Identification of proctolin in the central nervous system of the horseshoe crab, Limulus polyphemus

Groome, James R.; Tillinghast, Edward K.; Townley, Mark A.; Vetrovs, A.; Watson, Winsor H.; Hunt, Donald F.; Griffin, Patrick R.; Alexander, Janice E.; Shabanowitz, Jeffrey

doi:10.1016/0196-9781(90)90072-d

Cited by 18 publications

(11 citation statements)

References 21 publications

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“…Adult specimens of horseshoe crabs, Limulus polyphemus, of carapace width 10 to 15 cm, were collected locally from Great Bay, NH. Horseshoe crabs were maintained as described in Groome et al (1990). The Limulus midgut, or stomachintestine, extends from the prosomal foregut to the rectum of the opisthosoma (Groome et al 1992).…”

Section: Methodsmentioning

confidence: 99%

“…Isolated midguts were prepared for tension recordings as described previously (Groome et al 1990). Preparations were perfused with natural seawater or Limulus saline (445 mM NaCI, 12 mM KC1, 10 mM CaC12, 46 mM MgC12, with 10 mM Hepes buffer at pH 7.5).…”

Section: Methodsmentioning

confidence: 99%

“…Proctolin and OCT produce contracture and elicit rhythmic contractions of the Limulus midgut (Groome et al 1990;unpublished). DA, at 3 x 10 .6 M, was added to 8 preparations pretreated with either 3 x 10-7 M proctolin or 3 x 10 .6 M OCT.…”

Section: Effect Of Da On Proctolin-and Oct-induced Contracturesmentioning

confidence: 99%

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Inhibitory actions of dopamine on Limulus visceral muscle involve a cyclic AMP-dependent mechanism

Groome

Lent

1992

J Comp Physiol A

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1. The catecholamines dopamine, epinephrine and norepinephrine were detected in alumina extracts of Limulus midgut tissue using high performance liquid chromatography with electrochemical detection. Moderate levels of norepinephrine (28.2 +/- 2.1 ng/g) and dopamine (24.0 +/- 5.2 ng/g) were detected in the midgut, while epinephrine levels (7.4 +/- 0.9 ng/g) were less. Catecholamines were present in all regions along the longitudinal axis of the midgut, and norepinephrine and dopamine levels were highest in posterior regions. 2. Catecholamines decreased muscle tonus and inhibited spontaneous contractions of the Limulus midgut. Dopamine typically decreased spontaneous midgut activity at doses of 10(-8) M or greater, and produced inhibitory actions on all regions of the Limulus midgut. In some preparations epinephrine and norepinephrine elicited a secondary rhythmicity. The actions of dopamine opposed the excitatory effects produced by either proctolin or octopamine. 3. Catecholamines significantly elevated levels of cyclic AMP in Limulus midgut muscle rings. Dopamine (10(-5) M) increased cyclic AMP with a time course consistent with its physiological effects. Forskolin and several methyl xanthines increased Limulus midgut cyclic AMP levels and mimicked the inhibitory effects of dopamine on the isolated midgut preparation. Cyclic nucleotide analogues also produced dopamine-like effects on the isolated midgut preparation. Inhibition of cyclic nucleotide phosphodiesterase prior to addition of dopamine enhanced the effect of this amine to decrease baseline muscle tension. 4. The inhibitory effects of 10(-5) M dopamine on the midgut persisted in solutions of zero sodium and in the presence of tetrodotoxin. Zero calcium solutions gradually reduced spontaneous midgut activity and the effects of dopamine. Calcium channel blockers did not prohibit dopamine-induced relaxation.(ABSTRACT TRUNCATED AT 250 WORDS)

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Inhibitory actions of dopamine on Limulus visceral muscle involve a cyclic AMP-dependent mechanism

Groome

Lent

1992

J Comp Physiol A

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“…The pellets were solubilized with 100 ~tl Protosol (New England Nuclear, Boston, MA) and 1 ml Aquasol (New England Nuclear) was added to permit liquid scintillation counting (LKB 1214 Rackbeta Excel, Finland). Peptide standards and Limulus tissue extracts display parallel inhibition characteristics in these assays (Watson et al 1984;Groome et al 1990). We used dilutions of gut extracts which produced 50% or less inhibition of antibody to tracer binding to eliminate the possibility of extract interference with this assay.…”

Section: Animals Adult Horseshoe Crabs (Limulus Polyphemus Carapacementioning

confidence: 98%

“…The midgut, or intestine, of the horseshoe crab is supplied by a plexus of nerves which originate from the CNS (Patten and Redenbaugh 1899). The Limulus CNS contains immunoreactive proctolin (Groome et al 1990) and FMRFamide (Watson et al 1984), peptides which have been suggested as circulating or local modulators of muscle contractility and synaptic efficacy in Limulus. The isolated midgut exhibits an ongoing pattern of activity upon which the action of putative modulators may be assessed, and the quantity of this tissue (approximately I 0 g in adult Limulus) facilitates biochemical studies.…”

Section: Introductionmentioning

confidence: 99%

Peptidergic regulation of the Limulus midgut

Groome

deTschaschell

1992

J Comp Physiol A

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1. The morphology and innervation of the midgut (intestine) in the horseshoe crab, Limulus polyphemus was investigated. The organization of this tissue was examined with routine histology. Radioimmunoassay, immunohistochemistry and high performance liquid chromatography were employed to detect, localize and identify peptidergic innervation of the midgut. The actions of synthetic and native proctolin-like and FMRFamide-like peptides were compared on the isolated midgut preparation. 2. Levels of proctolin and FMRFamide were determined in extracts of Limulus midgut tissue using radioimmunoassay. High levels of proctolin-like immunoreactivity (69.5 +/- 11.3 ng/g) were detected, while levels of FMRFamide-like immunoreactivity (0.8 +/- 0.2 ng/g) were less. Proctolin levels were equally distributed, while the levels of FMRFamide-like immunoreactivity exhibited an anterior bias. 3. Proctolin- and FMRFamide-like immunoreactivities in the Limulus midgut were localized with immunohistochemistry. Proctolin- and FMRFamide-immunoreactive elements were detected in intestinal nerve branches and individual fibers running along the surface of the midgut in whole-mount preparations. In sectioned tissue, staining for these peptides was observed throughout the midgut, typically associated with muscle bands and fibers. Only a few immunoreactive cell bodies were observed. 4. Proctolin, and several FMRFamide-like peptides produced distinct and opposing actions on the isolated Limulus midgut preparation. Proctolin elicited contracture and rhythmic contractions of this tissue, while FMRFamide and N-terminally extended analogs of FLRFamide relaxed gut tension. FMRFamide-like peptides partially reversed the excitatory actions of proctolin. 5. Proctolin- and FMRFamide-like peptides in Limulus midgut extracts were partially characterized with high performance liquid chromatography. One peak of proctolin-like activity was detected on a linear gradient of 18 to 31.5% acetonitrile. The native proctolin-like peptide produced excitatory actions on the isolated midgut preparation which were indistinguishable from those produced by synthetic proctolin. Several peaks of FMRFamide-like bioactivity (Busycon radula protractor muscle assay) were detected with a linear gradient of 5 to 30% acetonitrile. Fractions from two distinct peaks produced FMRFamide-like inhibitory effects on the isolated Limulus midgut preparation. These findings suggest a role for proctolin-like and FMRFamide-like peptides as regulators of intestinal motility in Limulus.

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Immunocytochemical localization of FLRFamide-, proctolin-, and CCAP-like peptides in the stomatogastric nervous system and neurohemal structures of the crayfish,Cherax destructor

1999

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To compare the stomatogastric nervous system of the crayfish Cherax destructor with those of other decapod species, the distribution of FLRF (Phe‐Leu‐Arg‐Phe) amide‐, proctolin‐ and crustacean cardioactive peptide (CCAP)‐like immunoreactivities was studied in the stomatogastric nervous system and in neurosecretory structures by using wholemount immunocytochemical techniques and confocal microscopy. In addition, the number of cells in the stomatogastric ganglion (19–24) and axon profiles in the stomatogastric nerve (157–165) were counted. FLRFamide‐like immunoreactivity was present within numerous cell bodies and neuropil of the commissural ganglia, in the neuropil of the stomatogastric ganglion, and in one cell body of the esophageal ganglion. FLRFamide‐like immunoreactivity was also found in two cell bodies at the junction of the stomatogastric nerve with the superior esophageal nerve and in two cell bodies in the inferior ventricular nerve. Proctolin‐like immunoreactivity was present in numerous cell bodies and neuropil of the paired commissural ganglia and in the neuropil of the stomatogastric ganglion. CCAP‐like immunoreactivity was found in the neuropil and in one to four cell bodies in the commissural ganglia. Both proctolin‐ and CCAP‐immunoreactive varicosities occurred on the surface of the circumesophageal connectives and on the postesophageal commissure, indicating a neurohemal source within the stomatogastric nervous system, which was verified by electron microscopy. The pericardial organs showed FLRFamide‐, proctolin‐, and CCAP‐like immunoreactivity. This staining pattern suggests that FLRFamide‐like and proctolin‐like peptides are used as neurohormones and as neuromodulators in the stomatogastric nervous system of the crayfish C. destructor, whereas CCAP‐like peptides may only affect the stomatogastric ganglion as a neurohormone. J. Comp. Neurol. 414:511–532, 1999. © 1999 Wiley‐Liss, Inc.

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Identification of proctolin in the central nervous system of the horseshoe crab, Limulus polyphemus

Cited by 18 publications

References 21 publications

Inhibitory actions of dopamine on Limulus visceral muscle involve a cyclic AMP-dependent mechanism

Inhibitory actions of dopamine on Limulus visceral muscle involve a cyclic AMP-dependent mechanism

Peptidergic regulation of the Limulus midgut

Immunocytochemical localization of FLRFamide-, proctolin-, and CCAP-like peptides in the stomatogastric nervous system and neurohemal structures of the crayfish,Cherax destructor

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