David W. Wohlers

Processing of sound signals by six types of neurons in the prothoracic ganglion of the cricket,Gryllus campestris L.

Summary1.Six types of auditory neurons, arranged as mirror-image pairs, were characterized intracellularly within the prothoracic ganglion.2.Two of the six neuron types are restricted to the prothoracic ganglion and exhibit an omegashaped structure (Fig. 2, ON1, ON2). Two other neuron types have axons ascending to the head ganglion (Fig. 2, AN1, AN2). A neuron with a descending axon (Fig. 2, DN1) and a T-shaped neuron (Fig. 2, TN) having both an ascending and a descending axon were also characterized.3.Physiologically, these neuron types are characterized by their suprathreshold responses in the range of 2–20 kHz (Fig. 3), by the time course of epsps (Figs. 4 and 5), and by their responses related to monaural and binaural input (Fig. 6). ON1 is excited by the ear ipsilateral to the cell body and inhibited from the opposite ear (Fig. 6). AN1 and DN1 receive only excitatory input from the ear contralateral to the cell body (Fig. 6). ON2 and TN receive excitatory input from both ears (Fig. 6). Ipsilateral stimulation of AN2 at the calling song frequency results in a response ranging from weak excitation to strong inhibition; contralateral stimulation at moderate and high intensities is always excitatory, but can exhibit mixed excitation and inhibition (Figs. 6 and 7).4.Artificial calling songs excite all six types of neurons. ON1, AN1, and DN1 copy both syllables and pauses more precisely than ON2, AN2, and TN.5.All six neuron types copy the temporal sequence of calling songs containing syllables of 2.5 ms duration. Intersyllable pauses smaller than 5 ms are not copied by any of the neuron types (Fig. 8).6.Artificial calling songs with a duty cycle of 50% (equal duration of syllable and pause) are phonotactically effective only when syllable intervals lie within the range of 20–60 ms. None of the neurons reported here are tuned specifically to this range (Fig. 9).

Intracellular-recording and staining of cricket auditory interneurons (gryllus-campestris l, gryllus-bimaculatus degeer)

SummaryIntracellular recording and staining techniques have been used to investigate physiological and anatomical properties of two kinds of auditory interneurons in the prothoracic ganglion of the crickets,Gryllus campestris L. andGryllus bimaculatus DeGeer: a segmental interneuron (the omega cell) and an auditory interneuron with an ascending axon (AIAA).Using cobalt-nitrate-filled electrodes we were able to study physiological features for periods as long as 10 min following penetration and simultaneously stain the cell. With potassium-acetate electrodes, similar intracellular recordings lasted longer than an hour.1.The omega cell is a large segmental auditory neuron with four branching areas (fields A, B, C, D). Two such cells are found in the prothoracic ganglion, as mirror images of one another.2.The omega cell receives its auditory input entirely from the cell-body side. EPSPs as well as spike activity are seen in the large dendritic root of field A; the spikes are conducted to the contralateral side of the neuron and invade fields C and D, which are considered to be output areas of the neuron.3.When the two ears are isolated acoustically, allowing separate stimulation of the tympana, it appears that the two omega cells inhibit one another mutually. They may, therefore, participate in the neuronal mechanism involved in directional sensitivity.4.The omega cell is best tuned to the carrier frequency of the conspecific song, but exhibits a secondary threshold minimum in the region of the higher harmonics. The response is linearly related to the log of the sound intensity, as found in the auditory afferents.5.The temporal pattern of the cricket calling song is well copied by the omega cell. No optimal “tuning” to specific syllable durations, syllable periods or verse durations was observed. Maintained tones elicit a tonic response and sound envelopes are mimicked by both subthreshold and spike-activity.6.The auditory interneuron with ascending axon (AIAA) has a dendritic field and ascending axon restricted almost entirely to one hemisphere of the prothoracic ganglion; the cell body is located in the anterior contralateral hemisphere.7.The AIAA exhibits IPSPs highly synchronized in response to sounds presented at the carrier frequency. At sound frequencies above 10 kHz the AIAA is activated, but the responses to the syllables of the calling song are not as sharply separated as in the omega cell.8.Latencies of AIAA spike activity are comparable to those in the omega cell and suggest that both cells are first order interneurons.9.The frequency-dependence of excitation and inhibition in the AIAA make it a candidate element of a neuronal process that might modify response to the calling song in the presence of the courtship song.

Primary auditory neurons in crickets - physiology and central projections

Summary1.The tibial tympanal organs ofGryllus campestris L. andGryllus bimaculatus DeGeer were stimulated with sound signals simulating the natural calling song, produced by sinusoidal amplitude envelope, using carrier frequencies (CF) in the range of 2–20 kHz at various intensities.2.Glass microelectrodes (filled with 3 M potassium acetate, or with 1 M cobalt nitrate for marking the fibers) were used to record extracellularly or quasi intracellularly the spike activity of single sensory fibers in the leg nerve (Fig. 1) responding to acoustic stimuli and to stain their central projections within the prothoracic ganglion (Figs. 6–9).3.The temporal pattern of the sound stimulus —syllable and verse structure — is reflected in the discharge pattern of the fibers studied (Figs. 1, 6, 7).4.Five types of fibers tuned to specific CFs were found (Fig. 2). These had peak sensitivities (a) at 4–5 kHz (CF of the species calling song), (b) at both 4–5 and 10–12 kHz, (c) below 2 kHZ, (d) at 12 kHz, and (e) at 17 kHz (CF of the species courtship song).5.The tuning is also evident in the responsiveness (spikes/syllable) to stimuli of different frequencies at suprathreshold intensities (Fig. 3).6.In the range of their best frequencies (BF) the fibers tuned to 4–5 and to 10–12 kHz are similar in intensity characteristics and in relationship of latencies to sound intensity, with a linear relationship between responsiveness and latency (Fig. 4). Within a single calling-song verse, the spike response decreases, and the latency increases, from the first to the fourth syllable (Fig. 5). Even at the highest intensities the discharge rate nerver exceeds 350 Hz.7.The central arborization and projection areas of all the marked fibers are restricted to the ipsilateral half of the prothoracic ganglion. Assuming that fibers marked include those recorded physiologically, it appears that those tuned to 4–5 kHz terminate exclusively in the “crescent-shaped” neuropile, localized and termed the “auditory neuropile” (Figs. 6, 7). Fibers with a peak sensitivity below 2 kHz differ distinctly from the 4–5 kHz group in that they have additional arborizations outside the auditory neuropile (Fig. 8). As yet no unequivocal data are available regarding the spatial arrangement of the fibers with two sensitivity peaks (both 4–5 and 10–12 kHz) and those tuned to 12 kHz or 17 kHz (Fig. 9).

Lucifer Yellow Histology

Walter W. Stewart discovered and synthesized the fluorescent dye, Lucifer yellow, based on the commercial dye, brilliant sulphoflavine (Stewart 1978, 1981). Lucifer yellow has since become the most popular and widely used intracellular marker, largely replacing its forerunner, Procion yellow 4RAN, for identifying physiologically recorded neurons.

Topographical organization of the auditory pathway within the prothoracic ganglion of the cricket Gryllus campestris L.

SummaryThe topographical organization of the prothoracic ganglion of the cricket, Gryllus campestris L., is described from horizontal, transverse, and sagittal sections of preparations specially treated to elucidate longitudinal tracts, commissures, and areas of neuropil. These structures were compared to those reported from other insect thoracic ganglia, resulting in still further evidence for a common basic morphological pattern among insect central nervous systems.Six types of auditory interneurons, all existing as mirrorimage pairs, were identified through intracellular application of the dye Lucifer yellow, and then related to several morphological patterns. Two intrasegmental neurons (ON1, ON2) are similar in location of cell bodies and course of neurites and axons; three intersegmental neurons (AN1, AN2, TN1) are likewise similar to one another. The axons of the two intrasegmental neurons cross the midline of the ganglion in the newly described ‘omega commissure’. Axons of the other four types all course within the median portion of the ventral intermediate tract and project intersegmentally.All six neuron types arborize within the ventral portion of the ring tract, the same neuropilar region in which auditory sensory neurons terminate. The ring tract is therefore considered the most important region for auditory information processing within the cricket prothoracic ganglion.

Morphological and physiological changes in central auditory neurons following unilateral foreleg amputation in larval crickets

Summary1.In view of the surprising recent demonstration that developmentally one-eared female crickets can track sound sources (Huber et al. 1984), we have looked for correlates in the morphological and physiological properties of auditory interneurons of these animals. One foreleg was amputated in the 3rd/4th or penultimate (8/9th) larval instar; in both cases the leg regenerated without developing a functional ear. In the adult stage, these animals were studied first for their phonotactic behavior and then by intracellular recording and staining; three types of auditory interneurons in the prothoracic ganglion were identified: the omega neuron ON1, and the ascending neurons AN1 and AN2.2.Of these three neuron types, those that normally receive excitatory input from the side now deafferented send dendrites across the midline of the ganglion, along specific pathways, to end in the auditory neuropil of the intact side (Figs. 1–4).3.The new connections are functional, as shown by the responses of the neurons to synthesized calling songs presented to the remaining ear. With respect to the copying of chirp structure, threshold curves and intensity characteristics, these neurons respond like cells in intact animals that are presented with the same stimulus on the side ipsilateral to the main input region of the neurons (Figs. 2–4). The implication is that in animals with one ear missing, functional pathways within the central nervous system are reorganized, resulting in better orientation of one-eared animals.

Analysis of the Cricket Auditory System by Acoustic Stimulation Using a Closed Sound Field

SummaryA closed sound field system for independent stimulation of both cricket hearing organs is described. The system was used to measure acoustic parameters of the peripheral auditory system inGryllus campestris and to analyze inhibitory responses of the omega cell, a segmental auditory interneuron in the prothoracic ganglion.1.Best sound transmission in the tracheal pathway occurs at 5 kHz. Closing of the prothoracic spiracles results in increased sound transmission but does not influence the frequency of best transmission in most animals (Fig. 6 B). Sound transmission is modulated by abdominal contractions associated with the respiratory cycle (Fig. 7).2.AttenuationΔ and phase shift ϕ in the tracheal pathway have been determined for the frequency range of 2 to 10 kHz in animals with closed spiracles.Δ shows a minimum at 5 kHz and ϕ increases almost linearly with frequency (Fig. 11).3.Sound components acting on each side of the large tympanal membrane form a resultant sound pressure based on linear superposition. This resultant sound pressure represents the effective stimulus of the auditory sense organ (Fig. 12).4.The response of the omega cell is dependent upon both intensity and relative phase of sound signals applied to the tympanal membranes (Fig. 10).5.At 5 kHz, the response of the omega cell decreases linearly with increasing contralateral (inhibitory) stimulus intensity over a wide range of intensities. The latency between stimulus onset and response is nearly independent of contralateral inhibition (Figs. 15 and 16).6.Response (spike number) differences between an omega cell and its complementary mirror image cell due to different stimulus intensities at both ears are enhanced by the neuronal mechanism of contralateral inhibition. In one animal the gain in spike number difference at 5 kHz was calculated to be 60% relative to the response difference when contralateral inhibition was disabled.7.Evidence for a low frequency (f≦2 kHz) ipsilateral inhibition of the omega cell is presented (Fig. 17).

Central projections of fibers in the auditory and tensor nerves of cicadas (homoptera, cicadidae)

SummaryThe auditory and tensor nerves of cicadas are mixed nerves containing both afferent and efferent elements. In 17-year cicadas, and in Okanagana rimosa, the auditory nerve contains afferents from body hairs, from the detensor tympani-chordotonal organ, and some 1300–1500 afferents from the hearing organ. Within the fused metathoracic-abdominal ganglionic complex the receptors from both the auditory and tensor nerves form a neuropilar structure that reveals the metameric organization of this complex. A few fibers run anteriorly, projecting into the meso and prothoracic ganglia. Within the ganglionic complex a division of auditory nerve afferents into a dense intermediate and a more diffuse ventral neuropile is observed. In addition, a dorsal motor neuropile is outlined by arborizations of the timbal motor neuron. This neuron is one of several efferent cell types associated with the auditory nerve, and there is an indication that several efferent fibers innervate the timbal muscle. There is anatomical evidence for a possible neuronal coupling between the bilaterally symmetrical large timbal motor neurons. In general, central projections from the auditory and tensor nerves support evidence of a structural “layering” within the CNS of insects.

Tympanal membrane motion is necessary for hearing in crickets

Summary1.Sound guided through the tracheal tube to the internal tracheal spaces in the region of the cricket ear is capable of eliciting auditory neural responses in the prothoracic ganglion if the tympanal membrane is allowed to vibrate freely. If the tympanal membrane motion is prevented mechanically neural responses are abolished (Fig. 3) whereas the sound pressure in the tracheal air spaces behind the tympanum is increased.2.If the motion of the tympanum, as measured with laser vibrometry, is prevented by adjusting the internal and external sound pressure, then neural responses cease simultaneously (Fig. 5).3.These findings demonstrate that motion of the large tympanum is a necessary requisite in the sound transduction process of the cricket ear.

Auditory nerve and interneurone responses to natural sounds in several species of cicadas

ABSTRACT. The calling and courtship songs of 17‐year cicadas and of Says cicadas differ both in the sound frequency spectrum and in temporal pattern. Multiunit recordings with hook electrodes from the whole auditory nerve show that the hearing organs are especially sensitive to transient stimuli occurring in natural sounds. Artificially produced clicks elicit bursts of spikes synchronized among various primary sensory fibres. These fibres respond to natural calling and courtship songs with a specificity dependent on carrier frequency, rhythm and transient content of the sound, following sound pulses (i.e. tymbal actions) up to repetition rates of 200 Hz. An ascending, plurisegmental interneurone was characterized by intracellular recording and simultaneously stained with cobalt. Its main arborization spatially overlaps the anterior part of the sensory auditory neuropile, and the axon was traced as far as the prothoracic ganglion. Direct input from primary auditory fibres was suggested by latency measurements. Intracellular recordings from such neurons in different species show distinct auditory input, with phasic‐tonic spike responses to tones. In general, the interneurone response is more species‐specific to calling than to courtship songs, and the preferential response to the conspecific calling song is based primarily upon sound frequency content.