Nudibranch Mollusc Research Articles

Associations between the White Sea invertebrates and oxygen-evolving phototrophic microorganisms

Eleven species of White Sea invertebrates (sponges, actinians, hydroids, polychaetes, and nudibranch mollusks) were tested for the presence of associated oxygen-evolving phototrophic microorganisms (OPM) (microalgae and cyanobacteria). Two main approaches were applied: (a) light and electron microscopy of intact samples and fixed preparations of invertebrates, and (b) isolation of microorganisms from samples of invertebrates after mild surface sterilization. The obtained results lead to conclusions on the formation of multicomponent associations by White Sea invertebrates and OPM based on the following data: (1) isolation of 27 cultures of OPM from eight species of invertebrates (sponges, hydroids, polychaete trochophore larva), (2) specificity of association between epibiontic communities of microorganisms and macroorganisms within the same biotope, and (3) spatial integration of micro- and macropartners resulting in the formation of morphological structures within the interorganismic contact zones.

Cnidosac morphology in dendronotacean and aeolidacean nudibranch molluscs: from expulsion of nematocysts to use in defense?

Nudibranchs (Mollusca, Gastropoda) feeding on tentacles and polyps of Cnidaria thereby ingest both latent and discharged nematocysts (NCs) along with the food mass. In eolid nudibranchs (Opisthobranchia, Aeolidacea), many of the undischarged NCs are transported to terminal cnidosacs in their body appendages (cerata) and incorporated as kleptocnidae for defense. In the present report, the occurrence and fate of NCs in the digestive tracts of eolids is compared with hydrozoan-feeding dendronotacean nudibranchs (Opisthobranchia, Dendronotacea), which may show more basic stages in the evolution of cnidosacs. Tomographic reconstructions of the distal tips of cerata were composed from series of semithin light microscopic sections, utilizing 3D-surface rendering software. Doto acuta (Dendronotacea, Dotidae) does not have cnidosacs; transmission electron micrographs show that the NCs are digested in lysosomes of digestive gland cells. In contrast, species of the genus Hancockia (Dendronotacea, Hancockiidae) have several small cnidosacs in each ceras; they accumulate NCs in the digestive cells, as well as in the cnidosacs. Many of the cnidosacs were found open to the exterior with NCs in the process of expulsion. These and other structural details suggest assigning a function of expelling the NCs to the Hancockia spp. cnidosacs. It is proposed that cnidosacs similar to those of Hancockia spp. provide a clue to understanding how the defensive function of eolid cnidosacs may have evolved.

Comparative Study of TPep-Like Immunoreactive Neurons in the Central Nervous System of Nudibranch Molluscs

In the sea slug Tritonia diomedea, mucociliary crawling is controlled partly by two pairs of bilaterally symmetrical neurons located in the pedal ganglia. These neurons, known as the Pedal 5 and Pedal 6 cells, produce a class of neuropeptides called TPeps. Using immunohistochemistry we identified TPep-like immunoreactive (TPep-LIR) neurons in diverse nudibranch species. All species examined had 2–7 large, TPep-LIR cells located in each pedal ganglion. The absolute size of the largest TPep-LIR neuron was correlated with foot size. Species with a bigger foot size tended to have larger TPep-LIR cells. However, the number of cells in a given species was not correlated with the size of the adult foot. The presence of large, TPep-LIR cells across the nudibranchs suggests that part of the neural circuitry controlling mucociliary locomotion has been conserved, although the size and number of cells is variable across species. We conclude that the motor circuit underlying crawling might adapt to changes in foot size by changing the size of motor neurons in the circuit, but that changes in cell number are not directly related to foot size.

Sensory Regulation of Network Components Underlying Ciliary Locomotion inHermissenda

Ciliary locomotion in the nudibranch mollusk Hermissenda is modulated by the visual and graviceptive systems. Components of the neural network mediating ciliary locomotion have been identified including aggregates of polysensory interneurons that receive monosynaptic input from identified photoreceptors and efferent neurons that activate cilia. Illumination produces an inhibition of type I(i) (off-cell) spike activity, excitation of type I(e) (on-cell) spike activity, decreased spike activity in type III(i) inhibitory interneurons, and increased spike activity of ciliary efferent neurons. Here we show that pairs of type I(i) interneurons and pairs of type I(e) interneurons are electrically coupled. Neither electrical coupling or synaptic connections were observed between I(e) and I(i) interneurons. Coupling is effective in synchronizing dark-adapted spontaneous firing between pairs of I(e) and pairs of I(i) interneurons. Out-of-phase burst activity, occasionally observed in dark-adapted and light-adapted pairs of I(e) and I(i) interneurons, suggests that they receive synaptic input from a common presynaptic source or sources. Rhythmic activity is typically not a characteristic of dark-adapted, light-adapted, or light-evoked firing of type I interneurons. However, burst activity in I(e) and I(i) interneurons may be elicited by electrical stimulation of pedal nerves or generated at the offset of light. Our results indicate that type I interneurons can support the generation of both rhythmic activity and changes in tonic firing depending on sensory input. This suggests that the neural network supporting ciliary locomotion may be multifunctional. However, consistent with the nonmuscular and nonrhythmic characteristics of visually modulated ciliary locomotion, type I interneurons exhibit changes in tonic activity evoked by illumination.

Oxygen profiles in egg masses predicted from a diffusion–reaction model

We developed a novel diffusion-reaction model to describe spatial and temporal changes in oxygen concentrations in gelatinous egg masses containing live, respiring embryos. We used the model in two ways. First, we constructed artificial egg masses of known metabolic density using embryos of the Antarctic sea urchin Sterechnius neumayeri, measured radial oxygen profiles at two temperatures, and compared our measurements to simulated radial oxygen profiles generated by the model. We parameterized the model by measuring the radius of the artificial masses, metabolic densities (=embryo metabolic rate x embryo density) and oxygen diffusion coefficients at both ambient (-1.5 degrees C) or slightly warmer (+1.5-2 degrees C) temperatures. Simulated and measured radial oxygen profiles were similar, indicating that the model captured the major biological features determining oxygen distributions. Second, we used the model to analyze sources of error in step-change experiments for determining oxygen diffusion coefficients (D), and to determine the suitability of simpler, analytical equations for estimating D. Our analysis indicated that embryo metabolism can lead to large (several-fold) overestimates of D if the analytical equation is fitted to step-down-traces of central oxygen concentration (i.e. external oxygen concentration stepped from some high value to zero). However, good estimates of D were obtained from step-up-traces. We used these findings to estimate D in egg masses of three species of nudibranch molluscs: two Antarctic species (Tritonia challengeriana and Tritoniella belli; -1.5 and +2 degrees C) and one temperate Pacific species (Tritonia diomedea; 12 and 22 degrees C). D for all three species was approximately 8 x 10(-6) cm(2) s(-1), and there was no detectable effect of temperature on estimated D. For the Antarctic species, D in egg masses was 70-90% of its value in seawater of similar temperature.

Temperature–oxygen interactions in Antarctic nudibranch egg masses

The Southern Ocean is one of the coldest, most stable marine environments on Earth and represents a unique environment for investigating metabolic consequences of low temperature. Here we test predictions of a new diffusion-reaction model of O(2) distributions in egg masses, using egg masses of the Antarctic nudibranch mollusk, Tritonia challengeriana. When warmed from -1.5 degrees to +1.5 degrees C, embryos of T. challengeriana showed large increases in O(2) consumption (Q(10) values of 9.6-30.0). Oxygen electrode measurements in intact masses showed, however, that O(2) levels were high throughout and virtually unaffected by temperature. The model suggested that both effects stemmed from very low metabolic densities in egg masses. Detailed morphological measurements of egg masses of T. challengeriana and a temperate congener, T. diomedea, revealed large differences in structure that may be related to O(2) availability. Egg masses of T. challengeriana were approximately twice as thick. However, the most dramatic effects were observed in embryos: embryos of T. challengeriana were >32 times larger (by volume) than embryos of T. diomedea. Antarctic embryos also were contained singly in large egg capsules ( approximately 500 mum diameter). Consequently, Antarctic embryos occurred at much lower densities, with very low metabolic densities.

Granular Chitin in the Epidermis of Nudibranch Molluscs

Chitin is usually found in stiff extracellular coatings typified by the arthropod exoskeleton, and is not associated with the soft, flexible mollusc skin. Here, we show, however, that chitin in nudibranch gastropods (Opisthobranchia, Mollusca) occurs as intracellular granules that fill the epidermal cells of the skin and the epithelial cells of the stomach. In response to nematocysts fired by tentacles of prey Cnidaria, the epidermal cells of eolid nudibranchs (Aeolidacea) release masses of chitin granules, which then form aggregates with the nematocyst tubules, having the effect of insulating the animal from the deleterious action of the Cnidaria tentacles. Granular chitin, while protecting the animal, does not interfere with the suppleness and flexibility of the skin, in contrast to the stiffness of chitin armor. The specialized epidermis enables nudibranchs to live with and feed on Cnidaria.

Lipid Classes and Fatty Acid Composition of the Tropical Nudibranch Mollusks Chromodoris sp. and Phyllidia coelestis

Two nudibranch mollusks, Chromodoris sp. and Phyllidia coelestis, collected from tropical waters of the Northwestern Pacific, were analyzed for lipids. The aim of this study was to fill the gap in knowledge of lipid biochemistry of mollusks. Phospholipids (PL) were the dominating lipid class followed by sterols (13%). Neutral lipids were not detected in Chromodoris sp. By contrast, P. coelestis contained TAG, diacylglyceryl ether, long chain alcohol and esters of sterols. Among PL, PC was predominant (about 50%); PE, PS and CAEP were almost in equal proportions. Sixty five FA were identified as methyl esters and N-acyl pyrrolidides by GC-MS. The sea slugs exhibited a wide diversity of FA. The common marine n-3 PUFA, 20:5n-6 and 22:6n-3, constituted 0.6-1.3% of the total FA, whereas n-6 PUFA, 22:4n-6, 20:4n-6, and 18:2n-6, were the main (25%). Among monounsaturated FA, 7-21:1 was the main (up to 6.2%). The non-methylene-interrupted (NMI) FA were found (9.4 and 12.4%), including the known 5,11-20:2, 5,13-20:2, 7,13-22:2, 7,15-22:2 and a novel isomer 7,13-21:2 (up to 3.9%). The pathway of its biosynthesis was suggested. A series of very long chain FA (VLC FA), with the main 5,9-25:2 and 5,9-26:2, were identified. High level of VLC FA (8.7 and 11.7%) in sea slugs is apparently the result of predation on sponges. Another unique feature concerned a high abundance of various odd and branched FA (16.7 and 34%), which could have originated from the dietary origin or symbiotic bacteria. This is the first report on lipid and FA composition of nudibranchs.

Homologues of serotonergic central pattern generator neurons in related nudibranch molluscs with divergent behaviors

Homologues of a neuron that contributes to a species-specific behavior were identified and characterized in species lacking that behavior. The nudibranch Tritonia diomedea swims by flexing its body dorsally and ventrally. The dorsal swim interneurons (DSIs) are components of the central pattern generator (CPG) underlying this rhythmic motor pattern and also activate crawling. Homologues of the DSIs were identified in six nudibranchs that do not exhibit dorsal-ventral swimming: Tochuina tetraquetra, Melibe leonina, Dendronotus iris, D. frondosus, Armina californica, and Triopha catalinae. Homology was based upon shared features that distinguish the DSIs from all other neurons: (1) serotonin immunoreactivity, (2) location in the Cerebral serotonergic posterior (CeSP) cluster, and (3) axon projection to the contralateral pedal ganglion. The DSI homologues, named CeSP-A neurons, share additional features with the DSIs: irregular basal firing, synchronous inputs, electrical coupling, and reciprocal inhibition. Unlike the DSIs, the CeSP-A neurons were not rhythmically active in response to nerve stimulation. The CeSP-A neurons in Tochuina and Triopha also excited homologues of the Tritonia Pd5 neuron, a crawling efferent. Thus, the CeSP-A neurons and the DSIs may be part of a conserved network related to crawling that may have been co-opted into a rhythmic swim CPG in Tritonia.

Adaptation of Underwater Video for Near-Substratum Current Measurement

We describe a new method to measure current near the substratum. A variety of current meters are available to record flow data relevant to benthic organisms in their natural habitat. Several factors, including accuracy and precision, sample volume, sample rate, deployment constraints, and cost make them more or less appropriate to different applications. Our method, based on tracking particles in videos from a camera mounted near the substratum, provides an inexpensive option for measuring flow. We validate this “current camera” against a conventional electromagnetic meter, and then use it successfully to estimate flows experienced by the nudibranch Tritonia diomedea in its natural habitat. As we implement it, this current meter samples a large volume and provides a measure of bulk flow in the habitat. However, the method is adaptable to videos acquired from other cameras at a variety of size scales and locations. We therefore conclude that our method is an option that should be considered by biologists interested in measuring flow. In a recent field study (1), we used current direction to partially explain navigation behaviors of the nudibranch mollusc Tritonia diomedea Bergh. To avoid interference with the behaviors and our ability to observe them, we chose to measure flow some distance away from the slugs. As a consequence of this constraint, we needed integrated current headings (over time or space) to overcome the heterogeneous nature of flow in this habitat and thus allow us to estimate flow in nearby regions. Furthermore, we needed measurements made near the substratum, to correspond with flow experienced by the benthic slugs. Several current meter options are available. These include acoustic doppler velocimeters (ADVs), acoustic doppler current profilers (ADCPs), and electromagnetic meters. However, none of

Comparative mapping of serotonin‐immunoreactive neurons in the central nervous systems of nudibranch molluscs

The serotonergic systems in nudibranch molluscs were compared by mapping the locations of serotonin-immunoreactive (5-HT-ir) neurons in 11 species representing all four suborders of the nudibranch clade: Dendronotoidea (Tritonia diomedea, Tochuina tetraquetra, Dendronotus iris, Dendronotus frondosus, and Melibe leonina), Aeolidoidea (Hermissenda crassicornis and Flabellina trophina), Arminoidea (Dirona albolineata, Janolus fuscus, and Armina californica), and Doridoidea (Triopha catalinae). A nomenclature is proposed to standardize reports of cell location in species with differing brain morphologies. Certain patterns of 5-HT immunoreactivity were found to be consistent for all species, such as the presence of 5-HT-ir neurons in the pedal and cerebral ganglia. Also, particular clusters of 5-HT-ir neurons in the anterior and posterior regions of the dorsal surface of the cerebral ganglion were always present. However, there were interspecies differences in the number of 5-HT-ir neurons in each cluster, and some clusters even exhibited strong intraspecies variability that was only weakly correlated with brain size. Phylogenetic analysis suggests that the presence of particular classes of 5-HT-ir neurons exhibits a great deal of homoplasy. The conserved features of the nudibranch serotonergic system presumably represent the shared ancestral structure, whereas the derived characters suggest substantial independent evolutionary changes in the number and presence of serotonergic neurons. Although a number of studies have demonstrated phylogenetic variability of peptidergic systems, this study suggests that serotonergic systems may also exhibit a high degree of homoplasy in some groups of organisms.

Dopamine modulation of Ca2+ dependent Cl- current regulates ciliary beat frequency controlling locomotion inTritonia diomedea

The physiological mechanisms controlling ciliary beating remain largely unknown. Evidence exists supporting both hormonal control of ciliary beating and control via direct innervation. In the present study we investigated nervous control of cilia based locomotion in the nudibranch mollusc, Tritonia diomedea. Ciliated pedal epithelial (CPE) cells acting as locomotory effectors may be electrically excitable. To explore this possibility we characterized the cells' electrical properties, and found that CPE cells have large voltage dependent whole cell currents with two components. First, there is a fast activating outward Cl(-) current that is both voltage and Ca(2+) influx dependent (I(Cl(Ca))). I(Cl(Ca)) is sensitive to DIDS and 9-AC, and resembles currents of Ca(2+)-activated Cl(-) channels (CaCC). Ca(2+) dependence also suggests the presence of voltage-gated Ca(2+) channels; however, we were unable to detect these currents. The second current, a voltage dependent proton current (I(H)), activates very slowly and is sensitive to both Zn(2+) and changes in pH. In addition we identify a new cilio-excitatory substance in Tritonia, viz., dopamine. Dopamine, in the 10 mumol l(-1)-1 mmol l(-1) range, significantly increases ciliary beat frequency (CBF). We also found dopamine and Tritonia Pedal Peptide (TPep-NLS) selectively suppress I(Cl(Ca)) in CPE cells, demonstrating a link between CBF excitation and I(Cl(Ca)). It appears that dopamine and TPep-NLS inhibit I(Cl(Ca)) not through changing [Ca(2+)](in), but directly by an unknown mechanism. Coupling of I(Cl(Ca)) and CBF is further supported by our finding that DIDS and zero [Cl(-)](out) both increase CBF, mimicking dopamine and TPep-NLS excitation. These results suggest that dopamine and TPep-NLS act to inhibit I(Cl(Ca)), initiating and prolonging Ca(2+) influx, and activating CBF excitation.

Nervous control of ciliary beating by Cl-, Ca2+ and calmodulin inTritonia diomedea

In vertebrates, motile cilia line airways, oviducts and ventricles. Invertebrate cilia often control feeding, swimming and crawling, or gliding. Yet control and coordination of ciliary beating remains poorly understood. Evidence from the nudibranch mollusc, Tritonia diomedea, suggests that locomotory ciliated epithelial cells may be under direct electrical control. Here we report that depolarization of ciliated pedal epithelial (CPE) cells increases ciliary beating frequency (CBF), and elicits CBF increases similar to those caused by dopamine and the neuropeptide, TPep-NLS. Further, four CBF stimulants (zero external Cl(-), depolarization, dopamine and TPep-NLS) depend on a common mode of action, viz. Ca(2+) influx, possibly through voltage-gated Ca(2+) channels, and can be blocked by nifedipine. Ca(2+) influx alone, however, does not provide all the internal Ca(2+) necessary for CBF change. Ryanodine receptor (RyR) channel-gated internal stores are also necessary for CBF excitation. Caffeine can stimulate CBF and is sensitive to the presence of the RyR blocker dantrolene. Dantrolene also reduces CBF excitation induced by dopamine and TPep-NLS. Finally, W-7 and calmidazolium both block CBF excitation by caffeine and dopamine, and W-7 is effective at blocking TPep-NLS excitation. The effects of calmidazolium and W-7 suggest a role for Ca(2+)-calmodulin in regulating CBF, either directly or via Ca(2+)-calmodulin dependent kinases or phosphodiesterases. From these results we hypothesize dopamine and TPep-NLS induce depolarization-driven Ca(2+) influx and Ca(2+) release from internal stores that activates Ca(2+)-calmodulin, thereby increasing CBF.

Spongian Diterpenes from Australian Nudibranchs: An Anatomically Guided Chemical Study ofGlossodorisatromarginata

An Australian population of the nudibranch mollusk Glossodoris atromarginata has been found to contain furanoditerpenes of the spongian series. Spongia-13(16),14-dien-3-one (1) and 3alpha-acetoxy-19-hydroxyspongia-13(16),14-dien-2-one [corrected] (2) were isolated for the first time from a natural source, along with a series of known diterpenes (3-7). Anatomical dissection of the animals revealed the relative distribution and chemical variation of secondary metabolites. Structural studies have provided a basis for chemical comparisons between populations from different geographic locations.

Odours detected by rhinophores mediate orientation to flow in the nudibranch mollusc,Tritonia diomedea

Tritonia diomedea is a useful neuroethological model system that can contribute to our understanding of the neural control of navigation. Prior work on both sensory and locomotory systems is complemented by recent field experiments, which concluded that these animals primarily use a combination of odours and water flow as guidance cues. We corroborate these field results by showing similar navigation behaviours in a flow tank. Slugs crawled upstream towards both prey and conspecifics, and turned downstream after crawling into a section of the flow tank downstream of a predator. Controls without upstream odour sources crawled apparently randomly. We then tested whether these behaviours depend on odours detected by the rhinophores. Outflow from a header tank was used to generate prey, predator and unscented control odour plumes in the flow tank. Slugs with rhinophores crawled upstream towards a prey odour plume source, turned downstream in a predator odour plume, and showed no reaction to a control plume. Slugs without rhinophores behaved similarly to controls, regardless of odour plume type. Finally, we used extracellular recordings from the rhinophore nerve to demonstrate that isolated rhinophores are chemosensitive. Afferent activity increased significantly more after application of all three odour types than after unscented control applications. Responses were odour specific. We conclude that rhinophores mediate orientation to flow, and suggest that future work should focus on the integration of mechanosensation and chemosensation during navigation in T. diomedea.

Orientation and Navigation Relative to Water Flow, Prey, Conspecifics, and Predators by the Nudibranch MolluscTritonia diomedea

Progress in understanding sensory and locomotory systems in Tritonia diomedea has created the potential for the neuroethological study of animal navigation in this species. Our goal is to describe the navigational behaviors to guide further work on how the nervous system integrates information from multiple senses to produce oriented locomotion. Observation of T. diomedea in its habitat has suggested that it uses water flow to navigate relative to prey, predators, and conspecifics. We test these hypotheses in the field by comparing slug orientation in time-lapse videos to flow direction in circumstances with and without prey, predators, or conspecifics upstream. T. diomedea oriented upstream both while crawling and after turning. This trend was strongest before feeding or mating; after feeding or mating, the slugs did not orient significantly to flow. Slugs turned downstream away from an upstream predator but did not react in control situations without an upstream predator. These data support the hypothesis that T. diomedea uses a combination of odors (or some other cue transported downstream) and water flow to navigate relative to prey, predators, and conspecifics. Understanding the context-dependent choice between upstream and downstream crawling in T. diomedea provides an opportunity for further work on the sensory integration underlying navigation behavior.

Field Behavior of the Nudibranch MolluscTritonia diomedea

The nudibranch mollusc Tritonia diomedea has been a useful model system for studies of how the brain controls behavior. However, no broad study of T. diomedea field behavior exists--an important deficit since laboratory behaviors may differ from what occurs in nature. Here we report analysis of time-lapse video of the slugs in their natural habitat to describe behaviors and their relationships to sensory cues. We found that movements relative to conspecifics, prey, and predators correlated with direction of water flow. These observations lead to three new navigational hypotheses: regardless of the actual heading to the target, T. diomedea crawls (1) upstream toward potential mates, (2) upstream toward food, and (3) downstream away from predators. We also describe both the behavior and its sensory context for feeding, escape swims, mating, and egg-laying, among other behaviors. Field behaviors were similar to published descriptions of laboratory behavior. However, the field observations add contextual detail, including preceding and subsequent behaviors and interactions with suites of habitat features not present in the laboratory. For example, the escape swim, previously studied as an isolated behavior in response to a single stimulus, appears to be affected by multiple sensory modalities and coordinated with several other behaviors. Our work will provide a basis for future neuroethological experimentation and also is the first step in the study of navigation in T. diomedea.

Invertebrate studies and their ongoing contributions to neuroscience

Invertebrates have been deployed very successfully in experimental studies of the nervous system and neuromuscular junctions. Many important discoveries on axonal conduction, synaptic transmission, integrative neurobiology and behaviour have been made by investigations of these remarkable animals. Their advantages as model organisms for investigations of nervous systems include (a) the large diameter of neurons, glia and muscle cells of some invertebrates, thereby facilitating microelectrode recordings; (b) simple nervous systems with few neurons, enhancing the tractability of neuronal circuitry; and (c) well-defined behaviours, which lend themselves to physiological and genetic dissection. Genetic model organisms such as Drosophila melanogaster and Caenorhabditis elegans have provided powerful genetic approaches to central questions concerning nervous system development, learning and memory and the cellular and molecular basis of behaviour. The process of attributing function to particular gene products has been greatly accelerated in recent years with access to entire genome sequences and the application of reverse genetic (e.g. RNA interference, RNAi) and other post-genome technologies (e.g. microarrays). Studies of many other invertebrates, notably the honeybee (Apis mellifera), a nudibranch mollusc (Aplysia californica), locusts, lobsters, crabs, annelids and jellyfish have all assisted in the development of major concepts in neuroscience. The future is equally bright with ease of access to genome-wide reverse genetic technologies, and the development of optical recordings using voltage and intracellular calcium sensors genetically targeted to selected individual and groups of neurons.

Central pattern generator for swimming in Melibe

The nudibranch mollusc Melibe leonina swims by bending from side to side. We have identified a network of neurons that appears to constitute the central pattern generator (CPG) for this locomotor behavior, one of only a few such networks to be described in cellular detail. The network consists of two pairs of interneurons, termed 'swim interneuron 1' (sint1) and 'swim interneuron 2' (sint2), arranged around a plane of bilateral symmetry. Interneurons on one side of the brain, which includes the paired cerebral, pleural and pedal ganglia, coordinate bending movements toward the same side and communicate via non-rectifying electrical synapses. Interneurons on opposite sides of the brain coordinate antagonistic movements and communicate over mutually inhibitory synaptic pathways. Several criteria were used to identify members of the swim CPG, the most important being the ability to shift the phase of swimming behavior in a quantitative fashion by briefly altering the firing pattern of an individual neuron. Strong depolarization of any of the interneurons produces an ipsilateral swimming movement during which the several components of the motor act occur in sequence. Strong hyperpolarization causes swimming to stop and leaves the animal contracted to the opposite side for the duration of the hyperpolarization. The four swim interneurons make appropriate synaptic connections with motoneurons, exciting synergists and inhibiting antagonists. Finally, these are the only neurons that were found to have this set of properties in spite of concerted efforts to sample widely in the Melibe CNS. This led us to conclude that these four cells constitute the CPG for swimming. While sint1 and sint2 work together during swimming, they play different roles in the generation of other behaviors. Sint1 is normally silent when the animal is crawling on a surface but it depolarizes and begins to fire in strong bursts once the foot is dislodged and the animal begins to swim. Sint2 also fires in bursts during swimming, but it is not silent in non-swimming animals. Instead activity in sint2 is correlated with turning movements as the animal crawls on a surface. This suggests that the Melibe motor system is organized in a hierarchy and that the alternating movements characteristic of swimming emerge when activity in sint1 and sint2 is bound together.

An unusual dendrodorid: redescription of the tropical nudibranch Dendrodoris atromaculata (Alder & Hancock, 1864) (Anthobranchia: Doridoidea: Dendrodorididae)

Dendrodoris atromaculata (Alder & Hancock, 1864) is a large, tropical nudibranch mollusc living in the Indo-west Pacific Ocean. A detailed description of the external morphology of the living animal and an investigation of internal anatomy are presented for the first time. In addition the microstructure of certain organs (i.e., notal tissue and the vestibular gland associated with the reproductive system) are compared with other Dendrodoris species. Dendrodoris atromaculata is unique among currently described species of Dendrodoris in three respects, the gill structure is “cross-like”, the notum possesses prominent spicules and the notum is covered in finger-like processes. These findings necessitate a reassessment of previously documented generic features of Dendrodoris and reveal a need for changes to future phylogenetic analyses of both porostome and cryptobranch dorids.