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.

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.

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.

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.

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.

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.

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.

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.

Initial characterization of the transplant of immortalized chromaffin cells for the attenuation of chronic neuropathic pain.

Cultures of embryonic day 17 (E17) rat adrenal and neonatal bovine adrenal cells were conditionally immortalized with the temperature-sensitive allele of SV40 large T antigen (tsTag) and chromaffin cell lines established. Indicative of the adrenal chromaffin phenotype, these cells expressed immunoreactivity (ir) for tyrosine hydroxylase (TH), the first enzyme in the synthetic pathway for catecholamines. At permissive temperature in vitro (33 degrees C), these chromaffin cells are proliferative, have a typical rounded chromaffin-like morphology, and contain detectable TH-ir. At nonpermissive temperature in vitro (39 degrees C), these cells stop proliferating and express increased TH-ir. When these immortalized chromaffin cells were transplanted in the lumbar subarachnoid space of the spinal cord I week after a unilateral chronic constriction injury (CCI) of the rat sciatic nerve, they survived longer than 7 weeks on the pia mater around the spinal cord and continued to express TH-ir. Conversely, grafted chromaffin cells lost Tag-ir after transplant and Tag-ir was undetectible in the grafts after 7 weeks in the subarachnoid space. At no time did the grafts form tumors after transplant into the host animals. These grafted chromaffin cells also expressed immunoreactivities for the other catecholamine-synthesizing enzymes 7 weeks after grafting, including: dopamine-beta-hydroxylase (DbetaH) and phenylethanolamine-N-methyltransferase (PNMT). The grafted cells also expressed detectable immunoreactivities for the opioid met-enkephalin (ENK), the peptide galanin (GAL), and the neurotransmitters y-aminobutyric acid (GABA) and serotonin (5-HT). Furthermore, after transplantation, tactile and cold allodynia and tactile and thermal hyperalgesia induced by CCI were significantly reduced during a 2-8-week period, related to the chromaffin cell transplants. The maximal antinociceptive effect occurred 1-3 weeks after grafting. Control adrenal fibroblasts, similarly immortalized and similarly transplanted after CCI, did not express any of the chromaffin antigenic markers, and fibroblast grafts had no effect on the allodynia and hyperalgesia induced by CCI. These data suggest that embryonic and neonatal chromaffin cells can be conditionally immortalized and will continue to express the phenotype of primary chromaffin cells in vitro and in vivo; grafted cells will ameliorate neuropathic pain after nerve injury and can be used as a homogeneous source to examine the mechanisms by which chromaffin transplants reverse chronic pain. The use of such chromaffin cell lines that are able to deliver antinociceptive molecules in models of chronic pain after nerve and spinal cord injury (SCI) offers a novel approach to pain management.

Expression of galanin immunoreactivity in gonadotropin-releasing hormone neurons in mice: a confocal microscopic study.

The expression of galanin immunoreactivity (galanin-IR) in gonadotropin-releasing hormone (GnRH) neurons was investigated in mice using double label immunohistochemistry combined with confocal laser scanning microscopy. A large proportion of GnRH cells in proestrous mice and very few GnRH cells in male mice exhibited galanin-IR. These results are consistent with earlier reports in rats. Unlike in rats, the proportion of GnRH cells coexpressing galanin in mice was high following ovariectomy (OVX) and the treatment of OVX mice with estrogen decreased the number of GnRH cells with galanin-IR. The GnRH system can be considered more active during proestrous and following OVX since the output of luteinizing hormone is elevated during these phases in females. Since the induction of galanin-IR in GnRH cells is more pronounced in OVX and proestrous mice, the expression of galanin-IR in GnRH cells in mice appears to be an activation-dependent phenomenon rather than a direct effect of estrogen. However, in OVX mice treated with steroids to induce an LH surge the number of GnRH cells with galanin-IR was not proportionately increased. The possible reasons for this discrepancy are also discussed.

VIP-, galanin-, and neuropeptide-Y-immunoreactive fibers in the chicken carotid bodies after various types of denervation.

The chicken carotid body receives numerous branches from the vagus nerve, especially distal (nodose) ganglion, and the recurrent laryngeal nerve. Dense networks of peptidergic nerve fibers immunoreactive for substance P, calcitonin gene-related peptide (CGRP), galanin, vasoactive intestinal peptide (VIP) and neuropeptide Y are distributed in and around the carotid body. Substance-P- and CGRP-immunoreactive fibers projecting to the chicken carotid body mainly come from the vagal ganglia. In the present study, various types of denervation experiments were performed in order to clarify the origins of VIP-, galanin- and neuropeptide-Y-immunoreactive fibers in the chicken carotid bodies. After nodose ganglionectomy, midcervical vagotomy or excision of the recurrent laryngeal nerve, VIP-, galanin- and neuropeptide-Y-immunoreactive fibers were unchanged in the carotid body region. Furthermore, these peptidergic fibers remained unaffected even by removal of the nodose ganglion in conjunction with severance of the recurrent laryngeal nerve that induced a marked decrease in TuJ1-immunoreactive fibers in the carotid body region. VIP-, galanin- and neuropeptide-Y-immunoreactive fibers are densely distributed around the arteries supplying the carotid body in normal chickens. The peptidergic fibers around the arteries were also unaffected after the denervation experiments. However, after removal of the 14th cervical ganglion of the sympathetic trunk, which lies close to the vertebral artery on the root of the brachial plexus and issues prominent branches to the artery, VIP-, galanin- and neuropeptide-Y-immunoreactive fibers almost disappeared in the carotid body region. The ganglion contained many VIP-, galanin- and neuropeptide-Y-immunoreactive neurons. Thus it is clear that VIP-, galanin- and neuropeptide-Y-immunoreactive fibers in the chicken carotid body region are mainly derived from the 14th cervical sympathetic ganglion via the vertebral artery.

Galanin-like immunoreactive nerve fibres in the anterior pituitary of the normal and adrenalectomized rat.

Our previous studies have shown the presence of substantial amounts of substance P and calcitonin gene-related peptide-like immunoreactive nerve fibres in the anterior pituitary of the monkey, dog and rat. Furthermore, synaptic relationships have been demonstrated between these nerve fibres and the gland cells in the dog and rat. The substance P and calcitonin gene-related nerve fibres increase in number following adrenalectomy and ovariectomy, respectively. The present study was aimed to investigate the galanin-containing nerve fibres in the anterior pituitary of normal and adrenalectomized rats. The results showed only a small amount of galanin-like immunoreactive nerve fibres in the normal anterior pituitary, which were present among the gland cells as well as along the blood vessels. Following adrenalectomy, the number of galanin-like immunoreactive nerve fibres increased and ramification appeared more frequently. The results substantiate our hypothesis of a dual neural-humoral regulation of the mammalian anterior pituitary gland.

Colocalization of inhibitory mediators, NO, VIP and galanin, in canine enteric nerves.

The colocalization of three putative inhibitory mediators of enteric nerves, vasoactive intestinal peptide (VIP), galanin (GAL) and nitric oxide synthase (nNOS), was examined in the myenteric plexus of canine antrum, intestine and colon. Many ileal and colonic neurons contained nNOS-immunoreactive (nNOS-IR) activity with some also containing VIP-IR; only a few neurons also contained GAL-IR. Ileal and colonic VIP-IR nerves often appeared to be interneurons innervating nNOS nerves. Many antral neurons contained VIP-IR with nearly all also containing GAL-IR. A few also contained nNOS-IR. The predominance of nNOS-IR neurons relative to VIP-IR and GAL-IR neurons in the ileal and colonic, but not the antral, myenteric plexus is consistent with NO being the primary inhibitory mediator in the intestine but not in the antrum.

Pattern of distribution of neuropeptides in the camel lacrimal gland.

The pattern of distribution of neuropeptides, including neuropeptide-Y (NPY), vasoactive intestinal polypeptide (VIP), neurotensin (NT), serotonin (5-HT), galanin (GAL), leucine-enkephalin (LEU-ENK) and calcitoningene-related-peptide (CGRP), in the nerves of the camel lacrimal gland was investigated using immunohistochemical techniques. Fresh lacrimal gland segments, obtained from adult camels slaughtered in the local abattoir, were used for the immunohistochemical techniques. NPY and LEU-ENK immunoreactivity was observed in the nerve cell bodies and nerve fibers of the camel lacrimal gland. VIP, GAL and CGRP were demonstrated predominantly in fine varicose nerve fibers lying on the basolateral surfaces of the lacrimal acinar cells. NT and 5-HT were identified mainly in neurons situated in the periacinar regions, close to the basal surfaces of the acinar cells. It is concluded that the camel lacrimal nerves contain several neuropeptides including NPY, VIP, NT, 5-HT, GAL, LEU-ENK and CGRP which may modulate lacrimal fluid and protein secretion.

Galanin inhibits ACTH release in vitro and can be demonstrated immunocytochemically in dispersed corticotrophs.

Double labeling of dispersed anterior pituitary cells revealed the coexistence of galanin immunoreactivity with adrenocorticotropic hormone (ACTH) as well as prolactin. However, only laser confocal microscopy showed three different areas of immunoreactivity within the corticotroph cytoplasm, two of them for ACTH and galanin separately and the third containing both immunoreactivities. To determine a possible relation between ACTH cells and galanin, 4-day cultures of anterior pituitary cells from female rats were examined by cell blot assay, and they showed ACTH release inhibition by 10(-6)M galanin. Furthermore, after 17 beta-estradiol treatment to maximize lactotroph galanin release in vitro, the cell blotting-assessed secretory level of corticotroph cells was very similar to that of cells in the presence of 10(-6)M galanin. In fact, immunoneutralization with galanin antiserum quenched the inhibitory effect of galanin on ACTH secretion. Our study suggests that locally produced galanin can modulate corticotropin release.

Enteric neuropeptides in streptozotocin-diabetic rats; effects of insulin and aldose reductase inhibition.

The aim of the present study was to determine whether diabetes-induced changes in the distribution of enteric neuropeptides, could be prevented in 12-week streptozotocin-diabetic rats, by rigorous control of glycaemia, using daily adminstration of insulin, or an aldose reductase inhibitor (ponalrestat). The pattern of distribution of nerve fibres and cell bodies, containing immunoreactive vasoactive intestinal polypeptide (VIP), galanin (GAL), calcitonin gene-related peptide (CGRP) and substance P was examined in the myenteric plexus of ileum from control, untreated diabetic, insulin-treated diabetic and aldose reductase inhibitor-treated diabetic rats. The increase in VIP- and GAL-like immunoreactivity, seen in the myenteric plexus of untreated diabetic rat ileum, was not present in the myenteric plexus of ileum from insulin- and aldose reductase inhibitor-treated diabetic rats. With CGRP-like immunoreactive fibres, there was a clear decrease in the ileum of untreated diabetic rats. This was prevented by insulin treatment, but aldose reductase inhibitor treatment had no effect. No alterations in substance P-like immunoreactivity were seen in the myenteric plexus of ileum from any of the groups investigated. Generally, the similarity of effect of ponalrestat and insulin on VIP and galanin expression in this study supports a primary effect of insulin via glycaemic control. The dissimilarity of the effect of the two treatments on CGRP expression may imply a neurotrophic effect of insulin, although there are certainly consequences of hyperglycaemia other than exaggerated flux through the polyol pathway.