Chemoreception in the nudibranch gastropod Phestilla sibogae

ISSN 0300-9629/97/$17.00 PII SO300-9629(97)00014-5

Camp. B&hem. Physiol. Vol. 118A, No. 3, pp. 727-735, 1997 Copyright 0 1997 Elsevier Science Inc. All rights reserved.

ELSEVIER

Chemoreception in the Nudibranch Gastropod

Phestillu sibogue

Bernard F. Murphy and Michael G. Hadfield KEWALO MARINELABORATORY, UNIVERSITY OF HAWAII,41 AHUI STREET,HONOLULU, HI 96813, 1J.S.A.

ABSTRACT. The tropical nudibranch Phestillu sibogae feeds exclusively on corals of the genus Porites, which it locates and recognizes by chemical cues. Morphological and physiological analyses of chemosensory neural pathways in P. sibogae focussed on two pairs of cephalic tentacles, the rhinophores and the oral tentacles. Two nerves from each oral tentacle pass directly to the brain, whereas multiple nerves from the rhinophores converge on paired rhinophoral ganglia that connect to the cerebral ganglia. Chemical sensitivity, determined by changes in rates of discharge, was monitored by suction electrodes attached to cut ends of rhinophoral or oral-tentacle nerves while the excised structure was perfused with extracts of PO&S spp., non-food corals, L-amino acids or glutamate-receptor modulators. Although the oral tentacles were relatively insensitive to the substances tested, responses in rhinophores were positive to extracts of Porites compressa and a non-food coral, were biphasic to I_aspartic and r-glutamic acids and were perceptible but weak to several other amino acids. Rhinophores responded positively to the glutamate-receptor modulators D-glutamate, 6,7-dinitroquinoxaline-2,3-dione (DNQX) and kainic acid but not to N-methyl-D-aspartate (NMDA); the initlal inceased firing-rate response to L-glutamate disappeared when the glutamate was dissolved in magnesium- or cobalt-substituted calcium-free seawater. For the substances tested here, including food corals and a set of amino acids, the rhinophores are the main chemosensory organs for P. sibogae. Glutamate receptors pharmacologically similar to the kainic acid type of vertebrates are present. COMP BIOCHEM PHYSIOL 118A;3:727-735, 1997. 0 1997 Elsevier Science Inc. KEY WORDS.

Chemical senses, chemoreception,

glutamate receptors, Mollusca, nudibranch,

INTRODUCTION Marine different

gastropods

are a dietarily

diverse

taxonomic

group;

species feed on detritus, algae, coral or other seden-

tary or active animals. For generalist feeders, finding food is a minor problem, but for an organism that specializes in only one or very few prey species, finding food is clearly a life-or-death challenge. The nudibranch gastropod PhestiIIQ sibogae is such a specialist. Adults of P. sibogae feed exclusively on corals of the genus Purites (19), and larvae of the nudibranch are known to settle and metamorphose as a speciric response to Potites spp. (l&19). At approximately 3 days after hatching, planktonic veligers of P. sibogae become capable of metamorphosing into juvenile sea slugs. The stimulus for this transformation is a soluble substance secreted or otherwise given off by Porites spp. (15). This metamorphic inducer, whose structure has not been determined, is a small (~500 Da), polar, organic molecule of great morphogenetic potency (14). Despite the known chemosensory metamorphic dependencies of opisthobranch larvae (12,16) and despite the long Adress reprmt requests to: M.G. Hadfield, Kewalo Marine Laboratory, University of Hawaii, 41 Ahui St., Honolulu, HI 96813. Tel. 808-539-7319; Fax 808-599-4817; E-mail: [email protected]. Received 26 June 1996; accepted 30 December

1996.

sensory tentacles

and extensive use of some sea slugs as neurological models, especially Aplysia spp. (23), the chemosensory systems of nudibranchs have been little studied. Investigations of the notaspidean Pkurobranchaea californica (8,25), the anaspidean Aplysia califomica (11,29,33) and a few nudibranchs (summarized in 18) demonstrate that opisthobranch mollusts can find food at a distance and react positively to dissolved food cues when they are micro-applied to regions around the oral tentacles and rhinophores [reviewed in (6)]. Electrophysiological recordings have confirmed the ability of A. californica (11,22) and P. culifomica (3) to detect food odors and amino acids. The ability of nudibranch gastropods such as Hermissenda cmssicornis ( 1) and Tritonia diomedea (10) to detect food odors has also been confirmed. Harris (17) provided evidence that two species of the genus Phestilla, P. mehobrunchiu and P. sibogue, find their specific coral prey by distance chemoreception. However, there has been only a single electrophysiological study of P. sibogue, and chemoreception was not addressed (36). As in other more studied opisthobranchs, the chemosensory structures in P. sibogae consist of two sets of tentacles: oral tentacles that extend from the front margins of the head, lateral to the mouth, and rhinophores that rise above the dorsal surface of the head. As part of our explorations of pathways and mechanisms of metamorphic induction in

B. F. Murphy and M. G. Hadheic

728

P. sibogae [e.g., (14)], we have, in the present study, sought to clarify the chemosensory

mechanisms by which adults of

this species detect their prey corals. We have carried out both anatomical and physiological investigations to visualize major chemosensory

pathways and clarify the roles of

the oral tentacles and rhinophores

in the chemosensory

bi-

ology of I’. sibogue. We specifically explored sensitivities both tentacle

pairs to extracts

of

from several coral species,

some typically eaten by I’. sibogae and some not. Because studies of other opisthobranchs [e.g., (20)] and many other aquatic organisms indicated sensitivity to dissolved amino acids and preliminary exploration

of coral exudates revealed

a large diversity of amino acids, we included such tests in our investigations tentacular

of electrical

output of rhinophoral

nerves. After determining

and

that chemoreception

included sensitivity to L-glutamate, a compound of intense interest as a neurotransmitter,

we carried out pharmacnl~+$-

cal studies to further characterize ceptors on the rhinophores if possible,

apparent glutamate

re-

of P. sibogm and assign them,

to one or more of the recognized

glutamate-

receptor types.

MATERIALS

AND

METHODS

Colonies of P. sibogm are maintained continuously in constant-flow seawater tanks in our laboratory. Although most adult animals are offspring of previous laboratory-reared generations, the populations are periodically augmented with new animals collected from the cc& reefs of Hawaii. These stocks are fed by placing heads of locally collected Porites compressa coral in their tanks. All specimens of P. sibogae used in this study were taken from the laboratory colonies. Small specimens of P. sibogatl were fixed in B~~uin’s fluid, embedded in epon and sectioned to locate potential

serially with glass knives

chemosensory

pathways. Figure 1 was

prepared by reconstruction from these sections. Other morphological studies were made on freshly dissected specimens with the aid of a stereomicroscope. To test the chemical sensitivities of oral tentacles or rhinophores, the respective structure was isolated from the head, the main nerve(s)

leading to the structure was cut

and the cut nerve end was drawn into a suction electrode. For investigations of the rhinophore,

t

100 Mm

FIG. 1. Left lateral view of the brain of Phestilla sibogae, anterior toward the top of the page, showing br, base of the rhinophore; rg, rhinophoral ganglion; cg, cerebral ganglion; ot, oral tentacle nerves; bg, buccal ganglion; pg, pedal ganglion; st, statocyst; e, eyespot; ANT, anterior.

the nerve was typically

cut proximal to the rhinophoral ganglion, so that the combined output of multiple nerves emerging from the sensory epithelium of the rhinophore would be recorded. In the case of the oral tentacles, recordings were made from the larger of the two nerves emerging from one tentacle. Initial experiments were conducted with a petroleum jelly barrier hetween the body of the rhinophore and the rhinophoral ganglion, but when we compared the output with and without the barrier and found no difference, the barrier was no longer used. Afferent signals coming from the nerves in the rhinophore or oral tentacle were amplitied, displayed on an

oscilloscope and passed through a window discriminator (RAD-1 l-A, Winston Electronic Co., San Francisco, CA), which determined the number of spikes every 10 sec. The window discriminator

displayed the count and provided an

output signal whose voltage was proportional to the number of spikes detected in the time interval. This information was registered with a chart recorded. The excised oral tentacles or rhinophores were pinned to Silgard (Dow Corning Co., Midland, MI) in a shallow glass dish, perfused with filtered buffered (pH 8.3; Trizma, Sigma)

Chemoreception in a Nudibranch

seawater (FSW) for 10 min as a control, perfused with a test solution for 10 min and then perfused again with FSW for 10 min. A lo-min application period was chosen because chemosensory responses in molluscs had been shown to be substantially longer lived than those for other sensory modaliries (11). Test solutions included seawater conditioned by one of the living corals, P. compressa, Porites lobata, Pocillopura meandrinu or Montipora vewucosa (a piece of live coral was allowed to stand for 18-24 hr in a bowl of aerated seawater; the coral was removed and the seawater was passed through a 0.45pm Millipore filter; maximal responses to “coral water” typically occurred when seawater was conditioned at least overnight), and several L-amino acids and glutamate-receptor agonists were dissolved in FSW. We additipnally tested 5 mM glutamate in magnesium-substituted, calcium-free seawater [MBL artificial seawater (6) in which calcium was replaced with an osmolar equivalent of magnesium] and 5 mM glutamate in cobalt-substituted, calciumfree seawater (in which calcium was replaced by cobalt and magnesium concentration remained normal). In an effort to measure the chemosensory response of P. sibogae to conspecific individuals, we prepared a conditioned seawater by maintaining several adult animals in about 1 1 of seawater for 12 hr. In experiments with calcium-free seawater, the prewash solution was calcium-free seawater without glutamate; this provided a control for effects of calcium substitution alone. For all other experiments, the control solution was FSW alone. The salinity and pH of every perfusion solution were determined and adjusted, if necessary, to 35 parts per thousand and pH 8.3-8.4. A trial of a coral extract or an amino acid solution consisted of five or six perfusion experiments. The spike count for each IO-see period in each experiment was taken by hand from the chart record and entered into a computer, and a statistical program was used to calculate mean spike count and standard error for each lo-set interval across replicates. The computer program was then used to generate a time/firing-rate graph for each experiment.

RESULTS In P. sibogae, both oral tentacles and rhinophores are innervated from the brain. The brain of this sea slug, as in other gaatropods, is made up of three pairs of fused or partially fued cerebral, pleural and pedal ganglia (Fig. 1). Each ganglia contains large identifiable neurons (36) such as characterize the central nervous systems of all well-studied opisthobranch molluscs (23). Each oral tentacle is innervated by two small nerves that arise from the anteroventral region of each cerebral ganglion, without intervening ganglia (Fig. 1). On each side, a short, stout nerve trunk extends dorsoanteriorly from each cerebral ganglion to a rhinophoral ganglion from which five to six nerves run distally into each rhinophore (Fig. 1). The nerves extending from the distal surface of the rhinophoral ganglion divide into progressively

729

finer branches that extend throughout the length of the rhinophore. Experiments with iontophoretic injection of Lucifer Yellow into cell bodies in the rhinophoral ganglion showed that neurons from at least some of the cells in the rhinophoral ganglion synaptically interact with cells in an anterior region of the cerebral ganglion, which are characterized by the presence of many small neuronal somata. Continuous and regular discharge was recorded from the cut ends of the rhinophoral and oral-tentacle nerves when the organs were perfused with seawater. The oral-tentacle output was regular and slowly declined from a mean of about 40 counts/IO set to a mean of around 20 counts/l0 set at the end of a 30-min experiment (Fig. 2A). In the first perfusion experiments, Trizma buffer, added to seawater in all subsequent experiments, was tested and found to be without effect on the continuous output from the oral tentacles or rhinophores (Figs. 2A and 3A). When the oral tentacles were perfused with P. compressa-conditioned water, there was no response (Fig. 2B). When the oral tentacles were perfused with 5 mM L-aspartic (Fig. 2C) and L-glutamic acid in FSW (Fig. 2D), small positive responses were evident. The continuous afferent output of the rhinophores was more variable than that of the oral tentacles and appeared to decay less over the course of an experiment. Rhinophores responded with large, yet short-lived, positive responses when perfused with P. compressa-conditioned water (Fig. 3B). The rhinophores also responded to water conditioned by P. lobata, another food-source coral, with a smaller yet longer lived increase in firing rate (Fig. 4A). A small increase in firing rate was seen when a rhinophore was perfused with water conditioned by the non-food-source coral, P. meandrina (Fig. 4B). However, there was no or only a small effect when water from another non-food-source coral species, M. P)ewucosa, was applied (Fig. 4C). When rhinophores were perfused with solutions of 5 mM aspartic acid or glutamic acid, distinctively biphasic responses occurred (Fig. 3C and D). The response to aspartic acid was still biphasic at 1 mM (Fig. 5A) but only slightly positive at 0.1 mM. The rhinophoral response to glutamic acid was still biphasic at 1.0 mM (Fig. 5B) and at 0.1 mM (Fig. 5C) but barely detectable at 0.01 mM. Perfusion of the rhinophores with solutions of several other amino acids, all at a concentration of 5 mM, produced varying positive responses, all smaller in magnitude than those elicited by glutamate and aspartate. Alanine, lysine, glycine and serine elicited moderate responses in the rhinophores (Fig. 6AD), whereas GABA, arginine and betaine stimulated little or no response. There were significant firing-rate increases in the rhinophoral nerve when the rhinophore was perfused with the glutamate-receptor modulators o-glutamate (5 mM, Fig. 7A), 6,7-dinitroquinoxaline-2,3-dione (DNQX) (1 mM, Fig. 7B) and kainic acid (0.1 and 0.01 mM, Fig. 7C and D) but not with application of N-methyl-D-aspartate (NMDA). The positive part of the glutamate/aspartate re-

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FIG. 2. Firing rate/time plots of continuous oral tentacle neural output; test solution was applied between arrows. Each dot represents the mean tiring rate per 10 set calculated from five to six separate experiments; vertical lines through dots display SEM. Test solutions were (A) buffered seawater, (B) seawater conditioned with PO&es compressor, (C) 5 mM t-aspartic acid and (D) 5 mM L-glutamic acid.

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FIG. 3. Firing rate/time plots of continuous rhmophoral nerve output; test solution was applied between arrows. Each dot vertical lines through dots display represents the mean firing rate per 10 set calculated from five to six separate experiments; SEM. Test solutions were (A) buffered seawater, (B) seawater conditioned with Porites compressa, (C) 5 mM L-aspartic acid and (D) 5 mM L-glutamic acid.

Chemoreception

in a Nudibranch

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by perfusion of the rhinophore

ter alone did not affect the spontaneous

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afferent output of neither the oral tentacles

nor the rhino-

phores was affected by perfusion of the organ with seawater that had been conditioned

100 80 60 40 20

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either magnesium-substituted (Fig. 8A) or cobalt-substituted (Fig. 8B) calcium-free seawater. Calcium-free seawathe rhinophoral

120

with adults of P. sibogue.

DIGCUSSION

‘I”“,

I’. sibogae finds a very specific coral

prey by distance chemoreception (17 and data presented here). To further understand the location and nature of che-

30

5. Firing rate/time plots of continuous rhinophoral nerve output; test solution was applied between arrows. Each dot representsthe mean fuing rate per 10 set calculated from five to six separate experiments, vertical lines through dots display SEM. Test solutionswere (A) 1.O mM L-asparticacid, (B) 1.0 mM L-glutamicacid and (C) 0.1 mM L-glutamic acid. FIG.

many small cells. The function

The tropical nudibranch

25

Elapsed Tame (Min)

of this region of the brain

remains to be discovered. Because the recording electrodes in this study encompassed

the rhinophoral

nerve proximal

moreceptive capacities in P. sibogae, we focused our studies on the two pairs of tentacles arising from the head, cephalic

to the rhinophoral ganglion, they probably received a mixture of signals, some arising in distal receptors and some after synaptic transmission in the ganglion. Interpretation

tentacles

of the data is based on this assumption.

having been implicated

in chemoreception

of all

gastropods studied. As in other gastropods, the main sensory netves from the oral tentacles and rhinophores communicate

with the cerebral

ganglion

of P. sibogae

(Fig.

The main nerves originating rhinophores

of P. sibogue conduct

in the oral tentacles

and

large numbers of action

1). Oral-

potentials. Our recordings of neural output after the applica-

tentacle nerves pass directly to the cerebral ganglia, whereas those from the rhinophores pass first into a rhinophoral gan-

tion of a set of specific odorants demonstrate that at least part of the peripheral activity of the rhinophores (Fig. 3),

glion. Iontophoretic injection of cell bodies in the rhinophoral ganglion demonstrated that at least some of the

but not the oral tentacles

(Fig. 2), is chemosensory.

These

rhinophoral neurons synapse in this ganglion with intenneurons whose axons pass through a short tract to an area

findings are consistent with physiological and behavioral observations reported by Bicker et al. (3,4) for the notaspidian sea slug P. californica but contrast with behavioral obser-

in the anterior part of the cerebral ganglia characterized

vations on two other nudibranch

by

species, H. crassicomis

(1)

B. F. Murphy and M. G. Hadfield

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FIG. 6. Firing rate/time plots of continuous rhmophoral nerve output; test solution was applied between arrows. Each dot represents the mean firing rate per 10 set calculated from five to six separate experiments; vertical lines through dots display SEM. Test solutions were (A) 5 mM L-alanine, (B) 5 mM blysine, (C) 5 mM bglycine and (D) 5 mM L-serine.

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Chemoreception

733

in a Nudibranch

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and Tritonia diomedea (IO), whose oral tentacles were found to be the major peripheral organs involved in chemoreception and whose rhinophores were thought to be important only for rheotaxis. However, the same odorants were not used in all of these studies, making comparisons equivocal. Although we cannot be certain that the action potentials monitored by our suction electrodes arose directly as output from sensory endings, their persistence when petroleum jelly barriers were placed between the stimulus sites on the rhinophoral epithelium and the recording sites proximal to the rhinophoral ganglia demonstrates that, regardless of the degree of intermediate processing, the signals ultimately originated in distal regions of the rhinophores as responses to the stimulating substances. Bicker et al. (4), using nervebackfilling techniques, found that axons arising directly from sensory cells in the rhinophoral epithelium of P. califomica encountered no synapses distal to the rhinophoral ganglion. Additionally, Xin et al. (37) showed that chemostimuli applied to the head of Aplysia produced synaptic input to the cerebral ganglia even when peripheral synaptic activity was blocked and that sensory afferents with peripheral cell bodies provide much input to the central nervous system in this species. Also for Aplysia, Fredman and JahanParwar ( 11) showed that chemosensory responses from the tentacles to the cerebral ganglia were unaffected when peripheral synaptic activity was blocked.

The data presented here amply support the contention that P. sibogae detects its prey corals (Fig. 3) via chemosensory endings on its rhinophores and not its oral tentacles. The observation of physiological output in response to at least one non-food coral species (Fig. 4B) makes it likely that rhinophoral receptors are stimulated by a mixture of substances in our coral-conditioned waters, not all of them attractants to the nudibranch. Certainly, the slugs’ behavioral responses to these coral signals are vastly different (unpublished personal observations), and each species of coral may have a very different chemical signature, composed of different molecules or the same molecules in different ratios. Physiological data presented here demonstrate that P. sibogae, like many other vertebrate and invertebrate aquatic animals, detects dissolved amino acids. Furthermore, 19 different amino acids were detected by HPLC in seawater in which a head of P. com~ressu had been left standing for 1 hr, and a prominent peak occurred at the point where aspartic acid eluted in control samples (data collected in collaboration with D. Manahan). Of all the amino acids tested here, the largest and most complex responses of the rhinophores of P. sibogae were to aspartic and glutamic acids. Thus, the possibility exists that amino acids participate in the attraction of P. sibogae to its p’ey coral. The rhinophores of P. sibogae appear to bear receptors for more than one type of amino acid. At least one receptor group responds strongly to acidic amino acids and poorly to basic amino acids. The biphasic response to acidic amino acids (Figs 3 and 5) suggests the presence of at least two aspartate/glutamate receptors, although one of these may be at the site of a synapse in the rhinophoral ganglion. The similarity of responses to aspartic and glutamic acids, except at low concentrations (Figs 3 and 5), allows two alternate interpretations: only a single receptor is present, and it is more sensitive to glutamate, or two different receptors are present. Yarowsky and Carpenter (38) reported separate aspartate and glutamate receptors on the neurons of Aplysia and at least two classes for each. Research currently underway in out laboratory seeks to determine if aspartate and glutamate have separate receptors in I’. sibogae. Three types of synaptic glutamate receptors have been demonstrated in molluscan neurons (28,35), and it is apparent that the positive glutamate response in rhinophores of P. sibogae is due to a non-NMDA receptor of the kainic acid-sensitive subtype (i.e., the positive response to D-glut& mate [Fig. 7A] and kainic acid at 0.01 mM [Fig. 7D] and the lack of response to even high concentrations of NMDA). The large positive response to the kainatereceptor-subtype antagonist DNQX (Fig. 7B) provides more evidence for the presence of this particular subtype in the rhinophores of P. sibogae. However, the unexpected positive nature of the response to DNQX emphasizes again the lack of consistent pharmacological responses of glutamate t-eceptots across phyla [e.g., (21,24,31)]. Kainic acid had no effect on the chloride

or potassium

734

B. F. Murphy

currents generated by glutamate Aplysiu (24) nor did it suppress cultured

olfactory

rnaz maximus

interneurons

(30).

of the response

in cultured action-potential

neurons tiring

from the terrestrial

We interpret

to glutamate

the initial,

of in

slug Li-

positive

part

in P. sibogae to be synaptic,

spectrum

of aquatic

amino

acids.

tween

internal

should strated

animals,

Although

and

conservation

and external

and M. G. Hadtield

synaptic

receptors

of receptor

parts of animal

for

types be-

nervous

systems

not be surprising, it has not been broadly demon[however, see Carr et al. (S)]. Extensive data on glu-

generated in the rhinophoral ganglion after the initial olfactory transduction event, because it was eliminated by perfu-

tamate-receptor sequences from rats and the freshwater snail Lymnatla stag&s (20,32) may permit detinitive com-

sion with calcium-free

parisons.

and

B). The

seawater

subsequent

(compare

negative

part

aspartate response probably represents tion event in peripheral chemoreception, eliminated receptor

by calcium-free

seawater

may be of the ibotenic to coral extract

the initial transducbecause it was not (Fig. 8A and

8); its sub-

is positive

response

of the rhino-

is difficult

tc> interpret

light of this, but the initial olfactory transduction be negative and its sign subsequently reversed cessing

by interneurons

There sible

is apparent

for amino

A. californica,

Because

glutamate-

acid glutamate-receptor

type (3 1). The fact that the measured phore

Figs 3D and 8A of the

in the rhinophoral

lack of homology

acid detection

gastropods.

phores

In the latter

are more sensitive

cluding

alanine,

aspartic

two species,

to a number

to

acid,

glutamic

acid,

and, for P. californica but not P. sibogm, betaine personal

communication).

the rhinophores

The

is similar

the rhino-

of amino

anatomy

acids, giycine

leaves the possibility

that their

olfactory

conspecific

animals.

I-‘. sibogue routinely large reproductive

differs between

in-

(R. Gillette, ot

and I’. sibogae

the oral tentacles

converge

the two species

on a ganglion

in ccjntact

oral tentacle glia. There larvae

nerves

also pass directly

are also differences from

different

in the responses

taxonomic

from

orders

2

3

gan-

of gastn)poJ

to amino

4.

acids.

Amino acids have no morphogenetic affect on veliger larvae of P. sibogae (12,13), whereas larvae of the prosobranch gastropod

Haliotis rufescens

are readily

induced

aggregations.

remains

chemoreception

References 1 Agershorg,

two nerve>

the cerebral ganglia H. crussicornis (2), into the cerebral

In aquaria,

adult

follow each other’s slime trails and form However,

in our tests, nei-

unknown;

perhaps

of conspecific

to of

their role lies

slime trails.

This research was supported by Ofice of Naval Research pant> NO@O14-91-I-1533 and N00014-94-1-0524. We are indebted to Dr. Bernhard Ruthensteiner for preparing Figure 1 from serial sections and Lo Dr. Esther Leise for her critical reading of the manuscript. B F M thanks Dr. James Larimer for use of a computer for word processing.

distal to the brain

in P. califoomica (4) but not in I’. sibogm, where from each oral tentacle directly enter on each side]. In another nudibranch,

[nerves

role, if they have

ther the oral tentacles nor the rhinophores responded seawater conditioned with I’. sibogm, and the function

(rhinophoral nerves converge on rhinophoral ganglia, not directly on the cerebral ganglia), whereas innervation of the oral tentacles

lies in locating

(3)

of innervation

in P. californicu

only very slightly to amino to be in food detection. This

In

are much more sensitive

aspartic and glutamic acids than the rhinophnres (23), a condition that is reversed in P. sibogae and the notaspidean sea slug P. californica.

of P. sibogae do not respond

one,

the oral tentacles

ganglion.

in the organs respon-

in different

the oral tentacles

in

event may during pro-

the oral tentacles

to ct~~l extracts and respond acids, their function is unlikely

to metamc>r-

5.

phose by GABA (27). Extensive investigations of the metamorphosis-inducing pathways of larvae of the abalone Haliotis rufescem have demonstrated the presence of an external receptor that, although typically sensing an unidentified algal product, shows great functional similarities to GABA

6

synaptic receptors of vertebrates [e.g., (9,26,27,34)]. Our results support the interesting hypothesis that exter-

7.

nal receptors sensitive to glutamic acid may be structurally similar to at least one class of glutamate synaptic receptors

$.

that has been most extensively studied in vertebrates. It will be of considerable interest to learn if there are evolutionary links between olfaction of amino acids, broadly sensed in a

Y.

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