Pharmacological brain stimulation releases elaborate stridulatory behaviour in gomphocerine grasshoppers – conclusions for the organization of the central nervous control

Grasshoppers produce a variety of sounds generated by complex movements of the hindlegs. Stridulation, performed in the context of partner finding, mating and rivalry, can be released by pressure injection of cholinergic agonists into the protocerebrum. Particularly stimulation with muscarinic agonists induced long-lasting stridulation that resembled the natural behaviour to an astonishing degree, not only with respect to their temporal structure and right/left coordination, but also to changes in the song sequences according to the progress of courtship stridulation, even including accessory movements of other parts of the body. According to the complexity of their stridulatory behaviour ten gomphocerine species were chosen for this comparative study. The results indicate that the protocerebrum fulfils two important tasks in the control of stridulation: (1) it integrates sensory input relevant to stridulation that represents a certain behavioural situation and internal state of arousal, and (2) it selectively activates and deactivates the thoracic networks that generate the appropriate movement and sound patterns. With the knowledge of the natural behaviour and the accessibility to pharmacological and electrophysiological studies, the cephalic control system for stridulation in grasshoppers appears to be a suitable model for how the brain selects and controls appropriate behaviours for a given situation.     

Identified descending brain neurons control different stridulatory motor patterns in an acridid grasshopper

During courtship sequences male grasshoppers of the species Omocestus viridulus successively perform with their hindlegs three different stridulatory movement patterns: ordinary stridulation, hindleg shaking and precopulatory movements. Microinjection of acetylcholine into protocerebral neuropil regions can either elicit complete courtship sequences or evoke one of the three motor patterns. Intracellular recordings and stainings revealed three types of descending brain neurons: B-DC-3, B-DC-4 and B-DC-5. All three types of interneurons have a medial axon position in the connectives. They cross the midline of the protocerebrum and exhibit a profuse arborization pattern within the medial dorsal protocerebral neuropil. Stimulation of each type of interneuron specifically elicits one particular motor pattern of courtship behaviour. Courtship of the grasshopper O. viridulus may therefore be controlled by successive activation of these descending brain neurons. Accepted: 27 September 1996     

Pharmacological Activation of Stridulation in the Grasshopper Chorthippus albomarginatus (Orthoptera: Gomphocerinae)

Calling and courtship stridulatory behavior of Chorthippus albomarginatus was induced by injections of acetylcholine agonists into the protocerebrum. Pharmacologically induced stridulation, in many parameters, was quite similar to the natural behavior. However, the order of the courtship element alternation was different from that of the natural song. In some cases the pharmacologically induced stridulation included only one or two courtship elements. Based on the exclusive stimulation of a particular element of courtship songs and similarity of its movement pattern with the calling song, both patterns of Ch. albomarginatus stridulation appear to be homologous. The results obtained on this species confirm the idea of a hierarchic organization of the central nervous control of stridulation in gomphocerine grasshoppers and indicate participation of certain protocerebrum structures in this control.     

Pulses, patterns and paths: neurobiology of acoustic behaviour in crickets

Crickets use acoustic communication for pair formation. Males sing with rhythmical movements of their wings and the mute females approach the singing males by phonotaxis. Females walking on a trackball rapidly steer towards single sound pulses when exposed to split-song paradigms. Their walking path emerges from consecutive reactive steering responses, which show no temporal selectivity. Temporal pattern recognition is tuned to the species-specific syllable rate and gradually changes the gain of auditory steering. If pattern recognition is based on instantaneous discharge rate coding, then the tuning to the species-specific song pattern may already be present at the level of thoracic interneurons. During the processing of song patterns, changes in cytosolic Ca²⁺ concentrations occur in phase with the chirp rhythm in the local auditory interneurone. Male singing behaviour is controlled by command neurons descending from the brain. The neuropil controlling singing behaviour is located in the anterior protocerebrum next to the mushroom bodies. Singing behaviour is released by injection of cholinergic agonists and inhibited by γ-butyric acid (GABA). During singing, the sensitivity of the peripheral auditory system remains unchanged but a corollary discharge inhibits auditory processing in afferents and interneurons within the prothoracic auditory neuropil and prevents the auditory neurons from desensitisation.     

Muscarinic excitation in grasshopper song control circuits is limited by acetylcholinesterase activity

The species- and situation-specific sound production of grasshoppers can be stimulated by focal application of both nicotinic and muscarinic receptor agonists into the central body complex of the protocerebrum. Pressure injection of the intrinsic transmitter acetylcholine only elicits fast and short-lived responses related to nicotinic receptor-mediated excitation. Prolonged sound production that includes complex song patterns requires muscarinic receptor-mediated excitation. In addition, basal muscarinic excitation in the central body neuropil seems to determine the general motivation of a grasshopper to stridulate. To demonstrate that endogenous acetylcholinesterase limits the activation of muscarinic receptors by synaptically released acetylcholine in the central body of Chorthippus biguttulus, we investigated both its presence in the brain and effects on sound production resulting from inhibition of esterase activity. Acetylcholinesterase activity was detected in the upper and lower division of the central body. Both these neuropils known to be involved in the cephalic control of stridulation were also shown to contain muscarinic acetylcholine receptors expressed by columnar neurons suggested to serve as output neurons of the central complex. Pressure injection of the acetylcholinesterase inhibitor eserine into protocerebral control circuits of restrained male grasshoppers stimulated long-lasting stridulation that depended on scopolamine-sensitive muscarinic receptors. In restrained males, eserine released the typical response song by potentiating the stimulatory effect of the conspecific female song. Eserine-mediated inhibition of acetylcholinesterase in the central body prolongs the presence of synaptically released acetylcholine at its postsynaptic receptors and increases its potency to activate muscarinic receptor-initiated signaling pathways acting to promote grasshopper sound production.     

Separate localization of sound recognizing and sound producing neural mechanisms in a grasshopper

Summary In the two acridid speciesChorthippus parallelus andCh. montanus, the sound template by which females recognize male song varies with temperature, as does the song itself. At relatively high temperatures the females respond best to simulated songs with high syllable frequencies, and at lower temperatures songs with lower syllable frequencies are preferred.The temperature around the supraesophageal and metathoracic ganglia of female grasshoppers was monitored by implanted thermocouples, and either the head or the thorax was warmed selectively while the animal was free to move (within the imits of the wires). Then simulations of the conspecific song varying in syllable frequency corresponding to different song temperatures were presented, and the stridulatory responses of the animals were observed.The results were as follows. 1. Song recognition (in particular, the position of the peak of the response curve) depended on the temperature of the head. 2. The rate of stridulatory hindleg movement was determined by the temperature of the thoracic ganglia.This result provides strong evidence against the genetic coupling hypothesis.     

‘Switching-off’ of an auditory interneuron during stridulation in the acridid grasshopperChorthippus biguttulus L.

Summary In freely moving grasshoppers of the speciesChorthippus biguttulus compound potentials were recorded from the neck connectives with chronically implanted hook electrodes. The spikes of one large auditory interneuron, known as the G-neuron (Kalmring 1975a, b) were clearly distinguishable in the recordings and the neuron was identified by its physiology and morphology. In quiescent grasshoppers the G-neuron responds to auditory and vibratory stimuli, but responses to both stimuli are suppressed during stridulation in males (Fig. 1, top, Fig. 7). When a male's wings were removed so that the stridulatory movements of its hindlegs produced no sound, the suppression of the G-neuron response still occurred (Fig. 1, bottom). When proprioceptive feedback from the hindlegs was reduced, by forced autotomy of the legs, the switching-off remained incomplete (Fig. 3) (production of stridulatory patterns was inferred on the basis of electromyograms from the relevant thoracic musculature). Imposed movement of the hindlegs, on the other hand, suppressed the G-neuron response in a graded fashion, depending on the frequency of the movement (Figs. 4 and 5). These experiments suggest that the switching-off is brought about by a combination of proprioceptive feedback and central efferences. The switching-off phenomenon could either protect the grasshopper's auditory pathway from undesired effects of overloading by its own intense song (e.g. self-induced habituation as described by Krasne and Wine 1977) and should therefore apply for most auditory neurons. Alternatively it could prevent escape reflexes from being triggered by stridulatory self-stimulation and consequently might apply only for neurons involved in such networks (as the G-neuron might be).     

Stridulatory pattern generation in acridid grasshoppers: metathoracic interneurons in Stenobothrus rubicundus (Germar 1817)

Stridulation was elicited in tethered gomphocerine grasshoppers of the species Stenobothrus rubicundus in order to identify interneurons of the stridulation pattern generator, and describe their morphological and physiological properties. Nine types of such neurons could be found and characterized; eight of those could additionally be compared to corresponding neuron types previously known from other species. As shown in detail for one selected type, the neurons of the stridulation pattern generator are very similar in their anatomical appearance, and possess similar physiological qualities at least in two species with similar stridulation patterns. Stridulation interneurons of species with largely different stridulatory motor patterns have a similar morphology, but show a different activation timing throughout the stridulation. Nevertheless, special properties such as resetting or initiation capability of certain stridulation interneurons seem to be conserved throughout the species. The results suggest that the stridulation pattern generator of different species consists of a uniform set of interneurons that change their activity pattern to produce species-specific song movements.     

Central nervous control of hindleg coordination in stridulating grasshoppers

 Stridulation in many gomphocerine grasshoppers is characterized by specific phase shifts between the two hindlegs as well as different movement patterns produced by the left and the right leg. The underlying neuronal excitation patterns are generated by networks on either side of the metathoracic ganglion. The role of the intraganglionic commissures in right-left coordination and the production of differing movement patterns was investigated by transecting the metathoracic ganglion mediosagittally in Omocestus viridulus, Chorthippus biguttulus and Chorthippus mollis. In all three species, after this operation both hindlegs produced the same pattern and no longer different movements. The effects of transection on coordination differed: rapid movement rhythms, like those typical of Ch. biguttulus and the vibratory parts of the song of Ch. mollis, on the two sides drifted with respect to one another. In contrast, the slow rhythms characteristic of O. viridulus and the song subunits of Ch. mollis were completely synchronized. It is inferred that in intact animals the pathways for coordination of the rapid stridulatory rhythms are exclusively intraganglionic, whereas the phase relations of the slow rhythms are additionally influenced by way of anterior right-left connections, perhaps within the suboesophageal ganglion. Accepted: 15 October 1996     

Participation of GABA and glycine in the formation of somato-sympathetic reflexes]

Experiments were conducted on cats with intact spinal cord, and after spinalization a study was made of the action of strychnine and picrotoxin (synaptic GABA and glycine antagonists) on the components of somatosynaptic reflexes caused by stimulation of the skin and muscular intercostal afferents. Induced potentials were recorded in the white ramus of the third thoracic segment. The action of strychnine and picrotoxin on the values of different reflex components was not identical; this pointed to a different localization of the inhibitory action of GABA and glycine in the central link of the somatosympathetic reflex arch.     

Descending stridulatory interneurons in the suboesophageal ganglion of two grasshopper species

1. The activity of interneurons in the suboesophageal ganglion of the acridid grasshoppers Omocestus viridulus (L.) and Chorthippus mollis (Charp.), recorded intracellularly during stridulation, was found to conform to the rhythm of the singing movements. The arborizations of these neurons in this ganglion are largely bilaterally symmetrical; the axon descends contralaterally to the soma and passes at least into the metathoracic ganglion.
2. The anatomical and physiological characteristics of these neurons are similar in the two species and of four types. Three of them exhibit a tonic, spontaneous activity in the resting animal, which is modulated in the stridulatory rhythm as soon as singing begins. The fourth type has no resting activity and discharges only during the song, in a stridulation-specific pattern.
3. By transecting the connectives it was shown that the rhythmic activity of the neurons is not determined by input from the brain, nor is it generated in the suboesophageal ganglion itself. It is based on information about the song pattern that ascends from the thoracic ganglia.

     

Primitive nervous systems: Action of aminergic drugs and blocking agents on activity in the ventral nerve cord of the flatworm Notoplana acticola

Electrically evoked activity in the submuscular ventral longitudinal nerve cords of Notoplana acticola is depressed by GABA and glycine in the presence of high magnesium concentrations. This inhibition occurs with 0.001–0.01 millimolar concentrations of these putative aminergic neurotransmitters and is reversible when washed out. The action of GABA and glycine was reversed nonspecifically by picrotoxin, bucuculline, and strychnine. PTZ (Pentylenetetrazole) was shown to mimic the effects that these blocking agents had on evoked activity when they were tested alone. The release of inhibition by these blocking agents is similar to that of decerebration. Three possible mechanisms responsible for synaptic activity in high Mg²⁺ concentrations are discussed and the possibility that the effector site of interaction may be the chloride ionophore is explored.     

Serotonin in the leech central nervous system: anatomical correlates and behavioral effects

Summary Stridulation of grasshoppers is controlled by hemisegmental pattern generator subunits which probably are restricted to the metathoracic ganglion complex (TG3-complex). The coordination of left and right pattern generator subunits depends on commissures of the TG3-complex (Ronacher 1989). The coordination of the stridulatory movements was studied in Chorthippus dorsatus males with partial mediosagittal incisions in the TG3-complex.Animals bearing anterior incisions in the TG3-complex, by which all commissures of the metathoracic neuromere and the first abdominal neuromere were transected, were still able to produce bilaterally coordinated species-specific stridulatory movements. Commissures of the T3- and A1-neuromere, thus, are not necessary, and the A2-, A3-commissures are sufficient for this coordination (Figs. 3, 4).Animals with partial posterior incisions, extending until A1, had deficits in their stridulation pattern; the coordination between the hindlegs was impaired though not completely lost (Fig. 6). This is discussed in view of the structure of stridulation interneurons identified in a related grasshopper species (Omocestus viridulus).These results indicate an unexpected substantial contribution of the abdominal neuromeres A2 and A3 to the control of stridulatory movements. This constitutes an interesting parallel to the flight control system of locusts where interneurons located in the first 3 abdominal neuromeres also appear to contribute to the flight pattern generator (Robertson et al. 1982).Abbreviations A1–A3 abdominal neuromeres 1–3 - T3 metathoracic neuromere - TG3-complex metathoracic ganglion complex including A1–A3     

Abdominal ventilatory pattern in crickets depends on the stridulatory motor pattern

Abstract. Ventilatory motor patterns were recorded from abdominal muscles in crickets, Gryllus campestris L.and Teleogryllus commodus (Walker), at rest and during three types of stridulatory motor activity; calling, courtship and aggressive song.
Increases in ventilatory period were almost exclusively due to an increase of the pause between expiratory bursts, whereas abdominal ventilatory bursts remained constant at 200 ms.Ventilatory patterns depended on the stridulatory motor pattern and indicated that the same basic respiratory oscillator exists in both cricket species.
In G.campestris there was a strict 1:1 coupling between chirps and ventilatory bursts.In T.commodus such a relationship was also observed for the chirp part of the songs, but less strictly for the trill part of the calling song and not for the courtship song.In both species the onset of the ventilatory burst was within ± 100 ms of a stridulatory chirp.Ventilatory burst lasted longer the earlier they began before a stridulatory chirp.This suggests strongly that the stridulatory motor pattern terminates the expiratory burst, and thus influences the ventilatory motor pattern.     

The complex stridulatory behavior of the cricket Eneoptera guyanensis Chopard (Orthoptera: Grylloidea: Eneopterinae)

Crickets produce stridulated sounds by rubbing their forewings together. The calling song of the cricket species Eneoptera guyanensis Chopard, 1931 alternates two song sections, at low and high dominant frequencies, corresponding to two distinct sections of the stridulatory file. In the present study we address the complex acoustic behavior of E. guyanensis by integrating information on the peculiar morphology of the stridulatory file, the acoustic analysis of its calling song and the forewing movements during sound production. The results show that even if E. guyanensis matches the normal cricket functioning for syllable production, the stridulation involves two different closing movements, corresponding to two types of syllables, allowing the plectrum to hit alternately each differentiated section of the file. Transition syllables combine high and low frequencies and are emitted by a complete forewing closure over the whole file. The double-teeth section of the stridulatory file may be used as a multiplier for the song frequency because of the morphological multiplication due to the double teeth, but also because of an increase of wing velocity when this file section is used. According to available phylogenetic and acoustic data, this complex stridulation may have evolved in a two-step process.     

Studies of the role of gamma-aminobutyric acid in the hypothalamic control of feed intake in sheep

Fourteen sheep were used to study the role of gamma-aminobutyric acid (GABA) on the hypothalamic control of feed intake. Injections (1 microL) of pentobarbital (262 nmol) into preoptic and paraventricular areas induced feeding in satiated sheep. Injections of GABA into the same loci gave variable results, probably because the neuronal and glial uptake of GABA limits its effects. Muscimol, a GABA agonist with a higher affinity for postsynaptic GABA receptors than GABA, injected at doses from 0 to 0.750 nmol, gave a cubic dose-response curve; the highest feed intake was measured at 0.5 nmol. The response induced by muscimol was blocked by preinjections of two GABA antagonists, picrotoxin and bicuculline, with picrotoxin being more effective than bicuculline. Muscimol responsive loci were identified mainly in the preoptic, paraventricular, and anterior hypothalamus. The data suggests that neurons sensitive to gamma-aminobutyric acid may be implicated in the control of feed intake in sheep.     

A narrow hybrid zone between the grasshoppers Stenobothrus clavatus and Stenobothrus rubicundus: courtship song analysis

The closely related grasshopper species Stenobothrus rubicundus and Stenobothrus clavatus are known to hybridize in a very narrow contact zone on Mt. Tomaros in northern Greece. These species produce very different and complex courtship songs accompanied with visual display. We analyzed the courtship songs and underlying stridulatory movements of the hind legs in natural hybrids from Mount Tomaros. The two species were also hybridized in the laboratory and their songs were compared with the songs of the natural hybrids. Some hybrid songs were shown to have intermediate features between parental songs, whereas other hybrid songs comprised completely new elements. The clavatus‐like song elements were found to dominate in hybrid songs. These song features may influence the mating success of hybrid males in the contact zone. A comparison of hybrid songs with the song pattern of the north European S. rubicundus populations allowed us to suggest a scenario of S. rubicundus and S. clavatus origin. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ?? , ??–??.     

Identification of a novel residue within the second transmembrane domain that confers use-facilitated block by picrotoxin in glycine alpha 1 receptors.

The central nervous system convulsant picrotoxin (PTX) inhibits GABA(A) and glutamate-gated Cl(minus sign) channels in a use-facilitated fashion, whereas PTX inhibition of glycine and GABA(C) receptors displays little or no use-facilitated block. We have identified a residue in the extracellular aspect of the second transmembrane domain that converted picrotoxin inhibition of glycine alpha1 receptors from non-use-facilitated to use-facilitated. In wild type alpha1 receptors, PTX inhibited glycine-gated Cl(minus sign) current in a competitive manner and had equivalent effects on peak and steady-state currents, confirming a lack of use-facilitated block. Mutation of the second transmembrane domain 15'-serine to glutamine (alpha1(S15'Q) receptors) converted the mechanism of PTX blockade from competitive to non-competitive. However, more notable was the fact that in alpha1(S15'Q) receptors, PTX had insignificant effects on peak current amplitude and dramatically enhanced current decay kinetics. Similar results were found in alpha1(S15'N) receptors. The reciprocal mutation in the beta2 subunit of alpha1beta2 GABA(A) receptors (alpha1beta2(N15'S) receptors) decreased the magnitude of use-facilitated PTX inhibition. Our results implicate a specific amino acid at the extracellular aspect of the ion channel in determining use-facilitated characteristics of picrotoxin blockade. Moreover, the data are consistent with the suggestion that picrotoxin may interact with two domains in ligand-gated anion channels.     

GABA regulates the buccal motor output of Helisoma by two pharmacologically distinct actions

GABA was tested for its effects on patterned motor activity (PMA) underlying feeding. Using buccal motoneuron B19 to monitor PMA through intracellular recordings, GABA was found to exert effects at two levels. First, GABA stimulated rhythmic patterned activity resembling fictive feeding, which is under the control of the buccal CPG. In addition, GABA produced a direct inhibition of neuron B19. Both effects were observed when the buccal ganglia were studied in isolation from the rest of the central nervous system, suggesting local interactions with GABA receptors of buccal neurons. Furthermore, these two actions of GABA were found to be pharmacologically distinguishable. The direct hyperpolarization of neuron B19 was mimicked by muscimol, but not baclofen, and involved an increased chloride conductance, which was blocked by picrotoxin.Baclofen duplicated CPG activation by GABA. Picrotoxin had no effect on GABA- or baclofen-induced PMA.These results demonstrate that the Helisoma buccal ganglia have two GABA receptor types which resemble, pharmacologically, mammalian GABA_A and GABA_B receptors, and that GABA plays a key role in feeding patterned motor activity in Helisoma.Abbreviations CPG central pattern generator - GABA gammaamino butyric acid - HPLC high performance liquid chromatography - IPSP inhibitory postsynaptic potential - PMA patterned motor activity - SLRT supralateral radular tensor muscle     

Male singing behaviour and female presence in the territory in whitethroats Sylvia communis

Unmated male songbirds usually change their vocal behaviour when females enter their territories. Either the males court the females by changing the rate or pattern with which their normal long-ranging full songs are emitted, or they shift to special displays and long- or short-ranging vocalisations. In this study we quantified how female presence and behaviour affect the singing behaviour of male whitethroats. In the presence of a female the male frequently performed song flights, maybe to locate the female before it was courted, with sequences of diving-song displays. The courtship was interrupted by periods of perch songs. Female dscharp calls and short movements made the males initiate or resume courtship, whereas short horizontal jumps made the males intensify their courtship. Overall, the males changed their dual-function song activity in females' presence by emitting fewer perch songs and more flight songs. The quiet diving songs were only emitted during direct courtship of a female. The song types emitted immediately before, during, and after courtship are all highly variable, thus allowing for a quick assessment of the male's song repertoire. The courtship was also interrupted by periods of male woid calling, a call that is known to have a deterring effect on rival males. Bouts of woid calls were usually followed by song flights, again probably to locate the female that might have moved out of sight, or maybe to locate potential rival intruders. The latter was supported by an increased intrusion rate during female presence. Communicated by P. McGregor