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PubMed2003

Dopamine and histamine in the developing stomatogastric system of the lobster Homarus americanus.

Dopamine and histamine are neuromodulators found in the adult stomatogastric nervous system (STNS) of several crustacean species. We used antibodies against tyrosine hydroxylase (TH) and histamine to map the distribution and developmental acquisition of the dopamine and histamine neurons in the STNS of the lobster, Homarus americanus. Embryos, larvae, juvenile and adult animals were studied. TH labeling was present in the STNS as early as E80-85 (80-85% of embryonic development). A subset of preparations in embryos, larvae, juveniles, and adults contained 1-5 labeled somata in the stomatogastric ganglion. Histamine staining appeared in the STNS as early as E50. The distribution of both TH and histamine staining remained relatively constant through development. Electrophysiological recordings demonstrated that receptors for both amines are present in the embryo. Bath application of dopamine increased the frequency of the pyloric rhythm in embryos, and evidence for dopaminergic activation of peripherally initiated spiking in motor axons was seen. In embryos and adults, histamine inhibited the motor patterns produced by the stomatogastric ganglion (STG). These data suggest that the dopaminergic and histaminergic systems in H. americanus appear relatively early in development and that the effects of each are largely maintained through development.

Animals↗
PubMed2001

Serotonergic and octopaminergic systems in the squat lobster Munida quadrispina (Anomura, Galatheidae).

Immunocytochemical mapping of serotonergic and octopaminergic neurons in the central nervous system of the squat lobster Munida quadrispina reveal approximately 120 serotonin-immunoreactive cell bodies (distributed throughout the neuromeres except in abdominal ganglion 5) and 48 octopamine-immunoreactive cell bodies (in brain and thoracic neuromeres but none in the circumesophageal or abdominal ganglia). Immunopositive neuropils for both amines are distributed in multiple areas in each neuromere and overlap extensively. Serotonergic and octopaminergic neurons have extensive bilateral projections in abdominal ganglia, whereas the majority of projections in thoracic and subesophageal ganglia are unilateral (contralateral to soma). This difference correlates with typical differences between abdominal and thoracic motor system coordination. Processes of immunoreactive cells for both amines form extensive, peripheral, neurosecretory-like structures. Serotonin seems to be released peripherally in more segments, and from more nerves per segment, than octopamine. M. quadrispina has fewer serotonergic and octopaminergic immunoreactive cells, in particular, fewer segmentally repeated cells, than other species studied to date. Nevertheless, the general organization of the aminergic systems is similar, and several aminergic cells have locations and morphologies that strongly suggest homology with identified aminergic cells in other crustaceans. Among these are segmentally repeated neurons that, in M. quadrispina, form serotonin-immunopositive tubular structures in the thoracic hemiganglia innervating pereiopods 1-3 that are unlike anything reported previously for any species. Comparisons of immunocytochemical maps within one species and between species exhibiting different behaviors provide insights into possible sites of action, functional differences between, and evolution of biogenic aminergic systems.

Animals↗
PubMed2001

Neural pathways connecting the deutocerebrum and lateral protocerebrum in the brains of decapod crustaceans.

The olfactory and accessory lobes of eureptantian decapod crustaceans are bilateral brain neuropil regions located within the deutocerebrum. Although the olfactory lobe seems to receive only primary olfactory inputs, the accessory lobe receives higher-order multimodal (including olfactory) inputs. The output pathways from both the olfactory and accessory lobes are provided by the axons of a large population of projection neurons, whose somata lie adjacent to the lobes. The axons of these neurons form a large tract that projects bilaterally to the medulla terminalis and hemiellipsoid body in the lateral protocerebrum. To gain insights into the ways in which olfactory information is processed on leaving the deutocerebrum, we examined the neuroanatomy of the projection neuron pathways of three species of eureptantian decapod crustaceans: the freshwater crayfish, Procambarus clarkii and Orconectes rusticus, and the clawed lobster, Homarus americanus. Projection neurons were labeled by focal injections of the lipophilic tracers DiI and DiA into the olfactory and accessory lobes. In all three species, projection neurons innervating the accessory lobe were found to exclusively innervate the neuropils of the hemiellipsoid body. In contrast, projection neurons innervating the olfactory lobes primarily target neuropil regions of the medulla terminalis. The results of this study indicate, therefore, that the projection neuron pathways from the olfactory and accessory lobes project to separate, largely nonoverlapping regions of the lateral protocerebrum. The implications of these findings for our understanding of the processing of olfactory information in the brains of decapod crustaceans are discussed.

Animals↗
PubMed2001

Development and connectivity of olfactory pathways in the brain of the lobster Homarus americanus.

The main output pathways from the olfactory lobes (primary olfactory centers) and accessory lobes (higher-order integrative areas) of decapod crustaceans terminate within both of the main neuropil regions of the lateral protocerebrum: the medulla terminalis and the hemiellipsoid body. The present study examines the morphogenesis of the lateral protocerebral neuropils of the lobster, Homarus americanus, and the development of their neuronal connections with the paired olfactory and accessory lobes. The medulla terminalis was found to emerge during the initial stages of embryogenesis and to be the target neuropil of the output pathway from the olfactory lobe. In contrast, the hemiellipsoid body is first apparent during mid-embryonic development and is innervated by the output pathway from the accessory lobe. The dye injections used to elucidate these pathways also resulted in the labeling of a previously undescribed pathway linking the olfactory lobe and the ventral nerve cord. To increase our understanding of the morphology of the olfactory pathways in H. americanus we also examined the connectivity of the lateral protocerebral neuropils of embryonic lobsters. These studies identified several interneuronal populations that may be involved in the higher-order processing of olfactory inputs. In addition, we examined the neuroanatomy of ascending pathways from the antenna II and lateral antenna I neuropils (neuropils involved in the processing of chemosensory and tactile inputs). These studies showed that the ascending pathways from these neuropils innervate the same regions of the medulla terminalis and that these regions are different from those innervated by the olfactory lobe output pathway.

Animals↗
PubMed2006

Identification, physiological actions, and distribution of VYRKPPFNGSIFamide (Val1)-SIFamide) in the stomatogastric nervous system of the American lobster Homarus americanus.

In this study, the peptide VYRKPPFNGSIFamide (Val(1)-SIFamide) was identified in the stomatogastric nervous system (STNS) of the American lobster, Homarus americanus, using matrix-assisted laser desorption/ionization-Fourier transform mass spectrometry (MALDI-FTMS). When bath-applied to the stomatogastric ganglion (STG), synthetic Val(1)-SIFamide activated the pyloric motor pattern, increasing both burst amplitude and duration in the pyloric dilator (PD) neurons. To determine the distribution of this novel SIFamide isoform within the lobster STNS and neuroendocrine organs, a rabbit polyclonal antibody was generated against synthetic Val(1)-SIFamide. Whole-mount immunolabeling with this antibody showed that this peptide is widely distributed within the STNS, including extensive neuropil staining in the STG and commissural ganglia (CoGs) as well as immunopositive somata in the CoGs and the oesophageal ganglion. Labeling was also occasionally seen in the pericardial organ (PO), but not in the sinus gland. When present in the PO, labeling was restricted to fibers-of-passage and was never seen in release terminals. Adsorption of the antibody by either Val(1)-SIFamide or Gly(1)-SIFamide abolished all Val(1)-SIFamide staining within the STNS, including the STG neuropil, whereas adsorption by other lobster neuropeptides had no effect on immunolabeling. These data strongly suggest that the staining we report is a true reflection of the distribution of this peptide in the STNS. Collectively, our mass spectrometric, physiological, and anatomical data are consistent with Val(1)-SIFamide serving as a locally released neuromodulator in the lobster STG. Thus, our study provides the first direct demonstration of function for an SIFamide isoform in any species.

Animals↗
PubMed2007

Neuronal morphology and neuropil structure in the stomatogastric ganglion of the lobster, Homarus americanus.

The stomatogastric nervous system (STNS) has long been used as a model system for the study of central pattern generation, neuromodulation, and network dynamics. Anatomical studies of the crustacean stomatogastric ganglion (STG) in different species have mostly been restricted to subsets of neurons and/or general structural features. For the first time, we describe the morphology of all STG neurons belonging to the two circuits that produce the well-described pyloric and gastric rhythms in the lobster, Homarus americanus. Somata sit on the dorsal and lateral surface of the STG and send a single primary neurite into the core of the neuropil, which is mostly made up of larger lower order branches. The perimeter of the neuropil consists mostly of finer higher order branches. Immunohistochemical labeling for synaptic proteins is associated with the small diameter branches. Somata positions are not constant but show preferred locations across individuals. The number of copies is constant for all neuron types except the PY and GM neurons (PY neuron number ranges from 3 to 7, and GM neuron number ranges from 6 to 9). Branch structure is largely nondichotomous, and branches can deviate substantially from cylindrical shape. Diameter changes at branch points can be as large as 20-fold. Clearly, the morphology of a specific neuron type can be quite variable from animal to animal.

Animals↗
PubMed1976

Growth of lobster giant axons: correlation between conduction velocity and axon diameter.

The conduction velocity and axon diameter of medial and lateral giant axons in lobsters was followed during growth from early juvenile (13 mm) to adult (250 mm) forms. In both axons, conduction velocity increased more than 4-fold in the growth period examined. Correlated with the increase in conduction velocity is an increase in axon diameter and the relationship between these two parameters is expressed by the equation CV = krX, where X equals 0.5 for the lateral and 0.7 for the medial giant axons. The sheath surrounding the giant axons also increases in size during growth with that of the medial giant axon being considerably thicker than that of the lateral giant axon. The significance of the increase in axon diameters is to increase conduction velocity and thereby reduce the latency of the escape response mediated by these giant axons.

Animals↗
PubMed1980

Development of excitatory innervation in the lobster claw closer muscle.

The development of excitatory innervation to the claw closer muscles of 1st (larval), 4th (juvenile), and adult stage lobster was examined by thin serial section electron microscopy. This was possible since neuromuscular terminals of the excitatory axon are distinguishable from those of the inhibitory axon by the shape of their synaptic vesicles in all three stages. In the adult cutter claw closer muscle, innervation of a dorsal and a ventral fiber which is supplied by the fast closer excitor (FCE) and slow closer excitor (SCE) axons respectively, was qualitatively as well as quantitatively similar. Consequently no attempt was made to distinguish between innervation by FCE and SCE axons in the subsequent analysis of 1st, 4th, and adult stage claw closer muscles. Excitatory innervation in the 1st larval stage is limited to four discrete locations in the entire cross-sectional area of the closer muscle. It subsequently spreads to each individual muscle fiber in the adult, thus demonstrating the tremendous proliferation of innervation during development. Concomitantly the mean size of synapses increases significantly from the 1st stage to the 4th stage to the adult lobster. This increase in synaptic size may occur by both the fusion and enlargement of existing smaller synapses. In contrast, the mean size of presynaptic dense bars and their mean number per synapse remained fairly constant in each of the 1st, 4th, and adult stages. However, a relatively greater proportion of adult synapses possessed two or more dense bars compared to their larval and juvenile counterparts. Development of innervation of the lobster claw closer muscle therefore consists of a substantial proliferation of axonal tissue and enlargement of synaptic size.

Animals↗
PubMed1981

The distal limb motor neurons in the thoracic ganglion of the spiny lobster.

The 16 motor neurons which innervate the muscles in the limb distal to the autotomy plane have been found in the third thoracic ganglion of the southern rock lobster, Jasus novaehollandiae. They were traced from the autotomy plane to the ganglion by stripping them from the small and large leg nerves to the edge of the ganglion where they were filled with cobalt dye. Cell bodies of 11 of these neurons lie in an anterior cluster of somata, and the remainder lie in a ventral rind of somata. The neurites from these somata expand into enlarged central segments, 400 micrometers long and 10-30 micrometers in diameter. These segments run parallel and in apposition to each other as they course through the neuropile a few cell layers beneath the dorsal surface of the ganglion. Near their origin some central segments send branches both medially between fibers of the interganglionic connectives, and anteroventrally into the connective. Distally the segments give rise to side branches which project around the anterior and posterolateral borders of the neuropile. The central segments constrict near the lateral edge of the neuropile, rise to its surface, and form axons. Eight axons, including those which innervate the flexor muscle of the carpus, enter an anterior motor tract which exists the side of the ganglion in a first root, the source of the small leg nerve. The remaining eight axons, which include those that innervate the extensor muscle of the carpus, enter a posterior motor tract which exists the side of the ganglion in a second root, the source of the large leg nerve. All the motor fibers which innervate the muscles of the limb, with the exception of the retractor of the coxa, run in these two tracts within the ganglion. Fibers which innervate the retractor of the coxa and the muscles of the thorax leave the ganglion in a third root at the posterodorsal border of the ganglion. The central segment, as the common structure of confluence of neuropilar branches, is the likely candidate for the integrating region of each neuron.

Animals↗
PubMed1983

Fast and slow motoneurons with unique forms and activity patterns in lobster claws.

The form of the fast closer excitor (FCE) and the slow closer excitor (SCE) motoneurons to the closer muscle in the claw of the lobster Homarus americanus was determined by injecting cobalt chloride or Lucifer Yellow into their respective somata. Both neurons are monopolar with the single neurite rising vertically to the dorsal surface of the ganglion, then travelling along this surface to where it gives off its dendrites before entering the second nerve root as an axon. The FCE and SCE motoneurons, however, differ in their dendritic form in several respects. First, the FCE completely lacks an anterior dendritic field, which is well elaborated in the SCE. Second, the FCE has fewer large primary dendrites in its posterior field than the SCE. Third, the posterior dendritic field of the FCE is not as extensive as that of the SCE. Fourth, the axon of the FCE originates from one of the posterior primary dendrites while that of the SCE is an axial extension of its neurite. Thus the SCE has a more elaborate dendritic field than the FCE, which may account for its greater excitability. For instance, recordings from intact lobsters show that the SCE has a lower firing threshold and is active for longer periods of time and at higher frequencies than the FCE.

Action Potentials↗
PubMed1986

Mapping of proctolinlike immunoreactivity in the nervous systems of lobster and crayfish.

Whole-mount immunocytochemical techniques have been used to map candidate proctolin-containing cells in the central nervous systems of the lobster, Homarus americanus, and the crayfish, Procambarus clarkii. Proctolinlike immunoreactivity was detected in cell bodies and neuropil regions in all central ganglia, and immunoreactive axons were detected in most interganglionic connectives and nerve roots. Cell body staining was confined to fewer than 2% of all cells. Immunoreactive neurons include motoneurons, sensory neurons, neurosecretory cells, and interneurons. Colocalization of the proctolinlike antigen with other neurotransmitters was indicated in a number of cases. Many aspects of the distribution of immunoreactivity were similar in lobster and crayfish; however, staining differences were detected in a number of identified neurons and neural groups, including neurons that innervate the pericardial organs and hindgut motoneurons. Further studies of such neurons might provide interesting clues about the physiological functions of proctolin and the evolution of peptide transmission.

Animals↗
PubMed1986

Modulatory action and distribution of the neuropeptide proctolin in the crustacean stomatogastric nervous system.

Immunocytochemical methods were used to map the distribution of proctolinlike immunoreactivity in the stomatogastric nervous systems (stomatogastric ganglion (STG), paired commissural ganglia (CG), oesophageal ganglion (OG), and connecting nerves) of three crustacean species: Panulirus interruptus, Cancer borealis, and Homarus americanus. Although the patterns of proctolinlike staining were similar among the three species, some differences were also observed. Over 70% of the proctolinlike material in STGs, as measured by radioimmunoassay, was indistinguishable from authentic proctolin in reverse-phase high-performance liquid chromatography. Bath application of proctolin to STGs from Cancer and Panulirus induced characteristic and robust (though somewhat different) changes in their motor patterns. The threshold concentration was approximately 10(-9)M proctolin, and the effects were dose-dependent. These data suggest that the neuropeptide proctolin serves as a neuromodulator of the stomatogastric ganglion.

Animals↗
PubMed1987

Distribution and partial characterization of FMRFamide-like peptides in the stomatogastric nervous systems of the rock crab, Cancer borealis, and the spiny lobster, Panulirus interruptus.

The distribution of FMRFamide-like peptides was studied in the complete stomatogastric nervous system [the paired commissural ganglia, single oesophageal ganglion, and the single stomatogastric ganglion (STG)] of two decapod crustacean species, the spiny lobster Panulirus interruptus and the rock crab Cancer borealis, by using immunocytochemical techniques. Antiserum 231 from the O'Donohue laboratory and antiserum 671C (described here) gave essentially the same staining patterns. In the commissural ganglia of both species there were ten to 20 stained neurons and dense neuropilar staining. The oesophageal ganglion of the crab had four stained neurons. Lucifer Yellow backfills followed by immunostaining showed that the two larger stained neurons of the oesophageal ganglion sent processes into the inferior ventricular nerve. The two smaller neurons sent processes into the inferior oesophageal nerves. The oesophageal ganglion of the lobster had two stained neurons that sent processes into the inferior ventricular nerve as well. None of the somata of the STG stained in either species, but in both species stained fibers were seen in the stomatogastric nerve that entered the STGs and ramified profusely throughout the neuropil. In some preparations of the crab, a stained fiber was visible in the dorsal ventricular nerve. The amounts of the FMRFamide-like peptides found in all regions of the nervous system of P. interruptus and C. borealis were determined by radioimmune assay (RIA). Column chromatography and high-performance liquid chromatography suggest that, in both species, much if not all of the RIA-assayable material is accountable for by peptides that are larger and more hydrophobic than FMRFamide.

Animals↗
PubMed1987

FMRFamidelike peptides of Homarus americanus: distribution, immunocytochemical mapping, and ultrastructural localization in terminal varicosities.

The distribution of FMRFamidelike peptides was studied in the nervous system of the lobster Homarus americanus by using immunocytochemical and radioimmunological techniques. By radioimmunoassay FMRFamidelike immunoreactivity (FLI) was found in low levels (ca. 1 pmol/mg protein) throughout the ventral nerve cord and in much higher amounts (60-100 pmol/mg protein) in the neurosecretory pericardial organs. Immunocytochemical studies showed FLI in approximately 300-350 cell bodies, and in distinct neuropil regions, neuronal fiber tracts, and varicose endings. Specificity of the immunostaining was tested by preabsorbing the antiserum with FMRFamide, with peptides having similar carboxyl termini to FMRFamide (Met-enkephalin-Arg-Phe, Phe-Met-Arg-Tyr-amide), with several amidated peptides (alpha-melanocyte-stimulating hormone, substance P, oxytocin), and with proctolin, a peptide found widely distributed in the lobster nervous system. Of these substances, only FMRFamide blocked the staining. In addition to the pericardial organs, significant levels of FLI were found in neurosecretory regions associated with thoracic second roots and in the connective tissue sheath that surrounds the ventral nerve cord. In all three regions, immunocytochemical studies showed the FLI to be localized to fine fibers and associated terminal varicosities lying close to the surface of the tissue, with no obvious target in their immediate vicinity. When examined at the ultrastructural level, the immunoreactive varicosities of the thoracic second roots and of the ventral nerve cord sheaths were found a few microns from the surface of the tissue and contained electron-dense granules. In the immunoreactive nerve cord sheath endings, in addition to the large, dense granules, small, clear vesicles were found. The appearance and location of these terminals suggest a neurohormonal role for FMRFamidelike peptides in lobsters. The observation that low levels of FLI are found in the hemolymph supports this suggestion. In addition, the localization of FLI to particular neuronal somata, fiber tracts, and neuropil regions suggests possible functional roles for these peptides in (1) integration of visual and olfactory information, (2) function of the anterior and posterior gut, and (3) the control of exoskeletal muscles.

Animals↗
PubMed1987

Purification and characterization of FMRFamidelike immunoreactive substances from the lobster nervous system: isolation and sequence analysis of two closely related peptides.

In the preceding paper (Kobierski et al: J. Comp. Neurol. 266:1-15, '87) FMRFamidelike immunoreactivity (FLI) was localized to specific cells and processes in the nervous system of the lobster Homarus americanus. In an effort to establish a role for this material we have purified and characterized a variety of immunoreactive peptides that can be extracted from the secretory pericardial organs. By using gel-filtration chromatography and three different HPLC systems, it has been established that little or no authentic FMRFamide is present. Of the major immunoreactive components two peptides were purified in sufficient quantity for microsequence analysis and have been tentatively identified as the octapeptides Ser-Asp-Arg-Asn-Phe-Leu-Arg-Phe-amide (FLI 3) and Thr-Asn-Arg-Asn-Phe-Leu-Arg-Phe-amide (FLI 4). Both of these are novel neuropeptides with some sequence homology to the previously described FMRFamide family. The pericardial organs release FLI when depolarized with 100 mM K+ in the presence of calcium. Between 75 and 80% of this release is accounted for by FLI 3 and FLI 4. One of these peptides (FLI 4) has been synthesized and shown to cochromatograph with the endogenous immunoreactive material. Preliminary studies show that this peptide can act as a modulator of exoskeletal and cardiac neuromuscular junctions.

Amino Acid Sequence↗
PubMed1988

Highly active neuromuscular system in developing lobsters with programmed obsolescence.

The primary locomotory apparatus in the three larval stages of the lobster, Homarus americanus, are paddlelike structures on the thoracic appendages called exopodites, which beat almost continuously. Consequently their power and return-stroke muscles are examples of highly active but short-lived neuromuscular systems. The muscles, which are well vascularized, are of the fast type with 2-3-micron sarcomere lengths and 6 thin filaments surrounding a thick one. The most striking feature, however, is the large volume of mitochondria making up 40-50% of the fiber. They appear as simple cylinders packed several layers deep along the periphery of the fiber and as large, multibranched forms distributed throughout the fiber and subdividing it into smaller units. The motor innervation to the return-stroke muscle is via 3 excitatory axons, which generate large junctional potentials and twitch contractions. The muscle is densely populated with large neuromuscular synapses, most of which have a well-defined active site or dense bar denoting the site of transmitter release. Altogether this motor system is specialized for prolonged activity. Atrophy of the neuromuscular system occurs by the late larval third stage. The muscle fibers lose their identity, fuse, and become vacuolated. The myofibrils condense and erode and the mitochondria are lost. Atrophy of motor innervation is gradual with individual axons dropping out. The largest axon providing most of the innervation is the first to degenerate. Early degenerative changes affect the axon and neuromuscular terminals but not the synaptic contacts, dense bars, and vesicles, which appear intact. Continued atrophy in the postlarval fourth stage reduces the exopodites to vestiges. Thus the return-stroke muscle of the larval exopodites in which muscle fiber and motoneurons are identifiable permits study of the interaction between a neuron and its target muscle undergoing programmed obsolescence.

Animals↗
PubMed1990

Immunohistochemical and biochemical evidence for the putative inhibitory neurotransmitters histamine and GABA in lobster olfactory lobes.

As an initial effort to investigate possible inhibitory interactions in the olfactory system of the spiny lobster, studies were conducted to identify and localize the putative inhibitory neurotransmitters histamine and GABA in the olfactory lobe. Biochemical studies demonstrated that olfactory lobe tissue was capable of synthesizing histamine from radioactive histidine and GABA from glutamic acid. Immunohistochemistry was used to localize histamine and GABA in brain sections, by using either avidin-biotin conjugated peroxidase or fluorescein conjugated secondary antibody. Specific histamine-like and GABA-like immunoreactivity was found in soma clusters of olfactory interneurons, adjacent to the olfactory lobe. Small, putative glial cells displaying intense histamine-like immunoreactivity were found interspersed among the glomeruli of the lobe. The accessory lobe exhibited moderate immunostaining for both histamine and GABA. Positive immunostaining for histamine and GABA was also found in the olfactory lobes, with a predominance of staining in the outer caps of the glomeruli, which are thought to be the regions where the primary afferent terminals contact the processes of second-order olfactory neurons. These findings collectively implicate inhibition at the first synaptic level of the olfactory pathway in the spiny lobster.

Animals↗