperactivation of backing; this is supported by the following lines of evidence. First, innexin mutants exhibit an elevated backward premotor interneuron activity via calcium

imaging analyses. In innexin mutants, the level of calcium transients in AVA and AVE was significantly higher than that of wild-type animals, whether they

were imaged as a single ROI (Figures 6A–6A″) or separately (Figures S3A–S3A″ and S3B–S3B″), suggesting that premotor interneurons of the backward circuit become hyperactivated in the absence of UNC-7 or UNC-9 innexins. Consistent with an inverse activation between forward and backward premotor interneurons (Figure 1F), the calcium level of AVB was reduced in innexin mutants (Figures 6B–6B″). The change of cameleon signals was not due to a change in the expression level of these calcium sensors in innexin mutants (Figure S3C). The reciprocal change in the premotor interneuron activity, BAY 73-4506 price specifically an increase in AVA/AVE GPX6 (backward circuit) and a decrease in AVB (forward circuit), correlates with the shift of innexin mutants’ preference in directional motion to backing. When UNC-7 expression was specifically restored in AVA in unc-7 mutants, concurrent with restored continuous forward movement and reduced backing ( Figure 5B), the calcium level in AVA/AVE was also significantly reduced ( Figures 6A–6A″). However, an expression of UNC-7 in AVA of unc-9 unc-7

mutants did not result in a rescue of forward movement ( Figure 5A), implying that the reduction of AVA/AVE activity depends on restoring AVA-A coupling. Second, AVA exhibited an increased electrical activity and increased membrane input resistance in unc-7 mutants by in situ whole-cell recordings. AVA exhibited spontaneous excitatory electric activity ( Figure 7A). The peak amplitude ( Figure 7B), but not the frequency ( Figure 7C), of such activities was significantly increased in unc-7 animals; the increased amplitude was rescued when UNC-7 expression was specifically restored in AVA ( Figures 7A–7C). Although there was no significant change in the resting membrane potential of AVA ( Figure 7D), their input membrane resistance was significantly increased in unc-7 mutants ( Figure 7E). Such an increase was also rescued when UNC-7 expression was restored in AVA ( Figure 7E). These results indicate that UNC-7-mediated AVA-A coupling functions as shunts to dampen AVA’s excitability and activity.