sensory influences on the coordination of two leg joints during searching movements of stick insects

Biol. Cybern. 64, 329-335 (1991) Biological Cybernetics �9 Springcr-Verlag 1991

Sensory influences on the coordination of two leg joints during searching movements of stick insects G. Karg, G. Breutel, and U. Biissler

Fachbereich Biologie der Universit~it, W-6750 Kaiserslautern, Federal Republic of Germany

Received July 6, 1990/Accepted in revised form September 27, 1990

Abstract. Animals (Cuniculina impigra) possessing only one foreleg with restrained coxa perform very stereo- typed searching movements during which the move- ments of the femur-tibia and coxa-trochanter joints are well coordinated. After ablation of either hairfield BF1 (measuring the position of the coxa-trochanter joint)or the apodeme of the femoral chordotonal organ (mea- suring the position of the femur-tibia joint) each joint can still be moved but the coordination changes and becomes very labile. The consequences for the ideas about the construction principles of the pattern genera- tor for searching movements are discussed.

Introduction

Theoretical considerations (Grillner 1981) and experi- ments on stick insects (Nothof and Biissler 1990) led to the following hypothesis: The pattern generators for the different kinds of rhythmic movements of a particular leg consist of the same set of constant functional ele- ments (subunits). A certain constant functional element is responsible for the movements of a single joint. The pattern generators for different kinds of movements (e.g. forward or backward walking, searching move- ments) only differ in the variable elements, which inte- grate the constant dements to a pattern generator with a well coordinated motor output. The alternative hypo- thesis (separate pattern generators exist for each of the different kinds of movements) has most likely to be rejected (Nothof and Biissler 1990).

The experimental results leading to this hypothesis came from investigations of the neural system produc- ing the so-called "active reaction" upon stimulation of the femoral chordotonal organ in stick insects. This system is able to control velocity and endpoint of active movements of the femur-tibia joint (B/issler 1986, 1988; Weiland and Koch 1987). An analog model of this system showed that it is alone able to generate rhythmic movements of the joint (B/issler and Koch 1989). There is strong evidence that this neural system participates in

the generation of the motor output for three different kinds of movement: the motor output for forward walking, the one for backward walking and the one for searching movements (Nothof and B/issler, 1990).

Assuming that this hypothesis is correct raises the question: What is the nature of the variable elements? Do they only consist of central parts (e.g. direct con- nections between the central nervous parts of the differ- ent constant functional elements via local interneurones) or are sense organs also involved? In stick insects there exists some small evidence on the participation of sense organs from the following result (Biissler and Wegner 1983): In a denervated thoracic ventral nerve cord a rather irregular rhythm of protrac- tor and retractor motor neurone activity can still be produced. But no coordination between the activity of these muscles and the muscles moving other joints could be found in this denervated preparation. This could indicate that the different constant functional elements are still working but that they are not inte- grated to a whole pattern generator when sense organs are not present.

If sense organs are integral parts of the variable functional elements, it should be possible to obtain an un- or miscoordinated motor output simply by removal of the sense organs. If this could be verified it would show that the neural elements responsible for joint movements are separate from those responsible for coordination of joint movements. This would support the hypothesis that the pattern generators for different kinds of movement of a particular leg consist of a set of constant (unspecific) functional elements, which can be integrated in different ways by different variable ele- ments each specific for a certain pattern generator.

To investigate the problem how the movements of different joints are coordinated during active move- ments, a preparation is needed which fulfills the follow- ing criteria: (i) Only two joints should move to make the analysis simpler. ~/) The movement should be very stereotyped in order to detect alterations.

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(iii) The sense organs measuring the positions of these joints should be easily accessible.

Preliminary experiments showed that the following preparation fulfills these criteria: A stick insect (Cuniculina) possessing only one foreleg is fixed on a platform in such a way that its body and the coxa of the remaining leg cannot be moved. When such an animal becomes active, only the coxa-trochanter, femur-tibia and tibia-tarsus joints of the leg can be moved. From a first inspection by eye these movements are strong and stereotyped for coxa-trochanter and femur-tibia joints. Therefore we concentrated on these two joints which have the additional advantage of being monitored by sense organs which can easily be de- stroyed.

Material and methods

The experiments were carried out on adult female Cuni- culina impigra Redtenbacher (syn. Baeulum impigrum Brunner) from the colony at Kaiserslautern University.

All legs were removed mid-coxa (to remove trochanteral campaniform sensilla) except for the right foreleg. The animals were restrained dorsal side up on a foam plastic plate. The coxa of the right foreleg was situated exactly at the edge of the plate. It was immobi- lized by dental adhesive (Scutan or Protemp, Espe), so that the angle between body and leg (seen from above) was approximately 60 ~ Care was taken that the coxa- trochanter joint was totally free to move (Fig. 1).

The animal was touched on the abdomen to acti- vate it. The searching movements were recorded by a video-camera (Sony DXC 101e) and a video-recorder (JVC HR-D 530 E6) with 25 frames/s. The camera was positioned perpendicular to the plane of movement of femur and tibia. To facilitate later evaluation white dots were placed on coxa, end of femur and end of tibia. The records were evaluated partly frame by frame and partly only by playing them back at reduced rate.

Fig. I. Position of the animal during the experiments

The trochanteral hair-plate BF1 (Wendler 1964) (syn. trHP, Pfliiger et al. 1981) was shaved off using a chip of razor blade. The success of the operation was checked after the experiments by scanning electron microscopy.

In order to remove the influence of the femoral chordotonal organ, its receptor apodeme was cut after a small opening had been made in the distal third of the femur. The success of the operation was checked after the experiments by a careful dissection of the leg.

Results

Searching movements were recorded as complete cycles, i.e. down and up movement. A total of 497 cycles were recorded from 6 intact legs, while 400 cycles were recorded from 6 legs with hairfield BFI removed and 402 cycles from 6 legs with cut receptor apodeme of the femoral chordonal organ. The time between operation and beginning of experiments was at least 3 h. These 1299 searching cycles can be subdivided into 8 groups;

I. Searching movements normally performed by in- tact legs: A downward movement starts with a fast depression of the coxa-trochanter joint. The femur-tibia joint is fully extended and does not move at first or is even extended (finishing its extension movement). The coxa-trochanter joint movement slows down whereas the femur-tibia joint flexion starts slowly and becomes faster towards the end of downward movement. Figure 2a illustrates this behaviour. The kind of coordination becomes more obvious when the femur-tibia angle (an- gle between the longitudinal axes of femur and tibia) is plotted against the coxa-trochanter angle (angle be- tween the longitudinal axes of coxa and trochantero-fe- mur) (Fig. 2b). As the coxa cannot be seen because it was embedded in Protemp, the coxa-trochanter angle zero was set at the horizontal position of the femur.

The upward movement (Fig. 3) starts with a rapid extension of the femur-tibia joint normally accompa- nied by a slower levation of the coxa-trochanter joint.

The amplitudes of the movements are large (coxa- trochanter joint between 30 ~ and 60 ~ femur-tibia joint between 55 ~ and 100~ In the most elevated position the coxa-trochanter joint is far away from its anatomi- cally possible extreme position, but the femur-tibia joint is at or very close to full extension (femur-tibia angle 160~176 In the most depressed position both joints do not reach their anatomically possible extreme posi- tions.

In intact animals, this type of movement was per- formed in a very stereotyped manner. This is illustrated when several randomly selected movements of different animals are plottted together (Fig. 4). The values for downward movements are grouped within a very nar- row range, whereas the values for upward movements show a wider distribution.

2. Both joints move in the same coordination as in (1), i.e. the femur-tibia joint is flexed during a depres- sion of the coxa-trochanter joint and vice versa, but the amplitude of movement for both joints is markedly

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smaller than in (1). The coordination was not so stereo- typed as for (I) in intact legs. This type was rare in intact as well as in most operated legs (Fig. 9).

3. The femur-tibia joint remains in an extended position (femur-tibia angle 160~ ~ ) and only the coxa-trochanter joint moves up and down (Fig. 5). This type was especially frequent for legs with ablated hair- plate BF1.

4. Like (3), but the femur-tibia angle is smaller and changes only slightly or not at all (Fig. 6). This type was rather frequent in legs with cut receptor apodemes.

5. The coxa-trochanter joint remains immovable in its upper extreme position. The femur-tibia joint flexes and extends, in most cases with a rather high frequency (Fig. 7). This type was also rather frequent in legs with cut receptor apodemes.

6. The coxa-trochanter joint does not move, but is not fully elevated (sometimes fully depressed). Only the

femur-tibia joint moves. This type was rare under all circumstances.

7. There is a simultaneous coxa-trochanter depres- sion and femur-tibia extension (and vice versa). Figure 8 shows an example.

8. Although at least one joint moves, it is not possible to classify the movement under any of the types 1-7.

Figure 9 shows the frequency of occurrence of the different types of movements. Large differences between the animals are obvious except for intact legs. In legs with hairfield BF1 removed (BF1 measures position of coxa-trochanter joint), movements predominated dur- ing which only the coxa-trochanter joint moved and the femur-tibia joint remained still in its extended position. Cutting the receptor apodeme of the femoral chordo- tonal organ (which measures the position of the femur- tiba joint) produced more variable results. There was a

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certain tendency towards movements of only a single joint, either only the coxa-trochanter joint (and the femur-tibia joint remaining in a middle position) or the femur-tibia joint (and the femur maximally elevated). Except for animal 5 in the series with BFI removed, each of the operated animals was able to rhythmically move either only the coxa-trochanter or only the femur- tibia joint and also both joints simultaneously. When both joints were moved they moved with the same rhythm; differing movcmcnt frequencies of the two joints had never been observed.

Discussion

Construction principles of the pattern generator for searching movements

The results show that the movements produced by the intact foreleg with restrained coxa become very vari-

able when one of the sense organs which measure the position of the coxa-trochanter or the femur-tibia joint does not function. Both sense organs arc not the only ones for their respective joints. The position of the coxa-trochanter joint is also measured by a ventral hairfield (Tatar 1976, cited according to B/isslcr 1983) and two strand receptors with central cell bodies (Br[iu- nig, 1982 for Extatosoma; they also exist in Cuniculina, T. Hofmann, unpublished results). The position of the femur-tibia joint is also measured by six multipolar sense cells (B/issler 1977). When either hairfield BF1 or the femoral chordotonal organ no longer functioned a small percentage of searching movements were still performed like in intact legs and in all cases the rhythm was the same when both joints moved. This shows that parts of the neural system responsible for the coordination of the two joints are still functioning. The results do not allow to decide whether this is due to the remaining sense organs in the appropriate joint or to central connections.

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After ablation of one of the main sense organs both leg joints were still able to move rhythmically. Appar- ently, these sense organs are not necessary for the generation of rhythmic movements neither of the "own" joint nor of the adjacent joint. Whether these single-joint-movements are generated mainly centrally or mainly under the influence of the remaining sense organs, cannot be distinguished at the moment (but see below for the femur-tibia joint). The two main sense organs (BF1 and chordotonal organ) are apparently only necessary for the stereotyped coordination be- tween the movements of the two joints.

There are two possible explanations for this fact: (1) After the operation the animal no longer intended to perform searching movements. Instead it used indepen- dent neuronal "programmes" with a different kind of coordination between the joints. (2)The animal still "tries" to perform normal searching movements and the disturbed coordination is a result of a miscoordi- nated searching "programme",

If assumption (2) is correct, the results show that the systems generating the single-joint movements (which are not disturbed by the operation) and the system coordinating these movements (which is dis- turbed by the operation) are not identical. In other words, the neural system generating the searching movements in the intact leg with restrained coxa con- sists of elements which are able to generate the rhyth- mic movements of a single joint, and of elements which are responsible for the coordination. It is not a compact system which generates the searching motor output for all joints as a whole (Nothof and B/issler 1990) and can therefore not be divided up into movement-generating and coordinating elements. This conclusion is consis- tent with the hypothesis of Grillner (1981) and Nothof and B/issler (1990) that pattern generators for rhythmic leg movements consists of distinguishable functional elements. Some functional elements are responsible for the movements of a single joint. They are used for all kinds of movements (constant elements) and are not specific for a certain pattern generator. Others are responsible for the coordination of the constant ele- ments and differ from movement type to movement type (variable elements).

The ablation experiments show, that the variable elements, which coordinate the coxa-trochanter and the femur-tibia joints during searching movements under the conditions used, possess sense organs as integral parts. From our experiments it cannot be decided whether these variable elements are simply coordinating pathways between the constant functional elements (Nothof and B/issler 1990) or part of a superior struc- ture or programme which conducts the orchestra formed by the constant elements (comparable to the orchestration hypothesis of Sombati and Hoyle 1984).

If the operated legs no longer intended to perform normal searching movements (assumption 1) and in- stead used different neuronal "programmes", the "pro- gramme" in use must depend upon sensory feedback. If such a "programme" is build up of moduls (constant functional elements) this idea cannot be distinguished from the above orchestration idea. If there are separate "programmes" for each of the different movement types which share no common moduls, it is hard to imagine that so many different pattern generating sys- tems exist in one half-ganglion. In conclusion, the data fit the idea of Grillner (1981) and Nothof and B/issler (1990).

The high degree of necessary feedback is perhaps a peculiarity of stick insects. In many other cases stick insect motor programmes also depended to a much higher degree on sensory feedback than the motor programmes are generally thought to do in other ani-

mals (summary: B/issler 1983, 1988). Therefore the stick insect might be used as a model system for other animals as well, as far as the modular construction principles of its pattern generators are concerned. But perhaps it is no model for other animals, as far as the high degree of necessary feedback is concerned. Sense organs are much easier accessible than central nervous structures. Therefore construction principles of pattern generators are easier to investigate in this animal than in more "centralized" animals.

The role o f hairfieid B F I and chordotonal organ

Removal of the hairfield BF1 produced in most cases movements of only the coxa-trochanter joint whereas the femur-tibia joint remained fully extended. Appar- ently, either the signal f rom the intact BF1 induces the femur-tibia joint to flex or destruction of BF1 induces signals which inhibit the movement of the femur-tibia joint. The first interpretation is consistent with results on intact legs: here a downward movement starts with a depression of the coxa-trochanter joint and it takes some time until the femur-tibia joint is flexed (Figs. 2 and 4a). But the influences of BF1 on femur-tibia movement cannot be very strong, because in legs with cut receptor apodemes the movement of the coxa- trochanter joint alone is in many cases not able to induce femur-tibia movements (type 4). This might be interpreted that in the intact leg the signal from BF1 only triggers a mechanism, which includes the femoral chordotonal organ, and this mechanism is able to am- plify the trigger signal. A mechanism with these proper- ties has previously been described for the femur-tibia joint. It is the neural system producing the active reac- tion under open-loop conditions. I t accelerates any slow flexion movement of the femur-tibia joint and deter- mines the endpoint o f a flexion movement (B/issler 1986, 1988; Weiland and Koch 1987). It would be able to amplify a small signal coming f rom BF1 under natural conditions. I f it is the main cause for the strong flexion of the femur-tibia joint, the flexion should start slowly and should then be accelerated more and more. Exactly this behaviour was found in reality. This is the second indication, that this system is part of the pattern generator for searching movements (a stronger one came from N o t h o f and B/issler 1990).

Although the system producing the active reaction (which includes the femoral chordotonal organ) is able to produce movements of the femur-tibia joint, it is apparently not necessary for these movements, because they can be performed by legs with cut receptor apodeme when the coxa-trochanter joint is maximally elevated (movement type 5).

The results agree with the following idea: In each of the two joints there is a separate functional element (constant functional element) which is alone able to generate rhythmic movements. The element of the coxa- trochanter joint generates these movements in the active animal all the time. The hairfield BF1 possibly con- tributes to its function but it is not a necessary compo- nent. The functional element of the femur-tibia joint

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consists of the neural system producing the active reac- tion but has either also other components or the central nervous parts of it are alone able to generate the motor output. In most cases this element is overriden by a constant input which causes the joint to extend in the active animal. This constant input can be counterbal- anced by a signal f rom BF1. I t produces a slight flexion which is now accelerated by the system producing the active reaction. The flexion movement stops under the influence of this system when a certain position is reached (B/issler 1988) and an extension movement is started. The constant input can also be counterbalanced by signals from the trochanteral campaniform sensilla (they are strongly excited in the most elevated and the most depressed position of the coxa-trochanter j o i n t - F. Delcomyn, personal communication) which also allows rhythmic movements of the femur-tibia joint. They are only in part generated under the influence of the system producing the active reaction (see movement types 5 and 6 in legs with cut receptor apodeme).

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Prof. Dr. U. B/issler Fachbereich Biologic der Universit/it Postfach 3049 W-6750 Kalserslautern Federal Republic of Germany