The structure and function of serially homologous leg motor neurons in the locust. I. Anatomy

Twenty-one prothoracic and 17 mesothoracic motor neurons innervating leg muscles have been identified physiologically and subsequently injected with dye from a microelectrode. A tract containing the primary neurites of motor neurons innervating the retractor unquis, levator and depressor tarsus, flexor tibiae, and reductor femora is described. All motor neurons studied have regions in which their dendritic branches overlap with those of other leg motor neurons. Identified, serially homologous motor neurons in the three thoracic ganglia were found to have: (1) cell bodies at similar locations and morphologically similar primary neurites (e.g., flexor tibiae motor neurons), (2) cell bodies at different locations in each ganglion and morphologically different primary neurites in each ganglion (e.g., fast retractor unguis motor neurons), or (3) cell bodies at similar locations and morphologically similar primary neurites but with a functional switch in one ganglion relative to the function of the neurons in the other two ganglia. As an example of the latter, the morphology of the metathoracic slow extensor tibiae (SETi) motor neurons was similar to that of pro- and mesothoracic fast extensor tibiae (FETi) motor neurons. Similarly the metathoracic FETi bears a striking resemblance to the pro- and the mesothoracic SETi. It is proposed that in the metathoracic ganglion the two extensor tibiae motor neurons have switched functions while retaining similar morphologies relative to the structure and function of their pro- and mesothoracic serial homologues.     

Proprioceptive feedback in locust kicking and jumping during maturation

Campaniform sensilla monitor the forces generated by the leg muscles during the co-contraction phase of locust (Schistocerca gregaria) kicking and jumping and re-excite the fast extensor (FETi) and flexor tibiae motor neurones, which innervate the leg muscles. Sensory signals from a campaniform sensillum on the proximal tibia were compared in newly moulted locusts, which do not kick and jump, and mature locusts which readily kick and jump. The activity pattern of FETi during co-contraction was mimicked by stimulating the extensor tibiae muscle. Less force was generated and the spike frequency of the sensory neurone from the sensillum was significantly lower in newly moulted compared to mature locusts. Depolarisation of both FETi and flexor motor neurones as a result of sensory feedback was consequently less in newly moulted than in mature locusts. The difference in the depolarisation was greater than the decrease in the afferent spike frequency suggesting that the central connections of the afferents are modulated. The depolarisation could generate spikes in FETi and maintain flexor spikes in mature but not in newly moulted locusts. This indicates that feedback from the anterior campaniform sensillum comprises a significant component of the drive to both FETi and flexor activity during co-contraction in mature animals and that the changes in this feedback contribute to the developmental change in behaviour.Abbreviations aCS anterior campaniform sensillum - ETi extensor tibiae - FETi fast extensor tibiae motor neurone - FlTi flexor tibiae - pCS posterior campaniform sensillum     

Assisting components within a resistance reflex of the stick insect, Cuniculina impigra

ABSTRACT. Rapid relaxation (shortening) of the femoral chordotonal organ in Cuniculina impigra Redtenbacher induces a depolarization followed by hyperpolarization of the fast and slow extensor tibiae motor neurons (FETi and SETi). The initial depolarization is caused by acceleration-sensitive units of the chordotonal organ. The reverse sequence of responses is induced in flexor motor neurons. The common inhibitor neuron (CI) is depolarized by both lengthening (stretch) and relaxation of the chordotonal organ.
The initial depolarization of FETi and SETi and the initial hyperpolarization of flexor motor neurons produced by rapid relaxation of the chordotonal organ and the depolarization of CI produced by lengthening of the chordotonal organ all oppose the resistance reflex response. However, these assisting components are weak compared to the resisting ones.     

Excitatory interactions between antagonistic motor neurones underlying locust kicking and jumping during maturation after the adult moult

There is a change in the synaptic connections between motor neurones that underlie locust kicking and jumping during maturation following the adult moult. The fast extensor tibiae (FETi) motor neurone makes monosynaptic excitatory connections with flexor tibiae motor neurones that have previously been implicated in maintaining flexor activity during the co-contraction phase of jumping, in which energy generated by the muscles of a hind leg is stored. The amplitude of the FETi spike decreases when repetitively activated, and this decrement is larger in locusts immediately following the adult moult than in mature locusts. The decrement in␣the FETi spike is correlated with a greater decrease in the amplitude of the flexor excitatory postsynaptic potential (EPSP) in newly moulted locusts and in turn with the failure of these locusts to kick or jump. The results presented here indicate that the developmental change in the connections between the motor neurones contributes to the change in behaviour following the moult. Accepted: 28 April 1997     

Neuromuscular plasticity in the locust after permanent removal of an excitatory motoneuron of the extensor tibiae muscle

The capacity of the larval insect nervous system to compensate for the permanent loss of one of the two excitatory motoneurons innervating a leg muscle was investigated in the locust (Locusta migratoria). In the fourth instar, the fast extensor tibiae (FETi) motoneuron in the mesothoracic ganglion was permanently removed by photoinactivation with a helium-cadmium laser. Subsequently, the animals were allowed to develop into adulthood. When experimental animals were tested as adults after final ecdysis, fast-contracting fibers in the most proximal region of the corresponding extensor muscle, which are normally predominantly innervated by FETi only, uniformly responded to activity of the slow extensor tibiae (SETi) neuron. In adult operated animals, single pulses to SETi elicited large junctional responses in the fibers which resulted in twitch contractions of these fibers similar to the responses to FETi activity in control animals. The total number of muscle fibers, their properties as histochemically determined contractional types (fast and slow), and their distribution were not affected by photoinactivation of FETi. Possible mechanisms enabling the larval neuromuscular system to compensate for the loss of FETi through functionally similar innervation by a different motoneuron, i.e. SETi, are discussed.     

Embryonic development of the innervation of the locust extensor tibiae muscle by identified neurons: formation and elimination of inappropriate axon branches

Intracellular dye fills have been used to reveal the pattern of embryonic growth of each of the four neurons which innervate the extensor tibiae muscle (ETi) of the hind leg of the locust. The growth cone of the slow extensor tibiae motoneuron (SETi), the first of the four neurons to leave the central nervous system, pioneers nerve 3 (N3). The fast extensor motoneuron (FETi), the next neuron to grow out, follows earlier outgrowing motoneurons into the periphery in nerve 5 (N5) and then rejoins SETi in N3. As it transfers from N5 to N3, it is transiently dye-coupled to the Tr1 pioneer neuron which spans the gap between the two nerves. It then follows SETi onto the ETi muscle in the femur. The common inhibitory neuron and the dorsal unpaired median neuron (DUMETi) follow SETi and FETi in nerves 3B2 and 5B1, respectively. SETi's growth cone requires almost twice as long to reach ETi as those of the three later motoneurons, all of which follow preexisting neural pathways. At least three of the four developing motoneurons form one or more axon branches not found in the adult. These branches may occur (1) at segmental boundaries; (2) where the nerve, which the growth cone is following, itself branches or the growth cone encounters another nerve; or (3) when the axon continues to grow beyond its target muscle. These findings contrast with the apparent absence of inappropriate axon branches in another developing locust neuromuscular system and during the innervation of zebrafish myotomes, but resemble in some ways the transient production of inappropriate axonal branches reported for embryonic leech motoneurons.     

Axon growth from limb motorneurons in the locust embryo: the effect of target limb removal on the pattern of axon branching in the periphery

Metathoracic limb buds have been unilaterally ablated from locust embryos at 25 to 30% of embryonic development and the effect of this operation on the axon morphology of the motorneuron fast extensor tibiae (FETi) observed at later embryonic stages. In control embryos this neuron sends a single axon out the main leg nerve, nerve 5, to the extensor tibiae muscle in the femur. In limb ablated embryos the axon of FETi is found in a wide variety of aberrant peripheral nerve pathways and projects to a wide range of foreign muscles. There is a degree of apparent selectivity, but no rigid hierarchy, in the choice of pathway and muscle made by FETi. A high degree of variability is found between one embryo and another in the extent and pattern of axon branching. The axon of FETi is generally found in pathways that correspond to nerves in control embryos but on occasion grows along novel routes. An anteriorly directed dendritic branch, seldom seen in control FETi neurons, is frequently seen in experimental FETis. These findings are discussed in terms of the rules for specific axon growth in normal development.     

Effects of temperan a central synapse between identified motor neurons in the locust

Summary Changing the temperature from 10–40 °C modifies the transmission at an established monosynaptic connection between the fast extensor tibiae (FETi) and flexor tibiae motor neurons in the metathoracic ganglion of the locustSchistocerca gregaria (Forskål). Striking changes occur to the shape of the spikes, to membrane resistance, to the synaptic delay, and to the evoked synaptic potentials.In the presynaptic FETi motor neuron, raising the temperature reduces the amplitude of an antidromic spike recorded in the soma by a factor of 10 (40 mV to 4 mV), reduces the time taken to reach peak amplitude by 5 (3.5 to 0.7 ms) and decreases the duration at half maximum amplitude by 0.5. The conduction velocity of the spike in the axon is increased by 50% from 10 °C to 40 °C. Orthodromic spikes are affected by temperature in a similar way to the antidromic spikes.The membrane resistance of both pre- and postsynaptic motor neurons falls as the temperature is raised. The membrane resistance of FETi falls by a factor of 4 (about 4 M at 10 °C to 1 M at 40 °C). A contributory component to this fall could be the increase in the frequency of synaptic potentials generated as a result of inputs from other neurons. No temperature dependence could be demonstrated on the voltage threshold relative to resting potential for evoking orthodromic spikes, but because the resistance changes, the current needed to achieve this voltage must be increased at higher temperatures.The latency measured from the peak of the spike in the soma of FETi to the start of the EPSP in the soma of a flexor motor neuron decreases by a factor of 20 (10 ms at 10 °C to 0.5 ms at 40 °C).In a postsynaptic flexor tibiae motor neuron, the amplitude of the evoked synaptic potential increases by a factor of 3.4 (5 mV to 17 mV), its duration at half maximum amplitude decreases by 3 (7 ms at 12 °C to 2.3 ms at 32 °C) and its rate of rise increases by 3. An increased likelihood that spikes will occur in the flexor contributes to the enhanced amplitude of the compound EPSP at temperatures above 20 °C.Abbreviation FETi fast extensor tibiae motor neuron     

The neural basis of catalepsy in the stick insect cuniculina impigra

The femur-tibia control system which is responsible for catalepsy is studied in the open-loop configuration (input: stimulation of the femoral chordotonal organ; output: spike-frequency of FETi and SETi as well as the force produced by the extensor tibiae muscle). Comparison of motor neuron activities and muscle force reveals the input-output relationships of the extensor tibiae muscle. This muscle behaves like a low-pass filter with a small time constant for rising inputs and a large time constant for falling inputs. It forms the decisive low-pass filter for force production of the complete system. For freely moving tibia, the elastic properties of the muscles combined with the inert mass of the tibia contribute to the low-pass filter properties. The muscle does not contribute to the high-pass filter properties of the complete system. During repetitive stimulation FETi habituates quickly.Supported by DFG Ba 578     

Motor patterns during kicking movements in the locust

Locusts (Schistocerca gregaria) use a distinctive motor pattern to extend the tibia of a hind leg rapidly in a kick. The necessary force is generated by an almost isometric contraction of the extensor tibiae muscle restrained by the co-contraction of the flexor tibiae (co-contraction phase) and aided by the mechanics of the femoro-tibial joint. The stored energy is delivered suddenly when the flexor muscle is inhibited. This paper analyses the activity of motor neurons to the major hind leg muscles during kicking, and relates it to tibial movements and the resultant forces.During the co-contraction phase flexor tibiae motor neurons are driven by apparently common sources of synaptic inputs to depolarized plateaus at which they spike. The two excitatory extensor motor neurons are also depolarized by similar patterns of synaptic inputs, but with the slow producing more spikes at higher frequencies than the fast. Trochanteral depressors spike at high frequency, the single levator tarsi at low frequency, and common inhibitors 2 and 3 spike sporadically. Trochanteral levators, depressor tarsi, and a retractor unguis motor neuron are hyperpolarized.Before the tibia extends all flexor motor neurons are hyperpolarized simultaneously, two common inhibitors, and the levator trochanter and depressor tarsi motor neurons are depolarized. Later, but still before the tibial movement starts, the extensor tibiae and levator tarsi motor neurons are hyperpolarized. After the movement has started, the extensor motor neurons are hyperpolarized further and the depressor trochanteris motor neurons are also hyperpolarized, indicating a contribution of both central and sensory feedback pathways.Variations in the duration of the co-contraction of almost twenty-fold, and in the number of spikes in the fast extensor tibiae motor neuron from 2–50 produce a spectrum of tibial extensions ranging from slow and weak, to rapid and powerful. Flexibility in the networks producing the motor pattern therefore results in a range of movements suited to the fluctuating requirements of the animal.     

The neural basis of catalepsy in the stick insect

The known nonlinearities of the femur-tibia control loop of the stick insect Carausius morosus (enabling the system to produce catalepsy) are already present in the nonspiking interneuron E4: (1) The decay of depolarizations in interneuron E4 following slow elongation movements of the femoral chordotonal organ apodeme could be described by a single exponential function, whereas the decay following faster movements had to be characterized by a double exponential function. (2) Each of the two corresponding time constants was independent of stimulus velocity. (3) The relative contribution of each function to the total amount of depolarization changed with stimulus velocity. (4) The characteristics described in (1)–(3) were also found in the slow extensor tibiae motoneuron. (5) Single electrode voltage clamp studies on interneuron E4 indicated that no voltage dependent membrane properties were involved in the generation of the observed time course of decay. Thus, we can trace back a certain behavior (catalepsy) to the properties of an identified, nonspiking interneuron.Abbrevations FETi fast extensor tibiae motor neuron - FT-joint femur-tibia joint - FT-control loop femur-tibia control loop - SETi slow extensor tibiae motor neuron - R regression coefficient     

Glutamatergic central nervous transmission in locusts

It is generally believed that neural transmission in the central nervous systems of insects is cholinergic, on the basis of secondary evidence: the presence of cholinesterase, and sensitivity of a nonsynaptic region of the neuron, its cell body, to iontophoresed acetylcholine. In the present work a preparation has been developed which takes advantage of the availability of identified motor neurons in the locust metathoracic ganglion with known 3-dimensional geometry of dendritic fields. These neurons transmit at their peripheral neuromuscular junctions with glutamate. The fast extensor tibiae motor neuron also makes excitatory central connections onto its functional antagonists the flexor tibiae motor neurons. Unless Dale's principle is contravened, transmission at these central synapses should also be glutamatergic. This transmission onto flexor motor neurons was found to be attenuated 70% by a glutamatergic blocker. Glutamate iontophoresed into selected areas of neuropil into which the motor neurons have dendritic branches caused the neurons to be depolarized, in a dose-dependent manner. Individual motor neurons were directly excited to spike with suprathreshold iontophoretic current. With long durations of release they were desensitized, but recovered quickly with rest. The data provide evidence that central transmission onto motor neurons in the locust metathoracic ganglion is glutamatergic.     

Local innervation patterns of the metathoracic flexor and extensor tibiae motor neurons in the cricket Gryllus bimaculatus

To elucidate neural mechanisms underlying walking and jumping in insects, motor neurons supplying femoral muscles have been identified mainly in locusts and katydids, but not in crickets. In this study, the motor innervation patterns of the metathoracic flexor and extensor tibiae muscles in the cricket, Gryllus bimaculatus were investigated by differential back-fills and nerve recordings. Whereas the extensor tibiae muscle has an innervation pattern similar to that of other orthopterans, the flexor has an innervation unique to this species. The main body of the flexor muscle is divided into the proximal, middle and distal regions, which receive morphologically unique terminations from almost non-overlapping sets of motor neurons. The proximal region is innervated by about 12 moderate-sized excitatory motor neurons and two inhibitory neurons while the middle and distal regions are innervated by three and four large excitatory motor neurons, respectively. The most-distally located accessory flexor muscle, inserting on a common flexor apodeme with the main muscle, is innervated by at least four small excitatory (slow-type) and two common inhibitory motor neurons. The two excitatory and two inhibitory motor neurons that innervate the accessory flexor muscle also innervate the proximal bundles of the main flexor muscle. This suggests that the most proximal and distal parts of the flexor muscle participate synergistically in fine motor control while the rest participates in powerful drive of tibial flexion movement.     

Leg kinematics and muscle activity during treadmill running in the cockroach, Blaberus discoidalis: II. Fast running

We have combined kinematic and electromyogram (EMG) analysis of running Blaberus discoidalis to examine how middle and hind leg kinematics vary with running speed and how the fast depressor coxa (Df) and fast extensor tibia (FETi) motor neurons affect kinematic parameters. In the range 2.5–10 Hz, B. discoidalis increases step frequency by altering the joint velocity and by reducing the time required for the transition from flexion to extension. For both Df and FETi the timing of recruitment coincides with the maximal frequency seen for the respective slow motor neurons. Df is first recruited at the beginning of coxa-femur (CF) extension. FETi is recruited in the latter half of femur-tibia (FT) extension during stance. Single muscle potentials produced by these fast motor neurons do not have pronounced effects on joint angular velocity during running. The transition from CF flexion to extension was abbreviated in those cycles with a Df potential occurring during the transition. One effect of Df activity during running may be to phase shift the beginning of joint extension so that the transition is sharpened. FETi is associated with greater FT extension at higher running speeds and may be necessary to overcome high joint torques at extended FT joint angles. Accepted: 24 May 1997     

The structure and function of serially homologous leg motor neurons in the locust. II. Physiology

Simultaneous intracellular recordings were made from pairs of motor neurons in the pro- or mesothoracic ganglion of the locust. Though central connections were sought between pairs of motor neurons, none were found. This is in sharp contrast to the findings that flexor and extensor tibiae neurons in the metathoracic ganglion make certain connections between themselves (Hoyle and Burrows, 1973; Heitler and Burrows, 1977a). As the previously mentioned authors believed that the metathoracic flexor-extensor connections were used as part of the motor program for jumping and kicking, the present results strongly support their hypothesis. Common PSPs have been found in a variety of pairs of motor neurons. Of note are common PSPs of the same sign to antagonists. Different innervation patterns have been found for the flexor and extensor muscles. It is proposed that serially homologous motor neurons serving similar functions are, to a first approximation, similar in the locust. Serially homologous motor neurons serving different functions will, in most cases, have altered structures and/or functions.     

Pre-processing and transfer entropy measures in motor neurons controlling limb movements

Directed information transfer measures are increasingly being employed in modeling neural system behavior due to their model-free approach, applicability to nonlinear and stochastic signals, and the potential to integrate repetitions of an experiment. Intracellular physiological recordings of graded synaptic potentials provide a number of additional challenges compared to spike signals due to non-stationary behaviour generated through extrinsic processes. We therefore propose a method to overcome this difficulty by using a preprocessing step based on Singular Spectrum Analysis (SSA) to remove nonlinear trends and discontinuities. We apply the method to intracellular recordings of synaptic responses of identified motor neurons evoked by stimulation of a proprioceptor that monitors limb position in leg of the desert locust. We then apply normalized delayed transfer entropy measures to neural responses evoked by displacements of the proprioceptor, the femoral chordotonal organ, that contains sensory neurones that monitor movements about the femoral-tibial joint. We then determine the consistency of responses within an individual recording of an identified motor neuron in a single animal, between repetitions of the same experiment in an identified motor neurons in the same animal and in repetitions of the same experiment from the same identified motor neuron in different animals. We found that delayed transfer entropy measures were consistent for a given identified neuron within and between animals and that they predict neural connectivity for the fast extensor tibiae motor neuron.     

Glutamatergic transmission between antagonistic motor neurones in the locust

The pharmacology of the direct central connections between the fast extensor and flexor motor neurones of a locust (Schistocerca gregaria) hind leg was studied. A spike in the fast extensor produces an EPSP in the flexor motor neurones. Glutamate depolarized the flexor motor neurones when injected into the neuropil. Quisqualate, but not by kainate or NMDA, also depolarized the flexor motor neurones. The fast extensor was also depolarized by glutamate, and also by kainate, but not by quisqualate, AMPA or NMDA. The glutamate response in the flexor motor neurones and the EPSP evoked by a spike in FETi both had similar reversal potentials. The FETi-evoked EPSP was blocked by bath application of the glutamate antagonist glutamic acid diethyl ester. The responses of extrasynaptic somata receptors to glutamate were compared to the neuropil responses. Glutamate usually hyperpolarized the somata of FETi and the flexor motor neurones. The response of a flexor motor neurone to glutamate was abolished at potentials less negative than -90 mV. The results provide evidence for glutamate transmission at central synapses in the locust, and show that presumed synaptic receptors in the neuropil differ to the extrasynaptic soma response     

Identified nonspiking interneurons in leg reflexes and during walking in the stick insect

In the stick insect Carausius morosus identified nonspiking interneurons (type E4) were investigated in the mesothoracic ganglion during intraand intersegmental reflexes and during searching and walking.In the standing and in the actively moving animal interneurons of type E4 drive the excitatory extensor tibiae motoneurons, up to four excitatory protractor coxae motoneurons, and the common inhibitor 1 motoneuron (Figs. 1–4).In the standing animal a depolarization of this type of interneuron is induced by tactile stimuli to the tarsi of the ipsilateral front, middle and hind legs (Fig. 5). This response precedes and accompanies the observed activation of the affected middle leg motoneurons. The same is true when compensatory leg placement reflexes are elicited by tactile stimuli given to the tarsi of the legs (Fig. 6).During forward walking the membrane potential of interneurons of type E4 is strongly modulated in the step-cycle (Figs.8–10). The peak depolarization occurs at the transition from stance to swing. The oscillations in membrane potential are correlated with the activity profile of the extensor motoneurons and the common inhibitor 1 (Fig. 9).The described properties of interneuron type E4 in the actively behaving animal show that these interneurons are involved in the organization and coordination of the motor output of the proximal leg joints during reflex movements and during walking.Abbreviations CLP reflex, compensatory leg placement reflex - CI1 common inhibitor I motoneuron - fCO femoral chordotonal organ - FETi fast extensor tibiae motoneuron - FT femur-tibia - SETi slow extensor tibiae motoneuron     

Slow motor neuron stimulation of locust skeletal muscle: model and measurement

The isometric force response of the locust hind leg extensor tibia muscle to stimulation of a slow extensor tibia motor neuron is experimentally investigated, and a mathematical model describing the response presented. The measured force response was modelled by considering the ability of an existing model, developed to describe the response to the stimulation of a fast extensor tibia motor neuron and to also model the response to slow motor neuron stimulation. It is found that despite large differences in the force response to slow and fast motor neuron stimulation, which could be accounted for by the differing physiology of the fibres they innervate, the model is able to describe the response to both fast and slow motor neuron stimulation. Thus, the presented model provides a potentially generally applicable, robust, simple model to describe the isometric force response of a range of muscles.