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Enkephalin-like immunoreactivity in the pedal ganglion of Mytilus edulis (Bivalvia) and its proximity to dopamine-containing structures.

The presence of enkephalin-like immunoreactivity in the pedal ganglion of the bivalve mollusc Mytilus edulis has been demonstrated. Enkephalin-like immunoreactivity is highly localized in those intraganglionic regions rich in dopamine-containing structures. In addition, FMRF-NH2-like (Phe-Met-Arg-Phe-HN2) immunoreactivity was found to occur in intraganglionic regions that are devoid of enkephalin-like immunoreactivity. The differential topographic distribution of these peptidergic immunoreactivities suggests involvement in separate functions. By contrast, the closeness of enkephalin-like immunoreactivity to dopamine histofluorescence supports previous data demonstrating biochemical and physiological links between these two systems.

Animals↗

Identical immunoreactivity of afferents to the rat suprachiasmatic nucleus with antisera against avian pancreatic polypeptide, molluscan cardioexcitatory peptide and neuropeptide Y.

Avian pancreatic polypeptide (APP)-like, molluscan cardioexcitatory peptide (FMRF)-like and neuropeptide Y (NPY)-like immunoreactivities were studied in a secondary visual pathway in rat brain. The cell bodies of this pathway are located in the lateral geniculate nucleus and its terminal plexus is found in the suprachiasmatic hypothalamic nucleus (SCN). The neurons and terminal plexus demonstrated by antiserum to each peptide are identical, and immunoreactivity is blocked by preabsorption of each antiserum with a low concentration of the antigen against which it was raised. Immunoreactivity is also blocked by preabsorption of each antiserum with either NPY or APP. In contrast, APP- and NPY-like immunoreactivities are blocked only partially when these antisera are preabsorbed with concentrations of FMRF as high as 100 microM. Since NPY is the only one of these peptides that has been isolated from mammalian brain, we conclude that NPY is the endogenous CNS peptide produced by neurons of the lateral geniculate-SCN projection.

Afferent Pathways↗

FMRF-amide-like immunoreactivity in brain and pituitary of the hagfish Eptatretus burgeri (Cyclostomata).

Paraffin sections of brain and pituitary of the hagfish Eptatretus burgeri were immunostained with an antiserum to FMRF-amide. Immunoreactivity was visible in a large number of neurons in the posterior part of the ventromedial hypothalamus and in long neuronal processes extending cranially from the hypothalamus to the olfactory system and caudally to the medulla oblongata. FMRF-amide-like immunoreactivity was also found in cells of the adenohypophysis. These observations suggest that the hagfish possesses a brain FMRF-amide-like transmitter system and pituitary cells containing FMRF-amide-like material. Antisera to ACTH, alpha-MSH and pancreatic polypeptide gave no immunoreaction in hagfish brain or pituitary.

Animals↗

Immunocytological localization of a somatostatin-like substance in the brain of the giant slug, Limax maximus L.

Immunocytological tests reveal the presence of a somatostatin-like substance in perikarya and axons in the brain of the giant slug Limax maximus L. Controls carried out on adjacent sections with absorbed antiserum or different antibodies raised against several biologically active peptides of vertebrates (ACTH-17-39, alpha- and beta endorphin, alpha- and beta MSH, methionin-enkephalin, TRH) demonstrate the specificity of the "staining". However, some cells are both somatostatin- and FMRF-amide-positive. In the cerebral ganglia, the right Z-area cells, responsible for the synthesis of the maturation hormone (MH) are strongly somatostatin-positive. These results suggest a similarity between the MH and the somatostatin-like material contained in the Z-area cells. The simultaneous presence of two peptides in one and the same cell, the nature (elementary granules or soluble product) of the material, and its site of release are discussed.

Animals↗

Immunocytochemical demonstration of peptidergic neurons in the central and peripheral nervous systems of the flatworm Microstomum lineare with antiserum to FMRF-amide.

The central nervous system (CNS) and the peripheral nervous system (PNS) of the flatworm Microstomum lineare were studied by means of the peroxidase-antiperoxidase (PAP) immunocytochemical method, with the use of antisera to the molluscan cardioactive peptide FMRF-amide. FMRF-amide immunoreactive perikarya and nerve fibres are observed in the CNS and the PNS. In the CNS, immunoreactive perikarya and nerve fibres occur in the brain, in the epithelial lining and the mesenchymal surroundings of the ciliated pits, and positive fibres in the longitudinal nerve cords. In the PNS, immunoreactive fibre bundles with variocosities occur in the pharyngeal nerve ring, in symmetrical groups of perikarya on each side of the pharynx, and in the mouth area. Positive perikarya and meandering nerve fibres appear in the intestinal wall. A few immunoreactive cells and short nerve processes are observed at the male copulatory organ and on both sides of the vagina. Some immunoreactive peptidergic cells do not correspond to cells previously identified by histological techniques for neurosecretory cells. The distribution of immunoreactivity suggests that the FMRF-amide-like substance in CNS and PNS in this worm has roles similar to those of the brain-gut peptides in vertebrates. The status of FMRF-amide-like peptides as representatives of an evolutionarily old family of peptides is confirmed by the positive immunoreaction to anti-FMRF-amide in this primitive microturbellarian.

Animals↗

Immunocytochemical demonstration of neuropeptides and serotonin in the tapeworm Diphyllobothrium dendriticum.

The present immunocytochemical study concerns the distribution of four neuropeptides, FMRF-amide, vasotocin, leu-enkephalin and neurotensin, and of the bioamine serotonin in the plerocercoid larva of Diphyllobothrium dendriticum. Anti-FMRF-amide and vasotocin-reactivity occurs in perikarya and nerve fibres in the CNS and PNS of this worm. The peptide-containing fibres surround and seem to innervate the musculature and to terminate beneath the basal lamina of the tegument at the inner surface of the bothridia, suggesting a neurotransmitter function. Anti-leu-enkephalin reaction occurs in perikarya and fibres in the main nerve cords and in the PNS. Anti-neurotensin reactive fibres were observed in the neuropile of the nerve cords. Serotonin immunoreactivity was found in neurons in the ganglionic commissure of the brain and along the main nerve cords. This study is the first immunocytochemical identification of neuropeptides and serotonin in a parasitic flatworm and the information gained may be of importance for the development of new antihelminthics.

Animals↗

Immunocytochemical localization of a substance in the eyestalk of the prawn, Palaemon serratus, reactive with an anti-FMRF-amide rabbit serum.

By use of a specific antiserum against the molluscan cardio-excitatory tetrapeptide FMRF-amide in combination with the PAP-method it was possible to obtain positive immunocytochemical reactions in several neurosecretory regions of the eyestalk of the prawn Palaemon serratus. FMRF-amide-like material was found in perikarya and nerve fibers of the medulla terminalis and in neurons in the lamina ganglionaris. The immunoreactivity observed in the glandular tissue located at the basal insertion of the eyestalk muscles must be ascribed to a non-specific reaction. The identification of immunopositive nerve fibers, ending on a nerve bundle in the medulla terminalis, and the fact that immunoreactive material was absent in the neurohemal sinus gland seem to indicate a neurotransmitter/neuromodulator function.

Animals↗

Stretch activation of the Aplysia S-channel.

The S-channel, a receptor-mediated K+ channel of Aplysia sensory neurons which functions in neuromodulation, bears a strong resemblance to the ubiquitous stretch-activated channels of snail neurons. Snail neuron stretch channels are stretch sensitive only in the patch, not at the macroscopic level, a situation which leaves open the question of their physiological role. If S-channels resemble snail stretch channels because both belong to the same general class of channels, the S-channel, too, should display stretch sensitivity in the patch. We show, using single-channel recording, that the S-channel can be activated by stretch. Furthermore, we show that Aplysia neurons in general have stretch-activated K+ channels. We suggest that the stretch-sensitive K+ channels of molluscan neurons and other preparations (e.g., Drosophila muscle, snail heart) are S-like channels, i.e., receptor-mediated channels which adventitiously exhibit mechanosensitivity in the patch.

Abdomen↗

Immunocytochemical identification of neural elements in the central nervous systems of a snail, some insects, a fish, and a mammal with an antiserum to the molluscan cardio-excitatory tetrapeptide FMRF-amide.

With an antiserum to the molluscan cardio-excitatory tetrapeptide FMRF-amide neurons and/or nerve fibers were immunocytochemically identified in the central nervous systems of a snail (Lymnaea stagnalis), some insects (Leptinotarsa decemlineata, Periplaneta americana, Locusta migratoria, Pieris brassicae), a fish (Poecilia latipinna) and a mammal (mouse). The fact that immunoreactive material was observed in neurohaemal organs (corpora cardiaca of the insects) as well as in axon terminals ending on other neurons, seems to indicate that this peptide can function as a neurohormone and/or as a neurotransmitter. The results sustain the hypothesis that biologically active peptides have a wide distribution in the animal kingdom.

Animals↗

Localisation of sulfakinin neuronal pathways in the blowfly Calliphora vomitoria.

The distribution of neurones immunoreactive to antisera raised against the undecapeptide C-terminal fragment of drosulfakinin II (DrmSKII), Asp-Gln-Phe-Asp-Asp-Tyr(SO3H)-Gly-His-Met-Arg-Phe-NH2, has been studied in the blowfly Calliphora vomitoria. Antisera were preabsorbed with combinations of the parent antigen, the tetrapeptide Phe-Met-Arg-Phe-NH2 and cholecystokinin, the vertebrate sulfated octapeptide (CCK-8), Asp-Tyr(SO3H)-Met-Gly-Trp-Met-Asp-Phe-NH2, in order to ensure specificity for the sulfakinin peptides of C. vomitoria (the nonapeptide callisulfakinin I is identical to drosulfakinin I and callisulfakinin II differs from DrmSK II only by the presence of -Glu3-Glu4- in place of -Asp3-Asp4-). Only four pairs of sulfakinin-immunoreactive neurons have been visualised in the entire nervous system. These occur in the brain: two pairs of cells situated medially in the caudo-dorsal region close to the roots of the ocellar nerve and two other pairs at the same level but positioned more laterally. Despite the small number of sulfakinin-immunoreactive cells, there are extensive projections to many areas of neuropile in the brain and the thoracic ganglion. The pathway of the medial sulfakinin cells extends into each of the three thoracic ganglia and a metameric arrangement of sulfakinin neuronal projections is also seen in the abdominal ganglia. Neither the dorsal neural sheath of the thoracic ganglion, nor the abdominal nerves contain sulfakinin-immunoreactive material. These observations suggest that the sulfakinins of the blowfly function as neurotransmitters or neuromodulators. They do not appear to have a direct role in gut physiology, as has been shown by in vitro bioassays for the sulfakinins of orthopterans and blattodeans. In addition to the neurones that display specific sulfakinin immunoreactivity, other cells within the brain and thoracic ganglion are immunoreactive to cholecystokinin/gastrin antisera. There are, therefore, at least two types of dipteran neuropeptides with amino acid sequences that are similar to the vertebrate molecules cholecystokinin and gastrin.

Amino Acid Sequence↗

FMRF-amide immunoreactivity in the mammalian gastroenteropancreatic neuroendocrine system.

The presence of FMRF-amide, a cardioactive tetrapeptide, was studied by immunocytochemistry in human and rat gastric antrum and pancreas, and in the ovine, bovine, canine and rabbit pancreas. In human and rat gastric antrum, numerous cells contained FMRF-amide immunoreactive material. By staining of serial sections and by double staining, colocalization of immunoreactivity for gastrin and FMRF-amide was observed in part of the gastrin cells. In the pancreas of these and the other species, immunoreactivity for FMRF-amide was located both in acinar and islet endocrine cells. Colocalization of FMRF-amide and pancreatic polypeptide was found in a proportion of pancreatic polypeptide cells in the pancreas. FMRF-amide immunoreactivity never colocalized with the other neurohormonal peptides which occur in the gastric antrum and the pancreas. Our observations show that neuroendocrine cells occur in the gastric antrum and pancreas which are exclusively immunoreactive or gastrin and for pancreatic polypeptide respectively. In addition cells occur which show immunoreactivity for FMRF-amide as well as for gastrin in the gastric antrum and with antiserum to FMRF-amide as well as for pancreatic polypeptide in the pancreas. It is concluded that FMRF-amide antibodies probably recognize a substance in G and PP cells which is not identical but may be structurally related to gastrin and pancreatic polypeptide.

Animals↗

Immunocytochemical demonstration of neuropeptides and serotonin in the nervous systems of adult Schistosoma mansoni.

In the nervous system of adult Schistosoma mansoni, neuropeptides and serotonin were demonstrated immunocytochemically. Neurons and nerve fibers immunoreactive to anti-FMRF-amide, anti-substance P, anti-leu-enkephalin, and anti-growth hormone releasing factor were found in the central and peripheral nervous system. The peptidergic nerve cell bodies in the bilobed brain are large and often multipolar; those in the peripheral nerve net are smaller and often bipolar. Sensory receptors of two morphologically different types were found along the surface of the worm. They are mainly immunoreactive to anti-substance P and anti-leu-enkephalin, and, to a lesser degree, to anti-FMRF-amide. Serotonin-immunoreactive fibers are found in the central and peripheral nervous elements as well as in sensory endings.

Animals↗

Development of immunoreactivity to the invertebrate neuropeptide small cardiac peptide B in the tapeworm Diphyllobothrium dendriticum.

Neurons immunoreactive to small cardiac peptide B (SCPb) occur in the scolex and neck region of adult Diphyllobothrium dendriticum. The localisation of the SCPb-IR neurons in the peripheral nervous system is very pronounced; they are closely associated to the bothridial musculature in the scolex. SCPb-IR neurons were not observed in plerocercoid larvae but appeared after cultivation in vitro at 37 degrees C for 30 h. Functional and developmental aspects of the SCPb-IR neurons are discussed.

Animals↗

Distribution of FMRF-amide-like peptide in the nervous system of a mermithid nematode, Romanomermis culicivorax.

The nervous system of the mermithid nematode Romanomermis culicivorax was investigated at different developmental stages for the distribution of the neuropeptide FMRF-amide. An FMRF-amide-like peptide was detected by an indirect immunofluorescence technique. The peptidergic nervous system increased in complexity during development. In adults, FMRF-amide-like peptide was present in the nerve ring, cephalic papillary ganglia, cephalic nerves, amphids, ganglia posterior to the nerve ring, longitudinal nerve cords and several mid-body region ganglia. The distribution of peptidergic ganglia in the tail region was different for each sex. Four clusters of ganglia were present in the tail of females, whereas such ganglia were absent in the tail of males. The results suggest that FMRF-amide-like peptide is widely distributed within the nervous system of R. culicivorax and might function as a neurohormone or neurotransmitter in controlling physiological and developmental processes.

Aedes↗

The neurochemical basis of backward inhibition in a defense reaction reflex arc.

An investigation was carried out on a preparation of the isolated nervous system of the edible snail of the mechanism of habituation in the network of defense behavior neurons. It was established that the intracellular activation of one of the systems of the defense behavior command neurons, as in the case of the application of the neuropeptide FMRF-amide, leads to a decrease in the amplitude of the postsynaptic potentials in the remaining neurons of this class. An inference is reached, when account is taken of the presence of endogenous FMRF-amide in the defense behavior command neurons, regarding the possibility of active inhibition of the behavioral response to an appropriate stimulus through this neurochemical mechanism of the backward connection. The possible interrelationships with other modulating systems and the significance of such an association in the organization of behavior are discussed.

Animals↗

Immunocytochemical localization of neurosecretory amines and peptides in the free-living nematode, Goodeyus ulmi.

Mammalian antibodies to the neuroamines, serotonin and gamma-amino-butyric acid (GABA) and to the neuropeptides, adrenocorticotrophic hormone (ACTH) and FMRF-amide evoked a response to Goodeyus ulmi, a free-living nematode. Serotonin-like immunoreactivity was found in cell bodies in the nerve ring and in the ventral nerve cord in all developmental stages. Neurons in the vulva, implicated in egg-laying, were immunoreactive to anti-serotonin in G. ulmi females, while in males serotonergic nerve fibres was found in the spicular region. Immunoreactivity to ACTH was also seen to differ depending on the developmental stage of G. ulmi, being present only in the ventral cord from the late L3 stage. Anti-GABA immunoreactivity was localized in two cell bodies near the amphids in all life stages and FMRF-amide immunoreactivity was seen in the nerve ring in all developmental stages. No reactivity was found with antibodies to vasointestinal peptide and somatostatin-14.

Adrenocorticotropic Hormone↗

A review of the electrophysiological, pharmacological and single channel properties of heart ventricle muscle cells in the snail Lymnaea stagnalis.

Although a considerable body of information has accumulated describing the pharmacological properties of a wide range of molluscan muscle types, the physiological bases underlying these properties have not been thoroughly investigated. At present, little is known about the types of ion channels and their regulation in molluscan muscle cell membranes. Voltage-clamp, and more recently, patch-clamp techniques have revealed molluscan muscles possess a complex array of channel types with various pharmacological and electrophysiological properties. The gating properties of these channels and their modulation by chemical agents, however, are still poorly understood. This review summarizes some aspects of molluscan muscle function with particular reference to the heart ventricle muscle of the pond snail, Lymnaea stagnalis.

Animals↗

Phe-met-arg-phe (FMRF)-amide is a substrate source of NO synthase in the gastropod nervous system.

The possible involvement of the L-arginine-containing Phe-met-arg-phe (FMRF)-amide (FMRFa) in neuronal nitric oxide (NO) biosynthesis was studied in a gastropod species. We found NADPH-diaphorase-positive neurons and FMRFa-containing fibers in close proximity in the enteric nervous system. Administration of L-arginine and FMRFa induced quantitatively similar nitrite production in both intact intestinal tissues and tissue homogenates. These changes could be prevented by the presence of NOARG (an NO synthase inhibitor). Neither chemically modified FMRFa (D-arginine instead of L-arginine) nor amino acid constituents of FMRFa (methionine, phenylalanine) affected basal nitrite production. FMRFa-induced alterations were reduced in the presence of Na+ channel blockers (tetrodotoxin, amiloride, lidocaine), the Na+/K+ATPase inhibitor ouabain, or protease inhibitors (leupeptine, pepstatine-a). FMRFa and its amino acid constituents were analyzed by paper chromatography. When FMRFa was added to tissue homogenates, the peptide was eliminated within 1-2 min, whereas methionine, phenylalanine, arginine, and citrulline levels were elevated simultaneously. We tested the effects of FMRFa, L-arginine, and NOARG on intestinal contractile activity. FMRFa relaxed the intestine for 1-2 min and then induced contractions for 20-40 min. In the presence of NOARG, no relaxant effect of FMRFa was recorded. As administration of L-arginine strongly inhibits the mechanical activity of the intestinal muscle, NO production presumably plays a substantial role in the action of FMRFa, at least in the initial phase. Our biochemical data indicate a direct involvement of FMRFa in NO biosynthesis. FMRFa might be hydrolyzed by extracellular peptidases and then the locally released arginine might be transported into the cells and broken-down to produce NO. Depolarization-induced NO production attributable to the activation of amiloride-sensitive Na+ channels might also be involved.

Amiloride↗