Physiology of the auditory afferents in an acoustic parasitoid fly

Abstract

The fly, Ormia ochracea, possess a novel auditory organ, which allows it to detect airborne sounds. The mechanical coupling of its pair of tympanal membranes provides the basis for a unique means of sensing the direction of a sound source. In this study, we characterized the neuroanatomy, frequency tuning, and neurophysiological response properties of the acoustic afferents. Our experiments demonstrate that the fly's nervous system is able to encode and localize the direction of a sound source, although the binaural auditory cues available in the acoustic sound field are miniscule. Almost all of the acoustic afferents recorded in this study responded to short and long sound pulses with a phasic burst of one to four action potentials. A few afferents responded tonically for the duration of the sound stimulus. A prominent class of afferents responds to suprathreshold stimuli with only a single spike discharge, independent of stimulus level, frequency, or duration. We also tested the response of the afferents to speakers separated by 180 degrees along the azimuth of the fly. We found that the afferent responses have a shorter latency because of ipsilateral stimulation. This could be a temporal code of the direction of a sound source. The threshold frequency tuning for the afferents revealed a range of sensitivities to the frequency of the cricket host's calling song frequency. The difference in the number of afferents above threshold on either side of the animal is a population code, which can also be used for sound localization.

Fig. 1.

Anatomy of the acoustic afferents of the right bulba acustica in Ormiaochracea.A, This is a dorsal view of az-projection of 28, 1-μm-thick confocal optical sections. Texas Red dextran dye was applied to the bulba acustica, and >50 afferents were stained. Stained afferents project to three acoustic neuropil regions in the ventral region of the three neuromeres in the fused thoracic ganglion of the fly. B, A single auditory afferent stained with fluorescent dextrans. There is a dense projection into the T2 auditory neuropil area from this afferent. The T1 and T3 neuropil regions receive minimal projections from this afferent. C, A single auditory afferent stained with fluorescent Texas Red dextrans. There are dense projections into the T1 and T2 auditory neuropil area from this afferent. The T3 neuropil region does not receive any projections from this afferent.D, A single auditory afferent stained by Lucifer yellow injection. A single branch projects into the T1 auditory area from the primary neurite. This branch then continues projecting to the T2 auditory neuromere. There is a separate projection from the primary neurite into the T2 auditory area and a minimal projection to the T3 auditory area. AMN, Accessory mesothoracic neuromere;CN, cervical connective; FN, frontal nerve; HN, haltere nerve; T1, prothoracic neuromere; T2, mesothoracic neuromere;T3, metathoracic neuromere; VE, ventral ellipse; WN, wing nerve.

Fig. 2.

Summary of the frequency threshold response in the auditory afferents of Ormia ochracea. A,Average ± SDs for 28 afferents (type 1, type 2, and type 3) obtained with 10 msec pulses repeated at 1 Hz. B,Histogram of the number of afferents with best frequencies at a particular frequency. These data were compiled from the same tuning curves that are averaged in A. C,Comparison of the average tuning of the afferents in this study (solid line, left ordinate) with the membrane displacement measured with laser vibrometry measured in relative decibels (dotted line, right ordinate). The membrane response amplitude was measured with laser vibrometry, using a broad band white noise stimulus between 1 and 25 kHz (Robert et al., 1996).

Fig. 3.

A, Examples of three tuning curves for afferents with best frequencies <10 kHz. B,Examples of three tuning curves for afferents with best frequencies >10 kHz.

Fig. 4.

Physiological recordings of a type 1 phasic acoustic afferent. A, Response of a type 1 afferent to synthetic cricket song (50 pps, 5 kHz, 50% duty cycle, 85 dB SPL). Calibration: 2 mV, 10 msec. B, Phasic response of a type 1 afferent to a 100 msec sound pulse at its BF, 7 kHz at 80 dB SPL. Calibration: same as in A.C, Raster and PST histogram of a type 1 afferent in response to the repetition of the 100 msec stimulus at 1 Hz. Very little variation can be seen in the latency of the spike at this resolution (bin width, 20 μsec).

Fig. 5.

The proportion of phasic afferent spikes to sound pulses (10 msec, 5 kHz, 85 dB SPL) presented with varying interpulse intervals. Because the interpulse interval increases the phasic afferent is able to reset before the next sound pulse, so there is a higher proportion of responses. The time constant (τ) for the refractory period of this phasic afferent is defined as the interpulse interval that will allow the afferent to respond to 63% of the sound pulses.

Fig. 6.

Physiological recordings of a type 2 phasic acoustic afferent. A, Response to synthetic cricket song (50 pps, 4 kHz, 50% duty cycle, 95 dB SPL). Calibration: 2 mV, 10 msec. B, Phasic response to a 100 msec sound pulse at the BF of the cell, 5 kHz at 85 dB SPL. C, Raster and PST histogram of a type 2 afferent in response to the repetition of the 100 msec stimulus at 1 Hz. Four distinct clusters of spikes are seen in the response to this 100 msec sound pulse (bin width, 20 μsec).

Fig. 7.

Physiological recordings of a type 3 tonic acoustic afferent. A, Response to a single 10 msec cricket-like sound pulse (1 msec rise–fall, 5 kHz, 85 dB SPL). Calibration: 2 mV, 10 msec. B, Tonic response for the duration of the 100 msec at its BF, 7 kHz at 85 dB SPL. Calibration: 2 mV, 10 msec. C, Raster and PST histogram of a type 3 afferent in response to the repetition of the 100 msec stimulus at 1 Hz. The latency for the first two spikes is relatively invariant compared with the timing of the subsequent spikes during the rest of the stimulus (bin width, 20 μsec).

Fig. 8.

Direction-dependent physiology of the type 1 phasic afferents. A, There is a latency shift of the spikes elicited by stimuli of different intensities. In this experiment, sound was presented from 95–75 dB SPL in −5 dB steps. Calibration: 1 mV, 1 msec. B, There is a latency difference in the spikes elicited by sound on either side of the animal. This is caused by a decrease in the amplitude of the movement of the tympanal membrane contralateral to a sound stimulus. Thedashed reference line shows that a 10 dB difference in intensity would produce a latency shift consistent with the difference in the latency caused by ipsilateral versus contralateral stimulation.

Fig. 9.

Range fractionation of the all the afferent types. The cumulative histogram of the proportion of afferents above threshold in response to a 5 kHz stimulus at various intensities. The proportion of afferents above threshold on the contralateral side of the animal will be less because of the amplitude decrease in the movement of the contralateral tympanal membrane caused by the mechanical coupling of the tympana. The lines show the relative proportion of the afferents that would be above threshold in response to an 85 dB SPL ipsilateral stimulus. The contralateral side of the animal will be subject to a 10 dB attenuation of the stimulus, therefore 55% fewer afferents will be above threshold on that side.