Peptidergic regulation of the Limulus midgut

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.     

Evidence for the association of FMRFamide-related peptides with the spermatheca of Locusta migratoria

The association of FMRFamide-related peptides (FaRPs) with the spermatheca of Locusta migratoria was demonstrated using radioimmunoassay and immunohistochemical techniques. The physiological effects of various FaRPs on the neurally evoked contractions of the spermatheca were also examined. FMRFamide-like immunoreactivity (FLI) was demonstrated in processes and cell bodies situated in the VIIIth (terminal) abdominal ganglion. These included an anterior, central and posterior pair of ventral cell bodies positioned near the midline of the ganglion, in addition to two bilaterally paired dorsal cell bodies in the posterior region of the VIIIth abdominal ganglion. Two axons displaying FLI proceed down the ventral ovipositor nerve (VON) and into the receptaculum seminis nerve which innervates the anterior regions of the spermatheca. FLI was also noted in processes on the spermathecal muscle with the highest density occurring on the spermathecal sac and coil duct. FaRPs applied to the spermathecal muscle included GQERNFLRFamide, NFIRFamide, ADDRNFIRFamide, YGGFMRFamide, FMRFamide, ADVGHVFLRFamide and SchistoFLRFamide (PDVDHVFLRFamide). Dose-dependent physiological effects were only noted for FMRFamide, ADVGHVFLRFamide and SchistoFLRFamide. FMRFamide led to a dose-dependent increase in the amplitude of neurally evoked contractions with a threshold of approximately 5 x 10(-7) M. SchistoFLRFamide, and ADVGHVFLRFamide, had an inhibitory effect, decreasing the amplitude of neurally evoked spermathecal contractions.     

Physiological and morphological identification of photosensitive neurons in the opisthosomal ganglia of Limulus polyphemus

The motor outputs of the isolated opisthosomal ventral nerve cord in Limulus polyphemus are modulated by light. We have identified the photosensitive neurons and examined their physiological and morphological properties using intracellular recording and staining techniques. We found that photosensitive neurons are present in each ganglion of the opisthosomal ventral nerve cord. These neurons often discharged action potentials spontaneously in the dark, and they increased the frequency of this discharge in the light. The mean latency (+/-SD) of the light-induced action potential was 2.2 +/- 1.1 s. Cells responded in a graded fashion over a 2-log unit of light intensity. The peak spectral sensitivity was 425 nm or lower. The Lucifer-yellow-labeled photosensitive neurons had oval somata with mean (+/-SD) diameters of 102 +/- 3 microm (long axis) and 75 +/- 5 microm (short axis), and extended their axons to the contralateral region of the ventral nerve cord. The soma had no dendrites, and the axon had thin branches.     

Distribution of FMRFamide-like immunoreactivity in the brain and suboesophageal ganglion of the sphinx mothManduca sexta and colocalization with SCPB-, BPP-, and GABA-like immunoreactivity

Summary Using an antiserum against the tetrapeptide FMRFamide, we have studied the distribution of FMRFamide-like substances in the brain and suboesophageal ganglion of the sphinx mothManduca sexta. More than 2000 neurons per hemisphere exhibit FMRFamide-like immunoreactivity. Most of these cells reside within the optic lobe. Particular types of FMRFamide-immunoreactive neurons can be identified. Among these are neurosecretory cells, putatively centrifugal neurons of the optic lobe, local interneurons of the antennal lobe, mushroom-body Kenyon cells, and small-field neurons of the central complex. In the suboesophageal ganglion, groups of ventral midline neurons exhibit FMRFamide-like immunoreactivity. Some of these cells have axons in the maxillary nerves and apparently give rise to FMRFamide-immunoreactive terminals in the sheath of the suboesophageal ganglion and the maxillary nerves. In local interneurons of the antennal lobe and a particular group of protocerebral neurons, FMRFamide-like immunoreactivity is colocalized with GABA-like immunoreactivity. This suggests that FMRFamide-like peptides may be cotransmitters of these putatively GABAergic interneurons. All FMRFamide-immunoreactive neurons are, furthermore, immunoreactive with an antiserum against bovine pancreatic polypeptide, and the vast majority is also immunoreactive with an antibody against the molluscan small cardioactive peptide SCP_B. Therefore, it is possible that more than one peptide is localized within many FMRFamide-immunoreactive neurons. The results suggest that FMRFamide-related peptides are widespread within the nervous system ofM. sexta and might function as neurohormones and neurotransmitters in a variety of neuronal cell types.Abbreviations AL antennal lobe - BPPLI bovine pancreatic polypeptide-like immunoreactivity - FLI FMRFamide-like immunoreactivity - GLI GABA-like immunoreactivity - NSC neurosecretory cell - SCP _B LI small cardioactive peptide_B-like immunoreactivity - SLI serotonin-like immunoreactivity - SOG suboesophageal ganglion     

An FMRFamide antiserum differentiates between populations of antigens in the ventral nervous system of the locust,Schistocerca gregaria

Summary The distribution of FMRFamide immunoreactive neurones in the ventral nerve cord of the locust, Schistocerca gregaria, is described. These neurones are found only in the suboesophagael and thoracic ganglia, although immunoreactive processes are found in the neuropils of the abdominal ganglia. Many of these neurones also react with an antiserum raised against bovine pancreatic polypeptide (BPP), but this antiserum also reveals another population of cells in the abdominal ganglia. The staining obtained with the BPP antiserum is blocked by preabsorption of the antiserum with FMRFamide; the converse is not true: FMRFamide-like immunoreactivity is not suppressed by preincubation with BPP. These results suggest that there are at least two endogenous peptide antigens in the locust nerve cord: one is found in cells of the suboesophageal and thoracic ganglia, and the other is found in cells of the abdominal ganglia.     

Detection of FMRFamide-like immunoreactivities in the sea scallopPlacopecten magellanicus by immunohistochemistry and Western blot analysis

FMRFamide-like immunoreactivity was detected histochemically in the sea scallopPlacopecten magellanicus. Most immunoreactivity was concentrated in the cerebral, pedal, and parietovisceral ganglia, particularly in the cortical cell bodies and in their fibers which extend into the central neuropile. Whole-mount immunofluorescence studies were used to localize concentrations of immunoreactive cells on the dorsal and ventral surfaces of each ganglion. Immunoreactivity was also detected in nerves emanating from the ganglia. Strong immunoreactivity was localized in peripheral organs, including the gut and gills of juvenile and adult scallops. Weak immunoreactivity was detected in the gonads, heart, and adductor muscle of the adults. A broad FMRFamide-like immunoreactive band of 2.5–8.2 kDa was detected by Western blotting of acetone extracts of the parietovisceral ganglia. In the presence of protease inhibitors, two FMRFamide-like immunoreactive bands (7.2–8.2 kDa and >17 kDa) were obtained. Neither of these bands comigrated with the FMRFamide standard. It is concluded that peptides of the FMRFamide family are probably regulators of numerous central and peripheral functions inP. magellanicus.     

Characterization and distribution of FMRFamide immunoreactivity in the rat central nervous system

FMRFamide immunoreactive material (irFMRFamide) was studied in rat brain and gastrointestinal tract. Highest irFMRFamide concentrations were found in tissues of the gastrointestinal tract and, in the brain, highest concentrations were found in the hippocampus, midbrain, brainstem and hypothalamus. High pressure liquid chromatographic characterization of irFMRFamide demonstrated that the immunoreactive material in brain, pancreas and duodenum was different from molluscan FMRFamide but it was also distinct from any known neuropeptide.     

An identified dorsal unpaired median neurone and bilaterally projecting neurones exhibiting bovine pancreactic polypeptide-like/FMRFamide-like immunoreactivity in abdominal ganglia of the migratory locust

Summary Three antisera were used to study the distribution and anatomy of bovine pancreatic polypeptide (BPP)-like/FMRFamide-like immunoreactive neurones within the unfused abdominal ganglia of the migratory locust, Locusta migratoria. All the antisera used stained two or more clusters of perikarya, localized anteriorly and posteriorly near the midline within each unfused abdominal ganglion. Double labelling experiments with intracellular dye injection, or differential backfilling, combined with subsequent immunostaining were carried out to identify these neurones. Two of the antisera (antisera 1 and 2, both raised against FMRFamide) stained three groups of midline neurones, located anterior dorsal, anterior ventral and posterior dorsal within the ganglion. Neurones of the former of these two clusters projected via the anterior median nerve to a neurohaemal organ. The posterior cluster of midline cells comprised immunopositive perikarya all but one of which also projected via the anterior median nerve to innervate the neurohaemal organ. Double labelling with Lucifer yellow and antisera 1 and 2 showed that the remaining neurone was the previously identified doral unpaired median (DUM)heart1 neurone. The third antiserum (AK141), also raised against FMRFamide, stained neurones within an anterior dorsal cluster, and in a posterior cluster. Double labelling with differential Co²⁺/Ni²⁺-backfilling and the antiserum 3 (AK141) demonstrated that the large neurones of both clusters belonged to the population of bilaterally projecting neurones (BPNs), including the DUMheart1 neurone. Since the antisera cross-react with BPP and fail to label neurones when preadsorped with BPP or FMRFamide, we conclude that the labelled neurones contain polypeptides of the FMRFamide/BPP-family.     

Peripheral nerve connections influence the appearance of neurosecretary material in neural sheath of ventral ganglion of the fly Sarcophaga bullata: an immunocytochemical study

This study examined the role of the brain and peripheral connections with the target organs in the appearance of neurosecretary material within the dorsal neural sheath of the ventral ganglion of the fly S. bullata. Specifically, the accumulation of the neuropeptide FMRFamide (the neurosecretary material) was examined by immunocytochemistry. Immunoreactions were performed on: (1) a normal intact ventral ganglion, (2) an isolated ventral ganglion that was cultured in vivo, and (3) a ventral ganglion that was isolated by transection from the brain, but retained its peripheral nerve connections. The results demonstrate that (a) the neurons of the ganglia survive and exhibit FMRFamide immune reaction independent of their peripheral connections, and (b) the accumulation of neuropeptide in the dorsal neural sheath is controlled by intact peripheral nerve connections with the ganglion. It is suggested that in the absence of their peripheral connections, the axons of FMRFamide immunoreactive neurons fail to invade the neural sheath resulting in the accumulation of neurosecretary material.     

FMRFamide-like immunoreactivity in the nervous system of hydra

Summary FMRFamide-like immunoreactivity has been localized in different parts of the hydra nervous system. Immunoreactivity occurs in nerve perikarya and processes in the ectoderm of the lower peduncle region near the basal disk, in the ectoderm of the hypostome and in the ectoderm of the tentacles. The immunoreactive nerve perikarya in the lower peduncle region form ganglion-like structures. Radioimmunoassays of extracts of hydra gave displacement curves parallel to standard FMRFamide and values of at least 8 pmol/gram wet weight of FMRFamide-like immunoreactivity. The immunoreactive material eluted from Sephadex G-50 in several components emerging shortly before or after position of authentic FMRFamide. The presence of FMRFamide-like material in coelenterates shows that this family of peptides is of great antiquity.     

Action and immunoreactivity of neuropeptides in the buccal neuromuscular system of a prosobranch mollusc,Rapana thomasiana

Summary In a prosobranch mollusc, Rapana thomasiana, the catch-relaxing peptide H-Ala-Met-Pro-Met-Leu-Arg-Leu-NH₂ (CARP) was found to depress the contraction of the radula protractor and retractor elicited by electrical stimulations. The action of CARP was in contrast to that of other neuropeptides, H-Phe-Met-Arg-Phe-NH₂ (FMRFamide) and H-Phe-Leu-Arg-Phe-NH₂ (FLRFamide), which enhanced the contraction of the radula protractor and retractor, respectively. By immunohistochemical examinations, FMRFamide-like immunoreactive neurons were found on the rostral side of the right buccal ganglion and the caudal side of the left ganglion, where some CARP-like immunoreactive neurons were also distributed, indicating a possible coexistence of FMRFamide and CARP. FMRFamide- and CARP-like immunoreactivities were also detected in the neuropile of buccal ganglia, radula nerves arising from the ganglia, and nerve fibers in the radula muscles. The present results suggest that FMRFamide- and CARP-like peptides are involved in the regulation of the contraction of the radula muscles.     

Distribution of FMRFamide-like immunoreactivity in the brain and neurohypophysis of the lamprey,Lampetra japonica

Summary Distribution of molluscan cardio-excitatory tetrapeptide Phe—Met—Arg—Phe—NH₂ (FMRFamide) was determined by means of immunohistochemistry in the brain and neurohypophysis of the lamprey, Lampetra japonica. Many FMRFamide-like immunoreactive neurons were found in the periventricular nuclear region and in a region near the mammillary recess. Neurons situated in the former region were larger. The immunoreactive cell groups were shown to be located at sites differing from those of the AF-positive cell groups. The fibers of immunoreactive neurons extended in all directions within the brain and towards the spinal cord, some reaching the third ventricle and capillaries. Thus, FMRFamide-like immunoreactive peptides appear to function as neurotransmitters or neuromodulators and possibly also as neurohormones. FMRFamide-like immunoreactive material was rarely observed in the posterior neurohypophysis (neural lobe), but was noted to be present to a limited extent in the caudal part of the anterior neurohypophysis (median eminence). It would thus follow that FMRFamide-like immunoreactive neurons may not necessarily be related to the hypothalamo-neural lobe system, but may possibly be associated with the hypothalamoadenohypophysial system. The pineal body showed no FMRFamide-like immunoreactivity.     

The Larval Nervous System of Polygordius lacteus Scheinder, 1868 (Polygordiidae,Polychaeta): Immunocytochemical Data

The nervous system of the planktotrophic trochophore larva of Polygordius lacteus has been investigated using antibodies to serotonin (5-HT) and the neuropeptide FMRFamide. The apical ganglion contains three 5-HT-ir neurons, many FMRFamide-ir neurons and a tripartate 5-HT-ir and FMRFamide-ir neuropil. A lateral nerve extends from each side of the apical ganglion across the episphere and the ventral hyposphere, where the two nerves combine to form the paired ventral nerve cord. These nerves have both 5-HT-ir and FMRFamide-ir processes. Three circumferential nerves are associated with the ciliary bands: two prototroch and one metatroch nerve. All contain 5-HT-ir and FMRFamide-ir processes. An oral nerve plexus also contain both 5-HT-ir and FMRFamide-ir processes develops from the metatroch nerve, and an esophageal ring of FMRFamide-ir processes develops in later larval stages. In young stages the ventral ganglion contains two 5-HT-ir and two FMRFamide-ir perikarya; during development the ventral ganglion grows caudally and adds additional 5-HR-ir and FMRFamide-ir perikarya. These are the only perikarya that could be found along the lateral nerve and ventral nerve cord. The telotroch nerve develops from the ventral nerve cord. The 5-HT-ir and FMRFamide-ir part of the nervous system is strictly bilateral symmetric. and much of the system (i.e. apical ganglion, lateral nerves ventral nerve cord, dorsal nerve and oral plexus) is retained in the adult.     

FMRfamide-like immunoreactivity in the ventral nerve cord of the larval eastern spruce budworm,Choristoneura fumiferana (clemens) (Lepidoptera : Tortricidae)

Distribution of FMRFamide-like immunoreactivity was examined in the larval ventral nerve cord of the eastern spruce budworm, Choristoneura fumiferana (Lepidoptera : Tortricidae). Indirect immunofluorescent methods revealed the existence of 3 groups of FLI neurons in each ganglion. The neurons are distributed in a bilaterally symmetrical fashion at the anterodorsal, midlateral and posteroventral regions of the ganglia. There are 4 FMRFamide-like immunoreactive fiber tracts on the dorsal surface of the ganglia to which the anterodorsal FLI neurons project ipsilaterally, while the midlateral pair projects both ipsi-, and contralaterally. The last abdominal ganglion (AG8) has 4 additional pairs of FLI neurons; and axons from some of these extend into the median abdominal nerve, which suggests some role for this neuropeptide in the control of posterior structures of the larva.     

Colocalization of neuropeptide Y (NPY)-like and FMRFamide-like immunoreactivities in the brain of the Atlantic salmon (Salmo salar)

Summary The colocalization of the peptides neuropeptide Y (NPY) and Phe-Met-Arg-Phe-NH₂ (FMRFamide) in the brain of the Atlantic salmon was investigated with double immunofluorescence labeling and peroxidase-antiperoxidase immunocytochemical techniques. Colocalization of NPY-like and FMRE amide-like immunoreactivities was observed in neuronal cell bodies and fibers in four brain regions: in the lateral and commissural nuclei of the area ventralis telencephali, in the nucleus ventromedialis thalami, in the laminar nucleus of the mesencephalic tegmentum, and in a group of small neurons situated among the large catecholaminergic neurons in the isthmal region of the brainstem. All cell bodies in these nuclei were immunoreactive to both NPY and FMRF. We consistently observed larger numbers of FMRF-immunoreactive than NPY-immunoreactive fibers. In the nucleus ventromedialis thalami NPY- and FMRFamide-like immunoreactivities were colocalized in cerebrospinal fluid (CSF)-contacting neurons. NPY-immunoreactive, but not FMRF-immunoreactive, neurons were found in the stratum periventriculare of the optic tectum, and at the ventral border of the nucleus habenularis (adjacent to the nucleus dorsolateralis thalami). Neurons belonging to the nucleus of the nervus terminalis were FMRF-immunoreactive but not NPY-immunoreactive. The differential labeling indicates, as do our cross-absorption experiments, that the NPY and FMRFamide antisera recognize different epitopes. Thus, it is probable that NPY-like and FMRFamide-like substances occur in the same neurons in some brain regions.     

The distribution of pheromone-biosynthesis-activating neuropeptide (PBAN) immunoreactivity in the central nervous system of the corn earworm moth,Helicoverpa zea

Summary Production of sex pheromone in several species of moths has been shown to be under the control of a neuropeptide termed pheromone-biosynthesis-activating neuropeptide (PBAN). We have produced an antiserum to PBAN from Helicoverpa zea (Lepidoptera: Noctuidae) and used it to investigate the distribution of immunoreactive peptide in the brain-suboesophageal ganglion complex and its associated neurohemal structures, and the segmental ganglia of the ventral nerve cord. Immunocytochemical methods reveal three clusters of cells along the ventral midline in the suboesophageal ganglion (SOG), one cluster each in the presumptive mandibular (4 cells), maxillary (12–14 cells), and labial neuromeres (4 cells). The proximal neurites of these cells are similar in their dorsal and lateral patterns of projection, indicating a serial homology among the three clusters. Members of the mandibular and maxillary clusters have axons projecting into the maxillary nerve, while two additional pairs of axons from the maxillary cluster project into the ventral nerve cord. Members of the labial cluster project to the retrocerebral complex (corpora cardiaca and cephalic aorta) via the nervus corpus cardiaci III (NCC III). The axons projecting into the ventral nerve cord appear to arborize principally in the dorsolateral region of each segmental ganglion; the terminal abdominal ganglion is distinct in containing an additional ventromedial arborization in the posterior third of the ganglion. Quantification of the extractable immunoreactive peptide in the retrocerebral complex by ELISA indicates that PBAN is gradually depleted during the scotophase, then restored to maximal levels in the photophase. Taken together, our findings provide anatomical evidence for both neurohormonal release of PBAN as well as axonal transport via the ventral nerve cord to release sites within the segmental ganglia.Abbreviations A aorta - Br-SOG brain-suboesophageal ganglion complex - CC corpus cardiacum - PBS phosphate-buffered saline - PLI PBAN-like immunoreactivity - TAG terminal abdominal ganglion - VNC ventral nerve cord     

Immunohistochemical investigation of neuropeptides in the central nervous system of the amphioxus,Branchiostoma belcheri

The immunohistochemical localization of nine different neuropeptides was studied in the central nervous system of the amphioxus, Branchiostoma belcheri. In the brain, perikarya immunoreactive for urotensin I and FMRFamide were localized in the vicinity of the central canal. One of the processes of each of these perikarya was found to cross the dorso ventral slit-like lumen of the central canal. Oxytocin-immunoreactive short fibers, but not perikarya, were detected in the ventral part of the brain. Perikarya immunoreactive for arginine vasopressin/vasotocin, oxytocin and FMRFamide were widely distributed in the spinal cord. Arginine vasopressin/vasotocin-immunoreactive fibers often made contacts with Rohde cell axons. Angiotensin II-immunoreactive perikarya were observed in the posterior half of the spinal cord, and urotensin I-immunoreactive perikarya were found in the caudal region of the spinal cord. Cholecystokinin/gastrin-immunoreactive fibers, but not perikarya, were detected in the spinal cord; some extended as far as the ependymal layer of the cerebral ventricle. No colocalization of the peptides examined was observed. No immunoreactivity for atrial and brain natriuretic peptides nor for urotensin II was detected. The present study indicates that there are at least six separate neuronal systems that contain different peptides, respectively, in the central nervous system of the amphioxus. Their functions remain to be determined.Part of this investigation has previously been presented in abstract form (Uemura et al. 1989)     

Presence and distribution of FMRFamide-like immunoreactivity in the sea cucumber Holothuria scabra (Jaeger, 1833)

Since the discovery of FMRFamide, a cardioactive neuropeptide in the mollusk Macrocallista nimbosa (Lightfoot, 1786) by Price and Greenberg (Science 197:670–671, 1977), there has been a steady increase in our understanding of the distribution and diversity of FMRFamide and FMRFamide-related peptides (FaRPs) across phyla. The sea cucumber Holothuria scabra (Jaeger, 1833) (Echinodermata: Holothuroidea) is a high-premium tropical economic species that is overexploited globally and may be in imminent danger of extinction in some areas. Conservation of this species and its amenability to culture is however hampered by the limited knowledge about its biology, including the neurohormonal system. Using indirect immunofluorescence technique with antisera raised against the tetrapeptide FMRFamide as well as reverse-phase HPLC, we have demonstrated the widespread distribution of FMRFamide-like immunoreactivity in the tissues of H. scabra, including the coelomic fluid, suggesting that FaRPs might have neurohormonal activities in this species.     

Stimulation of brain Na+ channels by FMRFamide in Dahl SS and SR rats

Stimulation of brain Na+ channels by Phe-Met-Arg-Phe-NH2 (FMRFamide) increases sympathetic nerve activity and blood pressure (BP) in Wistar rats. Blockade of brain ouabain-like compounds (OLC) by specific antibody Fab fragments prevents these responses to intracerebroventricular FMRFamide. In the present study, we evaluated the effects of high-salt intake on brain FMRFamide levels and the responses of BP and brain OLC to intracerebroventricular infusion of FMRFamide in Dahl salt-sensitive (SS) and salt-resistant (SR) rats. FMRFamide and OLC content was measured with the use of RIA and ELISA, respectively. A high-salt diet (1,370 micromol Na+/g) for 2 wk significantly increased BP in Dahl SS but not in SR rats. On a regular salt diet, Dahl SS and SR rats showed similar FMRFamide levels in the whole hypothalamus, pons and medulla, and spinal cord. A high-salt diet for 2 wk did not affect FMRFamide levels in these tissues in both Dahl SS and SR rats. In Dahl SS but not in SR rats, chronic intracerebroventricular infusion of FMRFamide (200 nmol. kg(-1).day(-1)) for 2 wk significantly increased BP (mean arterial pressure: 116 +/- 5 vs. 100 +/- 2 mmHg; P < 0.01). Chronic intracerebroventricular infusion of FMRFamide significantly increased hypothalamic and pituitary OLC in Dahl SS but not SR rats. These results indicate that Dahl SS rats exhibit enhanced central responses to FMRFamide. In Dahl SS but not in SR rats on a high-salt diet, enhanced Na+ entry through FMRFamide-activated brain Na+ channels may increase brain OLC release, thereby leading to hypertension.     

Putative neurohemal areas in the peripheral nervous system of an insect,Gryllus bimaculatus,revealed by immunocytochemistry

The morphology and position of putative neurohemal areas in the peripheral nervous system (ventral nerve cord and retrocerebral complex) of the cricket Gryllus bimaculatus are described. By using antisera to the amines dopamine, histamine, octopamine, and serotonin, and the neuropeptides crustacean cardioactive peptide, FMRFamide, leucokinin 1, and proctolin, an extensive system of varicose fibers has been detected throughout the nerves of all neuromeres, except for nerve 2 of the prothoracic ganglion. Immunoreactive varicose fibers occur mainly in a superficial position at the neurilemma, indicating neurosecretory storage and release of neuroactive compounds. The varicose fibers are projections from central or peripheral neurons that may extend over more than one segment. The peripheral fiber varicosities show segment-specific arrangements for each of the substances investigated. Immunoreactivity to histamine and octopamine is mainly found in the nerves of abdominal segments, whereas serotonin immunoreactivity is concentrated in subesophageal and terminal ganglion nerves. Immunoreactivity to FMRFamide and crustacean cardioactive peptide is widespread throughout all segments. Structures immunoreactive to leucokinin 1 are present in abdominal nerves, and proctolin immunostaining is found in the terminal ganglion and thoracic nerves. Codistribution of peripheral varicose fiber plexuses is regularly seen for amines and peptides, whereas the colocalization of substances in neurons has not been detected for any of the neuroactive compounds investigated. The varicose fiber system is regarded as complementary to the classical neurohemal organs.