Peripheral multidendritic sensory neurons are necessary for rhythmic locomotion behavior in Drosophila larvae

Abstract

From breathing to walking, rhythmic movements encompass physiological processes important across the entire animal kingdom. It is thought by many that the generation of rhythmic behavior is operated by a central pattern generator (CPG) and does not require peripheral sensory input. Sensory feedback is, however, required to modify or coordinate the motor activity in response to the circumstances of actual movement. In contrast to this notion, we report here that sensory input is necessary for the generation of Drosophila larval locomotion, a form of rhythmic behavior. Blockage of all peripheral sensory inputs resulted in cessation of larval crawling. By conditionally silencing various subsets of larval peripheral sensory neurons, we identified the multiple dendritic (MD) neurons as the neurons essential for the generation of rhythmic peristaltic locomotion. By recording the locomotive motor activities, we further demonstrate that removal of MD neuron input disrupted rhythmic motor firing pattern in a way that prolonged the stereotyped segmental motor firing duration and prevented the propagation of posterior to anterior segmental motor firing. These findings reveal that MD sensory neuron input is a necessary component in the neural circuitry that generates larval locomotion.

Fig. 1.

MD sensory neurons' function is essential for larval crawling. (a) Locomotion speed of SN-Gal4/UAS-shi^ts larvae decreased with longer exposure to restrictive temperature (37°C) until they ceased crawling. n = 20 per column. (b) Percentage of SN-Gal4/UAS-shi^ts larvae immobilized after different incubation durations at restrictive temperature, reaching ≈100% in 20 min. n = 20 per column. No wild-type larvae were found to be immobile after the same incubation treatment. (c) Larval locomotion speed after inactivation of different types of sensory neurons. All experiments were done after a 10-min exposure to restrictive temperature (37°C). n = 15 per column. ∗, P < 0.001 versus wild-type. (d) Percentage of larvae immobilized after 20 min at restrictive temperature. n = 15 per column. All error bars indicate SE.

Fig. 2.

SN-Gal4 drives mCD8::GFP expression specifically in peripheral nervous system sensory neuron. (a) SN-Gal4 drives mCD8::GFP expression in all sensory neurons within one hemisegment of the larval body wall. (b) Schematized drawing of different types of sensory neurons. (c) Axonal projection of all sensory neurons (green) in larval ventral nerve cord (VNC) and brain (br) (red). Neuronal staining by anti-elav antibody was at a 1:50 dilution. (Scales bar: 100 μm.)

Fig. 3.

MD neuron function is required for the completion of locomotive peristalsis. (a) Peristalsis duration increased with the duration of exposing MD-Gal4/UAS-shi^ts larvae to 37°C. Peristalsis duration is not shown for the 20-min time point, because the larvae are immobile by then. n = 20 per symbol. (b) Peristalsis frequency decreased as the MD^Gal4/UAS-shi^ts larvae were exposed to 37°C. n = 20 per symbol.

Fig. 4.

MD sensory neuron input is crucial for the stereotyped firing duration of segmental nerves. (a) Diagram of segmental nerve and body wall muscles for an en passant nerve recording from two consecutive segments (L6 and L7). (b) Segmental nerve burst duration increased with prolonged inactivation of sensory neuron (n = 10). (c and d) Wild-type larvae retained rhythmic motor firing at 37°C in both segment L6 (c) and L7 (d). (e and f) Prolonged burst duration in MD^Gal4/UAS-shi^ts at 37°C in both segment L6 (e) and L7 (f). All error bars indicate SE. ∗, P < 0.001 versus wild type.

Fig. 5.

Blocking MD sensory neuron input prevents propagation of segmental nerve firing from L7 to L6. (a) Coordinated phase shift of segmental nerve firing in wild-type larvae after 15 min at 37°C. (b) Firing of L7, but not L6, segmental nerves in MD^Gal4/UAS-shi^ts larvae after 15 min at 37°C. (c) Hypothetical model for the role of MD neuron in generating the intersegmentally coordinated motor neuron (MN) firing for forward peristaltic locomotion. In the larval CNS, sensory inputs from L7 segment MD neurons is hypothesized to restrict the firing duration of motor neurons in the same segment (MN7) by inhibiting the CPG circuit activity and to activate motor neurons in the preceding segment (MN6) by exciting the CPG circuit activity in a process that likely involves the CPG but critically depends on MD sensory neuron activity.