Inhibition of metamorphosis by RFamide neuropeptides in planula larvae of Hydractinia echinata

The primitive nervous system in planula larvae of Hydractinia echinata (Cnidaria) has sensory neurons containing LWamide or RFamide neuropeptides. LWamides have been shown to induce metamorphosis of planula larvae into adult polyps. We report here that RFamides act antagonistically to LWamides. RFamides inhibit metamorphosis when applied to planula larvae during metamorphosis induction by treatment with LWamides (or other inducing agents such as CsCl ions, diacylglycerol and bacterial inducers). Our results show further that RFamides act downstream of LWamide release, presumably directly on target cells mediating metamorphosis. These observations support a model in which metamorphosis in H. echinata is regulated by sensory neurons secreting LWamides and RFamides in response to environmental cues.Edited by D. Tautz     

RFamide neuropeptide actions on molluscan proboscis smooth muscle: interactions with primary neurotransmitters

The potency (muscle force-generated) of a number of long-chain RFamide neuropeptides was examined in mechanical experiments with the radular-retractor and radular-sac muscles of gastropods Buccinum undatum and Neptunea antiqua. Many of the heptapeptides, octapeptides and the decapeptide LMS were found to induce greater contraction than FMRFamide in both smooth muscles and in both species. RFamide neuropeptides interacted with the neurotransmitter acetylcholine in an additive way and RFamide-induced contractions were inhibited by the neuromodulator serotonin. Pre-treatment with a calcium-free saline completely abolished acetylcholine-induced responses but only partially inhibited RFamide responses in the muscles, suggesting that acetylcholine acts to cause influx of extracellular calcium for contraction. In contrast, RFamide neuropeptides may mobilise intracellular calcium to maintain sustained tonic force in calcium-free conditions. This suggests that an additional involvement of a fast calcium channel may be present in the RFamide responses, since loss of the usual superimposed twitch activity is observed. Force regulation in these muscles appears to result from a complex interaction of RFamide neuropeptides with the primary transmitter acetylcholine and the neuromodulator serotonin.Abbreviations ACh acetylcholine - Ala alanine - Arg arginine - Asn asparagine - Asp aspartic acid - Cys cysteine - FLRFamide Phe-Leu-Arg-Phe-NH₂ - FMRFamide Phe-Met-Arg-Phe-NH₂ - Gln glutamine - Glu glutamic acid - Gly glycine - His histadine - Ile isoleucine - Leu leucine - LMS leucomyosuppressin - Met methionine - Nle norleucine - Phe phenylalanine - Pro proline - SCP_B (small cardioactive peptide B) Met-Asn-Tyr-Leu-Ala-Phe-Pro-Arg-Met-NH₂ - Ser serine - Val valine     

Examination of the role of FMRFamide-related peptides in the circadian clock of the cockroach Leucophaea maderae

The accessory medulla, the circadian clock of the cockroach Leucophaea maderae, is abundant in neuropeptides. Among these neuropeptides are the FMRFamide-related peptides (FaRPs), which generally share the C-terminal RFamide. As a first step toward understanding the functional role of FaRPs in the circadian clock of the cockroach, immunocytochemistry with antisera against various FaRPs, MALDI-TOF mass spectrometry, and injections of two FaRPs combined with running-wheel assays were performed. Prominent FMRFamide-like immunoreactivity was found in maximally four soma clusters associated with the accessory medulla and in most neuropils of the protocerebrum. By MALDI-TOF mass spectrometry, various extended FMRFamides of the cockroach L. maderae were partially identified in thoracic perisympathetic organs, structures known to accumulate extended FMRFamides in insects. By mass match, several of these peptides were also detected in the accessory medulla. Injections of FMRFamide and Pea-FMRFa-7 (DRSDNFIRF-NH₂) into the vicinity of the accessory medulla caused time-dependent phase-shifts of locomotor activity rhythms at circadian times 8, 18, and 4. Thus, our data suggest a role for the different FaRPs in the control of circadian locomotor activity rhythms in L. maderae.     

Fast Neurotransmission Related Genes Are Expressed in Non Nervous Endoderm in the Sea Anemone Nematostella vectensis

Cnidarian nervous systems utilize chemical transmission to transfer signals through synapses and neurons. To date, ample evidence has been accumulated for the participation of neuropeptides, primarily RFamides, in neurotransmission. Yet, it is still not clear if this is the case for the classical fast neurotransmitters such as GABA, Glutamate, Acetylcholine and Monoamines. A large repertoire of cnidarian Fast Neurotransmitter related Genes (FNGs) has been recently identified in the genome of the sea anemone, Nematostella vectensis. In order to test whether FNGs are localized in cnidarian neurons, we characterized the expression patterns of eight Nematostella genes that are closely or distantly related to human central and peripheral nervous systems genes, in adult Nematostella and compared them to the RFamide localization. Our results show common expression patterns for all tested genes, in a single endodermal cell layer. These expressions did not correspond with the RFamide expressing nerve cell network. Following these results we suggest that the tested Nematostella genes may not be directly involved in vertebrate-like fast neurotransmission.     

Evolutionary origin of rhopalia: insights from cellular‐level analyses of Otx and POU expression patterns in the developing rhopalial nervous system

SUMMARY In Cnidaria, the medusae of Scyphozoa and its sister‐group Cubozoa uniquely possess rhopalia at their bell margin. These sensory centers coordinate behavior and development. We used fluorescent in situ hybridization and confocal microscopy to examine mRNA expression patterns in Aurelia sp.1 (Cnidaria, Scyphozoa) during early medusa formation, while simultaneously visualizing the developing nervous system by immunofluorescence. The genes investigated include AurOtx1, and the POU genes, AurPit1, and AurBrn3, homologs of genes known to function in cephalar neural organization and sensory cell differentiation across Bilateria. Our results show that AurOtx1 expression defines the major part of the oral neuroectodermal domain of the rhopalium, within which distinct populations of AurBrn3‐ and AurPit1‐expressing sensory cells develop. Thus, despite the unique attributes of rhopalial evolution, we suggest that the rhopalial nervous system of scyphozoan medusae involves similar patterns of differential expression of genes that function in bilaterian cephalic structure and neuroendocrine system development. We propose that rhopalia evolved from preexisting sensory structures that developed distinct populations of sensory cells differentially expressing POU genes within Otx oral‐neuroectodermal domains. This implies some commonality of developmental genetic functions involving these genes in the still poorly constrained common ancestor of bilaterians and cnidarians.     

Immunohistochemical studies of GLWamides in Cnidaria

GLWamides are a recently described, novel family of neuropeptides in Cnidaria. Antibodies specific for the GLWamide terminus have been raised and used to evaluate the occurrence and localisation of immunopositive material in various Cnidaria in order to determine whether GLWamides are present and to obtain a first impression of the possible regulatory role of these neuropeptides. GLWamide immunoreactivity has been found in all species tested and is not confined to distinct life stages but is present during most of the life cycle of the Cnidaria. Additionally, GLWamides are expressed by different nerve cells at different life stages. GLWamide-immunoreactive cells constitute a subset of the neural equipment. Overall our data suggest that GLWamides generally occur in the nervous system of Cnidaria and that these peptides are multifunctional. Putative functions other than the control of development include the regulation of nematocyst discharge, muscle contraction and the regulation of gastric function.     

Antigenicity to neuropeptide F (NPF) in Stenostomum leucops and Microstomum lineare

The first native flatworm regulatory peptide, neuropeptide F (NPF) has recently been isolated and sequenced from the cestode Moniezia expansa (see Maule et al., 1991) and the turbellarian Artioposthia triangulata, (see Curry et al., 1992). NPF belongs to the neuropeptide Y (NPY) superfamily and the antiserum is known to show cross-reactivity to the vertebrate neuropeptides of the NPY superfamily. It terminates in RFamide, like the invertebrate neuropeptides FMRFamide and RFamide, and may cross-react with neuropeptides of the FMRFamide family. Strong immunoreactivity (IR) to FMRF- and RF-amide has been demonstrated in members of most flatworm groups. In the present study, IR to NPF (diluted 1:1000) is demonstrated in Stenostomum leucops (Catenulida) and Microstomum lineare (Macrostomida). The controls included: omitting primary antibody, using non-immune serum and liquid-phase absorption with the homologous antigen (1000 ng ml^–1). The NPF IR pattern was compared to the FMRF and RF-amide IR patterns in order to reveal differences or co-localization. In addition, the sequential appearance of NPF-positive cells in developing zooids was followed and double staining with a-5-HT made to complete the study.     

Neuropeptides in the Cnidaria

This chapter reviews experimental evidence for peptides actingas transmitters or morphogens in the Cnidaria. A wide rangeof brain/gut peptides have been localized immunohistochemicallyto specific neuronal populations in Hydra. These include gastrin/CCK,substance P, neurotensin, bombesin, oxytocin/vasopressin andFMRFamide. In most cases the amino-acid sequences of the cnidarianpeptides are somewhat different from their mammalian counterparts.The functions of these peptides in Hydra are unknown. A seriesof neuropeptides with the carboxy-terminus, Arg-Phe-amide, isubiquitous within the phylum. Immunoreactivity to antisera againstRFamide is seen in two characteristic neuronal subpopulations;a sensory and a ganglionic cell type. Two of these peptideshave now been isolated and sequenced. One has the structure,pGlu-Gly-Arg-Phe-amide (Antho-RFamide) and is found in severalanthozoans, a second with the sequence pGlu-Leu-Leu- Gly-Gly-Arg-Phe-amide(Poly A peptide) is found in the hydrozoan Polyorchis. Arg-Pheamidepeptides have excitatory effects on both muscles and neuronalsystems. In the anthozoans, RFamide peptides can increase thetone, contraction amplitude and frequency of a number of smoothmuscle systems. Additionally, in the anemone Calliactis, applicationof Antho-RFamide can dramatically increase the firing rate inone of the ectodermal conducting systems, the SSI. In Polyorchisseveral RFamides produce long duration spike trains in motorneurons that may or may not be associated with membrane depolarization.A peptide called ‘head activator’ (pGlu-(Pro)j-(Gly)8-Ser-Lys-Val-Ile-Leu-Phe)can induce the formation of a new head when it is released athigh concentrations at the cut surface of the column of Hydra.It acts by committing stem cells to become head specific neurons.     

Immunohistochemical identification of substance P in cutaneous sensory organs (electroreceptors) of gymnotid teleost fish

Summary The distribution of the neuropeptide substance P, which is considered to be a neurotransmitter or neuromodulator of the central nervous system, has been studied in the cutaneous electroreceptor organs (tuberous and ampullary organs) of 3 species of gymnotid fish: Apteronotus leptorhynchus, Eigenmannia virescens and Sternopygus sp. Immunohistochemical data have revealed that substance P is never present in the afferent fibers but is specifically localized in the electroreceptors of the three species examined. Substane P immunoreactivity is strictly localized in the sensory cells of the ampullary organs of all three species and in those of the tuberous organs of Apteronotus leptorhynchus and Sternopygus sp. In contrast, weak substance P immunoreactivity was observed only in certain tuberous sensory cells of Eigenmannia. Substance P immunoreactivity was also found in the accessory cells of certain organs: it was detected in the two types of accessory cells of the tuberous organs of Eigemmannia virescens, in the accessory cells type 2 of the tuberous organs of Sternopygus sp., and in all accessory cells of ampullary organs of Sternopygus sp. and Apteronotus leptorhynchus. In Sternopygus sp., positive staining was only evident if the substance P antibody was used at low concentration. Immunoreactivity for substance P in the sensory cells suggests that it has a transmitter or modulator function in these electroreceptors; the presence of substance P in the accessory cells remains to be explained.     

Comparative immunohistochemistry of the cephalic sensory organs in Opisthobranchia (Mollusca,Gastropoda)

The structure and function of the central nervous systems of opisthobranch gastropods have been studied extensively. However, the organisation and function of the peripheral nervous system are poorly understood. The cephalic sensory organs (CSOs) are known to be chemosensory structures in the head region of opisthobranchs. In the present study, we used immunohistochemical methods and confocal laserscanning microscopy to comparatively examine the CSOs of different opisthobranchs, namely Acteon tornatilis, Aplysia punctata, Archidoris pseudoargus and Haminoea hydatis. We wanted to characterise sensory epithelia in order to infer the function of sensory structures and the organs they constitute. Immunoreactivity against the three antigens tyrosine hydroxylase, FMRFamide and serotonin was very similar in the CSOs of all investigated species. Tyrosine hydroxylase-like immunoreactivity was detected primarily in subepidermal sensory cell bodies, which were much more abundant in the anteriorly situated CSOs. This observation indicates that these cells and the respective organs may be involved in contact chemoreception and mechanoreception. The dominant features of FMRFamide-like immunoreactivity, especially in the posterior CSOs, were tightly knotted fibres which reveal glomerulus-like structures. This suggests an olfactory role for these organs. Serotonin-like immunoreactivity was detected in an extensive network of efferent fibres, but was not found within any peripheral cell bodies. Serotonin-like immunoreactivity was found in the same glomerulus-like structures as FMRFamide-like immunoreactivity, indicating a function of serotonin in the efferent control of olfactory inputs. Besides this functional implication, this study could also add some knowledge on the doubtful homology of the CSOs in opisthobranch gastropods.     

Histamine is a major mechanosensory neurotransmitter candidate in Drosophila melanogaster

Histamine is known to be the neurotransmitter of insect photoreceptors. Histamine-like immunoreactivity is also found in a number of interneurons in the central nervous system of various insects. Here, we demonstrate by immunohistochemical techniques that, in Drosophila melanogaster (Acalypterae), most or all mechanosensory neurons of imaginal hair sensilla selectively bind antibodies directed against histamine. The histamine-like staining includes the cell bodies of these neurons as well as their axons, which form prominent fibre bundles in peripheral nerves, and their terminal projections in the central neuropil of head and thoracic ganglia. The specificity of the immunostaining is demonstrated by investigating a Drosophila mutant unable to synthesize histamine. Other mechanosensory organs, such as campaniform sensilla or scolopidial organs, do not stain. In the calypteran flies, Musca and Calliphora, we find no comparable immunoreactivity associated with either hair sensilla or the nerves entering the central nervous system, observations in agreement with earlier studies on Calliphora. Thus, histamine seems to be a major mechanosensory transmitter candidate of the adult nervous system of Drosophila, but apparently not of Musca or Calliphora.     

Role of a FMRFamide‐like family of neuropeptides in the pharyngeal nervous system of Caenorhabditis elegans

The nervous system of C. elegans has a remarkable abundance of flp genes encoding FMRFamide‐like (FLP) neuropeptides. To provide insight into the physiological relevance of this neuropeptide diversity, we have tested more than 30 FLPs (encoded by 23 flps) for bioactivity on C. elegans pharynx. Eleven flp genes encode peptides that inhibit pharyngeal activity, while eight flp genes encode peptides that are excitatory. Three potent peptides (inhibitory, FLP‐13A, APEASPFIRFamide; excitatory, FLP‐17A, KSAFVRFamide; excitatory, FLP‐17B, KSQYIRFamide) are encoded by flp genes, which, according to reporter gene constructs, are expressed in pharyngeal motoneurons. Thus, they may act through receptors localized on the pharyngeal muscle. The two other potent peptides, FLP‐8 (excitatory AF1, KNEFIRFamide,) and FLP‐11A (inhibitory, AMRNALVRFamide), appear to be expressed in extrapharyngeal neurons and are therefore likely to act either indirectly or as neurohormones. Intriguingly, a single neuron can express peptides that have potent but opposing biological activity in the pharynx. Only five flp genes encode neuropeptides that have no observable effect on the pharynx, but none of these have shown reporter gene expression in the pharyngeal nervous system. To examine the roles of multiple peptides produced from single precursors, a comparison was made between the bioactivity of different neuropeptides for five flp genes (flp‐3, flp‐13, flp‐14, flp‐17, and flp‐18). For all but one gene (flp‐14), the effects of peptides encoded by the same gene were similar. Overall, this study demonstrates the impressive neurochemical complexity of the simple circuit that regulates feeding in the nematode, C. elegans. © 2005 Wiley Periodicals, Inc. J Neurobiol, 2005     

Ocelli in a Cnidaria polyp: the ultrastructure of the pigment spots in Stylocoronella riedli (Scyphozoa,Stauromedusae)

Within the Cnidaria, the occurrence of ocelli at the polyp stage is only known in the species of Stylocoronella (Scyphozoa, Stauromedusae). The light-sensitive organs of S. riedli are ultrastructurally investigated. In this interstitial-living species, each of the up to 24 ocelli is composed of between seven and nine monociliary sensory cells and between one and four pigment cells. A striking feature of the photoreceptive cilia is their peculiar axonemal pattern. This is expressed (a) by the presence of a third central microtubule at a certain point and (b) by the balloon-like swelling of the distal portion of the cilium, with clearly scattered microtubules in this area. Although the polyps of S. riedli show no distinct reaction to light stimuli, the ultrastructural results corroborate the hypothesis that these organs are light-sensitive organs. The possible function of the pigment granules is discussed.Abbreviations bb basal body - c cilium - co collar - csv crescent-shaped vesicle - cv clear vesicle - dcv dense-core vesicles - k kinetosome - m mitochondrion - mvb multivesicular body - n nucleus - oc ocellus - pc piment cell - pg pigment granule - sc sensory cell - sr striated rootlet - v vesicle     

Distribution of SchistoFLRFamide-like immunoreactivity in the adult ventral nervous system of the locust,Schistocerca gregaria

SchistoFLRFamide (PDVDHVFLRF-NH₂) is one of the major endogenous neuropeptides of the FMRF-amide family found in the nervous system of the locust,Schistocerca gregaria. To gain insights into the potential physiological roles of this neuropeptide we have examined the distribution of SchistoFLRFamide-like immunoreactivity in the ventral nervous system of adult locusts by use of a newly developed N-terminally specific antibody. SchistoFLRFamide-like immunoreactivity in the ventral nerve cord is found in a subgroup of the neurones that are immunoreactive to an antiserum raised against bovine pancreatic polypeptide (BPP). In the suboesophageal ganglion three groups of cells stain, including one pair of large posterior ventral cells. These cells are the same size, in the same location in the ganglion and have the same branching pattern as a pair of BPP immunoreactive cells known to innervate the heart and retrocerebral glandular complex of the locust. In the thoracic and abdominal ganglia two and three sets of cells, respectively, stain with both the SchistoFLRFamide and BPP antisera. In the abdominal ganglia the immunoreactive cells project via the median nerves to the intensely immunoreactive neurohaemal organs.     

Phototactic responses in the gametes of the brown alga,Ectocarpus siliculosus

The action spectrum of phototaxis was determined and the photoreceptive mechanism was studied in Ectocarpus gametes (Ectocarpales, Phaeophyceae) using a computerized cell-tracking system. The fine structures of the stigma and the flagellar swelling were analyzed, and the reflective function of the stigma was demonstrated for the first time. Under monochromatic light stimulation, Ectocarpus gametes show mainly positive phototaxis between 370 nm and 520 nm. The action spectrum has a minor peak near 380 nm, and two major peaks at 430 nm and 450 nm or 460 nm and a shoulder at 470 nm adjoining a remarkable depression near 440 nm. Under unilateral stroboscopic illumination with more than four pulses per second, the gametes show clear phototaxis. However, the response is disturbed at lower frequencies. Addition of methyl cellulose, which increases the viscosity of the medium and slows down gamete rotation, decreases the threshold frequency. These results indicate that rotation of the gamete plays an essential role in the photoreceptive mechanism. Under equal intensities of bilateral illumination at an angle of 90°, most of the gametes swim on the resultant between the two light beams. This response is disturbed when the angle of the two light beams is as large as 120°. Observations by transmission electron microscopy show that the flagellar swelling fits precisely into a concave depression of the chloroplast at the central region of the stigma. Electron-dense material is present in that sector of the flagellar swelling which faces away from the stigma. Epifluorescence microscopy without a barrier filter and epipolarization microscopy reveal that stigmata reflect blue light. A hypothesis is formulated which discusses the possibility that the reflected light is focused onto the flagellar swelling.We are grateful to Jochen Schäfer and Elke Reinecke for their technical assistance and Dr. G. Konerman for access to epipolarization microscopy. We are also grateful to the Alexander von Humboldt Foundation for a Research Fellowship to H.K. and the Deutsche Forschungsgemeinschaft (University of Freiburg, Freiburg, FRG) for financial aid to D.-P.H.To whom correspondence should be addressed.     

Rethinking the Role of the Nervous System: Lessons From the Hydra Holobiont

Here we evaluate our current understanding of the function of the nervous system in Hydra, a non‐bilaterian animal which is among the first metazoans that contain neurons. We highlight growing evidence that the nervous system, with its rich repertoire of neuropeptides, is involved in controlling resident beneficial microbes. We also review observations that indicate that microbes affect the animal's behavior by directly interfering with neuronal receptors. These findings provide new insight into the original role of the nervous system, and suggest that it emerged to orchestrate multiple functions including host‐microbiome interactions. The excitement of future research in the Hydra model now relies on uncovering the common rules and principles that govern the interaction between neurons and microbes and the extent to which such laws might apply to other and more complex organisms.     

The presence and location of small cardioactive‐like peptides in larvae of Crassostrea virginica

We examined the presence and location of small cardioactive peptide (SCP)‐like neuropeptides in both the central and peripheral nervous systems of D‐hinge, newly eyed, and pediveliger larvae of Crassostrea virginica. Results indicate that SCP‐like substances are present early in development (D‐hinge larvae), and that presence in the central and peripheral nervous systems increases as the larva develops toward metamorphic competence (pediveliger larvae). In addition, in newly eyed and pediveliger larvae, SCP‐like labeling is found in a varying number of neurons within all central ganglia, with the possible exception of the accessory ganglia. Varicose labeling of axons is also documented within the ganglia, commissures, and connectives, as well as in peripheral nerves. Peripheral tissues innervated by axons exhibiting SCP‐like immunoreactivity include the velum, foot, esophagus, mantle, and various musculatures. As indicated by the location of SCP‐like labeling in various organs and tissues, it is likely that these neuropeptides modulate muscle contraction or ciliary beating in molluscan larvae such as those of C. virginica.     

Identification of the Drosophila and Tribolium receptors for the recently discovered insect RYamide neuropeptides

One year ago, we discovered a new family of insect RYamide neuropeptides, which has the C-terminal consensus sequence FFXXXRYamide, and which is widely occurring in most insects, including the fruitfly Drosophila melanogaster and the red flour beetle Tribolium castaneum (F. Hauser et al., J. Proteome Res. 9 (2010) 5296–5310). Here, we identify a Drosophila G-protein-coupled receptor (GPCR) coded for by gene CG5811 and its Tribolium GPCR ortholog as insect RYamide receptors. The Drosophila RYamide receptor is equally well activated (EC₅₀, 1 × 10⁻⁹ M) by the two Drosophila RYamide neuropeptides: RYamide-1 (PVFFVASRYamide) and RYamide-2 (NEHFFLGSRYamide), both contained in a preprohormone coded for by gene CG40733. The Tribolium receptor shows a somewhat higher affinity to Tribolium RYamide-2 (ADAFFLGPRYamide; EC₅₀, 5 × 10⁻⁹ M) than to Tribolium RYamide-1 (VQNLATFKTMMRYamide; EC₅₀, 7 × 10⁻⁸ M), which might be due to the fact that the last peptide does not completely follow the RYamide consensus sequence rule. There are other neuropeptides in insects that have similar C-terminal sequences (RWamide or RFamide), such as the FMRFamides, sulfakinins, myosuppressins, neuropeptides F, and the various short neuropeptides F. Amazingly, these neuropeptides show no cross-reactivity to the Tribolium RYamide receptor, while the Drosophila RYamide receptor is only very slightly activated by high concentrations (>10⁻⁶ M) of neuropeptide F and short neuropeptide F-1, showing that the two RYamide receptors are quite specific for activation by insect RYamides, and that the sequence FFXXXRYamide is needed for effective insect RYamide receptor activation. Phylogenetic tree analyses and other amino acid sequence comparisons show that the insect RYamide receptors are not closely related to any other known insect or invertebrate/vertebrate receptors, including mammalian neuropeptide Y and insect neuropeptide F and short neuropeptide F receptors. Gene expression data published in Flybase (www.flybase.org) show that the Drosophila CG5811 gene is significantly expressed in the hindgut of adult flies, suggesting a role of insect RYamides in digestion or water reabsorption.     

The presence and distribution of Antho-RFamide-like material in scyphomedusae

Summary The nervous systems of the scyphomedusae Chrysaora hysoscella, Cyanea capillata and Cyanea lamarckii (Phylum Cnidaria) were stained using an antiserum against the anthozoan neuropeptide Antho-RFamide. Staining was widespread in all three species. In Chrysaora, the antiserum revealed nerve nets in the subumrella and exumbrella ectoderm, in both faces of the oral lobes, and in the endoderm lining the subumbrella and exumbrella surfaces of the gastric cavity. The most prominent staining occurred in a dense plexus of neurons in the ectoderm at the base of the tentacles. This nerve net sent projections into the subumbrella ectoderm. For the most part, staining in the two species of Cyanea was similar to that in Chrysaora, with a few exceptions. These include the presence, in Cyanea, of an obvious tentacular nerve tract and nerve nets associated with clusters of cnidocytes in the tentacles. Radioimmunoassays of extracts from Chrysaora and Cyanea lamarkii revealed that both species contain large amounts of Antho-RFamide-like material (up to 55 nmol/animal). The results indicate that Antho-RFamide-like neuropeptides are widespread in scyphomedusae.     

Antisera to the sequence Arg-Phe-amide visualize neuronal centralization in hydroid polyps

Summary Antisera to the sequence Arg-Phe-amide (RF-amide) have a high affinity to the nervous system of fixed hydroid polyps. Whole-mount incubations of several Hydra species with RFamide antisera visualize the three-dimensional structure of an ectodermal nervous system in the hypostome, tentacles, gastric region and peduncle. In the hypostome of Hydra attenuata a ganglion-like structure occurs, consisting of numerous sensory cells located in a region around the mouth opening and a dense plexus of processes which project mostly radially towards the bases of the tentacles. In Hydra oligactis an ectodermal nerve ring was observed lying at the border of hypostome and tentacle bases. This nerve ring consists of a few large ganglion cells with thick processes forming a circle around the hypostome. This is the first direct demonstration of a nerve ring in a hydroid polyp.Incubation of Hydractinia echinata gastrozooids with RFamide antisera visualizes an extremly dense plexus of neuronal processes in body and head regions. A ring of sensory cells around the mouth opening is the first group of neurons to show RFamide immunoreactivity during the development of a primary polyp. In gonozooids the oocytes and spermatophores are covered with strongly immunoreactive neurons.All examples of whole-mount incubations with RF-amide antisera clearly show that hydroid polyps have by no means a diffuse nerve net, as is often believed, and that neuronal centralization and plexus formation are common in these animals. The examples also show that treatment of intact fixed animals with RFamide antisera is a useful technique to study the anatomy or development of a principal portion of the hydroid nervous system.