JOURNAL OF EXPERIMENTAL ZOOLOGY 286:219–230 (2000)

© 2000 WILEY-LISS, INC.

Morphology and Physiology of Auditory andVibratory Ascending Interneurones in Bushcrickets

BERND NEBELING*AG Neurobiologie, FB Biologie, Philipps Universität, 35033 Marburg,Germany

ABSTRACT Auditory/vibratory interneurones of the bushcricket species Decticus albifrons andDecticus verrucivorus were studied with intracellular dye injection and electrophysiology. The mor-phologies of five physiologically characterised auditory/vibratory interneurones are shown in thebrain, subesophageal and prothoracic ganglia. Based on their physiology, these five interneuronesfall into three groups, the purely auditory or sound neurones: S-neurones, the purely vibratory V-neurones, and the bimodal vibrosensitive VS-neurones. The S1-neurones respond phasically toairborne sound whereas the S4-neurones exhibit a tonic spike pattern. Their somata are located inthe prothoracic ganglion and they show an ascending axon with dendrites located in the protho-racic, subesophageal ganglia, and the brain. The VS3-neurone, responding to both auditory andvibratory stimuli in a tonic manner, has its axon traversing the brain, the suboesophageal gan-glion and the prothoracic ganglion although with dendrites only in the brain. The V1- and V2-neurones respond to vibratory stimulation of the fore- and midlegs with a tonic discharge pattern,and our data show that they receive inhibitory input suppressing their spontaneous activity. Theiraxon transverses the prothoracic ganglion, subesophageal ganglion and terminate in the brainwith dendritic branching.

Thus the auditory S-neurones have dendritic arborizations in all three ganglia (prothoracic,subesophageal, and brain) compared to the vibratory (V) and vibrosensitive (VS) neurones, whichhave dendrites almost only in the brain. The dendrites of the S-neurones are also more extensivethan those of the V-, VS-neurones. V- and VS-neurones terminate more laterally in the brain. Dueto an interspecific comparison of the identified auditory interneurones the S1-neurone is found tobe homologous to the TN1 of crickets and other bushcrickets, and the S4-neurone also can becalled AN2. J. Exp. Zool. 286:219–230, 2000. © 2000 Wiley-Liss, Inc.

In bushcrickets, the tympanal and the subgen-ual organs function in the perception of airbornesound and substrate borne vibration respectively(Schumacher, ’73; Rössler, ’92). The auditory andvibratory receptor cells involved in this processare located in the proximal parts of the anteriortibiae and are known to converge synaptically tohigher order auditory and/or vibratory neurones(Kalmring et al., ’79; Römer et al., ’88). Althoughin bushcrickets the number of receptor cells(Kalmring et al., ’93) and the number of ascend-ing neurones has been shown to be species spe-cific, receptor cells are always more numerousthan interneurones. Therefore, sensory inputmust converge at the level of the dendritic regionof a relevant interneurone.

Ascending interneurones in the ventral nervecord of bushcrickets have been subject to detailedmorphological and physiological research (Römeret al., ’88; Shen, ’93). In the brain and subeso-phageal ganglion (SEG), however, interneurones

have only been described physiologically (Rhein-laender and Kalmring, ’73). Very poor evidence isavailable on the morphology of the auditory inter-neurones in the brain, and so far no research hasbeen carried out to follow the projections of the ar-borizations of vibratory interneurones in the brain.By investigating both the physiology and morphol-ogy of ascending auditory and vibratory inter-neurones in the brain this study aimed to fill gapsin our knowledge of the organization of the audi-tory and vibratory pathways in Tettigoniids.

MATERIALS AND METHODSAnimals and preparation

Experiments were performed on sexually ma-ture male and female Decticus albifrons and

Grant sponsor: Sonderforschungsbereich Ökophysiologie (305);Grant number: Teilprojekt A7.

*Correspondence to: Bernd Nebeling, MZ für Augenheilkunde,Philipps Universität, Robert-Koch-Str. 4, 35033 Marburg, Germany.

Received 7 April 1999; Accepted 17 June 1999

220 B. NEBELING

Decticus verrucivorus obtained from their natu-ral habitat in southern France and Bavaria. Theanimals were kept in the laboratory for up to fourweeks under appropriate conditions (Rheinlaenderand Kalmring, ’73).

Prior to the experiments the animals wereanaesthetised with CO2 and dorsally attached toa holder. All legs were fixed, placing the forelegsto the wire holder of a mini-shaker in a naturalposition. The animals were then dissected fromthe ventral surface, the gut and the mouthpartswere removed, and the caudal part of the sub-oesophageal ganglion (SEG) together with theconnectives were exposed.

The experiments comply with the NIH “Prin-ciples of animal care” and current German laws.

Electrophysiology and stainingFor intracellular recordings, microelectrodes were

drawn from thick-walled borosilicate glass micro-capillaries. They were filled with 1.5 M NiCl2 andhad a resistance of about 20 MΩ. Alternatively, thetip of the microcapillaries was filled with 5% Luci-fer-Yellow (Sigma), backed up with 0.2 M LiCl so-lution (80 MΩ). In the latter, for iontophoreticinjection a constant hyperpolarizing current of 2–5nA was set for up to 1 hr. The NiCl2-technique ap-pears to be more suitable for recording frominterneurones of very small diameter, whereas themethod using Lucifer-Yellow is more advantageousfor stainings over long distances. Therefore, in thisstudy both techniques were applied.

The indifferent electrode was inserted into theabdomen. All experiments were carried out in ananechoic chamber which is acoustically insulatedagainst background noise.

Standard airborne-sound and substrate-bornesound stimuli (duration 20 ms and 100 ms, rise/fall time 1 ms, repetition rate 2/s) of different fre-quencies were used. The signals were generatedby two acoustic stimulators (Burchard II). Theacoustic stimuli were delivered ipsilaterally to thesite of penetration in a distance of 41 cm fromthe penetration via a HF-loudspeaker (Audax, TW8), perpendicular to the body axis of the animal.The vibration was passed to the fore- and midlegsby a minivibrator (Bruel + Kjaer, Typ 4810).

The intensities of the sound signals were cali-brated with a condenser microphone (Bruel +Kjaer 4135 or 4138) and a B + K measuring am-plifier (2209) using its “peak hold” function. Toensure comparability, all signal intensities aregiven in dB peak SPL, which is for sine waves 3dB above the respective RMS value. Sound mea-

surements were obtained on the preparation site,with no animal present.

The responses of each single cell were storedon magnetic tape (Racal 4DS) and later replayed,digitized (sampling frequency 200 or 400 kHz) andanalyzed by custom made computer programs.The responses are presented as spike-trains andperi-stimulus-time-histograms (PSTHs).

Threshold curve and response magnitude aregiven for each interneurone, and the neuroneswere classified physiologically according to Silveret al. (’80).

HistologyAfter the electrophysiological experiments, the

brain, SEG, and first two thoracic ganglia weredissected.

When NiCl2 had been employed, the nickel-ionswere precipitated with rubeanic acid. After fixa-tion, silver intensification, and dehydration, theganglia were cleared in styrene (Bacon andAltman, ’79). The projection patterns of theinterneurones were then reconstructed in wholemounts via camera lucida drawings.

For Lucifer-Yellow staining the ganglion chainwas fixed in phosphate-buffered 4% formalin for10 min and afterwards dehydrated and trans-ferred to 4% formalin in alcohol for at least 1 hr.The ganglia were rinsed in alcohol (100%) andstored in methyl salycilate. Whole mounts werephotographed in a series of focal planes in thefluorescence microscope. In addition, for perma-nent visualization the ganglia were embedded ingelatin/albumin and serial thick sections (20 µm)were obtained with a vibratome. Using the PAP-technique (Peroxidase-Anti-Peroxidase) the stain-ing of the interneurones could be intensified(Anti-Lucifer, rabbit, Sigma; second antibody:Goat Anti Rabbit; third step: rabbit PAP, intensi-fication with DiAmino-Benzidin; for more detailssee Nebeling, ’95).

A frontal view of the morphology of each neu-rone was drawn by using both whole mounts andserial-sections which allowed tracing very finebranches. A sagittal view was reconstructed us-ing only the serial sections and this allowed trac-ing only the main branches.

RESULTSThe nomenclature in this work follows the physi-

ological classification of Kühne et al. (’80) and Sil-ver et al. (’80) because vibrosensitive and bimodalneurones have so far not been characterised mor-phologically.

AUDITORY NEURONES IN THE BRAIN OF BUSHCRICKETS 221

As no sex specific differences were obvious, maleand females were not treated separately. Physi-ologically data of almost all types of interneuronesdescribed in literature could be recorded in bothspecies—no major differences were found. Addi-tionally, some neurones of the same type stainedin D. albifrons and in D. verrucivorus showed verysimilar arborization patterns. Therefore, the re-sults of both species were not treated separately.Based on their physiological responses the inter-neurones were divided into three groups: S, VSand V, and each is described separately.

S-neuronesThese interneurones are characterized by their

exclusive response to airborne sound (Silver et al.,’80) and comprise five different subtypes whichare distinguished by their specific spike patterns.Two of these subtypes were identified in this work.

S-neurone, type S1The S1-neurone has a large fiber diameter and

has therefore already been studied before by sev-eral authors (e.g., Kalmring et al., ’79). Followinga short habituation phase (not shown in the fig-ures) the S1-neurones (n = 21) responded to air-borne sound in a strictly phasic manner.

The responses remained phasic even to stimuliof long duration (100 ms). The neurones were sen-sitive to a broad band of frequencies (Fig. 1A–C).In the most sensitive frequency band between 12and 30 kHz the neurones responded to sound ofless than 40 dB SPL intensity. The response la-tency was short (12 ± 0.5 ms SD) in keeping withthe large diameter of these neurones and no spon-taneous activity was detected (Fig. 1A, B).

S1-neurones were injected with dye in the SEGfrom where the dye ascended to the terminationsin the cephalic ganglia and descended backwardsto the prothoracic ganglion, where it filled acontralaterally located somata (Fig. 1D). In theSEG, 8 to 10 collaterals form an extensive area ofend-branches ipsilaterally to the axon. In thebrain, S1-neurones terminate in the median partsof the ventro-lateral protocerebrum. In contrastto former investigations (Kalmring et al., ’79) anadditional arborization area was stained in themedian deutocerebrum. The spatial relation of theterminations to some major brain structures(mushroom body, central body, protocerebralbridge) of one representative S1-neurone is shownin both frontal and sagittal views (Fig. 1E, F) andshow that terminations of this neurone do not en-ter these brain structures.

S-neurone, type S4Neurones of the type S4 were recorded (n = 5)

in the median part of the neck-connectives, closelylateral to the S1-neurones. The diameter of theiraxon is large but smaller than that of the S1. Thephysiology of S4-neurones is demonstrated (Fig.2A–C). The most obvious difference to type S1-neurones is in their tonic spike-pattern followingstimuli of 20 ms and 100 ms duration. Almost nohabituation was found. The latency of theseneurones (19.5 ± 0.5 ms SD) was longer than thelatency of the S1-type; also the spike repetitionrate was higher. Like S1-neurones they respondedto sound over the whole frequency range from 3to 60 kHz with best frequencies from 10 to 25 kHz.

The morphology of S4-neurones closely re-sembles that of S1-neurones. Thus the somata arelocated contralaterally to the axon in the anteriorcortex of the TH1 (Fig. 2D) and a single axon as-cends to the brain.

Along the way the axon originates 5 to 7 (ipsi-lateral) collaterals in the SEG, oriented towardsthe midline similar to those in S1 although notas densely branched as those of S1. In the proto-cerebrum the axons of these S4-neurones form ex-tensive terminations extending into the optic stalk(Fig. 2E, F). Some collaterals terminate in themedian deutocerebrum building an additional,separate projection area. The terminations of theS4 neurones do not contact the mushroom bodyor the central body.

VS-neurones

These bimodal interneurones which show exci-tatory responses to both airborne sound and vi-bration were named VS-neurones by Silver et al.(’80) and five subtypes can be distinguished bytheir spike patterns. One of these types, the VS3-neurone, is described here. The physiological dataof VS3 neurones are shown in Figure 3A–C. Theyresponded tonically to sound and (phasic)-toni-cally to vibratory stimulation. In their responsesto airborne sound they resembled the S4 type. Incontrast to S4-neurones, VS3-neurones showedspontaneous activity and exhibited a greater la-tency of 29 ± 2 ms (SD). Spontaneous-activity wassuppressed for more than 200 ms after stimula-tion with stimuli of high efficiency (e.g., post-stimulus suppression occurs to vibration of 200Hz and high intensity). These neurones were sen-sitive to vibratory stimuli over the whole testedrange from 100 to 2000 Hz, having a thresholdacceleration of at least 0.03 m/s2.

222 B. NEBELING

Fig. 1. Morphology and physiological data of an S1-neu-rone (TN1). (A) Five responses to repetitive stimulations withwhite noise (WN, 20 ms duration, 73 dB SPL). (B) Spike trains(upper part) and PSTH (2 ms bin width) to white noise (WN,100 ms duration, 73 dB SPL). (C) Response magnitude tostimulation with airborne sound. (+) = tested without excita-tory effect. (D) Position and morphology within the head and

prothoracic ganglia, site of penetration (which is the samefor all experiments) is indicated. Arrow indicates an addi-tional arborization area, which was not stained in earlier in-vestigations. (E, F) Reconstruction of the endbranches withinthe brain and relationship to prominent structures; (E) fron-tal view, (F) sagittal view.

AUDITORY NEURONES IN THE BRAIN OF BUSHCRICKETS 223

Fig. 2. Morphology and physiological data of an S4-neu-rone (AN2). Spike trains (upper part) and PSTH (2 ms binwidth) to stimulation with (A) white noise (WN, 20 ms dura-tion, 73 dB SPL); (B) white noise (WN, 100 ms duration, 73dB SPL). (C) Response magnitude to stimulation with air-borne sound. (+) = tested without excitatory effect. (D) Posi-

tion and morphology within the head and prothoracic gan-glia. Arrow indicates an additional, separate projection area.(E, F) Reconstruction of the endbranches within the brainand relationship to prominent structures (E) frontal view, (F)sagittal view.

224 B. NEBELING

Fig. 3. Morphology and physiological data of a VS3-neu-rone. Spike trains (upper part) and PSTH (2 ms bin width)to stimulation with (A) white noise (WN, 20 ms duration, 73dB SPL); (B) vibration (200 Hz, 100 ms duration, 2.7 m/s2);arrows indicate the suppression of spontaneous activity. (C)Response magnitude to stimulation with airborne sound (up-

per diagram) and vibration. (+) = tested without excitatoryeffect. (D) Position and morphology within the head and pro-thoracic ganglia. (E, F) Reconstruction of the endbrancheswithin the brain and relationship to prominent structures;(E) frontal view, (F) sagittal view.

AUDITORY NEURONES IN THE BRAIN OF BUSHCRICKETS 225

Stainings of VS-neurones could be obtained inthe brain and the thoracic ganglia in four ani-mals. The reconstruction of a representative VS3-neurone in the brain, SEG and the TH1 isdemonstrated in Figure 3D and did not reveal asoma in any of those regions. Moreover, the axondid not reveal any collaterals in the SEG eventhough fine branches were stained in the brain.Thus the lack of collaterals in the SEG appearsreal. In the brain, the VS3-neurones terminatein the outermost layer of the ventro-lateralprotocerebrum in the form of a loop with somebranches running along the outer periphery of theganglion. The terminations of the VS3-neuronesdo not contact any of the central brain structuresshown (Fig. 3E, F).

V-neuronesThese neurones respond to vibrational stimula-

tion, but not to sound-stimuli at physiological in-tensities and may also be subdivided into fivetypes (Silver et al., ’80).

The type V1 neurone responded excitatorily tovibrational stimuli (n = 5). However, spontane-ous activity was inhibited after sound signals ofhigh intensity (Fig. 4A). Also vibration-stimuliinhibited the spontaneous activity. Spontaneousactivity was suppressed before and after the ex-citatory vibration response. The latency up tothe beginning of the inhibition (20 ± 2 ms SD)(measured up to the end of the last spontane-ous spike) was shorter than that of the excita-tory response to vibratory stimuli (52 ± 2 ms SD).The PSTHs show that the spontaneous activityreturned after 150 ms (Fig. 4A) and 300 ms (Fig.4B). These data demonstrate that the inhibitionoccurring before the excitatory response did notresult from poststimulus suppression of the pre-vious stimulus. The neurones did not habituateto repetitive stimuli. The responses of neuronesof this type are excitatory from 50 to 500 Hz (Fig.4C). Spontaneous activity was inhibited from 600to 2000 Hz.

In the SEG a few collaterals project towards themidline, but they do not cross it (Fig. 4D). In thebrain, terminations are located along the outerperiphery of the ventro-lateral protocerebrum. Onebranch describes a striking loop. A tiny collateralwas stained in the deutocerebrum. Although somebranches of these neurones lay in the same planeas the central body, none of the arborizations in-vade the brain structures shown (Fig. 4E, F).

The V2 type neurone has physiological data (n= 2) very similar to that of the V1-type (Fig. 5A–

C). If vibratory stimuli were presented, the spike-trains of this spontaneous active neurones showedtonic discharge patterns. Spontaneous activity wassuppressed before and after the excitatory vibra-tion response. The responses of the V2-neuroneshad a latency of 33 (±2 SD) ms to the beginningof the inhibition, or 61 (±2 SD) ms up to the firstspike (Fig. 5A). In contrast, vibration up to 2000Hz lead to excitatory responses instead of inhibi-tion (Fig. 5C).

The soma of the V2-neurone could not bestained, it is likely to be located posterior to theprothoracic ganglion. In the SEG, four tiny ipsi-lateral collaterals were stained. Two of them arelocated in the more caudal and two in the morefrontal part of the SEG. In the brain, the arboriza-tions project into the ventro-lateral protocerebrum(Fig. 5D). In relation to the central body andmushroom body the terminations lie more later-ally and caudally and do not overlap with thesebrain-structures (Fig. 5E, F).

Comparative morphologyThe identified interneurones in bushcrickets

show clear differences in the degree and locationof projections from their axons, that fall into twocategories. In the one category represented by theS-neurones there are projections in the SEG andin two locations (median deutocerebrum andventro-lateral protocerebrum) in the brain (Fig.6). In contrast, the other category represented bythe VS- and V-neurones have either no or littleprojections in the SEG and projections in the brainto a single location (median deutocerebrum) wherethese projections are more laterally placed.

The difference in the degree of the arborizationamong these two categories seen visually in thefigures, is also reflected in both the surface areaand volume measurements (Fig. 7). Thus the S-neurones have more extensive terminations thanthe VS- or V-neurones.

DISCUSSIONAlthough the morphology and physiology of au-

ditory and vibratory interneurones is known inthe prothoracic ganglion in a variety of bushcricketspecies (Rheinlaender and Kalmring, ’73; Kühneet al., ’80; Silver et al., ’80; Römer, ’87; Römer etal., ’88; Rheinlaender and Römer, ’86) their mor-phology in the SEG and the brain has been poorlyinvestigated (Kalmring et al., ’79; Shen, ’93). Thislack of knowledge is surprising as vibratory sig-nals are important in acoustic behaviour (Latimerand Schatral, ’83; Stiedl and Kalmring, ’89).

226 B. NEBELING

Fig. 4. Morphology and physiological data of a V1-neu-rone. Spike trains (upper part) and PSTH (2 ms bin width)to stimulation with (A) white noise (WN, 20 ms duration, 73dB SPL); (B) vibration (200 Hz, 100 ms duration, 2.7 m/s2);arrows indicate the suppression of spontaneous-activity. (C)Response magnitude to stimulation with vibration. (+) = tested

without excitatory effect. (D) Position and morphology withinthe head ganglia (TH1 was lost). Arrow indicates a branchdescribing a striking loop. (E, F) Reconstruction of theendbranches within the brain and relationship to prominentstructures; (E) frontal view, (F) sagittal view. Arrow see (D).

AUDITORY NEURONES IN THE BRAIN OF BUSHCRICKETS 227

Fig. 5. Morphology and physiological data of a V2-neu-rone. Spike trains (upper part) and PSTH (2 ms bin width)to stimulation with (A) vibration (800 Hz, 100 ms duration,2.7 m/s2); (B) vibration (100 Hz, 100 ms duration, 0.27 m/s2);arrows indicate the suppression of spontaneous activity. (C)

Response magnitude to stimulation with vibration. (+) = testedwithout excitatory effect. (D) Position and morphology withinthe head and prothoracic ganglia. (E, F) Reconstruction ofthe endbranches within the brain and relationship to promi-nent structures; (E) frontal view, (F) sagittal view.

228 B. NEBELING

Fig. 6. Positions of two representative neurones, one ofthe auditory (S1 = TN1) and one of the vibratory subsystem(V1) put together into one shape of the brain. For a bettercomparison, the V1-neurone was mirror-imaged.

Fig. 7. Size of the terminations of the neurone-types inthe brain. Areas are measured with the computer programAuto-CAD. Volumes are calculated by multiplication of theareas with the numbers and the thickness of the slides (20µm) in which terminations appear; relative units to be multi-plied by 102 (area/µm2), respectively, 103 (volume/µm3).

Cokl et al. (’77) suggested that informationabout airborne sound and substrate-borne vibra-tion may converge from the receptor cells to thesame auditory/vibratory interneurones. How thisconverged information may subdivide in the brainand SEG could be reflected in the morphology.

S-neuronesBased on their physiology, morphology, and

soma position the S1-neurone is found to be syn-onymous to the TN1 and the S4-neurone is syn-onymous to the AN2 in Tettigonia viridissima(Römer and Marquart, ’84; Römer et al., ’88). Thenames TN and AN were first given for a homolo-gous t-shaped neurone (TN) and an ascendingneurone (AN) by Wohlers and Huber (’82) in crick-ets. The present study brings together differentnomenclatures which so far have been used to de-scribe synonymous neurones either morphologi-cally or physiologically.

In the prothoracic ganglion staining of the AN2in various cricket and bushcricket species revealvery similar morphological features (Wiese, ’81;Römer and Marquart, ’84; Boyan, ’84; Römer etal., ’88; Mason and Schildberger, ’93). On the con-trary, the morphology of the AN2 in the brain ofDecticus albifrons differs from that of crickets(Boyan and Williams, ’82; Schildberger, ’84). Thetermination field of ascending interneuronesseems to be specific to the crickets, for ascendingauditory neurones of Locusta also project to thelateral protocerebrum (Eichendorf and Kalmring,’80; Boyan, ’83). In general, because of the simi-larities in the TH1 one may conclude that the den-dritic region of interneurones could reflect “old”morphological solutions, whereas in the brain thetermination fields may vary depending on the evo-lutionary necessities.

The present study supports the hypothesis thatoptical and auditory information may converge inthe optic stalk within the termination areas ofAN2 (S4) neurones. This pathway has alreadybeen suggested by Horridge (’64) in Acridids. Simi-larly, Eichendorf and Kalmring (’80) have reportedthat in Acridids auditory B1 neurones project intothe lateral horn, close to the optic lobes.

VS3-neuronesThe soma of a VS3-neurone was not stained in

any of the histological preparations, therefore nohomonymous morphological name can be given.Most likely the somata of these neurones are pos-terior to the prothoracic ganglion. Shen (’83)stained some giant interneurones with somata lo-

AUDITORY NEURONES IN THE BRAIN OF BUSHCRICKETS 229

cated in the terminal abdominal ganglion inTettigonia cantans and one of these neurones isvery similar in morphology to the VS3-neurone.Moreover, these giant interneurones respond towind input from the cerci of the same frequenciesused for vibratory stimulation in our work. As thecerci are seen to be homologous to legs, the giantinterneurones based in the abdominal ganglionmight be regarded as homologous to the VS3-neurones.

The VS3-interneurones may receive sensory in-put in the more caudal part of the CNS (e.g., themesothoracic or metathoracic ganglion) becauseof the relatively long latency (29 ± 2 ms) and thefact that there were no dendrites stained in theprothoracic ganglion. In this case information onairborne sound might have its input via anotherdescending interneurone to the VS3-neurone.

The fact that spontaneous activity is inhibitedby either sound or vibrational stimuli could be ex-plained by the influence of inhibitory inter-neurones. Stumpner and Ronacher (’91) found aninhibitory influence of airborne sound stimuli tospontaneous auditory neurones in Acridids, andSchul (’97) has reported inhibitory effects to theAN1 of Tettigonia viridissima.

V-neuronesFor both of the V-neurones, a soma could not

be located, although the axon of V2 stained in theprothoracic ganglion. Most likely the somata ofboth V-neurones are located in the more caudalpart of the CNS. In both neurones spontaneousactivity is inhibited even before the excitatory re-sponse occurs. One explanation could be that theseneurones receive input from inhibitory neuroneswith shorter latency than the excitatory input.

Comparison of auditory andvibrosensitive neurones

From the morphological characteristics revealedin this study it becomes obvious that the two au-ditory interneurones (S1, S4) differ from thevibrosensitive cells (VS, V). In the SEG, the S1and S4-neurones (TN1 and AN2) build extensiveterminations, whereas the VS3-, V1-, and V2-neurones have none or only weak arborizations(Figs. 1–5). Consequently, the influence of the VS-and V-neurones on the activity of higher orderinterneurones would be limited. On the otherhand, the SEG is known to receive the nerves in-nervating the mouthparts in its mandibular,maxillar and labiate neuromeres (Tyrer and Gre-gory, ’82). Also in Acridids and crickets, numer-

ous motoneurones innervating the neck-musclesare located in the SEG (Honegger et al., ’84). Sincethese animals protect themselves from predatorsby aggressive biting, there is likely to be a corre-lation between the behavioural element of defenceand the activation of the mouthparts. However,whether that the S-neurones are involved in thesebehaviour is still to be investigated, although ear-lier investigations found the S1-neurone being re-lated with the fight- or flight-reaction (McKay, ’70;Kalmring et al., ’79).

The S-neurones build more extensive termina-tions than the VS and V-neurones do in the brainof bushcrickets (Fig. 7). The bigger spatial repre-sentation of the auditory interneurones may rep-resent larger numbers of synaptic contacts onhigher order interneurones, and this may compen-sate for the smaller number of auditory receptorcells. Moreover, both S-neurones build a secondarborization area in the median deutocerebrum,which is missing in all V- or VS-neurones. Thelatter neurones also terminate more laterally inthe brain than the S-neurones. The collective dif-ferences between the S-neurones and the VS-, V-neurones dealing respectively with auditory andvibratory stimuli suggest that these two sensorymodalities are separated by different locations ofthe neuronal terminations in the brain and SEG.

ACKNOWLEDGMENTSThe author thanks Prof. Dr. K. Kalmring for

supporting this work and Dr. T. Sickmann for criti-cally reading the concept. C. Meyer improved theEnglish text. I am also grateful to Drs. W. Rössler,M. Jatho, and J. Schul for the friendly gift of thecomputer software for analytical purposes. Theauthor thanks Prof. Dr. U. Homberg and M.Müller for assistance with the antibody-procedure,and Mrs. S. Völk and H. Brandtner for technicalhelp. Dr. R. Lakes-Harlan and Prof. Dr. Schild-berger introduced me to intracellular recordings.

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