A simple neuronal system characterized by a monoclonal antibody to SCP neuropeptides in embryos and larvae of Tritonia diomedea (Gastropoda, Nudibranchia)

SCP-like antigenicity is first present in Tritonia diomedea in small cells of the cerebral ganglia and a single axon crossing the cerebral commissure of 8-day-old embryos. Other axons and neurons become antigenic as the larva develops. At 4-9 days after larvae hatch from the egg mass, 2 additional pairs of neurons are labeled. Axons extend from one pair to the left cerebral ganglion and from the other to the right. A second labeled axon is present across the cerebral commissure. In metamorphically competent larvae the cerebral and pedal neuropils, as well as two neurons in the buccal ganglia with axon(s?) across the commissure, are antigenic. The change in antigenicity as the larva becomes competent is presumably preparatory for juvenile life. The labeled buccal neurons may be B12, which are known to contain SCPs, extend an axon across the buccal commissure, and function in adult feeding behavior. The two large neurons strongly labeled by rabbit polyclonal antibodies against FMRFamide are clearly different from neurons labeled by monoclonal antibody against SCPs. This result supports the contention that different antigens are labeled by these two immune probes.     

Multilevel inhibition of feeding by a peptidergic pleural interneuron in the mollusc<Emphasis Type="Italic"> Lymnaea stagnalis</Emphasis>

The pleural interneuron PlB is a white neuron in the pleural ganglion of the snail Lymnaea. We test the hypothesis that it inhibits neurons at all levels of the feeding system, using a combination of anatomy, physiology and pharmacology. There is just one PlB in each pleural ganglion. Its axon traverses the pedal and cerebral ganglia, running into the buccal ganglia. It has neuropilar branches in the regions of the cerebral and buccal ganglia where neurons that are active during feeding also branch. Activation of the PlB blocks fictive feeding, whether the feeding rhythm occurs spontaneously or is driven by a modulatory interneuron. The PlB inhibits all the neurons in the feeding network, including protraction and retraction motoneurons, central pattern generator interneurons, buccal modulatory interneurons (SO, OC), and cerebral modulatory interneurons (CV1, CGC). Only the CV1 interneuron shows discrete 1:1 IPSPs; all other effects are slow, smooth hyperpolarizations. All connections persist in Ca²⁺/Mg²⁺-rich saline, which reduces polysynaptic effects. The inhibitory effects are mimicked by 0.5 to 100 mol l^–1 FMRFamide, which the PlB soma contains. We conclude that the PlB inhibits neurons in the feeding system at all levels, probably acting though the peptide transmitter FMRFamide.Electronic Supplementary Material Supplementary material is available in the online version of this article at http://dx.doi.org/10.1007/s00359-004-0503-x     

Morphology and physiology of pleural-to-buccal neurons coordinating defensive retraction with feeding arrest in the pond snail Lymnaea stagnalis

1. We have studied morphology, physiology and chemistry of a bilateral pair of pleural-to-buccal projecting neurons (PlB cells) of the pond snail Lymnaea stagnalis. Intracellular dye fills revealed axon arborization within neuropiles of ipsilateral pedal and cerebral ganglia, as well as in both buccal ganglia. Terminal axons of the left and right PlBs showed close proximity within the buccal commissure. 2. The left and right PlB neurons have been found electrotonically coupled and, sometimes, generating synchronous spikes. 3. The results show that two PlB cells operate as a single unit, and that paired buccal networks responsible for feeding rhythm are treated by the PlBs as a single target.     

Electrophysiological study of serotoninergic neuron Cl in the pteropod mollusk Clione

Functional characteristics of cerebral serotoninergic neuron Cl, axons of which terminate at the buccal ganglia [7], were investigated in the pteropod molluskClione. Stimulating neuron Cl induced activation of the feeding rhythm generator located in the buccal ganglia — an effect arising after a long latency and persisting for some tens of seconds once stimulation had ended. Neuron Cl receives feedback from buccal ganglion cells and this brings about periodic modulation in ganglia activity during the generation of feeding rhythm. Activity of neuron Cl is correlated with operation of the locomotor rhythm generator located in the pedal ganglia. The firing rate of Cl neurons increased upon activation of the locomotor generator (whether spontaneous or induced by stimulating certain command neurons). The correlation found between workings of the locomotor generator and activity of Cl neurons is thought to be one of the manifestations of feeding synergy involving simultaneous activation of the locomotor and buccal apparatus.Institute for Research on Information Transmission, Academy of Sciences of the USSR, Moscow. M. V. Lomonosov State University, Moscow. Translated from Neirofiziologiya, Vol. 23, No. 1, pp. 18–25, January–February, 1991.     

Coordinated motor neuron axon growth and neuromuscular synaptogenesis are promoted by CPG15 in vivo

We have used in vivo time-lapse two-photon imaging of single motor neuron axons labeled with GFP combined with labeling of presynaptic vesicle clusters and postsynaptic acetylcholine receptors in Xenopus laevis tadpoles to determine the dynamic rearrangement of individual axon branches and synaptogenesis during motor axon arbor development. Control GFP-labeled axons are highly dynamic during the period when axon arbors are elaborating. Axon branches emerge from sites of synaptic vesicle clusters. These data indicate that motor neuron axon elaboration and synaptogenesis are concurrent and iterative. We tested the role of Candidate Plasticity Gene 15 (CPG15, also known as Neuritin), an activity-regulated gene that is expressed in the developing motor neurons in this process. CPG15 expression enhances the development of motor neuron axon arbors by promoting neuromuscular synaptogenesis and by increasing the addition of new axon branches.     

Neuronal mechanisms for bilateral coordination of salivary gland activity in Helisoma

The salivary neuroeffector system of Helisoma consists of the paired salivary glands and buccal ganglia. Previous work demonstrated that neuronal control was required for coordination of activity in the two salivary glands. This neuronal control is provided by a pair of identified buccal ganglion neurons, 4R and 4L. This study examines the organization of this neuronal control and addresses the questions of monosynaptic vs. polysynaptic pathways as well as the bilateral effects of each neuron 4. Action potentials in neuron 4 elicit one-for-one EPSPs in a subpopulation of the salivary cells. These EPSPs can, in some cases, be increased by TEA injection into a neuron 4 and are unaffected by the addition of six-times normal calcium. These data coupled with the constancy of synaptic transmission, as well as morphological evidence, further indicate the monosynaptic nature of the connection between neurons 4 and salivary secretory cells. Three different mechanisms exist to insure that activity in 4R and 4L result in coordinated activation of the salivary glands: (1) Lucifer Yellow injection and direct intracellular recording and stimulation demonstrate that both 4R and 4L can send axons to and innervate both salivary glands; (2) both 4R and 4L receive virtually identical synaptic input from higher-order buccal ganglion neurons; and (3) 4R and 4L are electrically coupled. Thus, the system is organized with a high degree of redundancy, and bilateral synchrony of glandular activity is assured by mechanisms at various levels of neuronal organization.     

Serotonin-like immunoreactivity in the stomatogastric nervous systems of crayfishes from four genera

We used whole-mount immunocytochemistry to characterize the distribution of serotonin in the stomatogastric nervous systems of seven species of crayfish representing three genera from the family Cambaridae (Orconectes, Cambarus, and Procambarus) and one from the family Astacidae (Pacifastacus). In all species, we observed serotonin-like immunoreactivity in four gastropyloric receptor (GPR) neurons located in the lateral ventricular nerves, with one pair of neurons in each nerve. As in other crustaceans, the GPR axons project to the stomatogastric ganglion and to the bilateral commissural ganglia. In three crayfishes, we observed the GPR axons crossing the commissural ganglia, and extending toward the thoracic nervous system. This feature was most clearly and consistently seen in Pacifastacus leniusculus. The number of stained somata in the commissural ganglia varied among crayfish species from two (in Procambarus clarkii) to five (in Pacifastacus leniusculus). The largest soma (the L cell) displayed both serotonin- and tyrosine hydroxylase-like immunoreactivity in all species, suggesting that serotonin and dopamine are cotransmitters in this cell. The inferior esophageal nerve and a branch of this nerve (the inner labral nerve) contained several axons with serotonin-like immunoreactivity. These axons were clearly present in only one species (Procambarus clarkii). Serotonin acts as a neuromodulator of rhythms produced by circuits in the crab and lobster stomatogastric ganglion, and is likely to play a similar role in crayfish. Differences are apparent in the distribution of serotonin among crayfish species and between crayfish and other crustaceans, and could result in differences in the physiological action of this modulator.     

Responses of cerebral GABA-containing CBM neuron to taste stimulation with seaweed extracts in Aplysia kurodai

Aplysia kurodai distributed along Japan feeds well on Ulva pertusa but rejects Gelidium amansii with distinctive patterned movements of the jaws and radula. On the ventral side of the cerebral M cluster, four cell bodies of higher order neurons that send axons to the buccal ganglia are distributed (CBM neurons). We have previously shown that the dopaminergic CBM1 modulates basic feeding circuits in the buccal ganglia for rejection by firing at higher frequency after application of the aversive taste of seaweed such as Gelidium amansii. In the present experiments immunohistochemical techniques showed that the CBM3 exhibited gamma-aminobutyric acid (GABA)-like immunoreactivity. The CBM3 may be equivalent to the CBI-3 involved in changing the motor programs from rejection to ingestion in Aplysia californica. The responses of the CBM3 to taste stimulation of the lips with seaweed extracts were investigated by the use of calcium imaging. The calcium-sensitive dye, Calcium Green-1, was iontophoretically introduced into a cell body of the CBM3 using a microelectrode. Application of Ulva pertusa or Gelidium amansii extract induced different changes in fluorescence in the CBM3 cell body, indicating that taste of Ulva pertusa initially induced longer-lasting continuous spike responses at slightly higher frequency compared with that of Gelidium amansii. Considering a role of the CBM3 in the pattern selection, these results suggest that elongation of the initial firing response may be a major factor for the CBM3 to switch the buccal motor programs from rejection to ingestion after application of different tastes of seaweeds in Aplysia kurodai.     

Pleural-buccal interneurons in the pteropod mollusc Clione limacina

Paired, Phe-Met-Arg-Phe-NH₂-ergic pleural-to-buccal projecting neurons of the pleural ganglia were suggested to be responsible for feeding arrest associated with defensive withdrawal in freshwater and terrestrial pulmonate molluscs. In the present study, the pleural-to-buccal projecting cells were, for the first time, identified in a representative opisthobranch, the carnivorous marine pteropod Clione limacina. Two symmetric neurons of its pleural ganglia were found to be similar to the pulmonate pleural-to-buccal projecting neurons in the number of neurons, positions of their cell bodies in the central nervous system, a unique, indirect route of their axon, electrotonic coupling of the left and right cells, and expression of Phe-Met-Arg-Phe-NH₂-like immunoreactivity and inhibitory action on neurons participating in the motor program for feeding. In their turn, pleural-to-buccal projecting neurons receive excitatory inputs from the protractor interneurons involved in the feeding rhythm generation. Also, it was demonstrated that the pleural-to-buccal projecting cells activity positively correlates with spontaneous and induced acceleration of the locomotor rhythm. Accordingly, stimulation of the cerebral command neuron for locomotion, cell CPA1, excited pleural-to-buccal projecting neurons. We conclude that the neuronal network underlying feeding behavior in both pulmonate and opisthobranch molluscs is similarly linked to defensive behavior by pleural Phe-Met-Arg-Phe-NH₂-ergic neurons, thus indicating evolutionary conservation of these pleural-buccal projections. Accepted: 22 June 1999     

Microtubule-associated protein 2 within axons of spinal motor neurons: associations with microtubules and neurofilaments in normal and beta,beta'-iminodipropionitrile-treated axons

We have examined the distribution of microtubule-associated protein 2 (MAP2) in the lumbar segment of spinal cord, ventral and dorsal roots, and dorsal root ganglia of control and beta,beta'-iminodipropionitrile- treated rats. The peroxidase-antiperoxidase technique was used for light and electron microscopic immunohistochemical studies with two monoclonal antibodies directed against different epitopes of Chinese hamster brain MAP2, designated AP9 and AP13. MAP2 immunoreactivity was present in axons of spinal motor neurons, but was not detected in axons of white matter tracts of spinal cord and in the majority of axons of the dorsal root. A gradient of staining intensity among dendrites, cell bodies, and axons of spinal motor neurons was present, with dendrites staining most intensely and axons the least. While dendrites and cell bodies of all neurons in the spinal cord were intensely positive, neurons of the dorsal root ganglia were variably stained. The axons of labeled dorsal root ganglion cells were intensely labeled up to their bifurcation; beyond this point, while only occasional central processes in dorsal roots were weakly stained, the majority of peripheral processes in spinal nerves were positive. beta,beta'- Iminodipropionitrile produced segregation of microtubules and membranous organelles from neurofilaments in the peripheral nervous system portion and accumulation of neurofilaments in the central nervous system portion of spinal motor axons. While both anti-MAP2 hybridoma antibodies co-localized with microtubules in the central nervous system portion, only one co-localized with microtubules in the peripheral nervous system portion of spinal motor axons, while the other antibody co-localized with neurofilaments and did not stain the central region of the axon which contained microtubules. These findings suggest that (a) MAP2 is present in axons of spinal motor neurons, albeit in a lower concentration or in a different form than is present in dendrites, and (b) the MAP2 in axons interacts with both microtubules and neurofilaments.     

Descending unpaired median neurons with bilaterally symmetrical axons in the suboesophageal ganglion of Manduca sexta larvae

Three large median cell bodies with a diameter between 40 and 70 μm that exhibit octopamine immunoreactivity were identified in the posterior part of the suboesophageal ganglion of the tobacco hawkmoth larva, Manduca sexta. These neurons possess bilaterally symmetrical axons in the posterior neck connectives, and at least one of them extends through the whole ventral nerve cord to the terminal abdominal ganglion. Therefore, these neurons belong to the class of descending ventral unpaired median neurons. From each cell body, a primary neurite ascends anteriorly, which after bending dorsally turns posteriorly and then bifurcates to give rise to two descending axons. From the primary neurite two main dendritic branches ascend anteriorly, and four characteristic branches can be distinguished originating from them: two descending dendritic branches and two ascending dendritic branches. Dense arborizations from all these branches exist in all neuromeres of the suboesophageal ganglion. Intracellular recordings from these neurons show that in contrast to the ventral unpaired median neurons of thoracic and abdominal ganglia, they do not produce overshooting action potentials but exhibit passive soma spikes only. During pharmacologically evoked fictive motor patterns these neurons show coupling to various motor patterns such as crawling, feeding and molting.     

Microanatomy of the subumbrellar motor innervation inAglantha digitale (Hydromedusae: Trachylina)

Summary The hydrozoan medusaAglantha digitale (Müller 1776) has eight syncytical giant motor axons, up to 40 m in diameter, running from the margin, up the inside of the bell towards the apex. Giant motor axons injected with Lucifer Yellow CH are connected with lateral neurons running circumferentially across the subumbrellar muscle. These processes fill with the dye. Bundles of 20 to 50 small dye-coupled neurons extend circumferentially along the margin for up to 0.85mm. Giant motor axons injected with horseradish peroxidase divide into a few short branches on entering the inner nerve ring. Here the giant motor axon forms both chemical synapses and gap junctions with neurons that also send their axons into the inner nerve ring. In this region the inner and outer nerve ringe are connected by axons passing through openings in the intervening mesoglea.     

Morphological and physiological properties of the giant interneuron of the hermit crab (Pagurus pollicaris)

The physiological and morphological properties of the giant interneurons in the hermit crab Pagurus pollicaris are described. The cell bodies are located anteriorly in the supraesophageal ganglion, close to the mid-line. Each cell sends a neurite posteriorly and then laterally, so that they cross over in the center of the ganglion. Each axon then branches: one branch runs laterally while the other travels posteriorly and leaves the ganglion in the circumesophageal connective on the side contralateral to the cell body. The giant axons travel in the circumesophageal connectives and through the thoracic and abdominal ganglia without branching. Each giant axon makes synaptic contact with its ipsilateral giant abdominal flexor motor neuron and with a second flexor motor neuron that has its axon in the contralateral third root. In the supraesophageal ganglion there is a bidirectional synapse between the two giant interneurons. Intracellular recordings from the giant axons show that there is a delay of 0.5 to 0.75 ms that cannot be accounted for by spike propagation along the axons, and may be accounted for by a chemical synapse between the giant interneurons.     

Localization of the myomodulin-like immunoreactivity in the leech CNS

The distribution of myomodulinlike immunoreactivity in the leech CNS was determined using an antiserum raised against Aplysia myomodulin. Segmental ganglia contained approximately 60 immunoreactive neurons. In addition, numerous fibers containing immunoreactive varicosities were found throughout the neuropil. Using a combination of Lucifer Yellow injections and immunocytochemistry, we identified neurons including the anterior Pagodas (AP), annulus erector (AE), motor neurons, Leydig, longitudinal muscle motoneurons (L), S cells, and coupling interneurons, all of which are active during the touch-elicited shortening reflex. FMRF-amide-like immunoreactivity in three of these cells (L, AP, and AE) was previously demonstrated. Specific staining for myomodulin was abolished by preadsorption of the antiserum with synthetic myomodulin, but not with FMRF-amide. These results suggest a potential role for myomodulin in both intrinsic and extrinsic modulation of the leech touch-elicited shortening reflex. Further, it is possible that several neurons mediating this reflex contain multiple neuromodulatory peptides. © 1996 John Wiley & Sons, Inc.     

Morphological properties of neuron RPD1 in lymnaea stagnalis and its involvement in processing of polymodal sensory information

Characteristics of the right partietal ganglion neuron of the gastropod molluskLymnaea stagnalis (RPD1) were investigated by intracellular staining with Lucifer Yellow. Branches proceeding from this neuron are found in nerves of the right parietal, visceral, cerebral, and pedal ganglia of the central nervous system (CNS) as well as along peripheral nerves. Concentrations of RPD1 neurite branches were revealed in the distal, right parietal, and pleural ganglia. Electrophysiological techniques were used to investigate neuronal response to adequate stimulation of different sensory organs and cutaneous coverings (tentacles, lips, mantle, and so on). It was found that RPD1 has wide-ranging polymodal sensory input and responds to adequate stimulation of mechano-, chemo-, and photoreceptors of cutaneous coverings of the head of mantle. Stimulus application produced either subthreshold summated EPSP or a spike response in the neuron. Response in this unit during blockade of chemical synaptic transmission at peripheral and central regions of the nervous system is analyzed.A. A. Ukhtomskii Physiological Institute, A. A. Zhdanov State University, Leningrad. Translated from Neirofiziologiya, Vol. 20, No. 6, pp. 785–793, November–December, 1988.     

Prey capture phase of feeding behavior in the pteropod mollusc,clione limacina: neuronal mechanisms

The prey capture phase of feeding behavior in the pteropod mollusc Clione limacina consists of an explosive extrusion of buccal cones, specialized structures which are used to catch the prey, and acceleration of swimming with frequent turning and looping produced by tail bend. A system of neurons which control different components of prey capture behavior in Clione has been identified in the cerebral ganglia. Cerebral B and L neurons produce retraction of buccal cones and tightening of the lips over them — their spontaneous spike activities maintain buccal cones in the withdrawn position. Cerebral A neurons inhibit B and L cells and produce opening of the lips and extrusion of buccal cones. A pair of cerebral interneurons C-BM activates cerebral A neurons and synchronously initiates the feeding motor program in the buccal ganglia. Cerebral T neurons initiate acceleration of swimming and produce tail bending which underlies turning and looping during the prey capture. Both tactile and chemical inputs from the prey produce activation of cerebral A and T neurons. This reaction appears to be specific, since objects other than alive Limacina or Limacina juice do not initiate activities of A and T neurons.     

The morphology and electrophysiology of the neurones of the paired pedal ganglia of Lymnaea stagnalis (L.)

1. A morphological and electrophysiological map of the identifiable neurones and neuronal clusters of the paired pedal ganglia has been prepared. 2. Neuronal morphology was investigated using the fluorescent dye, Lucifer Yellow CH, whilst electrophysiological properties were studied using conventional intracellular recording techniques and the phase plane technique. 3. The paired pedal ganglia are largely symmetrical and giant neurones usually have contralateral homologues. 4. Neuronal clusters are also paired, but minor asymmetries, both of identifiable neurones and neuronal clusters have been found to exist. 5. These asymmetries are thought to be related to asymmetries of body form. 6. Most of the individually identifiable neurones possess obligatory axon branches which are invariant from one preparation to the next, but variant branches also occur. 7. Within the neuronal clusters, morphology appears to be more variable. 8. Individually identifiable neurones and neuronal clusters were characterized electrophysiologically according to the criteria of action potential shape, spontaneous activity pattern, electrical coupling and common synaptic inputs. 9. Homologous pairs of neurones usually have similar electrophysiological properties, as do those within clusters. 10. A number of wide-acting synaptic inputs have been identified on neurones of the pedal, buccal, visceral and parietal ganglia.