UVB irradiation induces contralateral changes in galanin, substance P and c-fos immunoreactivity in rat dorsal root ganglia, dorsal horn and lateral spinal nucleus.

The selection of control group is crucial, as the use of an inadequate group may strongly affect the results. In this study we examine the effect on contralateral tissue protein levels, in a model of unilateral UVB irradiation, as the contralateral side is commonly used as a control. Previous studies have shown that UVB irradiation increases immunoreactivity for inflammatory regulated neuropeptides. Unilateral UVB irradiation of rat hind paw was performed and corresponding contralateral spinal cord and dorsal root ganglia (DRG) were collected 2-96 h after and investigated for changes in galanin, substance P and c-fos immunoreactivity. Control tissue was collected from naïve rats. Measurement of skin blood flow from contralateral heel hind paws (Doppler), revealed no change compared to naïve rats. However, UVB irradiation caused a significant reduction in the contralateral proportion of galanin immunopositive DRG neurons, at all-time points, as well as an increase in the contralateral spinal cord dorsal horn, around the central canal and in the lateral spinal nucleus (2-48 h). The contralateral proportion of SP positive DRG neurons and dorsal horn immunoreactivity was unchanged, whereas the lateral spinal nucleus area showed increased immunoreactivity (48 h). UVB irradiation also induced a slight contralateral upregulation of c-fos in the dorsal horn/central canal area (24 and 48 h). In summary, unilateral UVB irradiation induced contralateral changes in inflammatory/nociceptive neuropeptides in spinal cord and afferent pathways involved in pain signaling already within 24 h, a time point when also ipsilateral neurochemical/physiological changes have been reported for rats and humans.

Review: Occurrence and Distribution of Galanin in the Physiological and Inflammatory States in the Mammalian Gastrointestinal Tract.

Galanin (GAL) is a broad-spectrum peptide that was first identified 37 years ago. GAL, which acts through three specific receptor subtypes, is one of the most important molecules on an ever-growing list of neurotransmitters. Recent studies indicate that this peptide is commonly present in the gastrointestinal (GI) tract and GAL distribution can be seen in the enteric nervous system (ENS). The function of the GAL in the gastrointestinal tract is, inter alia, to regulate motility and secretion. It should be noted that the distribution of neuropeptides is largely dependent on the research model, as well as the part of the gastrointestinal tract under study. During the development of digestive disorders, fluctuations in GAL levels were observed. The occurrence of GAL largely depends on the stage of the disease, e.g., in porcine experimental colitis GAL secretion is caused by infection with Brachyspira hyodysenteriae. Many authors have suggested that increased GAL presence is related to the involvement of GAL in organ renewal. Additionally, it is tempting to speculate that GAL may be used in the treatment of gastroenteritis. This review aims to present the function of GAL in the mammalian gastrointestinal tract under physiological conditions. In addition, since GAL is undoubtedly involved in the regulation of inflammatory processes, and the aim of this publication is to provide up-to-date knowledge of the distribution of GAL in experimental models of gastrointestinal inflammation, which may help to accurately determine the role of this peptide in inflammatory diseases and its potential future use in the treatment of gastrointestinal disorders.

Validation of antibody-based tools for galanin research.

Antibodies are an integral biomedical tool, not only for research but also as therapeutic agents. However, progress can only be made with sensitive and specific antibodies. The regulatory (neuro)peptide galanin and its three endogenous receptors (GAL1-3-R) are widely distributed in the central and peripheral nervous systems, and in peripheral non-neuronal tissues. The galanin system has multiple biological functions, including feeding behavior, pain processing, nerve regeneration and inflammation, to name only a few. Galanin could serve as biomarker in these processes, and therefore its receptors are potential drug targets for various diseases. For that reason, it is of paramount interest to precisely measure galanin peptide levels in tissues and to determine the cellular and subcellular localization of galanin receptors. A plethora of antibodies and antibody-based tools, including radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA) kits, are commercially available to detect galanin and its receptors. However, many of them lack rigorous validation which casts doubt on their specificity. A goal of the present study was to raise awareness of the importance of validation of antibodies and antibody-based tools, with a specific focus on the galanin system. To that end, we tested and report here about commercially available antibodies against galanin and galanin receptors that appear specific to us. Furthermore, we investigated the validity of commercially available galanin ELISA kits. As the tested ELISAs failed to meet the validation requirements, we developed and validated a specific sandwich ELISA which can be used to detect full-length galanin in human plasma.

UVB irradiation induces rapid changes in galanin, substance P and c-fos immunoreactivity in rat dorsal root ganglia and spinal cord.

Recent studies have shown that UVB irradiation induces primary and secondary hyperalgesia in rats and humans peaking about 24h after UVB exposure. In the present study we investigated the changes in galanin, substance P and c-fos immunoreactivity in rat DRG and spinal cord at the L5 level 2-96h after UVB irradiation. UVB irradiation of the heel area in rats almost increased the skin blood flow two-fold 24h after irradiation as measured by laser Doppler technique. UVB irradiation induced a significant reduction of the proportion of galanin positive DRG neurons for all time points, except at 12h. In the spinal cord, UVB irradiation induced increased immunoreactivity for galanin in the dorsal horn, the area around the central canal and interestingly also in the lateral spinal nucleus 12-96h after exposure. For substance P the proportion of substance P positive neurons was unchanged but UVB irradiation induced increased substance P immunoreactivity in the dorsal part of the spinal cord 48h after irradiation. UVB irradiation also induced c-fos immunoreactivity in the dorsal horn and the area around the central canal 24 and 48h after exposure. This translational model of UVB irradiation will induce rapid changes of neuropeptides implicated in nociceptive signaling in areas known to be of importance for nociception in a time frame, about 24h after exposure, where also neurophysiological alteration have been described in humans and rats.

Distribution of cocaine- and amphetamine-regulated transcript (CART), neuropeptide Y (NPY) and galanin (GAL) in the pterygopalatine ganglion of the domestic duck (Anas platyrhynchos f. domestica).

INTRODUCTION: Cocaine- and amphetamine-regulated transcript (CART), neuropeptide Y (NPY) and galanin (GAL) act as neurotransmitters and neuromodulators in both the central and peripheral nervous systems. Their presence has been found in different taxonomic groups, in particular in mammals. However, only few investigators have studied these neuropeptides in the class Aves (birds). The aim of the present study was to describe the distribution of CART, NPY and GAL in the pterygopalatine ganglion (PPG) of the domestic duck (Anas platyrhynchos f. domestica). MATERIAL AND METHODS: The experiment was conducted on 16 one-year-old domestic ducks of the Pekin breed of both sexes (8 males and 8 females). Frozen sections of the PPG were subjected to immunofluorescence staining using primary mouse monoclonal antibodies directed against CART and GAL and rabbit polyclonal antibody directed against NPY. Secondary antibodies were conjugated with Cy3 and FITC fluorochromes. RESULTS: CART, NPY, and GAL were present in the PPG of the domestic duck. The highest immunoreactivity (IR) in the ganglionic cells was found for CART in the majority (83-85%) of neurons of both superior (SPPG) and inferior (IPPG) PPG. CART-IR was also found in small aggregations of neurons on the medial surface of the Harderian gland, and on the course of the palatine branch of the facial nerve. CART-IR was also observed in the nerve fibers of these neurons' aggregations; however, it was low in comparison to the immunoreactivity of the perikarya. Immunoreactivity of NPY was found in ganglionic neurons, but above all in numerous fibers of the SPPG and IPPG and within aggregations on the surface of the Harderian gland. NPY-IR cells were distributed irregularly over the cross-sections of the tested aggregations, and constituted from 36% to 43% of the SPPG and from 37% to 40% of the IPPG of all cross-sectioned neurons. GAL-immunoreactive perikarya, distributed irregularly across the sections, were observed in the SPPG, where they constituted 61-65%, and in the IPPG, where they made up 50-57% of all neurons. All immunoreactive neurons were characterized by immunopositive neuroplasm and immunonegative cell nuclei. CONCLUSIONS: The presence of CART, NPY, and GAL in the PPG of the domestic duck suggests that these peptides may contribute to the secretory innervation of the glands of the mucosa of the palate and nasal cavity, the Harderian gland, and the lacrimal gland.

Identification of cell-penetrating peptides that are bactericidal to Neisseria meningitidis and prevent inflammatory responses upon infection.

Meningococcal disease is characterized by a fast progression and a high mortality rate. Cell-penetrating peptides (CPPs), developed as vectors for cargo delivery into eukaryotic cells, share structural features with antimicrobial peptides. A screen identified two CPPs, transportan-10 (TP10) and model amphipathic peptide (MAP), with bactericidal action against Neisseria meningitidis. Both peptides were active in human whole blood at micromolar concentrations, while hemolysis remained negligible. Additionally, TP10 exhibited significant antibacterial activity in vivo. Uptake of SYTOX green into live meningococci was observed within minutes after TP10 treatment, suggesting that TP10 may act by membrane permeabilization. Apart from its bactericidal activity, TP10 suppressed inflammatory cytokine release from macrophages infected with N. meningitidis as well as from macrophages stimulated with enterobacterial and meningococcal lipopolysaccharide (LPS). Finally, incubation with TP10 reduced the binding of LPS to macrophages. This novel endotoxin-inhibiting property of TP10, together with its antimicrobial activity in vivo, indicates the possibility to design peptide-based therapies for infectious diseases.

Galanin and α-MSH autoantibodies in cerebrospinal fluid of patients with Alzheimer's disease.

BACKGROUND: Neuropeptides galanin and α-melanocyte-stimulating hormone (α-MSH) are involved in the regulation of memory and appetite. Increased galanin and decreased α-MSH levels were reported in postmortem brains of patients with Alzheimer's disease (AD) but the underlying mechanisms are uncertain. Here we studied if autoantibodies (autoAbs) reacting with galanin and α-MSH are altered in AD. METHODS: Levels of free and total IgG autoAbs reacting with galanin and α-MSH were measured in sera and cerebrospinal fluid (CSF) of 18 subjects with AD and in 15 age-matched non-demented controls. Values were correlated with Mini-Mental State Examination (MMSE) score, body mass index (BMI) and CSF levels of AD biomarkers. RESULTS: CSF levels of total but not free IgG autoAbs against galanin were increased in AD, resulting in increased percentage of galanin autoAbs present as immune complexes. CSF levels of galanin total autoAbs and α-MSH free autoAbs correlated negatively with the severity of cognitive impairment as measured by MMSE. Both total and free autoAbs against galanin and α-MSH in CSF correlated negatively with age in AD patients but not in controls. CSF levels of galanin autoAbs and free α-MSH AutoAbs negatively correlated with CSF levels of t-Tau, p-Tau and ratios of t-Tau/Aß42 or p-Tau/Aß42 in AD patients but not in controls. CONCLUSIONS: AutoAbs reacting with galanin and α-MSH are present in CSF and are associated with clinical characteristics of AD patients. The functional significance and therapeutic potential of these autoAbs should be further clarified.

Cell-penetrating peptides, PepFects, show no evidence of toxicity and immunogenicity in vitro and in vivo.

Cell-penetrating peptide based vehicles have been developed for the delivery of different payloads into the cells in culture and in animals. However, several biological features, among which is the tendency to trigger innate immune response, limit the development of highly efficient peptide-based drug delivery vectors. This study aims to evaluate the influence of transportan 10 (TP10) and its chemically modified derivatives, PepFects (PFs), on the innate immune response of the host system. PFs have shown high efficiency in nucleic acid delivery in vitro and in vivo; hence, the estimation of their possible toxic side effects would be of particular interest. In this study, we analyzed cytotoxic and immunogenic response of PF3, PF4, and PF6 peptides in monocytic leukemia and peripheral blood mononuclear cell lines. In comparison with amphipathic PFs, TP10, TAT, stearyl-(RxR)(4) peptides, and the most widely used transfection reagents Lipofectamine 2000 and Lipofectamine RNAiMAX were also analyzed in this study. IL-1ß, IL-18, and TNF-α cytokine release was detected using highly sensitive enzyme-linked immunosorbent assay (ELISA). Cell viability was detected by measuring the activity of cellular enzymes that reduce water-soluble tetrazolium salts to formazan dyes and apoptosis was evaluated by measuring the levels of caspase-1 and caspase-3/7 over untreated cells. All peptides were found to be nontoxic and nonimmunogenic in vitro at the concentrations of 10 µM and 5 µM, respectively, and at a dose of 5 mg/kg in vivo, suggesting that these CPPs exhibit a promising potential in the delivery of therapeutic molecules into the cell without risks of toxicity and inflammatory reactions.

The galanin peptide family in inflammation.

The immune system defends the organism against invading pathogens. In recent decades it became evident that elimination of such pathogens, termination of inflammation, and restoration of host homeostasis all depend on bidirectional crosstalk between the immune system and the neuroendocrine system. This crosstalk is mediated by a complex network of interacting molecules that modulates inflammation and cell growth. Among these mediators are neuropeptides released from neuronal and non-neuronal components of the central and peripheral nervous systems, endocrine tissues, and cells of the immune system. Neuropeptide circuitry controls tissue inflammation and maintenance, and an imbalance of pro- and anti-inflammatory neuropeptides results in loss of host homeostasis and triggers inflammatory diseases. The galanin peptide family is undoubtedly involved in the regulation of inflammatory processes, and the aim of this review is to provide up-to-date knowledge from the literature concerning the regulation of galanin and its receptors in the nervous system and peripheral tissues in experimental models of inflammation. We also highlight the effects of galanin and other members of the galanin peptide family on experimentally induced inflammation and discuss these data in light of an anti-inflammatory role for this family of peptides.

Galanin-like immunoreactivity in the brain of the snake Bothrops jararaca.

The distribution of galanin-like immunoreactive perikarya and nerve fibers in the brain of the snake Bothrops jararaca was studied by means of immunohistochemistry using an antiserum against porcine galanin. Immunoreactive neurons were only detected in the infundibular recess nucleus. Immunoreactive fibers were found in the telencephalic, diencephalic and mesencephalic areas such as the dorsal cortex, nucleus accumbens, lamina terminalis, preoptic area, mediodorsal region of the supraoptic nucleus, subfornical organ, nucleus of the paraventricular organ, subcommisural organ and periventricular grey region. The habenula, paraventricular nucleus, infundibular recess nucleus and hypothalamo-hypophyseal tract presented denser innervations. The outer layer of the median eminence displayed numerous fibers located close to the portal system, while scarce fibers were seen in the inner median eminence and neural lobe of the hypophysis. The distribution of labelled neurons in the brain of this snake was more restricted than that described in a turtle. The wide hypothalamic and extrahypothalamic distribution of labelled fibers suggests that galanin peptides may have hypophysiotropic, neuromodulator and neurotransmitter roles in the snake B. jararaca.

Galanin in adjuvant arthritis in the rat.

OBJECTIVE: To study the concentration changes of galanin in the ankles and spinal cord and to detect the distribution of galanin in different tissues in arthritic rats. METHODS: Adjuvant arthritis was induced by intradermal injection of Mycobacterium butyricum in Freund's incomplete adjuvant at the base of the tail. The concentrations of galanin were measured by radioimmunoassay (RIA) and the distributions of galanin were detected by immunoelectron microscopy (iEM). RESULTS: Measurements were taken on Day 28 after injection. RIA results showed that the concentration of galanin was significantly lower in the ankles and spinal cords of rats with adjuvant arthritis compared to controls. Our iEM results showed a heterogeneous distribution of galanin labelling in different cells and tissue compartments. In arthritic rats, we observed decreased galanin labelling in the sciatic nerve and in macrophage-like cells in the synovial membrane and increased labelling in monocyte lineage cells, polymorphonucleated lineage cells in the bone marrow, fibroblasts in the periosteum, osteoclasts and osteocytes, and lower labelling in osteoblasts compared to controls. CONCLUSION: Galanin is involved in the response to inflammation in adjuvant arthritis and might play a role in the regulation of inflammation and nociception. These findings are in accordance with a biological role of galanin in the development of inflammatory arthritis.

Activation of a subset of lumbar spinothalamic neurons after copulatory behavior in male but not female rats.

The precise pathways that convey copulation-related information to forebrain regions activated during male and female sexual behavior are poorly understood. Previous work from our laboratory and others has demonstrated the existence of a spinothalamic pathway that is a candidate to relay information to these areas. This pathway originates from a population of spinothalamic neurons in the lumbar spinal cord containing several neuropeptides including galanin, located in laminas 7 and 10 of the lumbar segments 3 and 4. To investigate the involvement of these lumbar spinothalamic neurons in conveying copulation-related information, we tested the hypothesis that these cells are activated after ejaculation in male rats and vaginocervical stimulation in female rats. This was assessed using galanin or cholecystokinin as a marker for this subset of spinothalamic neurons and Fos-immunoreactivity as a marker for neuronal activation. The results demonstrated that activation of these spinothalamic neurons is triggered by stimuli associated with ejaculation. Fos induction was specifically associated with ejaculation, because mounts or intromissions did not trigger expression. Moreover, these spinothalamic neurons were not activated by vaginocervical stimulation in female rats. Spinothalamic neurons have generally been associated with signaling pain and temperature information. The present findings demonstrate that a specific subpopulation of spinothalamic neurons signals information associated with ejaculation.

Innervation of the uvea by galanin and somatostatin immunoreactive axons in macaques and baboons.

The neuropeptide galanin has not been localized previously in the primate uvea, and the neuropeptide somatostatin has not been localized in the uvea of any mammal. Here, the distribution of galanin-like and somatostatin-like immunoreactive axons in the iris, ciliary body and choroid of macaques and baboons using double and triple immunofluorescence labeling techniques and confocal microscopy was reported. In the ciliary body, galanin-like immunoreactive axons innervated blood vessels and the ciliary processes, particularly at their bases. In the iris, the majority of these axons was associated with the loose connective tissue in the stroma. Somatostatin-like immunoreactive axons were found in many of the same areas of the uvea supplied by cholinergic nerves. In the ciliary body, there were labelled axons within the ciliary processes and ciliary muscle. They were also found alongside blood vessels in the ciliary stroma. In the iris, somatostatin-like immunoreactive axons were abundant in the sphincter muscle and less so in the dilator muscle. A unilateral sympathectomy had no effect on the distribution of somatostatin-like or galanin-like immunoreactive axons, and these axons did not contain the sympathetic marker tyrosine hydroxylase. They did not contain the parasympathetic marker choline acetyltransferase, either. The galanin-like immunoreactive axons contained other neuropeptides found in sensory nerves, including calcitonin gene-related peptide, substance P and cholecystokinin. Somatostatin-like immunoreactive axons did not contain any of these sensory neuropeptides or galanin-like immunoreactivity, and they were neither labelled with an antibody to 200kDa neurofilament protein, nor did they bind isolectin-IB(4). Nevertheless, they are likely to be of sensory origin because somatostatin-like immunoreactive perikarya have previously been localized in the trigeminal ganglion of primates. Taken together, these findings indicate galanin and somatostatin are present in two different subsets of sensory axons in primate uvea.

Peptidergic innervation of blood vessels and interstitial cells in the testis of the cat.

We studied the innervation of the cat testis using a panel of antisera against the following neuronal markers: protein gene product 9.5 (PGP), neuropeptide Y, C-terminal peptide of neuropeptide Y, galanin, vasoactive intestinal peptide (VIP), calcitonin gene-related peptide, and substance P. Immunoreactivity against PGP, a general neuronal label, demonstrated the arrangement of fibers from the superior spermatic nerve (SSN) in the testicular pedicle and the cephalic testicular pole, and those of the inferior spermatic nerve (ISN) along the vas deferens and the inferior testicular ligament. The testicular parenchyma exhibited a very rich innervation, mainly distributed to blood vessels and Leydig cell nests, but also in close association with seminiferous tubules. Numerous peptidergic fibers were present in the SSN and ISN, albeit in different proportions. Thus, VIP-immunoreactive fibers were almost absent in the SSN, but were the most abundant subpopulation of the ISN. The testicular interstitium contained numerous peptidergic fibers, associated with blood vessels, interstitial Leydig cells, and seminiferous tubules. Similar fibers were related to the rete testis. Parenchymatous VIP-immunoreactive nerves disappeared after bilateral vasectomy. Stimulation of the ISN under experimental conditions was associated with an increase of blood flow, and induced a large release of VIP into the spermatic vein. The extensive and selective distribution of nerve fibers within the cat testicular parenchyma supports the importance of spermatic nerves for testicular function. Furthermore, the differences in the fiber composition of the SSN and ISN can be correlated with their opposing effects on testosterone secretion and testicular blood flow.

Large reduction in the number of galanin-immunoreactive cells in pancreatic islets of diabetic rats.

Although galanin has been shown to be present in pancreatic islet cells, there is no literature available on the pattern of distribution and the effect of galanin in the pancreas of diabetic animals or human models. The aim of this study was to examine whether galanin immunoreactivity changes after the onset of diabetes mellitus in the rat model. The present study used immunohistochemical techniques to examine the pattern of distribution of galanin-like immunoreactive cells in the pancreas of rats with streptozotocin-induced diabetes. The effect of galanin on insulin secretion from intact rat pancreatic tissue fragments was also investigated using a radioimmunoassay technique. Numerous galanin-like immunoreactive cells were observed in both the peripheral and central regions of the islet of Langerhans of normal rat pancreas. By contrast, the islets of diabetic rat pancreas contained significantly (P < 0.0001) fewer galanin-like immunoreactive cells than nondiabetic rats. Galanin was colocalized with insulin in the islets of normal and diabetic rats. Galanin had an inhibitory effect on insulin secretion from the isolated pancreatic tissue fragments of normal and diabetic rats at all concentrations (10(-12) to 10(-6) M) employed. Galanin at 10(-9) M caused a significant (P < 0.02) decrease in insulin secretion from normal rat pancreatic tissue fragments compared to basal. These observations indicate that galanin may play a significant role in the regulation of insulin secretion.

Intrinsic choroidal neurons in the duck eye receive sympathetic input: anatomical evidence for adrenergic modulation of nitrergic functions in the choroid.

Intrinsic choroidal neurons (ICN) in the duck eye form an intramural ganglionic plexus that may subserve complex integrative functions. A key feature of such ganglia is an innervation by sympathetic postganglionic neurons. The present study was thus aimed at determining the sympathetic postganglionic innervation of ICN. Choroids were processed for double immunofluorescence labelling with the following markers: tyrosine-hydroxylase (TH)/nitric oxide synthase (nNOS), TH/galanin (GAL), dopamine-beta-hydroxylase (DBH)/vasoactive intestinal polypeptide (VIP), TH/DBH and DBH/alpha-smooth-muscle actin (alphaSMA), and for triple immunofluorescence labelling with VIP/DBH/TH. Epifluorescence and confocal laser scanning microscopy were used for evaluation. Immunoperoxidase staining for TH or DBH in combination with NADPH-diaphorase histochemistry was applied for electron microscopy. ICN spread over the entire choroid but were concentrated in an equatorial zone passing obliquely from naso-cranial to temporocaudal. More than 80% of nNOS-positive ICN showed close appositions of TH/DBH-immunoreactive varicose nerve fibres at the light-microscopic level, as could be confirmed by confocal laser scanning microscopy. Ultrastructurally, these appositions could be defined as both synapses or close contacts without synaptic specialisation. Vascular and non-vascular smooth muscle fibres also received TH/DBH-immunopositive innervation. Our findings suggest that most ICN receive a sympathetic input that might modulate their nitrergic effects upon vascular and non-vascular smooth muscle fibres in the choroid and that they may have more complex functions than merely being a simple parasympathetic relay.

Monoclonal antibody to rat galanin: production, characterization, and in vivo immunoneutralization activity.

A monoclonal antibody (MAb) to galanin was prepared by cell fusion of myeloma Fox-NY and spleen cells from Robertsonian mice immunized with rat galanin. Hybridomas producing high-affinity antibodies were cloned in pristine-primed Balb/c mice. The antibody was purified by affinity chromatography and concentrated to 12 mg IgG/mL by dialysis. Immunoreactivity of the antibody was screened by radioimmunoassay. Ascites fluid contained approximately 10 mg/mL IgG that belong to the subclass of IgG2a as determined by enzyme-linked immunoadsorbent assay (ELISA). The titer of this IgG2a antibody entitled #G65G was 1:10,000 and the ID50 for rat galanin was 1000 fmol/mL as determined by liquid phase radioimmunoassay. Immunohistochemistry showed that this galanin MAb stains densed, beaded processes distributed to the enteric plexuses, where they appear to encircle neuronal cell bodies, to the muscle layer, where they are particularly abundant in the circular muscle layer and in the deep muscular layer, and to the mucosa. In vivo capacity of immunoneutralization by this antibody was tested in male Sprague-Dawley rats fasted for 24 h and anesthetized with urethane. Systemic injection of protein A purified galanin antibody (6 mg/rat) decreased by 70% of the inhibitory effect of intravenous galanin (2 nmol/kg/h i.v.) on gastric acid secretion induced by intracisternal TRH analog. These results show that galanin antibody #G65G is useful for in vivo immunoneutralization of galanin effects and is a valuable tool for immunohistochemical localization of galanin in gastrointestinal tissues.

Sexually dimorphic distribution of galanin in the preoptic area of red salmon, Oncorhynchus nerka.

A sexually dimorphic distribution of galanin in the preoptic region of the molly and goldfish has previously been demonstrated. Females of these species lack galanin-immunoreactive perikarya in the preoptic nucleus. In contrast, we have found, in female red salmon, galanin-immunoreactive neurons in the magnocellular preoptic nucleus, located far lateral to the preoptic recess, whereas many immunoreactive fibers are present in the preoptic area in both genders. In addition, many immunoreactive neurons have been seen in the nucleus preopticus periventricularis and nucleus lateralis tuberis, also in both sexes. These findings support the notion that galanin may play a gender-specific role in red salmon.

Radioimmunoassay for rat galanin: immunochemical and chromatographic characterization of immunoreactivity in tissue extracts.

Galanin is a regulatory peptide with wide distribution in the central and peripheral nervous system and with numerous biological effects. Several radioimmunoassays based on antisera raised against porcine galanin have been used to measure immunoreactivity in rat tissues. However, considerable lack of parallelism has been observed between the porcine standard and rat tissue extracts, which may decrease the reliability of the quantitative data. The purpose of the present study was therefore to raise antibodies against rat galanin and establish a competitive radioimmunoassay for rat galanin. Two antisera, RatGal4 and RatGal5, were characterized in detail. The homogeneity of the immunoreactive material from several tissues was also investigated with column chromatography. At reverse-phase high-pressure liquid chromatography more than 95% of the immunoreactive material from rat CNS eluted as a single peak in the position of synthetic rat galanin, whereas almost half of the immunoreactive material from the intestine eluted in positions different from the synthetic peptide. Extracts of rat brains as well as jejunum diluted in parallel with the standard curve for both antisera. We conclude that measurements of rat galanin based on these antisera are therefore more reliable than those based on antisera raised against porcine galanin.

Intrinsic choroidal neurons in the duck eye express galanin.

Recently, it has been shown that the choroid of the duck eye harbours approximately 1,000 intrinsic choroidal neurons positive for vasoactive intestinal polypeptide and neuronal nitric oxide synthase. Their connections and functional significance are largely unknown. This study was performed to establish a typical chemical code for these neurons and to define their targets by using immunocytochemistry and confocal laser scanning microscopy. Almost all intrinsic choroidal neurons coexpressed galanin (GAL), vasoactive intestinal polypeptide (VIP), and neuronal nitric oxide synthase (nNOS)/NADPH-diaphorase. A few stained for GAL and/or nNOS only. Among extrinsic ganglia, GAL/VIP/nNOS coexpressing neurons were only found in the pterygopalatine ganglion where they accounted for approximately 30% of the neuronal population. Thus, GAL/VIP/nNOS-positive nerve fibres around branches of the ciliary artery and within the nonvascular smooth muscle stroma of the choroid may originate mainly from intrinsic neurons and to some extent in a subpopulation of pterygopalatine ganglionic neurons exhibiting the same chemical coding. Close contacts of GAL-positive fibres upon intrinsic choroidal neurons may indicate reciprocal connections between them. Thus, intrinsic choroidal neurons may represent peripherally displaced pterygopalatine ganglion neurons forming a local network for regulation of vascular and nonvascular smooth muscle tone in the duck choroid. They may be integrated in the neuronal circuitry controlling intraocular pressure, choroidal thickness, accommodation, and axial bulbus length.