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Neuronal pathways mediating head turning behavior in Aplysia
ELSEVIER
Neuroseience Letters 186 (1995) 197-199
IIMOZIIN~ I[II[R8
Neuronal pathways mediating head turning behavior in Aplysia Yuanpei Xin, IrvingKupfermann* Center for Neurobioh~gy and Behaviour, College of Physicians and Surgeons, Columbia University, 722 West 168th Street, Research Annex, New York, NY 10032, USA Received 19 October 1994; revised version received 22 December 1994; aecepted 3 January 1995
Abstract
Seaweed applied to the head of Aplysia elicits a head tuming response in the direction of the stimulus. The major motor neurons involved in head turning appear to be located largely in the pedal ganglion. Using an autoradiographic technique, we obtained evidenee that there are afferents in the skin of the head that project directly to the pedal ganglion by way of the cerebral-pleural conneetives. Bilateral lesioning of the cerebral-pleural connectives, however, did not interfere with head turning, and unilateral lesions of either a cerebral-pedal or cerebral-pleural connective did not differentially affect turns evoked by stimuli applied ipsilaterally or contralaterally to the lesion. However, a combined unilateral lesion of a cerebral-pedal and cerebral-pleural connective resulted in elimination or significant reduction of ipsilateral tums. The data suggest that head turns elicited by food, are mediated by the combined information travelling in both the pedal and pleural connectives.
Keywords:Aplysia; Head turning response; Autoradiography; Food stimulus
Head turning in Aplysia is a component of a variety of behaviors including egg-laying, 'exploratory' behavior and the appetitive phat»e of feeding behavior [1,11]. When food stimuli are applied to an anterior tentacle, animais turn their neck and head towards the stimulus. The cerebrai ganglion, which receives the nerves innervating the head, is connected to the pedal-pleural ganglia (Fig. 1), which contain motor neurons that appear to mediate head turning [1,4,8]. Previous observations [3,5,12] indicate that the cerebral gang)Jon contains a number of interneurons that project to the pedal ganglion by way of the bilateral cerebral-pedal (C-P) and cerebral-pleural (C-PL) connectives. We have used autoradiography [2,9] to determine whether there might be direct afferent inputs to the pedal ganglion from sensory cells with peripherally located cell bodies in the head, and if so, determine which connectives carry this input. Using Aplysia californica (200-300 g, Marinus, Inc., Long Beach, CA), presumptive peripheral cell bodies were radio-labelled by removing small pieces of skin with attached nerves and ganglia, and placing the skin in a subchamber containing seawater with [35S]methionine. After a 2-h incubation, the tissue * Corresponding author, Tel.: +1 212 9602397; Fax: +1 212 9602410.
was thoroughly washed and the preparation was then incubated in tissue culture medium for 1-2 days to allow axonal transport of labelled macromolecules into the central nervous system. Following fixation in paraformaldehyde (1 h), the nervous system was embedded in wax, and sectioned (16/+m). Sections were coated in Ilford K5D emulsion, and exposed for 5-7 days, developed in Kodak D-19 developer, and post-stained with Toluidine Blue. The results obtained from 15 preparations suggest that there is a dense projection of direct afferents from the lips and tentacles that tavels to the cerebral ganglion (Fig. 2), and continues to the pedal-pleural ganglia. Surprisingly the projection was mediated exclusively by the C-PL connectives, with the strongest projection present in the ipsilateral C-PL connective. There was no sign of labelled fibers in the C-P connectives. Further details of control experiments and the distribution of label within specific ganglia will appear elsewhere [13]. We next examined the role of the pedal or pleural connectives in head turning responses. In preliminary experiments (N = 5), we confirmed the previous report [6] that animais with bilateral section of the C-P connectives, do not exhibit either spontaneous or evoked turns. However, it was difficult to ascertain the specific role of the
031M-3940/95/$09.50 © 1'995 Elsevier Science Ireland Ltd. Ail rights reserved SSDI 0 3 0 4 - 3 9 4 0 ( 9 5 ) 1 I ~ 3 1 2 - H
198
1I. Xin, L Kupfermann / Neuroscience Letters 186 (1995) 197-199
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Fig. 1. Schematic diagram of the major ganglia and eonnectives mediating head turning. The field of view shown in Fig. 2 is indicated by the rectangle. Abbreviations: AT, anterior tentacular nerve; cereb., cerebrai; C-B, cerebral-buecal connective, C-P, cerebral-pedal connective; C-PI, cerebral-pleural connective; g., ganglion; LLAB, lower labial nerve; P, pedal, pleur., pleural; ULAB, upper labial nerve.
pedal connectives, since these animais exhibited profound behavioral deficits not directly related to head turning. The animais were difficult to arouse with food; they failed to raise into the head-up position, and they did not extend their head when stimulated by food. Given the observation that there may be direct afferents travelling in the pleural connectives and that in addition, interneurons may project to the pedal ganglion via the C-PL connectives, we examined whether the C-PL connectives were necessary for head turning. It was puzzling that previous studies involving bilateral section of the pleural connecrives [6,10] failed to report any effects on head turning. In the present study, however, when we examined animais with bilateral C-PL lesions (N = 3), we confirmed that their head turning was indistinguishable from normal animais. We postulated that failure to detect effects of bilaterai sectioning of the C-PL connectives may be due to the fact that the resulting deficits may be relarively subtle. We therefore studied animais in which only one connective was sectioned, so that turns ipsilateral or contralateral to the section could be compared in the same animal. Surgery was performed using previously described techniques [7]. Care was taken to minimize damage to blood vessels when the connectives were cut. For sham operated animais, the nervous systems was exposed, but the connectives were not cut. Animais were tested 24 h after surgery. For ail data involving statistical analysis, testing was donc by one or two observers that were unaware of the specific experimental treatment. Pieces of seaweed (Laver) were used to arouse the animal into a head up posture, and horizontal head turns were elicited by
touching seaweed to the left or right anterior tentacle in a counterbalanced order. For each stimulus application, animais were given a score of 0, 50 or 100, based on an estimate of how the response compared to normal turns: 0 for no turn, 50 for a turn that was weaker than normal (less than approximately 30°), and 100 for normal turns (greater than 30°). Following unilateral lesion of the C-PL connectives, none of rive animais showed any sign of a deficit for turns elicited by stimuli presented to the tentacle ipsilaterai compared to that contraiaterai to the sectioned connective, confirming a previous preliminary observation [10]. Ail animais received a maximum score, for ai1 turns. Similarly sham operated animais (N = 2) had no deficits. We next reasoned that the failure to observe that turn magnitude was not differentially affected by a unilateral lesion of the C-PL connective may be due to the possibility that either the C-PL or C-P connective may be sufficient to compensate for any deficits produced by sectioning the other. We therefore compared the effects of a unilateral C-P connective lesion (N = 12 ) or C-PL lesion (N = 11), with that of a combined unilaterai C-PL and C-P connective lesion (N = 14). We found that ipsilateral versus contraiateral turns were not significantly different in unilateral C-P or C-PL connective lesioned animais (Fig. 3). For animais with the combined unilateral lesions, however, turns towards the side of the lesion were significantly reduced compared to turns towards the contralaterai side. Six of fourteen of these animais totally lost their ability to exhibit turns of their neck when ipsilaterai stimuli were presented, although smail movements of the anterior head were sometimes observed. A variety of explanations of our findings are possible. For example, a unilaterai C-PL connective lesion may produce a mild deficit that cannot be detected, and a uni-
Fig. 2. Autoradiogram (darkfield) of a horizontal section of the cerebral ganglion following radiolab¢lling of a piece of skin taken from the left anterior tentacle. The section is odented with the anterior part of the ganglion toward the top of the figure, and th• field covered is indicated by the rectangle in Fig. l. Dense lab¢lling was observed in the various regions of the ganglion, including the C-PL connectives (indicated by filled triangles), but not in the C-P connectives. This section shows the fibers as they exit from the cerebral ganglion.
Y. Xin, I. Kupfermann / Neuroscience Letters 186 (1995) 197-199
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tutning is mediated both by the pedal c o n n e c t i v e , which p r o v i d e s a route by w h i c h interneurons can act, as well as by the pleural connective, w h i c h p r o v i d e s a p a t h w a y either for interneurons, for direct afferents or both. W e thank S.C. R o s e n and A.J. S u s s w e i n for c o m m e n t s on the paper. Supported by P H S grants M H - 3 5 5 6 4 and G M 320099.
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Fig. 3. Mean turn score following unilateral lesions of the cerebralpodal (C-P) connective alone (N = 12), cerebral-pleural (C-PL) connective alone (N = 11), or of the combined lesion of the C-P connective and the C-PL connective (iV= 14). Animais were tested 'blind' with pieces of seaweed presented to the tentacle ipsilateral and conIalateral to the lesioned side (turns ral:ed as 0, 50 or 100). There was a significant overall effeet of the lesions (Kruskal-Wallis analysis of variante on ipsilateral-contralateral differenee scores, P < 0.01). Individual MannWhitney U-tests revealed that there was no significant difference of the mean ipsilateral-contralateral tutu scores for either the animais with the C-P or C-PL connective lesion (P > 0.05). For animais with the combined unilateral lesions, the turns ipsilateral to the lesions were significantly reduced (P < 0.05), and the ipsilaterai-contralateral difference scores were signifieantly gleater than that of either the pedal or the pleural connective lesion groups (P < 0.05, two-tailed Mann-Whitney U-tests). Six of the fourteen animais with eombined lesions made no neck-head turns to the ipsila,leral Side, but made full turns to the contralateral side. In the pedal connective lesion group, ail animais tumed both to the ipsilateral side and to the conoalateral side, but 8 of 12 exhibited less than full turns (with no consistent pattem of whether the reduced turn was ipsilateral or contralateral). The overall results were very similar for the same animais tested bya second blind observer. lateral C - P c o n n e c t i v e lesion m a y p r o d u c e a non-specific effect that results in a w e a k e n i n g o f both ipsilateral and contralateral turning, thereby b r i n g i n g the unilateral deficit o f the C - P L c o n n e c t i v e lesioned animals to a level at which it can be detected. Alternatively, effects specific to head turning m a y be p r o d u c e d via pathways that inv o l v e both connective:», but the p r e s e n c e o f either connective m a y c o m p e n s a t e for the deficit p r o d u c e d by the other. O n e reasonable possibility is that normal head
[1] Cook, D.G. and Carew, T.J. Operant conditioning of head-waving in Aplysia. III. Cellular analysis of possible reinforcement pathways, J. Neurosci., 9 (1989) 3115-3122. [2] Droz, B. Autoradiography as a tool for visualizing neurons and neuronal processes. In W.M. Cowan and M. Cuénod (Eds.), The Use of Axonal Transport for Studies of Neuronal Conncctivity, Elsevier, Amsterdam, 1975, pp. 127-154. [3] Fredman, S.M. and Jahan-Parwar, B. Command neurons for locomotion in Aplysia, J. Neurophysiol., 49 (1983) 1092-1117. [4] Hening, W.A., Walters, E.T., Carew, T.J. and Kandel, E.R. Motorneuronal control of locomotion in Aplysia, Brain Res., 179 (1979) 231-253. [5] Jahan-Parwar, B. and Fredman, S.M. Cerebral ganglion of Aplysia: cellular organization and origin of nerves, Comp. Biochem. Physiol., 54A (1976) 347-357. [6] Jahan-Parwar, B. and Fredman, S.M. Neural control of locomotion in Aplysia: mie of the central ganglia, Behav. Neural. Biol. 27 (1979) 39-58. [7] Kupfermann, 1. Dissociation of the appedtive and consummatory phases of feeding behavior in Aplysia: a lesion study, Behav. Biol., 10 (1974) 89-97. [8] Nagahama, T., Weiss, K.R. and Kupfermann, I. Body postural muscles active during food arousal in Aplysia are modulated by diverse neurons that receive monosynaptic excitation from the neuron C-PR, J. Neurophysiol., 72 (1994) 314-325. [9] Oison, L.M. and Jacklet, J.W. The circadian pacemaker in the Aplysia eye sends axons throughout the central nervous system, J. Neurosei., 5 (1985) 3214-3227. [10] Teyke, T., Weiss, K.R. and Kupfermann, I. A subpopulation of cerebral B cluster neurones of Aplysia californica is involved in defensive head withdrawal but hOt appetitive head movements, J. Exp. Biol., 147 (1989) 1-20. [11] Teyke, T., Weiss, K.R. and Kupfermann, I. Appetitive feeding behavior of Aplysia: behavioral and neural analysis of direeted head turning, J. Neurosci., l0 (1990) 3922-3934. [12] Teyke, T., Weiss, K.R. and Kupfermann, I. An idenUfied neuron (CPR) evokes neuronal responses reflecting food arousal in Aplysia, Science, 247 (1990) 85-87. [13] Xin, Y., Weiss, K.R. and Kupfermann, I., Distribution in the central nervous system of Aplysia of afferent fibers arising from cell bodies located in the periphery, J. Comp. Neurol., 1995, in press.
Neuroseience Letters 186 (1995) 197-199
IIMOZIIN~ I[II[R8
Neuronal pathways mediating head turning behavior in Aplysia Yuanpei Xin, IrvingKupfermann* Center for Neurobioh~gy and Behaviour, College of Physicians and Surgeons, Columbia University, 722 West 168th Street, Research Annex, New York, NY 10032, USA Received 19 October 1994; revised version received 22 December 1994; aecepted 3 January 1995
Abstract
Seaweed applied to the head of Aplysia elicits a head tuming response in the direction of the stimulus. The major motor neurons involved in head turning appear to be located largely in the pedal ganglion. Using an autoradiographic technique, we obtained evidenee that there are afferents in the skin of the head that project directly to the pedal ganglion by way of the cerebral-pleural conneetives. Bilateral lesioning of the cerebral-pleural connectives, however, did not interfere with head turning, and unilateral lesions of either a cerebral-pedal or cerebral-pleural connective did not differentially affect turns evoked by stimuli applied ipsilaterally or contralaterally to the lesion. However, a combined unilateral lesion of a cerebral-pedal and cerebral-pleural connective resulted in elimination or significant reduction of ipsilateral tums. The data suggest that head turns elicited by food, are mediated by the combined information travelling in both the pedal and pleural connectives.
Keywords:Aplysia; Head turning response; Autoradiography; Food stimulus
Head turning in Aplysia is a component of a variety of behaviors including egg-laying, 'exploratory' behavior and the appetitive phat»e of feeding behavior [1,11]. When food stimuli are applied to an anterior tentacle, animais turn their neck and head towards the stimulus. The cerebrai ganglion, which receives the nerves innervating the head, is connected to the pedal-pleural ganglia (Fig. 1), which contain motor neurons that appear to mediate head turning [1,4,8]. Previous observations [3,5,12] indicate that the cerebral gang)Jon contains a number of interneurons that project to the pedal ganglion by way of the bilateral cerebral-pedal (C-P) and cerebral-pleural (C-PL) connectives. We have used autoradiography [2,9] to determine whether there might be direct afferent inputs to the pedal ganglion from sensory cells with peripherally located cell bodies in the head, and if so, determine which connectives carry this input. Using Aplysia californica (200-300 g, Marinus, Inc., Long Beach, CA), presumptive peripheral cell bodies were radio-labelled by removing small pieces of skin with attached nerves and ganglia, and placing the skin in a subchamber containing seawater with [35S]methionine. After a 2-h incubation, the tissue * Corresponding author, Tel.: +1 212 9602397; Fax: +1 212 9602410.
was thoroughly washed and the preparation was then incubated in tissue culture medium for 1-2 days to allow axonal transport of labelled macromolecules into the central nervous system. Following fixation in paraformaldehyde (1 h), the nervous system was embedded in wax, and sectioned (16/+m). Sections were coated in Ilford K5D emulsion, and exposed for 5-7 days, developed in Kodak D-19 developer, and post-stained with Toluidine Blue. The results obtained from 15 preparations suggest that there is a dense projection of direct afferents from the lips and tentacles that tavels to the cerebral ganglion (Fig. 2), and continues to the pedal-pleural ganglia. Surprisingly the projection was mediated exclusively by the C-PL connectives, with the strongest projection present in the ipsilateral C-PL connective. There was no sign of labelled fibers in the C-P connectives. Further details of control experiments and the distribution of label within specific ganglia will appear elsewhere [13]. We next examined the role of the pedal or pleural connectives in head turning responses. In preliminary experiments (N = 5), we confirmed the previous report [6] that animais with bilateral section of the C-P connectives, do not exhibit either spontaneous or evoked turns. However, it was difficult to ascertain the specific role of the
031M-3940/95/$09.50 © 1'995 Elsevier Science Ireland Ltd. Ail rights reserved SSDI 0 3 0 4 - 3 9 4 0 ( 9 5 ) 1 I ~ 3 1 2 - H
198
1I. Xin, L Kupfermann / Neuroscience Letters 186 (1995) 197-199
¢~~~~
/ ~
ÆÆ/7
C-BConn.
"~~~ ~-
Pleur.g. Pedalg.
Fig. 1. Schematic diagram of the major ganglia and eonnectives mediating head turning. The field of view shown in Fig. 2 is indicated by the rectangle. Abbreviations: AT, anterior tentacular nerve; cereb., cerebrai; C-B, cerebral-buecal connective, C-P, cerebral-pedal connective; C-PI, cerebral-pleural connective; g., ganglion; LLAB, lower labial nerve; P, pedal, pleur., pleural; ULAB, upper labial nerve.
pedal connectives, since these animais exhibited profound behavioral deficits not directly related to head turning. The animais were difficult to arouse with food; they failed to raise into the head-up position, and they did not extend their head when stimulated by food. Given the observation that there may be direct afferents travelling in the pleural connectives and that in addition, interneurons may project to the pedal ganglion via the C-PL connectives, we examined whether the C-PL connectives were necessary for head turning. It was puzzling that previous studies involving bilateral section of the pleural connecrives [6,10] failed to report any effects on head turning. In the present study, however, when we examined animais with bilateral C-PL lesions (N = 3), we confirmed that their head turning was indistinguishable from normal animais. We postulated that failure to detect effects of bilaterai sectioning of the C-PL connectives may be due to the fact that the resulting deficits may be relarively subtle. We therefore studied animais in which only one connective was sectioned, so that turns ipsilateral or contralateral to the section could be compared in the same animal. Surgery was performed using previously described techniques [7]. Care was taken to minimize damage to blood vessels when the connectives were cut. For sham operated animais, the nervous systems was exposed, but the connectives were not cut. Animais were tested 24 h after surgery. For ail data involving statistical analysis, testing was donc by one or two observers that were unaware of the specific experimental treatment. Pieces of seaweed (Laver) were used to arouse the animal into a head up posture, and horizontal head turns were elicited by
touching seaweed to the left or right anterior tentacle in a counterbalanced order. For each stimulus application, animais were given a score of 0, 50 or 100, based on an estimate of how the response compared to normal turns: 0 for no turn, 50 for a turn that was weaker than normal (less than approximately 30°), and 100 for normal turns (greater than 30°). Following unilateral lesion of the C-PL connectives, none of rive animais showed any sign of a deficit for turns elicited by stimuli presented to the tentacle ipsilaterai compared to that contraiaterai to the sectioned connective, confirming a previous preliminary observation [10]. Ail animais received a maximum score, for ai1 turns. Similarly sham operated animais (N = 2) had no deficits. We next reasoned that the failure to observe that turn magnitude was not differentially affected by a unilateral lesion of the C-PL connective may be due to the possibility that either the C-PL or C-P connective may be sufficient to compensate for any deficits produced by sectioning the other. We therefore compared the effects of a unilateral C-P connective lesion (N = 12 ) or C-PL lesion (N = 11), with that of a combined unilaterai C-PL and C-P connective lesion (N = 14). We found that ipsilateral versus contraiateral turns were not significantly different in unilateral C-P or C-PL connective lesioned animais (Fig. 3). For animais with the combined unilateral lesions, however, turns towards the side of the lesion were significantly reduced compared to turns towards the contralaterai side. Six of fourteen of these animais totally lost their ability to exhibit turns of their neck when ipsilaterai stimuli were presented, although smail movements of the anterior head were sometimes observed. A variety of explanations of our findings are possible. For example, a unilaterai C-PL connective lesion may produce a mild deficit that cannot be detected, and a uni-
Fig. 2. Autoradiogram (darkfield) of a horizontal section of the cerebral ganglion following radiolab¢lling of a piece of skin taken from the left anterior tentacle. The section is odented with the anterior part of the ganglion toward the top of the figure, and th• field covered is indicated by the rectangle in Fig. l. Dense lab¢lling was observed in the various regions of the ganglion, including the C-PL connectives (indicated by filled triangles), but not in the C-P connectives. This section shows the fibers as they exit from the cerebral ganglion.
Y. Xin, I. Kupfermann / Neuroscience Letters 186 (1995) 197-199
I00
I
~
IP~i @ON'I'P~
q-
199
tutning is mediated both by the pedal c o n n e c t i v e , which p r o v i d e s a route by w h i c h interneurons can act, as well as by the pleural connective, w h i c h p r o v i d e s a p a t h w a y either for interneurons, for direct afferents or both. W e thank S.C. R o s e n and A.J. S u s s w e i n for c o m m e n t s on the paper. Supported by P H S grants M H - 3 5 5 6 4 and G M 320099.
"??~-? :~:I:~:i: .:,:.:.:.
iiiiiiii
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PED.~.
Fig. 3. Mean turn score following unilateral lesions of the cerebralpodal (C-P) connective alone (N = 12), cerebral-pleural (C-PL) connective alone (N = 11), or of the combined lesion of the C-P connective and the C-PL connective (iV= 14). Animais were tested 'blind' with pieces of seaweed presented to the tentacle ipsilateral and conIalateral to the lesioned side (turns ral:ed as 0, 50 or 100). There was a significant overall effeet of the lesions (Kruskal-Wallis analysis of variante on ipsilateral-contralateral differenee scores, P < 0.01). Individual MannWhitney U-tests revealed that there was no significant difference of the mean ipsilateral-contralateral tutu scores for either the animais with the C-P or C-PL connective lesion (P > 0.05). For animais with the combined unilateral lesions, the turns ipsilateral to the lesions were significantly reduced (P < 0.05), and the ipsilaterai-contralateral difference scores were signifieantly gleater than that of either the pedal or the pleural connective lesion groups (P < 0.05, two-tailed Mann-Whitney U-tests). Six of the fourteen animais with eombined lesions made no neck-head turns to the ipsila,leral Side, but made full turns to the contralateral side. In the pedal connective lesion group, ail animais tumed both to the ipsilateral side and to the conoalateral side, but 8 of 12 exhibited less than full turns (with no consistent pattem of whether the reduced turn was ipsilateral or contralateral). The overall results were very similar for the same animais tested bya second blind observer. lateral C - P c o n n e c t i v e lesion m a y p r o d u c e a non-specific effect that results in a w e a k e n i n g o f both ipsilateral and contralateral turning, thereby b r i n g i n g the unilateral deficit o f the C - P L c o n n e c t i v e lesioned animals to a level at which it can be detected. Alternatively, effects specific to head turning m a y be p r o d u c e d via pathways that inv o l v e both connective:», but the p r e s e n c e o f either connective m a y c o m p e n s a t e for the deficit p r o d u c e d by the other. O n e reasonable possibility is that normal head
[1] Cook, D.G. and Carew, T.J. Operant conditioning of head-waving in Aplysia. III. Cellular analysis of possible reinforcement pathways, J. Neurosci., 9 (1989) 3115-3122. [2] Droz, B. Autoradiography as a tool for visualizing neurons and neuronal processes. In W.M. Cowan and M. Cuénod (Eds.), The Use of Axonal Transport for Studies of Neuronal Conncctivity, Elsevier, Amsterdam, 1975, pp. 127-154. [3] Fredman, S.M. and Jahan-Parwar, B. Command neurons for locomotion in Aplysia, J. Neurophysiol., 49 (1983) 1092-1117. [4] Hening, W.A., Walters, E.T., Carew, T.J. and Kandel, E.R. Motorneuronal control of locomotion in Aplysia, Brain Res., 179 (1979) 231-253. [5] Jahan-Parwar, B. and Fredman, S.M. Cerebral ganglion of Aplysia: cellular organization and origin of nerves, Comp. Biochem. Physiol., 54A (1976) 347-357. [6] Jahan-Parwar, B. and Fredman, S.M. Neural control of locomotion in Aplysia: mie of the central ganglia, Behav. Neural. Biol. 27 (1979) 39-58. [7] Kupfermann, 1. Dissociation of the appedtive and consummatory phases of feeding behavior in Aplysia: a lesion study, Behav. Biol., 10 (1974) 89-97. [8] Nagahama, T., Weiss, K.R. and Kupfermann, I. Body postural muscles active during food arousal in Aplysia are modulated by diverse neurons that receive monosynaptic excitation from the neuron C-PR, J. Neurophysiol., 72 (1994) 314-325. [9] Oison, L.M. and Jacklet, J.W. The circadian pacemaker in the Aplysia eye sends axons throughout the central nervous system, J. Neurosei., 5 (1985) 3214-3227. [10] Teyke, T., Weiss, K.R. and Kupfermann, I. A subpopulation of cerebral B cluster neurones of Aplysia californica is involved in defensive head withdrawal but hOt appetitive head movements, J. Exp. Biol., 147 (1989) 1-20. [11] Teyke, T., Weiss, K.R. and Kupfermann, I. Appetitive feeding behavior of Aplysia: behavioral and neural analysis of direeted head turning, J. Neurosci., l0 (1990) 3922-3934. [12] Teyke, T., Weiss, K.R. and Kupfermann, I. An idenUfied neuron (CPR) evokes neuronal responses reflecting food arousal in Aplysia, Science, 247 (1990) 85-87. [13] Xin, Y., Weiss, K.R. and Kupfermann, I., Distribution in the central nervous system of Aplysia of afferent fibers arising from cell bodies located in the periphery, J. Comp. Neurol., 1995, in press.