J Comp PhysiolA (1990) 166:721-734

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9 Springer-Verlag1990

Neural coding in the chick cochlear nucleus Mark E. Warchol* and Peter Dallos Auditory PhysiologyLaboratoryand Department of Neurobiologyand Physiology,Northwestern University, Francis Searle Bldg., 2299 Sheridan Rd., Evanston, IL 60208, USA Accepted October 5, 1989

Summary. Physiological recordings were made from single units in the two divisions of the chick cochlear nucleus - nucleus angularis (NA) and nucleus magnocellularis (NM). Sound evoked responses were obtained in an effort to quantify functional differences between the two nuclei. In particular, it was of interest to determine if nucleus angularis and magnocellularis code for separate features of sound stimuli, such as temporal and intensity information. The principal findings are: 1. Spontaneous activity patterns in the two nuclei are very different. Neurons in nucleus angularis tend to have low spontaneous discharge rates while magnocellular units have high levels of spontaneous firing. 2. Frequency tuning curves recorded in both nuclei are similar in form, although the best thresholds of NA units are about 10 dB more sensitive than their NM counterparts across the entire frequency range. A wide spread of neural thresholds is evident in both NA and NM. 3. Large driven increases in discharge rate are seen in both NA and NM. Rate intensity functions from NM units are all monotonic, while a substantial percentage (22%) of NA units respond to increased sound level in a nonmonotonic fashion. 4. Most NA units with characteristic frequencies (CF) above 1000 Hz respond to sound stimuli at CF as 'choppers', while units with CF's below 1000 Hz are 'primary-like'. Several 'onset' units are also seen in NA. In contrast, all NM units show 'primary-like' response. 5. Units in both nuclei with CF's below 1000 Hz show strong neural phase-locking to stimuli at their CF. Above 1000 Hz, few NA units are phase-locked, while phase-locking in NM extends to 2000 Hz. 6. These results are discussed with reference to the hypothesis that NM initiates a neural pathway which Abbreviations: A V C N anteroventral cochlear nucleus; C F characteristic frequency; D C N dorsal cochlear nucleus; N A nucleus angularis; N M nucleus magnocellularis; P S T post-stimulus-time; P V C N posteroventral cochlear nucleus; S R spontaneous rate * Present address: Department of Neuroscience, Box 396, University of Virginia School of Medicine, Charlottesville, Va 22908, USA

codes temporal information while NA is involved primarily with intensity coding, similar in principle to the segregation of function seen in the cochlear nucleus of the barn owl (Sullivan and Konishi 1984). Key words: Hearing - Auditory system - Cochlear nucleus - Chicken

Introduction The auditory nerve provides a relatively homogeneous set of neural response types to the first auditory brainstem center, the cochlear nucleus. Yet, single unit recordings from the various divisions of the cochlear nucleus in higher vertebrates reveal a wide diversity of response classes. Corresponding anatomical observations from this region point to a similar pattern. Differences are found in morphology of cell types both within a given region of the cochlear nucleus and between different divisions. Also, individual divisions in the cochlear nucleus give rise to ascending projections which terminate in differing higher centers within the brainstem and midbrain. Thus, a likely function of the cochlear nucleus appears to be the initiation of separate parallel pathways for the processing of different features of auditory stimuli. Determining the functions of these separate pathways remains an important goal in the study of auditory physiology. The animal species in which they are perhaps best understood is the barn owl. The cochlear nucleus of the owl, as in all birds, is composed of two distinct divisions: nucleus angularis and nucleus magnocellularis. Combined anatomical, physiological, and behavioral studies with this species reveal that nucleus angularis is the beginning of a pathway involved in the coding of sound intensity while nucleus magnocellularis initiates a pathway given to processing the temporal features of a sound stimulus (Sullivan and Konishi 1984; Takahashi et al. i 984). Information from these two pathways converges in the auditory midbrain, where it is used to syn-

722

M.E. Warchol and P. Dallos: Neural coding in the chick cochlear nucleus

thesize a n e u r a l t o p o g r a p h i c a l m a p o f a u d i t o r y space ( K n u d s e n a n d K o n i s h i 1978). A l t h o u g h the n e u r a l c o n n e c t i o n s a m o n g the c e n t r a l a u d i t o r y nuclei o f the b a r n owl a r e similar to those in o t h e r a v i a n species, m a n y details o f the o w l ' s c e n t r a l a u d i t o r y s y s t e m a p p e a r quite specialized. A l s o , the o w l ' s u n i q u e e x t e r n a l a u d i t o r y features (facial r u f f a n d a s y m m e t r i c a l e a r o p e n i n g s ) m a y a l l o w it to use a c o u s t i c cues differently f r o m o t h e r a v i a n species. M o s t b i r d s possess small h e a d d i a m e t e r s a n d s y m m e t r i c a l e a r o p e n i n g s , a n d m u s t t h e r e f o r e use s h o r t i n t e r a u r a l t i m i n g differences a n d small i n t e n s i t y differences ( b o t h o f w h i c h are enh a n c e d b y m e a n s o f a n air filled p a t h w a y t h r o u g h the skull c o n n e c t i n g the two m i d d l e e a r spaces - see Lewis a n d Coles 1980; C a l f o r d a n d P i d d i n g t o n 1988). Thus, it is n o t i m p l a u s i b l e to expect t h a t different p h y s i o l o g i c a l s p e c i a l i z a t i o n s will be f o u n d in the r e s p o n s e s o f n e u r o n s in the a u d i t o r y b r a i n s t e m nuclei. O n the o t h e r h a n d , it is e q u a l l y p o s s i b l e t h a t the s e g r e g a t i o n o f t e m p o r a l a n d intensity i n f o r m a t i o n f o u n d in the b a r n owl a u d i t o r y b r a i n s t e m is a c o m m o n f e a t u r e o f the a v i a n a u d i t o r y system. We have r e c o r d e d f r o m the t w o divisions o f the cochlear nucleus in the d o m e s t i c c h i c k to d e t e r m i n e h o w t i m ing a n d intensity i n f o r m a t i o n is c o d e d a n d c o n v e y e d to higher a u d i t o r y nuclei. N u c l e u s m a g n o c e l l u l a r i s a n d a n g u l a r i s also h a v e h o m o l o g u e s in the m a m m a l i a n a u d i t o r y system. N u c l e u s m a g n o c e l l u l a r i s is c o n s i d e r e d h o m o l o g o u s to the s p h e r i c a l o r b u s y cell r e g i o n o f the a n t e r o v e n t r a l c o c h l e a r nucleus ( J h a v a r i a n d M o r e s t 1982). T h e precise h o m o l o g u e o f nucleus a n g u l a r i s is less certain, b u t includes the p o s t e r o v e n t r a l c o c h l e a r nucleus a n d p e r h a p s the d o r s a l c o c h l e a r nucleus as well ( B o o r d 1969). W h i l e e v o l u t i o n a r y r e l a t i o n s h i p s m u s t ult i m a t e l y be a s c r i b e d on m o r p h o l o g i c a l g r o u n d s , k n o w l edge o f the r e s p o n s e types p r e s e n t in the divisions o f the a v i a n c o c h l e a r nucleus c a n p o i n t t o w a r d f u n c t i o n a l equivalences b e t w e e n the a v i a n c o c h l e a r nuclei a n d their mammalian counterparts.

Me~o~ Animals. Male domestic chicks (Gallus domesticus, white leghorn strain) were used in all experiments. They were obtained as day-old hatchlings from local suppliers and raised in brooders. Animals used in experiments were age P10-P28 (approximately 80-150 g). Surgical preparation. Animals were anesthetized with a single dose of pentobarbital (20 mg/kg) followed by ketamine (80 mg/kg). Anesthesia was maintained throughout the course of an experiment (4-12 h) on ketamine alone, with supplements given as needed. Cloacal temperature was monitored and maintained at 40-41 ~ The trachea was cannulated but the animals were not artificially ventilated. A patch of skin covering most of the top of the skull was removed and the bone cleared and dried. A metal rod was cemented to the top of the skull using cyanoacrylate followed by dental cement. This rod was used to fix the head rigidly, in a natural upright position, into a specially designed headholder. Further head stabilization was provided by two ear-bar supports, one of which contained the acoustic driver (described below).

A piece of covering most of the right dorsal hemisphere of the cerebellum was removed. Care was taken not to disturb the large venous sinus running along the dorsal midline, which was left unligated throughout the experiment. A slit was cut in the dura overlying the cerebellum to allow electrode penetration. Far lateral openings provided access to nucleus angularis while more medial openings were used to reach nucleus magnocellularis. The cerebellum was not aspirated or moved and no further surgery was performed.

Sound stimulus. Sound was generated with a Beyer DT-48 driver and delivered to the ear canal through a closed acoustic system. Stimulus sound levels were calibrated at the beginning of every experiment using a concentrically mounted probe-tube attached to a Knowles BT-1751 microphone. Typical maximum intensities were approximately 110-120 dB SPL across the frequency range studied (100-8000 Hz). Acoustical isolation. Sound calibration and neural recordings were carried out with the animal placed in a sound isolation booth (Industrial Acoustics). To reduce vibration inside the sound booth, the animal was mounted on an inertial table resting on piston air springs. Recording. Neural recordings were made with micropipette electrodes filled with 2 M NaC1 (tips: 0.5-1.0 ~m, impedance: 510 M~). Extracellular spikes were processed by a WPI 7000 series electrometer followed by 1000 • amplification. Spikes of a given amplitude could be selected by setting the trigger level of a custombuilt discriminator. Electrodes were positioned over the appropriate location of the cerebellum and advanced ventrally in approximately the dorsal-ventral plane. The auditory nuclei were reached at depths of 4500-6500 ~m, depending on the age and size of the bird. Units in both nuclei have large biphasic spikes and are found in a distinctive tonotopic arrangement (Konishi 1970). In any given animal, recordings were made from only a single nucleus. Auditory units were detected by noting responses to wide-band noise bursts (100 ms bursts, repeated 3/s) of approximately 80 dB SPL. An effort was also made to observe units within the cochlear nucleus whose firing was either unaffected or suppressed by the noise stimuli. After isolation of an auditory unit, a tuning curve was obtained using an automated online procedure. Stimulus frequencies (approximately 12 points/octave, 100-8000 Hz) are presented in ascending order. At a given frequency, neural threshold is determined as follows: Tone bursts (100 ms, 2.5 ms rise/fall time) are presented at a rate of 3/s. A spike count is made during a 50 ms period beginning 10 ms after the start of the tone burst. This count is compared with the number of spikes detected during a time period of equal length situated between tone burst presentations and beginning at least 20 ms after the conclusion of the previous tone burst (to eliminate effects of postmasking). Sound intensity is varied until the level that produces an increase of 2 spikes in the 50 ms sampling period during the tone burst is reached (relative to the spontaneous rate recorded between tone bursts). This sound level is defined as the neural threshold and is equivalent to the intensity which results in a neural discharge rate increase of 40 spikes/s. After a complete tuning curve has been obtained, a rate-intensity function is collected at the frequency of lowest threshold (characteristic frequency or CF), over an intensity range spanning from about 15-20 dB below neural threshold to about 115-120 dB SPL, divided into 5 dB steps. These sound levels are presented in random order. Tone bursts are repeated at a rate of 1/s and the sustained discharge rate (average firing rate during the last 80 ms of the 100 ms tone burst) is recorded. Each sound level is presented 5 times. At any intensity level, the reported discharge rate is the average value from these 5 successive presentations. Classification of a unit's response type and assessment of its phase-locking capabilities is carried out by collecting a post-stimu-

M.E. Warchol and P. Dallos: Neural coding in the chick cochlear nucleus lus-time (PST) histogram using a stimulus frequency equal to the unit's CF with a sound pressure 20 dB above threshold. Spikes are collected over 100 presentations (I/s) of tune bursts whose onset phase (positive going zero crossing at the stimulus generation site) is identical. The resulting histogram is classified by reference to types observed in previous studies of the cochlear nucleus in higher vertebrates (primary-like, chopper, onset etc. - Pfeiffer 1966). Neural phase-locking is quantified by computing a vector strength value (Goldberg and Brown 1969) for this histogram at the stimulus frequency. Histograms of both spontaneous activity and tone evoked activity at other stimulus levels were collected for many units as holding time permitted.

Histology. Verification of recording sites was carried out in many animals after all physiological recording was completed9 Electrodes filled with fast green dye dissolved in saline (Woolf 1980) were positioned near the location of previous tracks and lowered into the appropriate nucleus. The placement of the electrode tip could then be verified by recording an evoked multi-unit response ('neurophonic') to sound stimuli. The dye was deposited either by current injection (1-10 ~tA negative current) or by passive diffusion from large-tip electrodes. In all cases, the markings could be localized to a single nucleus (angularis or magnocellularis). Results

Results presented here are based on recordings from 103 units localized to nucleus angularis and 81 units from nucleus magnocellularis. In any individual animal, recordings were made from only a single nucleus. The animals used were 10-28 days post-hatch (P10-P28). No age related differences in any response property f r o m either nucleus were noted. Thus, the chick cochlear nucleus appears physiologically mature by age P10, in agreement with anatomical observations (Rubel and Parks 1988). The various types of response parameters are described separately.

Spontaneous activity. The profile of spontaneous rate (SR) differs substantially for the two nuclei. The average SR o f units isolated in nucleus angularis is 25.3 spikes/s (103 units, range 0-84.0 spikes/s). A histogram showing the distribution of SR's for the N A unit population is shown in Fig. 1 A. While this distribution can be described as unimodal, it is highly skewed toward the lower SR's. Most N A units have SRs of less than 20 spikes/s and m a n y have no spontaneous discharge. Spontaneous activity in nucleus magnocellularis follows a very different pattern from that seen in NA. The distribution of SRs across the N M unit population is shown in Fig. 1 B. The mean SR is 94.3 spikes/s (81 units, range 31.3-231.2 spikes/s). No magnocellular units have spontaneous rates below 31 spikes/s. Further, the mean spontaneous rate in N M is higher than the maxim u m SR found in N A units. The pattern of distribution of N M rates is quite wide and unimodal although slightly skewed toward lower SR's. Tuning properties. Tuning curves were obtained for all units described here in both nuclei. The frequency at which the neural threshold is lowest is defined as the characteristic frequency or CF. Unit thresholds at CF

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are widely scattered in both nuclei. The distribution of neural thresholds for N A units is shown in Fig. 2. The range of thresholds at most frequencies is 45-55 dB. A similarly wide spread is often seen in unit thresholds obtained from individual animals. The very lowest thresholds agree well with the published behavioral thresholds for the chick at these frequencies (Gray and Rubel 1984). Neural threshold does not appear to be related to spontaneous rate. Neurons with both high and low thresholds are found to have spontaneous rates of either near zero or exceeding 80 spikes/s. The distribution of thresholds for N M neurons as a function of CF is shown in Fig. 3. While the number of units with C F ' s below 800 Hz is limited, several trends are evident when this population is c o m p a r e d to that from NA. First, the lowest thresholds of N M neurons are about 10 dB higher than their N A counterparts of similar CF. Also, the spread of thresholds for N M units across frequency is about 30 dB, which is narrower than that found for N A units. As is the case in NA, no relation is apparent between threshold and SR for N M units. M a n y units were encountered in both nuclei which respond to very low frequency sound (10-200 Hz) with

724

M.E. Warchol and P. Dallos: Neural coding in the chick cochlear nucleus

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Fig. 5. Tuning curves from neurons in nucleus magnocellularis. Almost all units in both NA and NM have tuning curves resembling a simple 'V' shape

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taneous and driven discharge responses and are described fully in a separate publication (Warchol and Dallos 1989). All data presented here are from neurons with CF's above 100 Hz. The highest CF among cochlear nucleus units is 4840 Hz (unit isolated in NA). Examples of tuning curves obtained from NA units are shown in Fig. 4 and tuning curves from NM are shown in Fig. 5. Tuning exhibited by neurons in the two nuclei can be compared in several ways. Sharpness of tuning at CF is quantified by computing a Q~o value (Kiang et al. 1965). Plots of Qlo for NA and NM neurons are shown in Figs. 6A and B, respectively. In both nuclei, Qlo increases directly with increasing CF. Further, the Qlos in NA and NM cover virtually the same range across frequency. At low CFs (<500 Hz), Q~os range from 0.8-2.0. At higher CFs a wider spread of

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Qlo values is seen, but most units have Q~os between 2.0-5.0. The morphology of tuning curves obtained from both nuclei is generally similar. The overwhelming majority of tuning curves are 'V' shaped, although the slope of the low frequency portion of the tuning curve is often less than the slope of the high frequency half. The low and high frequency slopes can be quantified by representing each as the slope of a line drawn through points 5 and 25 dB above threshold on the appropriate side of each tuning curve. The resulting estimates of tuning curve slopes for units in both nuclei as a function of CF are shown in Fig. 7. Low frequency slopes in both NA and N M are generally clustered between - 5 0 to - 1 5 0 dB/octave and are relatively constant with CF, although a small increase with CF may be present in the N M population (Fig. 7 A, C). High frequency slopes, on the other hand, tend to increase with frequency and can exceed 300 dB/octave at higher CFs (Fig. 7B, D). Data on tuning curve slope offers further evidence that tuning in NA and N M is essentially similar. Finally, 5 units recorded in nucleus angularis have tuning curves whose morphology differs from the remaining population. Rather than having a simple 'V' shaped form, these units have tuning curves which possess long low frequency tails. The CF's of these units were all above 1800 Hz and their thresholds do not differ

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Tonotopic arrangement. Electrode passes through both nuclei were made in approximately the dorsal-ventral plane. The pattern of C F ' s of successive units encountered in each nucleus is very distinctive. In N M , units are arranged in nearly isofrequency columns, in agreement with a previous study of this nucleus in the chick (Rubel and Parks 1975). Thus on any single electrode pass through N M , only units with similar CFs are encountered. By contrast, units in N A are found to possess decreasing CFs as the electrode is m o v e d ventrally through the nucleus. The dorsal-most units have the highest CFs ( > 3000 Hz) while more ventral neurons m a y be tuned to frequencies below 200 Hz. This distribution of CFs in both N M and N A is probably unique enough to allow identification of electrode position within the cochlear nucleus complex if a sufficient n u m b e r of units are encountered to establish a clear tonotopic pattern. An example of the change in CF with increasing (ventral-ward) electrode depth from tracks through both nuclei is shown in Fig. 9.

Fig. 9. Examples of CFs of units encountered on electrode tracks through NA (top row) and NM (bottom row). A different and distinctive tonotopie pattern is found in each nucleus. Neurons in NA show a decrease in best frequency as the electrode is moved from dorsal to ventral, while units in NM are arranged in dorsoventral isofrequency columns

Rate/intensity relationships. The effect of sound intensity on discharge rate was studied in both nuclei. Rate/intensity functions were collected by stimulating at a unit's CF with m a n y intensity levels, varied in 5 dB steps. The various sound levels were presented randomly, rather than in ascending order. The steady state or adapted discharge rate was measured since this quantity is likely to be used by central auditory centers in the discrimination o f ongoing intensity differences. Monotonic rateintensity functions show saturating rates at the highest sound levels. The saturated rate for monotonically re-

sponding units is estimated by taking the simple mean of the measured discharge rate of the points on the ' flat' part of the rate-intensity curve9 The dynamic range is then defined as the total sound pressure range over which discharge increase occurs (i.e., the range between SPLs corresponding to spontaneous and saturated rates). Neurons from N A and N M are contrasted according to the following quantities: monotonicity, dynamic range, and saturated or m a x i m u m discharge rate.

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In nucleus angularis, 78% of units studied (21/27) responded to increasing sound intensity with monotonic increases in sustained discharge rate which saturates at high sound levels. Examples of such rate/intensity functions are shown in Fig. 10. Their dynamic ranges (defined as above) vary between 15-65 dB, and average 44 dB. Thus, most NA neurons can code intensity over a wide range of sound levels. The saturated or maximum discharge rates vary from 50-415 spikes/s. The remaining 22% of NA units have nonmonotonic rate/intensity functions at CF. Two examples are shown in Fig. 11. These units have excitatory regions of 2030 dB after which their discharge rate peaks. Further increases in sound level result in decreased firing rate. This decline can continue until the unit fires near or even below its spontaneous rate. A single unit (shown in Fig. 11) responded in a more complex fashion, with its discharge rate increasing again at the very highest

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intensities. Nonmonotonic units have lower SRs than the general NA population, averaging 9.2 spikes/s (range 0-25.2 spikes/s). Rate/intensity functions were obtained in the same manner for 16 neurons in nucleus magnocellularis. These units also tend to show large driven rate increases when

728

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stimulated at CF. Examples of NM rate/intensity relations are shown in Fig. 12. Eleven of the 16 NM units responded in a clearly monotonic fashion. Dynamic ranges average 38 dB (range: 15-55 dB) and saturated rates range from 110-480 spikes/s. Maximum discharge rates frequently exceed 400 spikes/s. One NM unit did not saturate at the highest sound level used (110 dB SPL) even though it was firing at a rate of 490 spikes/s. Four NM units have rate/intensity functions which are slightly nonmonotonic. These display moderate declines in discharge rate at high sound levels, although these high level rates are still substantially greater than spontaneous rates. Such response is likely to be a result of adaptation during data collection at high intensities. One example is shown in Fig. 12. No NM units respond with the exaggerated nonmonotonic patterns seen in NA. For this reason, all NM neurons are classified as monotonic. In both nuclei, the saturated discharge rate appears to increase directly with the dynamic range of the rateintensity function. Plots of saturated rate as a function of dynamic range for NA and NM neurons are shown in Fig. 13A and B, respectively. Such a pattern would result if rate-intensity functions of all units in a given nucleus have the same slope regardless of dynamic range.

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Response types. Post stimulus time (PST) histograms were obtained at a unit's CF with a sound level 20 dB above threshold. In nucleus angularis, 3 distinct PST histogram morphologies are seen. Examples of these are shown in Fig. 14. About half of the NA unit population studied (13/25 units) respond to tone bursts with an initially regular discharge, resulting in PST histograms which resemble the 'chopper' pattern common in the mammalian cochlear nucleus (e.g., Rhode and Smith 1986a). Most NA units with CFs above 1000 Hz display chopper type response. Also, all 6 nonmonotonic NA units have chopper PST histograms. Two of the 25 NA units responded by firing only one or two spikes at the onset of a tone burst. These 'onset' units are identical in form to PST histograms reported for the mammalian cochlear nucleus (Rhode and Smith 1986a). The remaining NA neurons (10/25) have PST histograms which are 'primary-like'. Most of these units have CFs below 1000 Hz. They usually fire with high rate during the initial segment (about 10 ms) of a tone burst after which their response adapts to a more moderate discharge rate. However, some low frequency primary-like units show no adaptation and respond in a simple phase-locked manner when stimulated at their CF.

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Only these 3 histogram types - chopper, onset, and primary-like - are seen in NA. Other types reported for the mammalian PVCN and DCN, such as 'pauser' and 'build-up' were not encountered. By contrast, the PST histogram morphology among the unit population in N M is much more homogeneous. All 24 N M units studied, when stimulated at their CFs, respond with a primary-like discharge, with only a slight diversity in form. Most N M units respond with an initially high discharge rate which adapts after 5-10 ms to a lower sustained firing. In a few units this adaptation is less pronounced, with a relatively constant discharge level maintained throughout the duration of the 100 ms tone burst. Examples of PST histograms, recorded in N M units are shown in Fig. 15.

Neural phase-locking. Synchrony of discharge to tones at CF can be assessed by computing a vector strength value (Goldberg and Brown 1969) for each of the histograms described in the previous section. Vector strengths for N A and N M neurons, plotted as a function o f CF, are shown in Fig. 16. Phase-locking to frequencies below about 800 Hz is strong for units in both nuclei. Above

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this frequency, vector strength begins to decline and differences in the phase-locking capabilities of neurons in N A and N M become apparent. Above 1000 Hz, most N A neurons respond as choppers, whose initial regular firing is independent of the stimulus frequency. Computed vector strength values for these units are very low and they likely convey little information about the temporal character of the stimulus waveform to higher auditory centers. Phase-locking ability in N M is somewhat better at high frequencies. About half o f the N M neurons with CFs above 1000 Hz still show significant phase-locking (defined as vector strength exceeding 0.2). Phase-locking becomes negligible in N M neurons for stimulus frequencies above 2000 Hz. A summary o f the physiological results obtained from N A and N M is shown in Table 1.

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M.E. Warchol and P. Dallos: Neural coding in the chick cochlear nucleus

Discussion

This study was undertaken in an effort to quantify the physiological responses of neurons in nucleus angularis and magnocellularis in the domestic chick in a way which might suggest functional differences between the two nuclei and provide comparisons with results obtained from other avian species. The development and anatomy of the chick auditory system has been extensively studied (Rubel and Parks 1988), and because the system has few features which appear highly specialized, it is likely a good model for the auditory systems of most birds. Also, the chicken produces a large repertoire of vocal signals (Collias and Joos 1953; Konishi 1965) and the importance of vocal communication between hen and chicks has been demonstrated (Gottlieb 1971). Thus, the chicken must both recognize many complex signals and be able to accurately localize their source. This discussion of physiological results from the two divisions of the chick cochlear nucleus will be aimed at exploring functional differences inferred from the single unit data. One plausible hypothesis is that the functional segregation evident in the two divisions of the barn owl cochlear nucleus (nucleus magnocellularis temporal information, nucleus angularis - intensity information; Sullivan and Konishi 1984) is a general feature of the avian auditory system. This suggestion is partially supported by anatomical considerations. Nucleus magnocellularis in all avian species so far examined projects (bilaterally) to a third-order brainstem center, nucleus laminaris. This nucleus, thought to be homologous to the mammalian medial superior olive (Boord 1969), appears to form a neural delay line (Young and Rubel 1983; Hyson etal. 1989) which could function in sound localization by processing timing differences present in spikes from both the ipsi- and contralateral magnocellular nuclei, similar in principle to the coincidence detection mechanism first proposed by Jeffress (1948). In the barn owl, nucleus laminaris has been shown to function in this way by direct physiological measurement (Sullivan and Konishi 1986; Carr and Konishi 1988). The neural projections of nucleus angularis are more complex. Some fibers travel to the contralateral nucleus of the lateral lemniscus (Boord 1969). Nucleus angularis also sends a direct projection to the (contralateral) nucleus of the auditory midbrain, nucleus mesencephali lateralis pars dorsalis (nMLD). The physiology of the nMLD has not been extensively studied in bird species other than the barn owl, but Coles and Aitkin (1979), recording in the chicken nMLD, found that the majority of units appeared to function as detectors of interaural intensity difference, being excited by sound sources located on one side of the head and inhibited by sounds originating from the other. Although nucleus laminaris also projects to the nMLD, data on how the chick midbrain processes interaural timing differences are not available. The physiological responses of NA and N M units in the chick are somewhat consistent with this hypothesis

of segregated function, although the situation does not appear as clear-cut as in the barn owl. Each of the various response parameters will now be discussed separately, and compared with data collected from other avian species as well as relevant results from the mammalian cochlear nucleus in an effort to understand how they might convey timing and intensity information to higher auditory centers.

Spontaneous activity. Clear differences are seen in both the mean spontaneous discharge rate and in the distribution of spontaneous rates in both nuclei. The mean SR of the NA unit population is 25.3 spikes/s (range: 084.0spikes/s) while the mean for N M units is 94.3 spikes/s (range: 31.3-231.2 spikes/s). So the mean SR for one division of the avian cochlear nucleus falls outside the range of SR seen in the other division. A similar pattern has also been reported for the cochlear nucleus of the redwing blackbird (Sachs and Sinnott 1978). While the mean SRs in both NA and NM of the blackbird are slightly higher than those found in the chick, the overall distribution of SR for these two species is very similar. Quantitative SR data are not available for these nuclei in the barn owl, but Sullivan and Konishi (1984) do state that spontaneous activity in nucleus angularis is lower than that found in nucleus magnocellularis. It is significant that the average rate and overall distribution of spontaneous activity in N M appears more similar to that reported for primary auditory neurons from several avian species (Sachs et al. 1974; Manley et al. 1985) than is the case for NA. We have recorded from 32 primary auditory afferents in the apical region of the chick cochlear ganglion with CFs ranging from 139-1269Hz (Warchol, unpublished observations). These neurons have a mean spontaneous rate of 63.9 spikes/s with a range of 18.8-149.8 spikes/s. The distribution of SRs is similar to that seen from N M neurons, although the mean and range are somewhat lower. Viewed in this way, the spontaneous firing of NM appears more 'primary-like' than NA, albeit with moderately elevated rates. This difference in the spontaneous firing patterns has functional implications which are relevant to the segregation of function considered here. Temporal information about a sound stimulus can be expressed by modulating the pattern of discharge of a unit if it has a relatively high background rate. This process can be thought of as a ' sampling' of the sound stimulus at time intervals equal to the mean spacing of a unit's firing. A high spontaneous rate will allow more accurate temporal sampling of a stimulus at low intensity levels where driven discharge increases are not great. If the primary function of N M is to convey timing information to nucleus laminaris, a high spontaneous rate would seem desirable. On the other hand, sound intensity is coded most directly by a absolute change in discharge rate over a fixed time interval. The most easily detected rate change would be from 'zero' spikes at a given time to 'some' spikes at a later time. In other words, a change from

M.E. Warcholand P. Dallos: Neural coding in the chick cochlear nucleus a very low or zero discharge rate to a moderately high rate should be more readily detected by more central auditory nuclei than a comparable absolute rate change imposed on a high background discharge. In NA we find that most units have spontaneous discharge rates below 20 spikes/s. Further, the wide distribution of neural thresholds will result in some subpopulation of NA units that are always able to increase their discharge from a very low to more moderate spike rate at all realistic sound levels. This pattern of spontaneous activity could make the population of NA neurons ideal for coding small intensity differences.

Neural thresholds and tuning. The best neural thresholds across the entire frequency range in nucleus angularis agree quite well with the behavioral audiogram reported for the domestic chick by Gray and Rubel (1984). It appears, then, that the chick is about 15-20 dB less sensitive to sounds at all frequencies in its range of hearing than are many mammalian species. In general, birds tend to have higher absolute sound thresholds than do mammals, with behavioral studies in a number of avian species revealing best thresholds of 5-15 dB SPL (Dooling 1980). The wide range of neural thresholds found in both nuclei is consistent with previous single unit studies of the avian peripheral auditory system. Manley and associates (1985) report results from 500 primary auditory units in the cochlear ganglion of the starling. A similar range (50 dB) in threshold across frequency is found. These authors state that high and low threshold units frequently occur in the same animal in random order, a pattern which is also apparent in the chick cochlear nucleus. The two nuclei differ with respect to neural sensitivity across the entire excitatory frequency range. Best neural thresholds in NA are consistently 10-15 dB lower than those in NM at the same frequency. This finding is also in agreement with at least one previous study. Konishi (1970) reports that, in a number of song bird species, thresholds of single units in NA are more sensitive than their NM counterparts. However, any differences in neural thresholds between these two nuclei which may have been noted in other avian species studied (pigeon, redwing blackbird, barn owl) have not been reported. The apparent difference in neural thresholds between NA and NM may be interpreted as a result of their serving distinct functions in the coding of auditory information. Neural threshold is defined by reference to a sound evoked increase in discharge rate. However, it is a consistent finding of auditory physiology (birds: Sachs et al. 1980; Manley et al. 1985; mammals: Rose et al. 1974) that the onset of neural phase-locking occurs at sound levels about 10-20 dB below the measured (rate) threshold. Thus, if we view the primary function of NM as providing temporal information to a coincidence detection network (nucleus laminaris), a more realistic definition of'threshold' would be the sound level at which spikes become entrained to temporal aspects

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of the stimulus (= phase-locking). Thought of in this way, the 'functional thresholds' of both nuclei may, in fact, be similar. The sharpness of tuning and morphology of tuning curves appear very similar in both nuclei. When quantified as a Qlo value or by measuring the slopes of the two halves of tuning curves, little difference between NA and NM is seen. It is likely that tuning curve shape is determined entirely by the primary units of the auditory nerve, which branch to innervate secondorder cells in both NA and NM (Whitehead and Morest 1981). A consequence of similar tuning in both nuclei is that the place representation of frequency across the entire population of auditory units is maintained.

Rate/intensity responses. In nucleus angularis, 78% of neurons studied (21/27) respond to increased sound intensity (at CF) with monotonic increases in discharge rate. Magnocellular units are all monotonic. The populations of monotonic units in both nuclei appear very similar in terms of average dynamic range (44 dB in NA vs. 38 dB in NM) and in saturated discharge rate. This is in clear contrast to the very different rate-intensity responses reported for NA and NM in the barn owl (Sullivan and Konishi 1984; Sullivan 1985). In NA of the owl, large driven rate increases are seen, while the principal response in NM neurons is a modification of the temporal pattern of firing without a corresponding rate increase. It is possible, then, that information on the intensity of sound stimuli is coded in both NM and NA of the chicken. How intensity information from NM could be utilized by nucleus laminaris is unclear and, unfortunately, no relevant physiological data from NL are available. It is notable that in the mammalian homologue of NL, the medial superior olive, neurons which appear to function as simple coincidence detectors are limited to low CFs, while higher CF units can display behavior which is more consistent with analysis of interaural intensity differences (Goldberg and Brown 1969). Since phase-locked information from the chick NM is limited to frequencies below 2000 Hz, the higher frequency regions of NL appear to lack sufficient temporal information to construct a place representation of interaural timing differences. Thus, it is possible that NL in the chick (and perhaps other non-owl avian species) may not simply detect temporal disparities but may also be involved in the detection of interaural intensity differences. Nonmonotonic rate-intensity functions are also seen in NA, where 22% (6/27) of the unit population shows an inhibition of firing at the highest sound levels. Such response patterns are limited to higher CF neurons, as all nonmonotonic units have CFs above 1200 Hz. Nonmonotonic rate responses are not seen in the barn owl cochlear nucleus (Sullivan 1985) but are present in about the same proportion in nucleus angularis of the redwing blackbird (Sachs and Sinnott 1978). The nonmonotonic units seen in the chick are probably equivalent to the 'type IV' class described by Sachs and Sinnott (1978) in the blackbird NA and in the mammalian DCN (Young and Brownell 1976). Since detailed response

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M.E. Warcholand P. Dallos: Neural codingin the chick cochlear nucleus

maps were not obtained for chick neurons, it impossible to determine how they fit into the classification scheme used by Sachs and Sinnott (1978). The presence of a moderate percentage of nonmonotonic units in the chick NA suggests an additional role for this nucleus beyond simple coding of intensity information. In many avian species (of which the chick and blackbird are good examples) a large number of conspecific vocalizations must be picked out and recognized from an often considerable background noise. Nonmonotonic units may be involved in this behavior, since they are selective for both particular frequency bands and specific intensity levels. Although the barn owl also possesses a rich vocal repertoire (Johnscard 1988) it is primarily nocturnal and may have to contend with much less background noise during communication and localization of prey. Extraction of a meaningful signal from background sound would not, then, be a likely function of nucleus angularis in the owl. It should be emphasized that these points are highly speculative. Only continued physiological study of many avian species with varied vocal behavior will reveal the behavioral significance of these response types present in the central auditory nuclei.

Histogram response patterns. Units from both nuclei are also classified by the morphology of their PST histogram response when stimulated at CF. Forty percent of NA units and all units from NM respond with a primary-like discharge pattern. In general, these units fire at a high rate during the initial part of a tone burst and then reduce their discharge to a more moderate rate (which is still substantially higher than SR). Primary-like response in NA is mainly limited to frequencies below 1000 Hz. Histogram responses are more homogeneous in NM and temporal information on higher frequency stimuli is preserved. Just over half (52%) of the PST histograms collected from NA units resemble a chopper pattern. More importantly, all but two NA units with CFs above 1000 Hz are choppers. Almost all N A units in the barn owl (whose CFs all exceed 1000 Hz) are choppers (Sullivan and Konishi 1984). Significantly, no units in the NA of the redwing blackbird respond as choppers (Sachs and Sinnott 1978). In the mammalian PVCN, the presumed homologue of at least part of the avian NA, about 40% of all units are classified as choppers (Rhode and Smith 1986 a). In mammalian species, true' chopper' responses are seen only with stimulus frequencies above 1000 Hz. So, in comparing response types across species, the chick NA appears functionally closer to its homologue in the barn owl (if the low CF units are excluded) and to the mammalian PVCN than to the angular nucleus of the blackbird. The varying percentage of chopper response types in homologous nuclei of different species is difficult to interpret. At issue is whether the chopper pattern serves any unique function in the auditory system which is directly relevant to behavior. Opinions on this matter vary. On one hand, Sullivan (1985) speculates that the initial regular firing of choppers is a means

of conveying intensity information in a highly economical fashion, since fewer intervals of a regular discharge pattern have to be sampled to produce an estimate of discharge rate than if the firing was more irregular. Consistent with this notion, Sullivan's data show that the initial portion of the PST histogram (which brackets the most regular discharge) undergoes the steepest rate increase as the sound level is raised. A differing (but not incompatible) view is provided by Rhode and Smith (1986a), who suggest that the regular firing of choppers serves only to provide a consistent neural response to higher auditory centers (i.e., to remove effects of the stimulus frequency on the pattern of discharge). A further role for choppers in the mammalian cochlear nucleus may to be serve as inhibitory interneurons connecting the different cochlear nucleus subdivisions (Voigt and Young 1980). This is unlikely to be a general function of choppers in the avian auditory system, since higher CF neurons in NA of some species are almost entirely choppers (owl, chick) while no choppers are found in the redwing blackbird, even though prominent inhibitory sidebands are a common feature of many units (Sachs and Sinnott 1978). Three of the PST histogram response types common in the mammalian cochlear nucleus - primary-like, chopper, and onset - are found in the chick NA. Significantly, in the 3 avian species where NA responses have been studied (chick, redwing blackbird, and barn owl) substantial differences are found in both the types of responses seen in NA and in their relative percentages. All neural response types in nucleus angularis of the barn owl, for example, resemble those found in the mammalian PVCN (Sullivan 1985). A very different situation is seen in the redwing blackbird, where DCN responses (build-up and pauser PSTH patterns and 'type IV' response maps) are relatively abundant. In the chick, all PST histogram response types resemble those found in the mammalian PVCN. However, nonmonotonic rateintensity functions are also common in the chick NA. In the mammal, nonmonotonic rate responses are only found in the DCN. For example, Rhode and Smith (1986b) find that 14% of choppers in the cat DCN possess nonmonotonic rate-intensity functions. It is likely, then, that nucleus angularis is not the simple functional equivalent of one of the divisions of the mammalian cochlear nucleus. But it is reasonable to expect that similar response types (chopper, onset, pauser, etc.) serve similar purposes in the coding of auditory information in both bird and mammal. Thus, it would be interesting to compare the response patterns obtained from the cochlear nucleus of various avian species which have differing auditory behaviors. Such studies may begin to explain the functional significance of these various PST histogram and rate response patterns.

Phase-locking. There is a clear distinction between NA and NM in phase-locking ability to stimulus frequencies above 1000 Hz. In part, this is a consequence of most NA units with CFs in this frequency range responding as choppers. It would appear that higher frequency tem-

M.E. Warchol and P. DaUos: Neural coding in the chick cochlear nucleus poral information is available to higher auditory centers only through the pathway initiated by NM. Even in NM, neural phase-locking extends only up to about 2000 Hz. This is somewhat lower than values obtained from the auditory nerve in other species, both avian and m a m m a lian. Phase-locking in the auditory nerve of the redwing blackbird, for example, extends to about 4000 Hz (Sachs et al. 1980). However, it is a c o m m o n finding in the mammalian cochlear nucleus that neural synchrony is less accurate in (primary-like) neurons of the cochlear nucleus than in the primary afferents themselves (Rhode and Smith 1986a; Kettner et al. 1985). The disparity in maximum vector strength between auditory nerve and cochlear nucleus units at a given frequency is 0.14).2 (vector strength units). In this light, the synchrony data obtained from the chick cochlear nucleus and those reported for the blackbird are quite similar. Obviously, neural synchrony in the chick is limited to much lower frequencies than have been shown for the owl. Phase-locking in barn owl cochlear nucleus neurons has been shown to extend to 8000 Hz (Sullivan and Konishi 1984), which is at least an octave higher than any other vertebrate species so far examined. This extraordinary finding almost certainly results from neural specializations underlying the owl's unique ability to accurately localize sound sources. It is not surprising that synchrony at very high frequencies is not a general feature of the avian auditory system. The upper frequency limit of phase-locking certainly has functional implications for the chick's ability to either localize sound or identify complex signals. In a separate study, we have estimated the minimum resolvable interaural time disparity in the chick (which is enhanced by the presence of the interaural canal connecting the two middle ear spaces see Calford 1988; Calford and Piddington 1988) using the physiological response properties reported here (Warchol and Dallos, unpublished). These values are then used to calculate minimum resolvable angles in auditory space along the horizontal plane. We have shown that, for frequencies below about 2000 Hz, temporal cues (as inferred from neural phaselocking ability) are adequate to account for minimum resolvable azimuthal angles o f 10-20 ~ This accuracy is comparable to that reported for other small avian species which presumably rely on similar cues (Klump et al. 1986; Lewald 1987a, b; Park et al. 1987).

Acknowledgements. We thank Drs. Don Born, Dianne Durham, and Ed Rubel for initiation into the art of chicken electrophysiology. Thanks also to M.A. Cheatham, S. Echteler, R. Hallworth, K. Ohlemiller, and J. Siegel for technical assistance and helpful discussions. Research was supported by NIH grant NS 08635.

References Boord RL (1969) The anatomy of the avain auditory system. Ann New York Acad Sci 167:186-198 Calford MB (1988) Constraints on the coding of sound frequency imposed by the avian interaural canal. J Comp Physiol A 162 :491-502

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