Distribution and characterization of neuropeptide Y-like immunoreactivity in the brain and pituitary of the goldfish

Summary The distribution of neuropeptide Y (NPY) immunoreactivity has been studied by means of immunocytochemistry and radioimmunoassay in the brain of the goldfish. It was found that NPY had a widespread distribution in the entire brain in particular in the telencephalon, diencephalon, optic tectum and rhombencephalon. In the pituitary gland, positive type-B fibers were observed in the various lobes frequently in direct contact with secretory cells, in particular the gonadotrophs, somatotrophs and MSH (melanocyte-stimulating hormone) secreting cells. When measured by radioimmunoassay, the highest NPY concentrations were found in the pituitary and telencephalon, confirming the results of immunocytochemistry. The displacement curves obtained with serial dilutions of brain extracts were parallel to that of synthetic porcine NPY. Following high performance liquid chromatography, the NPY-like material extracted from goldfish brain co-eluted as a single peak with synthetic porcine NPY. These data demonstrate the presence of an NPY-like substance widely distributed in the goldfish brain. The observation of NPY-immunoreactive fibers in the pituitary gland suggests that, among its other functions, NPY may play a role in the neuroendocrine regulation of pituitary function.     

Serotonin immunoreactive neurons in the central nervous system of an insect (Periplaneta americana)

Serotonin-like immunoreactivity was mapped in the central nervous system (CNS) of the cockroach, Periplaneta americana. Immunoreactive staining occurred in every ganglion of the CNS. The largest numbers of immunoreactive somata were detected in the optic lobes and the brain, and lowest numbers in the first and second thoracic ganglia. Dense stained fibers, ramifications, and varicosities were found in all ganglia, and numerous axon like processes occurred in all interganglionic connectives. Immunoreactive processes were not, however, detected in most of the peripherally projecting nerve roots. Processes were found only in roots of the suboesophageal ganglion and the tritocerebral lobes of the brain. A comparison of the map for serotonin immunoreactivity with one generated for the pentapeptide transmitter proctolin suggests that the two systems overlap only in the suboesophageal ganglion and the tritocerebrum. The amine and peptide may co-occur in neurons in these regions. The serotonin immunoreactive system appeared significantly different from the octopaminergic system of the ventral nerve cord. Seventy-two potentially identifiable immunoreactive cells were located in the cockroach CNS. Some of these may be suitable for physiological study of the functional role of serotonin.     

Organization of neuropeptide tyrosine-like immunoreactive system in the brain of the Antarctic fish, Trematomus bernacchii

Antarctic notothenioids have developed unique freezing-resistance adaptations, including brain diversification, to survive in the subzero waters of the Southern Ocean surrounding Antarctica. In this study we have investigated the anatomical distribution of neuropeptide tyrosine (NPY)-like immunoreactive elements in the brain of the Antarctic fish Trematomus bernacchii, by using an antiserum raised against porcine NPY. Perikarya exhibiting NPY-like immunoreactivity were observed in distinct regions of the brain. The most rostral group of immunoreactive perikarya was found in the telencephalon, within the entopeduncular nucleus. In the diencephalon, three groups of NPY-like immunoreactive perikarya were found in the hypothalamus. Two groups of positive cell bodies were found in distinct populations of the preoptic nucleus, whereas the other group was found in the nucleus of the lateral recess. More caudally, NPY immunoreactivity was detected in large neurons located in the subependymal layers of the dorsal tegmentum of the mesencephalon, medially to the torus semicircularis. NPY-like immunoreactive nerve fibres were more widely distributed throughout the telencephalon to the rhombencephalon. High densities of nerve fibres and terminals were observed in several regions of the telencephalon, olfactory bulbs, hypothalamus, tectum of the mesencephalon and in the ventral tegmentum of the rhombencephalon. The distribution of NPY-like immunoreactive structures suggests that, in Trematomus, this peptide may be involved in the control of several brain functions, including olfactory activity, feeding behaviour, and somatosensory and visual information. In comparison with other neuropeptides previously described in the brain of Antarctic fish, NPY is more widely distributed. Our data also indicate the existence of differences in the brain distribution of NPY between Trematomus and other teleosts. In contrast with previous results reported in other fish, Trematomus contains positive fibres in the olfactory bulbs and immunoreactive perikarya in the nucleus of the lateral recess, whereas NPY-immunopositive cell bodies are absent in the thalamus and rhombencephalon, and no NPY immunoreactivity is present in the pituitary. These differences could be related to the Antarctic ecological diversity of notothenioids living at subzero temperatures.     

Localization and identification of Neuropeptide Y (NPY)-like immunoreactivity in the frog brain

The distribution of neuropeptide Y (NPY) in the central nervous system of the frog Rana ridibunda was determined by immunofluorescence using a highly specific antiserum. NPY-like containing perikarya were localized in the infundibulum, mainly in the ventral and dorsal nuclei of the infundibulum, in the preoptic nucleus, in the posterocentral nucleus of the thalamus, in the anteroventral nucleus of the mesencephalic tegmentum, in the part posterior to the torus semicircularis, and in the mesencephalic cerebellar nucleus. Numerous perikarya were also distributed in all cerebral cortex. Important tracts of immunoreactive fibers were found in the infundibulum, in the preoptic area, in the lateral amygdala, in the habenular region, and in the tectum. The cerebral cortex was also densely innervated by NPY-like immunoreactive fibers. A rich network of fibers was observed in the median eminence coursing towards the pituitary stalk. Scattered fibers were found in all other parts of the brain except in the cerebellum, the nucleus isthmi and the torus semicircularis, where no immunoreactivity could be detected. NPY-immunoreactive fibers were observed at all levels of the spinal cord, with particularly distinct plexus around the ependymal canal and in the distal region of the dorsal horn. At the electron microscope level, NPY containing perikarya and fibers were visualized in the ventral nuclei of the infundibulum, using the peroxidase-antiperoxidase and the immunogold techniques. NPY-like material was stored in dense core vesicles of 100 nm in diameter. A sensitive and specific radioimmunoassay was developed. The detection limit of the assay was 20 fmole/tube. The standard curves of synthetic NPY and the dilution curves for acetic acid extracts of cerebral cortex, infundibulum, preoptic region, and mesencephalon plus thalamus were strictly parallel. The NPY concentrations measured in these regions were (pmole/mg proteins) 163±8, 233±16, 151±12 and 60±13, respectively. NPY was not detectable in cerebellar extracts. After Sephadex G-50 gel filtration of acetic acid extracts from whole frog brain, NPY-like immunoreactivity eluted in a single peak. Reverse phase high performance liquid chromatography (HPLC) and radioimmunoassay were used to characterize NPY-like peptides in the frog brain. HPLC analysis revealed that infundibulum, preoptic area and telencephalon extracts contained a major peptide bearing NPY-like immunoreactivity. The retention times of frog NPY and synthetic porcine NPY were markedly different. HPLC analysis revealed also the existence, in brain extracts, of several other minor components cross-reacting with NPY antibodies. These results provide the first evidence for the presence of NPY in the brain of a non-mammalian chordate and indicate that the structure of NPY is preserved among the vertebrate phylum. The abundance of NPY producing neurons in the hypothalamus and telencephalon suggests that this peptide may play both neuroendocrine and neurotransmitter functions in amphibians.     

An active principle from the corpora cardiaca,corpora allata and suboesophageal ganglion of the locust,inducing egg diapause in Bombyx mori

The activity of the substance(s) which are contained in the cephalic endocrine organs of the locust which induce egg diapause in Bombyx mori was examined by implantation and injection of saline extracts of these organs. Extracts from the median and lateral neurosecretory parts of the locust brain were not effective in inducing egg diapause. Extracts of the corpora cardiaca, corpora allata, and suboesophageal ganglion of the locust induced diapause eggs in Bombyx pharate adults from which the suboesophageal ganglion had been removed. The first two extracts could induce egg diapause even in isolated abdomens of pharate adults of Bombyx. In the locust corpora cardiaca, the activity was present only in the glandular lobe and not in the nervous region. This activity decreased when the nervi corporis cardiaci I and II and of nervi corporis allati I were cut. Allatectomy also brought about a decrease in the activity in the glandular lobe which could not be restored by the injection of juvenile hormone. The activity in the corpora allata was enhanced slightly by the disconnection though not significantly.From these results, it is assumed that the corpora cardiaca, corpora allata and suboesophageal ganglion of the locust contain and active principle(s) capable of inducing egg diapause in Bombyx mori. The nervous connections between the brain, corpora cardiaca, and corpora allata are essential for the accumulation of the active substance(s) in the glandular lobes of the corpora cardiaca.     

Distribution of SchistoFLRFamide-like immunoreactivity in the adult ventral nervous system of the locust,Schistocerca gregaria

SchistoFLRFamide (PDVDHVFLRF-NH₂) is one of the major endogenous neuropeptides of the FMRF-amide family found in the nervous system of the locust,Schistocerca gregaria. To gain insights into the potential physiological roles of this neuropeptide we have examined the distribution of SchistoFLRFamide-like immunoreactivity in the ventral nervous system of adult locusts by use of a newly developed N-terminally specific antibody. SchistoFLRFamide-like immunoreactivity in the ventral nerve cord is found in a subgroup of the neurones that are immunoreactive to an antiserum raised against bovine pancreatic polypeptide (BPP). In the suboesophageal ganglion three groups of cells stain, including one pair of large posterior ventral cells. These cells are the same size, in the same location in the ganglion and have the same branching pattern as a pair of BPP immunoreactive cells known to innervate the heart and retrocerebral glandular complex of the locust. In the thoracic and abdominal ganglia two and three sets of cells, respectively, stain with both the SchistoFLRFamide and BPP antisera. In the abdominal ganglia the immunoreactive cells project via the median nerves to the intensely immunoreactive neurohaemal organs.     

The vasopressin-like immunoreactive (VPLI) neurons of the locust,Locusta migratoria. I. Anatomy

Summary Antiserum to arginine-vasopressin has been used to characterise the pair of vasopressin-like immunoreactive (VPLI) neurons in the locust. These neurons have cell bodies in the suboesophageal ganglion, each with a bifurcating dorsal lateral axon which gives rise to predominantly dorsal neuropilar branching in every ganglion of the ventral nerve cord. There are extensive beaded fibre plexuses in most peripheral nerves of thoracic and abdominal ganglia, but in the brain, the peripheral plexuses are reduced while neuropilar branching is more extensive, although it generally remains superficial. An array of fibres runs centripetally through the laminamedulla chiasma in the optic lobes. Lucifer Yellow or cobalt intracellular staining of single VPLI cells in the adult suboesophageal ganglion shows that all immunoreactive processes emanate from these two neurons, but an additional midline arborisation (that was only partially revealed by immunostaining) was also observed. Intracellularly staining VPLI cells in smaller larval instars, which permits dye to reach the thoracic ganglia, confirms that there is no similar region of poorly-immunoreactive midline arborisation in these ganglia. It has been previously suggested that the immunoreactive superficial fibres and peripheral plexuses in ventral cord ganglia serve a neurohaemal function, releasing the locust vasopressin-like diuretic hormone, F₂. We suggest that the other major region of VPLI arborisation, the poorly immunoreactive midline fibres in the suboesophageal ganglion, could be a region where VPLI cells receive synaptic input. The function of the centripetal array of fibres within the optic lobe is still unclear.Abbreviations AVP arginine vasopressin - DIT dorsal intermediate tract - FLRF Phe-Leu-Arg-Phe - FMRF-amide Phe-Met-Arg-Phe-amide - LDT lateral dorsal tract - LVP lysine vasopressin - MDT median dorsal tract - MVT median ventral tract - SEM scanning electron microscopy - SOG suboesophageal ganglion - VIT ventral intermediate tract - VNC ventral nerve cord - VPLI vasopressin-like immunoreactive     

Neuropeptide Y (NPY)-like immunoreactive amacrine cells in retinas of frog and goldfish

Summary The distribution of neuropeptide Y (NPY)-like immunoreactivity in rat, rabbit, chick, frog and goldfish retinas was investigated by immunohistochemistry. Positive results were observed only in the frog and goldfish retinas. NPY immunoreactivity was associated with a small population of amacrine cell bodies in the inner nuclear layer and cell processes in the inner plexiform layer of both retinas. In the frog retina, three distinct layers containing immunoreactivity were observed in the inner plexiform layer. In contrast, the immunoreactivity in the same area of the goldfish retina was more or less separated into two layers. Convincing evidence could not be found for the co-existence of NPY-like material with other putative transmitter-like substances in the two retinas.Radioimmunoassay revealed the presence of small amounts of NPY-like immunoreactivity in the rabbit retina; the goldfish and frog retinas contained significantly more immunoreactive material. High performance liquid chromatography of the immunoreactive material in frog and goldfish retinas showed each retina containing different molecular forms of NPY-like proteins, neither of which resembled porcine NPY or PYY.The endogenous NPY-like material of the frog retina can be released by potassium depolarisation in a calciumdependent way. In view of all these data an NPY-like protein must now be considered a potential retinal transmitter.     

Neuropeptide Y in the intermediate lobe of the frog pituitary acts as an alpha-MSH-release inhibiting factor

The presence of neuropeptide tyrosine (NPY) in the intermediate lobe of the frog pituitary was demonstrated using indirect immunofluorescence, the immunogold technique and a specific radioimmunoassay combined with high pressure liquid chromatography (HPLC). A high density of NPY-containing fibers, was found among the parenchymal cells of the intermediate lobe. These fibers originated from the ventral infundibular nucleus, travelled via the median eminence to the pars intermedia. At the electron microscopic level, NPY-like material was found exclusively in nerve fibers where the product of the immunoreaction was associated to dense-core vesicles. High concentrations of NPY-like peptide were found in neurointermediate lobe extracts. After Sephadex G-50 gel filtration the major peak of immunoreactive material appeared to co-elute with synthetic porcine NPY. Conversely, HPLC analysis revealed that the NPY-like peptide of the frog pituitary had a retention time shorter than the porcine NPY. The localization of NPY-like material in the pars intermedia suggested a possible role of NPY in the regulation of melanotropic cell secretion. In fact, graded concentrations of synthetic NPY induced a dose-dependent inhibition of alpha-melanotropin (alpha-MSH) release in vitro. The lack of effect of a dopaminergic antagonist on NPY-induced alpha-MSH release inhibition demonstrated that the local dopaminergic system could not account for the NPY action. These results indicate that NPY located in the hypothalamo-hypophyseal system of the frog may act as a melanotropin-release inhibiting factor.     

Serotonin-immunoreactive neurons in the antennal lobes and suboesophageal ganglion of the honeybee

Summary We have used immunohistochemical methods to investigate the morphology of identified, presumptive serotonergic neurons in the antennal lobes and suboesophageal ganglion of the worker honeybee. A large interneuron (deutocerebral giant, DCG) is described that interconnects the deutocerebral antennal and dorsal lobes with the suboesophageal ganglion and descends into the ventral nerve chord. This neuron is accompanied by a second serotonin-immunoreactive interneuron with projections into the protocerebrum. Two pairs of bilateral immunoreactive serial homologues were identified in each of the three suboesophageal neuromeres and were also found in the thoracic ganglia. With the exception of the frontal commissure, no immunoreactive processes could be found in the peripheral nerves of the brain and the suboesophageal ganglion. The morphological studies on the serial homologues were extended by intracellular injections of Lucifer Yellow combined with immunofluorescence.     

Splanchnic Nerve Transection Abolishes the Age-Dependent Increase of Neuropeptide Y-Like Immunoreactivity in Rat Adrenal Gland

Using an antiserum directed against porcine neuropeptide Y (NPY), a high concentration of NPY immunoreactivity (NPY-IR) was detected in rat adrenal gland. The level of NPY-IR in the adrenal gland was found to increase considerably with age. Biochemical characterization by reverse-phase HPLC indicated that this increase was due to accumulations of NPY and another molecular form of NPY-like immunoreactive peptide. Chronic denervation of the splanchnic nerve abolished this age-dependent increase of NPY-IR rat adrenal gland.     

Localization of corticotropin-releasing factor-immunoreactive nervous tissue and colocalization with neuropeptide Y-like substance in the optic lobe and peduncle complex of the octopus (<Emphasis Type="Italic">Octopus vulgaris</Emphasis>)

The distribution of corticotropin-releasing factor (CRF)-like immunoreactivity and its colocalization with neuropeptide Y (NPY)-like substances were investigated in the optic lobe and peduncle complex of the octopus (Octopus vulgaris) using immunohistochemical techniques. In the optic lobe cortex, CRF-immunoreactive (CRF-IR) and NPY-immunonegative varicose fibers were observed in the plexiform layer. In the medulla, CRF-IR somata were seen in the cell islands, and CRF-IR varicose fibers were observed in the neuropil. About half of the CRF-IR structures in the medulla showed NPY-like immunoreactivity. In the peduncle lobe, no CRF-IR somata but abundant CRF-IR varicose fibers were observed, and about half of them showed NPY-like immunoreactivity. In the olfactory lobe, CRF-IR somata and abundant CRF-IR varicose fibers were observed. Almost all the CRF-IR somata located in the posterior olfactory lobule showed NPY-like immunoreactivity, whereas those seen in the median olfactory lobule were immunonegative for NPY. About half of the CRF-IR fibers in the anterior lobule neuropil were immunopositive for NPY, but those in the median and posterior lobule neuropils were immunonegative for NPY. In the optic gland, almost all the CRF-IR varicose fibers were immunoreactive for NPY. Western blot analysis of the optic lobe and peduncle complex indicated that anti-CRF antiserum labeled approximate 16.4- and 14.6-kDa bands and that anti-NPY antiserum labeled an approximate 16.2-kDa band. CRF-IR and NPY-immunoreactive neurons in the optic lobe may participate in the modulation of visual information and those in the optic gland may be involved in the regulation of endocrine function.     

Gastrin/cholecystokinin-like immunoreactivity in the nervous system of Aeschna cyanea (Insecta,Odonata)

Summary Gastrin/cholecystokinin (gastrin/CCK)-like immunoreactivity has been detected in the brain, suboesophageal ganglion and corpora cardiaca of the larva of Aeschna cyanea by radioimmunoassay and immunohistochemistry, by use of two antisera raised against the sulfated (CCK-8S) and the unsulfated form (CCK-8NS) of the carboxyl terminal octapeptide. Numerous immunoreactive neurons were demonstrated in the protocerebrum (exclusive of optic lobes) and suboesophageal ganglion where 20 and 15 symmetrical clusters of reactive cells, respectively, were observed. Immunoreactive cells also occurred in the tritocerebrum, the optic lobes and the frontal ganglion. In the corpora cardiaca, gastrin/CCK-like material was found both within intrinsic cells and axon terminals. RIA measurements support the immunohistochemical results in so far as large amounts of gastrin/CCK-like material were detected in the brain, corpora cardiaca and suboesophageal ganglion complex. Both boiling water-acetic acid- and methanol-extraction procedures were performed. Comparisons of the results lead to the conclusion that a large part of the gastrin/CCK-like material occurs as small molecules. Immunohistochemical procedures performed on material fixed in a solution of picric acid-paraformaldehyde demonstrated differences in the immunoreactivity of the tested antisera. First, the immunohistochemical reaction was always more pronounced when the CCK-8NS antiserum was used instead of the CCK-8S antiserum, which may be interpreted by a lower affinity of the latter. In the second place, some neurons strongly stained by the CCK-8NS antiserum were only very faintly if at all stained by the CCK-8S antiserum, which may mean that different peptides or at least distinct forms of the same precursor are detected.     

The distribution of pancreatic polypeptide in the nervous system and gut of the blowfly,Calliphora vomitoria (Diptera)

Summary The distribution of a neuropeptide, previously shown to have the same or a very similar amino acid composition as vertebrate pancreatic polypeptide (PP), has been studied in the nervous system and gut of the blowfly, Calliphora vomitoria. Neurones immunoreactive to a bovine PP antiserum occur in the thoracic and abdominal ganglionic components of the central nervous system, in addition to the brain and suboesophageal ganglion. Pancreatic polypeptide appears to be relayed from its cells of origin to a neurohaemal organ in the dorsal sheath of the thoracic ganglion. PP immunoreactivity is also found in cells of the hypocerebral ganglion of the stomatogastric nervous system and in associated nerve fibres. The mid-gut contains PP-positive material in flask-shaped cells of its epithelial lining.     

The presence and distribution of crustacean cardioactive peptide in the central and peripheral nervous system of the stick insect, Baculum extradentatum

Crustacean cardioactive peptide (CCAP)-like immunoreactivity was localized and quantified in the central and peripheral nervous system of the Vietnamese stick insect, Baculum extradentatum, using immunohistochemistry and enzyme-linked immunosorbent assay (ELISA). The brain, frontal ganglion, suboesophageal ganglion and ventral nerve cord displayed neurons and processes with CCAP-like immunoreactivity. The brain, in comparison to the other parts of the central nervous system, contained the greatest amount of CCAP (167 +/- 18 fmol), and showed CCAP-like staining in neurons, neuropil regions and the central complex. There were also CCAP-like varicosities and processes associated with the corpus cardiacum. The alimentary canal of B. extradentatum contained CCAP with the largest amount localized in the midgut (1110 +/- 274 fmol CCAP equivalents). The midgut contained numerous endocrine-like cells which stained positively for CCAP, whereas the foregut and hindgut revealed an extensive network of CCAP-like immunoreactive axons and varicosities. Based on physiological assays, the hindgut of the stick insect was found to be sensitive to CCAP, showing dose-dependent increases in contractions with threshold at 10(-10) M CCAP and maximal response at 5 x 10(-7) M CCAP. There were negligible quantities of CCAP in the oviducts and no CCAP-like immunoreactivity was associated with the oviducts. CCAP had no effect on spontaneous contractions of the oviducts. The presence of CCAP in the central nervous system, the stomatogastric nervous system, the corpus cardiacum and the alimentary canal, suggest broad ranging roles for CCAP in B. extradentatum.     

Localization of corazonin in the nervous system of the cockroach Periplaneta americana

Antisera raised to the cardioactive peptide corazonin were used to localize immunoreactive cells in the nervous system of the American cockroach. Sera obtained after the seventh booster injection were sufficiently specific to be used for immunocytology. They recognized a subset of 10 lateral neurosecretory cells in the protocerebrum that project to, and arborize and terminate in the ipsilateral corpus cardiacum. They also reacted with bilateral neurons in each of the thoracic and abdominal neuromeres, a single dorsal unpaired median neuron in the suboesophageal ganglion, an interneuron in each optic lobe, and other neurons at the base of the optic lobe, in the tritocerebrum and deutocerebrum. The presence of corazonin in the abdominal neurons and the lateral neurosecretory cells was confirmed by HPLC fractionation of extracts of the abdominal ganglia, brains and retrocerebral complexes, followed by determination of corazonin by ELISA, which revealed in each tissue a single immunoreactive peak co-eluting with corazonin in two different HPLC systems. Antisera obtained after the first three booster injections recognized a large number of neuroendocrine cells and neurons in the brain and the abdominal nerve cord. However, the sera from the two rabbits reacted largely with different cells, indicating that the majority of this immunoreactivity was due to cross-reactivity. These results indicate that the production of highly specific antisera to some neuropeptides may require a considerable number of booster injections.     

Insect myotropic peptides: differential distribution of locustatachykinin- and leucokinin-like immunoreactive neurons in the locust brain

Locustatachykinin I is one of four closely related myotropic neuropeptides isolated from brain and corpora-cardiaca complexes of the locust Locusta migratoria. Antiserum was raised against locustatachykinin I for use in immunocytochemistry. It was found that the antiserum recognizes also locustatachykinin II and hence probably also the other two locustatachykinins due to their similarities in primary structure. Locustatachykinin-like immunoreactive (LomTK-LI) neurons were mapped in the brain of the locust, L. migratoria. A total of approximately 800 Lom TK-LI neurons were found with cell bodies distributed in the proto-, deutoand tritocerebrum, in the optic lobes and in the frontal ganglion. Processes of these neurons innervate most of the synaptic neuropils of the brain and optic lobes, as well as the frontal ganglion and hypocerebral ganglion. The widespread distribution of LomTK-LI neurons in the locust brain indicates an important role of the locustatachykinins in signal transfer or regulation thereof. As a comparison neurons were mapped with an antiserum against the cockroach myotropic peptide leucokinin I. This antiserum, which probably recognizes the native peptide locustakinin, labels a population of about 140 neurons distinct from the LomTK-LI neurons (no colocalized immunoreactivity). These neurons have cell bodics that are distributed in the proto- and tritocerebrum and in the optic lobe. The processes of the leucokinin-like immunoreactive (LK-LI) neurons do not invade as large areas in neuropil as the Lom TK-LI neurons do and some neuropils, e.g. the mushroom bodies, totally lack innervation by LK-LI fibers. In some regions, however, the processes of the Lom TK-LI and LK-LI neurons are superimposed: most notably in the central body and optic lobes. A functinal relation between the two types of neuropeptide in the locust brain can, however, not be inferred from the present findings.     

Characterization of aPY-like peptides in anglerfish brain using a novel radioimmunoassay for aPY-Gly

Anglerfish peptide YG (aPY) was isolated from pancreatic islets of the anglerfish. Subsequent immunohistochemical and biochemical analyses demonstrated that anglerfish islet cells synthesize aPY. We have now developed and characterized a radioimmunoassay (RIA) for aPY and have examined extracts of anglerfish brain for aPY-like peptides. Brain extracts were subjected to gel filtration and high performance liquid chromatography (HPLC). Fractions from HPLC eluates were analyzed in the aPY RIA and also in a neuropeptide Y (NPY) RIA. A single peak of aPY-like immunoreactivity eluted from HPLC columns. The elution position of this aPY-like peptide coincided exactly with the aPY-Gly marker under several gradient conditions. Results from the NPY RIA confirmed the presence of several molecular forms of NPY-like immunoreactive peptides in the anglerfish brain. These results demonstrate the utility of the newly developed aPY RIA for studies of anglerfish brain peptides and extend our previous immunohistochemical demonstration of aPY-like staining in the anglerfish brain.     

FMRFamide-related peptides in the gut of Locusta migratoria L.: a comprehensive map and developmental profile

The gut tissues and associated nervous system of the African migratory locust, Locusta migratoria, were found to contain FMRFamide-like immunoreactive (FLI) material throughout the five larval instars and 2 weeks into the adult stage in both males and females. FMRFamide-like immunoreactivity associated with the locust gut was described using camera lucida techniques. FMRFamide-like immunoreactivity is observed in the frontal connectives, recurrent nerve, and oesophageal nerves; projections from the ingluvial ganglion onto the anterior midgut, and from the proctodeal nerve onto the hindgut and posterior midgut; in the neuropils of the frontal ganglion, hypocerebral ganglion and ingluvial ganglia; 30 cell bodies in the frontal ganglion; multipolar sensory cells on the foregut; and endocrine-like cells in the gastric caecae and midgut. Radioimmunoassay (RIA) was used to determine the quantities of FLI material in foreguts, gastric caecae, anterior and posterior midguts, and hindgut of first-fifth instar larvae, 1-3- and 14-17-day male and female adult locusts. As expected, as the tissue size (assessed by total protein content) increases, so does the amount of FLI material in each tissue. Normalizing for tissue size reveals significant differences in FLI content among the stages for each tissue tested. Reversed phase-high pressure liquid chromatography (RP-HPLC) followed by RIA has identified four groups of FLI fractions present in the gut, and different members of these groups are present in the various gut tissues.     

Distribution of PER protein,pigment-dispersing hormone,prothoracicotropic hormone,and eclosion hormone in the cephalic nervous system of insects

Investigations performed on adult insects revealed that putative components of the central pacemaker, the protein Period (PER) and the pigment-dispersing hormone (PDH), are immunocytochemically detectable in discrete sets of brain neurons throughout the class of Insecta, represented by a bristletail, mayfly, damselfly, 2 locust species, stonefly, 2 bug species, goldsmith beetle, caddisfly, honeybee, and 2 blowfly species. The PER-positive cells are localized in the frontal protocerebrum and in most species also in the optic lobes, which are their only location in damselfly and goldsmith beetle. Additional PER-positive cells occur in a few species either in the deuto- and tritocerebrum or in the suboesophageal ganglion. The PER staining was always confined to the cytoplasm. The PDH immunoreactivity consistently occurs in a cluster of perikarya located frontoventrally at the proximal edge of the medulla. The mayfly and both locust species possess additional PDH neurons in 2 posterior cell clusters at the proximal edge of the medulla, and mayfly, waterstrider, and 1 of the blowfly species in the central brain. PDH-positive fibers form a fanlike arrangement over the frontal side of the medulla. Two or just 1 bundle of PDH-positive fibers run from the optic lobe to the protocerebrum, with collaterals passing over to the contralateral optic lobe. Antisera to the prothoracicotropic (PTTH) and the eclosion (EH) hormones, which in some insects regulate the molting and ecdysis rhythms, respectively, typically react with a few neurons in the frontal protocerebrum. However, the PTTH-positive neurons of the mayfly and the damselfly and the EH-positive neurons of the caddisfly are located in the suboesophageal ganglion. No PTTH-like antigen was detected in locusts, and no EH-like antigens were detected in the damselfly, stonefly, locusts, and the honeybee. There are no signs of co-localization of the PER-, PDH-, PTTH-, and EH-like antigens in identical neurons.