Physiological Characterization of Supramedullary/Dorsal Neurons of the Cunner,Tautogolabrus adspersusAuthor(s): S. J. Zottoli, F. R. Akanki, N. A. Hiza, D. A. Ho-Sang, Jr., M. Motta, X. Tan and K.M. WattsSource: Biological Bulletin, Vol. 197, No. 2, Centennial Issue: October, 1899-1999 (Oct., 1999),pp. 239-240Published by: Marine Biological LaboratoryStable URL: http://www.jstor.org/stable/1542624 .

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PISCINE NEUROBIOLOGY AND BEHAVIOR

Reference: Biol. Bull. 197: 239-240. (October 1999)

Physiological Characterization of Supramedullary/Dorsal Neurons of the Cunner, Tautogolabrus adspersus

S. J. Zottoli, F. R. Akanki, N. A. Hiza, D. A. Ho-Sang Jr., M. Motta, X. Tan, K. M. Watts (Department of Biology, Williams College, Williamstown, Massachusetts 01267) and E.-A. Seyfarth1

The somata of large neurons can be found on the dorsal surface of the medulla oblongata (supramedullary neurons) and spinal cord (dorsal cells) in many teleost fish (1, 2). Thirty-five to forty of these neurons are arranged in a single, median, longitudinal row, from the posterior end of the fissura rhomboidalis through the anterior portion of the spinal cord of the cunner, Tautogolabrus adspersus (3). Comparative physiological studies have indicated that these cells in the cunner and other teleost fish make electro- tonic connections with one another, and that they respond to tactile

inputs from the skin (4). We report on some physiological prop- erties of these cells with the ultimate goal of determining their function.

Cunner, 9.8-11.5 cm in body length, were secured in a holding chamber and a respiratory current of chilled water was passed through the mouth and over the gills. D-tubocurarine was injected intramuscularly. After the application of local anesthetic (20% benzocaine, Ultradent, Inc.), the medulla oblongata and rostral

spinal cord were exposed and single microelectrode recordings (KCl-filled; 8-11 Mfl) were made from dorsal cell somata. The skin was stimulated either electrically with bipolar stainless-steel electrodes, or mechanically with a probe mounted on the face of an audio speaker. A sphere, 2-mm in diameter, located at the end of the probe contacted the skin.

In 17 neurons from 6 fish, resting potentials ranged from -56 to -71 mV (mean = -64.5), and evoked action potentials ranged in amplitude from 92-115 mV (mean = 103.7). Intra- cellular depolarization caused repetitive firing with no apparent change in spike height (Fig. 1A). Electrical stimulation of the skin resulted in depolarizations that, in some cases, could activate the dorsal cells (Fig. lB1). Repetitive stimulation of the skin at rates of about 1 per 15 s reduced the depolarizations (Fig. 1B2, arrow) revealing long-duration post-synaptic poten- tials (PSPs). PSPs were graded and were depressed at stimula- tion rates of 1 per s (Fig. 1B3). PSPs could be evoked from

many but not all skin locations in an individual neuron, sug- gesting that each cell may have a different pattern of skin innervation. Mechanical stimulation of the skin just caudal to the posterior limit of the dorsal fin produced PSPs in many cells. An action potential was elicited when these PSPs were combined with subthreshold intracellular depolarization of the somata (Fig. 1C). The apparent firing level in Figure 1C is much lower than that in Figure lB. Thus, the impulse is most likely arising at a distance from the soma. This interpretation is consistent with results from the puffer that indicate that syn- apses and the site of physiological impulse initiation are far from the soma (5).

I Zoologisches Institut, J. W. Goethe-Universitat, D-60054, Frankfurt a.M., Germany.

Supramedullary neurons in puffer and other teleost fish are electrotonically coupled and discharge synchronously to tactile stimulation (6). These neurons also respond to visual stimulation and saccular movement in the toadfish and boxfish (7). The activity of these cells in puffer is correlated with an efferent discharge in the dorsal roots (5). Immunohistological studies of the puffer supramedullary cells indicate that they project to cutaneous mu- cous glands and the epidermal layer of the skin (8). We speculate that these neurons in cunner and puffer discharge in response to skin stimulation that is associated with a threat. We also suggest that the output of supramedullary/dorsal neurons affects mucous secretion that may aid in protection from predation or may help prevent infection from a wound that would follow a predatory strike.

This work was supported in part by Howard Hughes Medical Institute and Essel Foundation grants to Williams College. We thank Liz Jonas and Len Kaczmarek for their support during this

study.

A B

I

10m 10 ms

2 -- 1 per 15s

3 ---1- Iper s I _

20ms

C

I J|

2 E 20 ms

Figure 1. Physiological responses of supramedullary/dorsal neurons to electrical and mechanical stimulation. A. Response of a neuron to two levels of intracellular current injection (bottom traces). Current was in-

jected for 60 ms and membrane voltage was recorded with a single electrode using bridge-balanced mode of an Axoclamp 2A amplifier. With increased current the cell shows increased rates of discharge from the middle to the upper trace. B (1) Electrical stimulation of the skin excites the cell. (2) Discrete depolarizations (arrow) are lost with repetitive stimula- tion (1 per 15 s) revealing a long-lasting PSP (every other stimulus is

shown). (3) The PSP amplitude is depressed at stimulation rates of 1 per s (intervening stimuli between the last stimulus in 2 and the first in this

panel are not shown). C. Mechanical stimulation causes a PSP that, when combined with a subthreshold intracellular stimulation, elicits an action

potential. Cl. Superimposed traces of a subthreshold intracellular stimu- lation (no action potential) and a mechanical stimulus (10 ms delay) combined with the subthreshold intracellular stimulation (action poten- tial). C2. PSP, shown at higher gain, was evoked by mechanical stimula- tion alone.

239

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REPORTS FROM THE MBL GENERAL SCIENTIFIC MEETINGS REPORTS FROM THE MBL GENERAL SCIENTIFIC MEETINGS

Literature Cited 1. Marini, M., and I. Benedetti. 1992. Pp. 217-236 in Neurology

Today. I. Benedetti, B. Bertolini, and E. Capanna, eds. Selected Sym- posia and Monographs U.Z.I., 7, Mucchi, Modena.

2. Zottoli, S. J., E.-A. Seyfarth, C. J. Tyler, and D. S. Palmer. 1999. Neurosci. Abstr. 25: 104.

3. Sargent, P. E. 1899. Anat. Anz. 15: 212-225. 4. Bennett, M. V. L. 1960. Biol. Bull. 119: 303.

Literature Cited 1. Marini, M., and I. Benedetti. 1992. Pp. 217-236 in Neurology

Today. I. Benedetti, B. Bertolini, and E. Capanna, eds. Selected Sym- posia and Monographs U.Z.I., 7, Mucchi, Modena.

2. Zottoli, S. J., E.-A. Seyfarth, C. J. Tyler, and D. S. Palmer. 1999. Neurosci. Abstr. 25: 104.

3. Sargent, P. E. 1899. Anat. Anz. 15: 212-225. 4. Bennett, M. V. L. 1960. Biol. Bull. 119: 303.

5. Bennett, M. V. L., S. M. Crain, and H. Grundfest. 1959. J. Gen.

Physiol. 43: 221-250. 6. Bennett, M. V. L., Y. Nakajima, and G. D. Pappas. 1967. J. Neu-

rophysiol. 30: 161-179. 7. Barry, M. A., M. Weiser, R. Baker, and M. V. L. Bennett. 1986.

Biol. Bull. 171: 490-491. 8. Funakoshi, K., T. Kadota, Y. Atobe, M. Nakano, R. C. Goris, and

R. Kishida. 1998. Neurosci. Letters 258: 171-174.

5. Bennett, M. V. L., S. M. Crain, and H. Grundfest. 1959. J. Gen.

Physiol. 43: 221-250. 6. Bennett, M. V. L., Y. Nakajima, and G. D. Pappas. 1967. J. Neu-

rophysiol. 30: 161-179. 7. Barry, M. A., M. Weiser, R. Baker, and M. V. L. Bennett. 1986.

Biol. Bull. 171: 490-491. 8. Funakoshi, K., T. Kadota, Y. Atobe, M. Nakano, R. C. Goris, and

R. Kishida. 1998. Neurosci. Letters 258: 171-174.

Reference: Biol. Bull. 197: 240-241. (October 1999)

Sharpening of Directional Auditory Input in the Descending Octaval Nucleus of the Toadfish, Opsanus tau

R. R. Fay and P. L. Edds-Walton (Parmly Hearing Institute, Loyola University Chicago, Chicago, Illinois 60626)

Reference: Biol. Bull. 197: 240-241. (October 1999)

Sharpening of Directional Auditory Input in the Descending Octaval Nucleus of the Toadfish, Opsanus tau

R. R. Fay and P. L. Edds-Walton (Parmly Hearing Institute, Loyola University Chicago, Chicago, Illinois 60626)

This investigation concerns the fate of acoustic directional en-

coding in the medulla of Opsanus tau. The descending octaval nucleus (DON) is the nucleus receiving the majority of auditory input from the saccule. Previous work has shown that saccular afferents encode directional auditory information that could be used to determine the location of a sound source (1, 2). Each saccular afferent projects to multiple sites along the rostral-caudal axis of the DON (3). Most saccular afferents show a cosinusoidal

response pattern to motional stimuli in the horizontal and mid-

sagittal planes (2). Recently, we reported that directional auditory responses were present in the DON (4). In this study, we examine directional response properties of cells in the DON that were

sharpened compared to primary saccular afferents. Prior to surgery, the toadfish was anesthetized lightly (3 ami-

nobenzoic acid), and the tail muscles were paralyzed (pancuro- nium bromide). The dorsal surface of the cranium was removed to

expose the left lateral surface of the medulla between the posterior ramus of the eighth cranial nerve (VIIIp) and cranial nerve IX, near the rostral and mid-region of the DON, respectively. The fish's head was secured in a circular dish (containing seawater) that is mounted on a three-dimensional shaker table described in detail elsewhere (2). Minishakers produced sinusoidal, translatory mo- tion of the dish in the horizontal and mid-sagittal planes at 30? intervals. This stimulus simulates the particle motion component of underwater sound.

Extracellular recordings were made using woodsmetal-filled electrodes (4, 5), with 10-12 g,m tip diameters and NaCl-filled

capillary tubes with 5-7 /um tips and resistances of 3-8 mfl. These electrodes produced comparable data. A three-axis micromanipu- lator was used to position the electrode, and recording sites were documented by noting micromanipulator position during record-

ing. Neurobiotin was injected (4% in 2M NaCl, 1900-2000 nA for 10-20 min) once per animal at an auditory site. This, procedure helped document recording location for use in future neuroana- tomical analyses.

The 88 recordings made in the DON contain two categories of directional responses: primary-like and sharpened. Primary-like DON cells have cosinusoidal directional response patterns similar to those seen in saccular afferents (1, 2). Many cells recorded in the DON had directional response patterns that differed from

This investigation concerns the fate of acoustic directional en-

coding in the medulla of Opsanus tau. The descending octaval nucleus (DON) is the nucleus receiving the majority of auditory input from the saccule. Previous work has shown that saccular afferents encode directional auditory information that could be used to determine the location of a sound source (1, 2). Each saccular afferent projects to multiple sites along the rostral-caudal axis of the DON (3). Most saccular afferents show a cosinusoidal

response pattern to motional stimuli in the horizontal and mid-

sagittal planes (2). Recently, we reported that directional auditory responses were present in the DON (4). In this study, we examine directional response properties of cells in the DON that were

sharpened compared to primary saccular afferents. Prior to surgery, the toadfish was anesthetized lightly (3 ami-

nobenzoic acid), and the tail muscles were paralyzed (pancuro- nium bromide). The dorsal surface of the cranium was removed to

expose the left lateral surface of the medulla between the posterior ramus of the eighth cranial nerve (VIIIp) and cranial nerve IX, near the rostral and mid-region of the DON, respectively. The fish's head was secured in a circular dish (containing seawater) that is mounted on a three-dimensional shaker table described in detail elsewhere (2). Minishakers produced sinusoidal, translatory mo- tion of the dish in the horizontal and mid-sagittal planes at 30? intervals. This stimulus simulates the particle motion component of underwater sound.

Extracellular recordings were made using woodsmetal-filled electrodes (4, 5), with 10-12 g,m tip diameters and NaCl-filled

capillary tubes with 5-7 /um tips and resistances of 3-8 mfl. These electrodes produced comparable data. A three-axis micromanipu- lator was used to position the electrode, and recording sites were documented by noting micromanipulator position during record-

ing. Neurobiotin was injected (4% in 2M NaCl, 1900-2000 nA for 10-20 min) once per animal at an auditory site. This, procedure helped document recording location for use in future neuroana- tomical analyses.

The 88 recordings made in the DON contain two categories of directional responses: primary-like and sharpened. Primary-like DON cells have cosinusoidal directional response patterns similar to those seen in saccular afferents (1, 2). Many cells recorded in the DON had directional response patterns that differed from

simple cosine functions: they are sharpened (see Fig. 1A) with

torpedo-shaped patterns (63% of 88). Sharpened cells were further

categorized as slightly (23%), moderately (28%), or highly (12%) sharpened based on the magnitude of their deviation from a cosine function. Some cells were sharpened in only one plane (n = 15, with only mid-sagittal plane sharpening in 11 cells, and only horizontal plane sharpening in 4 cells). The rest (n = 40) were

sharpened in both planes to some extent. Although we cannot

simple cosine functions: they are sharpened (see Fig. 1A) with

torpedo-shaped patterns (63% of 88). Sharpened cells were further

categorized as slightly (23%), moderately (28%), or highly (12%) sharpened based on the magnitude of their deviation from a cosine function. Some cells were sharpened in only one plane (n = 15, with only mid-sagittal plane sharpening in 11 cells, and only horizontal plane sharpening in 4 cells). The rest (n = 40) were

sharpened in both planes to some extent. Although we cannot

A: 16 - DON A: 16 - DON

-30 -30

Front 0

Front 0

Cosine Excitation + Cosine Excitation +

B: MODEL

Front

B: MODEL

Front Cosine Inhibition .... Cosine Inhibition ....

30 30

-60 -60

-90 -90

-120 -120

-1 ou ---- iu-150 150 180 / 180

5, 10, 15, 20 dB re: 1 nm Excitation & Inhibition = o , ~ ?

-1 ou ---- iu-150 150 180 / 180

5, 10, 15, 20 dB re: 1 nm Excitation & Inhibition = o , ~ ?

Figure 1. A: Directional responses for cell 6 (animal I) at four iso-displacement levels in the horizontal plane in polar coordinates. The

angular axis is the orientation of linear translatory motion. The radial axis is response magnitude (the Z-statistic: Z = R2N, where R is vector

strength-a phase-locking metric (2), where 0 < R - 1, and N is the number of spikes recordedfor eight repetitions of a 500 ms sinusoid at 100 Hz). The circular axis indicates a Z value of 350. In this polar plot, each directional stimulus axis is represented on the circle in degrees with

respect to straight ahead (0?). For each stimulus axis, response magnitude (Z) was plotted twice (at the nominal axis angle, and at this angle plus 180?). Thus, the distance between the polar plot origin and the plotted points represent the value of Z at each stimulus axis. B: Hypothetical excitatory (solid lines, + symbol) and inhibitory (dotted lines) cosine-

shaped directional inputs to a DON cell. The amplitudes and orientations

of the inputs were adjusted to form a difference function (gray lines and D

symbols, negative values set to zero) closely modeling the directional

response in Figure lA for the 15 dB level. The difference function is also

plotted in Figure 1A, which overlaps the cell's response completely.

Figure 1. A: Directional responses for cell 6 (animal I) at four iso-displacement levels in the horizontal plane in polar coordinates. The

angular axis is the orientation of linear translatory motion. The radial axis is response magnitude (the Z-statistic: Z = R2N, where R is vector

strength-a phase-locking metric (2), where 0 < R - 1, and N is the number of spikes recordedfor eight repetitions of a 500 ms sinusoid at 100 Hz). The circular axis indicates a Z value of 350. In this polar plot, each directional stimulus axis is represented on the circle in degrees with

respect to straight ahead (0?). For each stimulus axis, response magnitude (Z) was plotted twice (at the nominal axis angle, and at this angle plus 180?). Thus, the distance between the polar plot origin and the plotted points represent the value of Z at each stimulus axis. B: Hypothetical excitatory (solid lines, + symbol) and inhibitory (dotted lines) cosine-

shaped directional inputs to a DON cell. The amplitudes and orientations

of the inputs were adjusted to form a difference function (gray lines and D

symbols, negative values set to zero) closely modeling the directional

response in Figure lA for the 15 dB level. The difference function is also

plotted in Figure 1A, which overlaps the cell's response completely.

240 240

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