Short Communications Initiation of Flight By an Identified Wind Sensitive Neurone (Tcg) in The Locust

When the tarsi are not in contact with the ground, brief wind stimuli applied to the wind sensitive hairs on the head elicit flight in the locust (Weis-Fogh, 1949; Camhi, 1969). The descending interneurones mediating this reaction have not been identified but some likely candidates have been reported (Bacon & Tyrer, 1978; Simmons, 1980, 1981). The most extensively studied is the tritocerebral commissure giant (TCG) (Bacon & Tyrer, 1978; Bacon & Möhl, 1979). This large wind sensitive interneurone descends from the brain, crosses to the contralateral side of the nerve cord via the tritocerebral commissure, and makes excitatory connections with some flight motor neurones in the thoracic ganglia (Bacon & Tyrer, 1979). The location of the TCG’s axon in the tritocerebral commissure makes this neurone easily accessible for recording and stimulation in almost intact, freely moving animals. Recordings during flight, for example, have shown that the TCG discharges one or two spikes during each cycle (Bacon & Möhl, 1979). The spike activity results from rhythmic air turbulences caused by flapping of the wings and head movements during flight. This rhythmic activity may play a role in regulating motor output during flight manoeuvres, such as yaw (B. Möhl & J. Bacon, in preparation). It seems unlikely however that this is the only function for the TCG since it is capable of discharging in very high frequency bursts quite unlike those occurring in flying animals. Here we present evidence that the TCG also functions to facilitate the initiation of flight following a jump.

Experiments were performed on female and male Locusta migratoria obtained from a long established colony at the University of Alberta. In animals where the wings had been amputated, the mouthparts at the ventral side of the head were removed and the tritocerebral commissure exposed. A fine steel needle was inserted under the posterior part of the tritocerebral commissure (Fig. 1 A) and waxed to the cuticle on both sides of the head. This needle served as an electrode to record the activity of the TCG and the arrangement was stable enough to allow recording from the TCG during the jump without movement artifact. A ground electrode was inserted into the head through a hole in the cuticle near the recording electrode. After the electrode implantation, excess haemolymph in the head was removed and replaced with vaseline. The opening in the head capsule was then covered with Parafilm, which was sealed to the cuticle with wax.

The locust jump is characterized by an initial phase of co-contraction of hindleg flexor and extensor tibiae muscles followed by an inhibition of flexor muscle activity to release the jump (Heitler & Burrows, 1977). The inhibition of flexor muscle activity thus provided a convenient marker for the onset of a jump. In order to record flexor muscle activity, a pair of 50 μm copper wires, insulated except at the tip, were inserted into the femur of one hindleg. All wires leading to recording electrodes were glued to a thread by which the animal was tethered. Tethered animals could move freely in an arena of 50 cm diameter and jump up to heights of 70 cm. Immediately following the onset of the jump (indicated by the cessation of activity in the flexor myogram) the TCG discharged with a high frequency burst lasting at least 50 ms with a frequency of more than 200 Hz (Fig. 1B). Similar results were obtained in nine preparations. During vigorous jumps in thesç preparations burst durations ranged from 50 to 120 ms and the discharge rate ranged from 100 to 360 Hz. This activity is similar to that observed for wind stimulation in fixed animals. Stimulation of fixed animals with a wind velocity of 3 m/s causes a TCG discharge of approximately 200 Hz. The take off velocity at the beginning of the jump was measured in high speed cinematographic studies as 3 m/s, which is in accordance with results of Alexander & Bennet-Clark (1977).

The high frequency burst in the TCG during a jump suggests it might function to facilitate the initiation of flight. This was investigated by studying the effects of electrical stimulation of TCG with a pattern similar to that occurring during a jump. Following removal of the wings and amputation of all six legs, animals were pinned ventral side up on a cork board. The pins were placed through the cuticle of the first proracic segment. The animals were raised slightly from the cork board to allow unrestricted movements of the wing stumps. Flight activity was monitored by EMG recordings from the forewing first basalar muscle. The EMG electrodes (100/xm copper wire, insulated except for the tip) were inserted through the ventral cuticle at the site of muscle attachment. The tritocerebral commissure and the suboesophageal ganglion were exposed by removing the mouthparts. The TCG was stimulated by a monopolar silver wire electrode (75 μm) placed under the posterior commissure with the indifferent placed into the haemolymph. Recruitment of the TCG with increasing stimulus strength was monitored by recording from the ipsilateral connective between the suboesophageal and prothoracic ganglia (Fig. 2A). Wind stimuli delivered to the head elicited spikes in the TCG which could be recorded at the tritocerebral commissure. The lower trace of Fig. 2B shows the corresponding descending TCG unit recorded posterior to the suboesophageal ganglion when the sweep is triggered by the TCG spike in the commissure. When we stimulated the tritocerebral commissure with current pulses of 0·1 ms duration the threshold for activation of the TCG unit (Fig. 2C) was 0·44 V. This threshold varied only ± 0·04 V from preparation to preparation. We confirmed this threshold value in experiments where we recorded intracellularly from the TCG in a thoracic ganglion while stimulating the TCG extracellularly at the commissure.

High frequency (200 Hz) stimulation of a posterior commissure at threshold intensity for TCG activation reliably induced flight in 11 out of’ 17 animals. The six unresponsive animals all flew in response to TCG stimulation if they were first aroused by a wind stimulus to the head. Thus flight was evoked in all experimental animals by stimulation of one TCG. Only short duration trains of TCG activity (40–200 ms) were necessary for initiating flight. In the example shown in Fig. 2D a train of 50 ms duration elicited about 80 flight cycles which far outlasted the period of stimulation. According to Bacon & Tyrer (1978) the distal part of the tritocerebral commissure contains only two axons: the axon of the TCG and the axon of a far smaller neurone. We doubt that our results are due to activation of the smaller neurone since our stimulus was delivered just at threshold and we never observed a second unit in the recording from the connective. It is also unlikely our results are due to activation of other neurones either by current spread to the adjacent anterior commissure or by inputs to the brain via the two axons in the anterior commissure. Stimulation of one posterior commissure was still sufficient to induce flight when both connectives were either cut or crushed between the points of fusion of the anterior and posterior tritocerebral commissures, and when the other posterior commissure was also cut.

To test the extent to which the TCG might normally contribute to initiation of flight we investigated the response to wind stimulation to the head following transsection of both posterior commissures through which the axons of the TCGs pass. The onset of flight following the start of a wind stimulus was monitored by an EMG recorded from flight muscles with the EMG electrodes positioned to record both elevator and depressor activity (Fig. 3 insets). The wind velocity was 3 m/s and the stimulus onset was controlled by an electromagnetically activated valve. Cutting both posterior commissures significantly increased the latency of the initial elevator burst (Fig. 3), but had little effect on the latency of initial depressor activity. The average latency increase in elevator activity (34 trials in 7 animals) was 30 ms. This increase m latency to elevator activity presumably resulted in a longer latency to wing opening, since wing opening at the start of flight is produced by a burst of activity in elevator motoneurones (Pond, 1972). In this experiment we cut the axons of only four neurones, two of which were TCGs. We have no evidence that wind stimulation on the head activates the other two neurones while the TCGs are discharging. Therefore we suggest that it is the loss of the TCGs which caused the delayed activation of the elevator motoneurones. It should be noted that transection of the TCGs never prevented flight initiation. Therefore additional pathways must be involved in initiating flight in response to wind on the head.

Taken together the results of these experiments suggest that the TCGs facilitate the initiation of flight following a jump. The main results supporting this proposal are: (1) the TCGs discharged in a high frequency burst when an animal jumped, (2) electrical stimulation with stimulus parameters mimicking TCG activity during a jump induced flight in aroused animals, and (3) cutting the posterior tritocerebral commissures resulted in an increase of the latency for the initial elevator muscle burst in response to wind stimulation of the head. If the TCG does facilitate the initiation of flight then the functional organization of descending wind sensitive interneurones resembles the organization of interneurones in the escape systems of the goldfish and crayfish. In these animals activity in single large identified interneurones can initiate locomotion (Eaton, Lavender & Wieland, 1982; Reichert & Wine, 1982) yet parallel neural pathways are able to mediate this behaviour when the large interneurones are either deleted or inactive.

We thank R. Franklin, D. N. ReyeandR. M. Robertson for their helpful comments on the manuscript. This work was supported by a grant from the Canadian Medical Research Council.