RESUMEN
Substance P and hemokinin-1 were predominantly examined by immunoassays with their limitation to differentiate appropriately between both peptides. The use of liquid chromatography coupled with tandem mass spectrometry is a promising, highly selective alternative. Adsorption processes have been identified in preliminary experiments to play a crucial role in the loss of mass spectrometry intensity of both peptides. Therefore, a design of experiments concept was created to minimize nonspecific peptide adsorption. For this purpose, the most critical influencing parameters-(1) the composition of the injection solvent as well as (2) the most suitable container material-were systematically and concordantly investigated. The addition of modifiers, such as formic acid, dimethyl sulfoxide, and organic solvents, to the injection solvent led to a substantial gain of intensity of substance P and hemokinin-1 compared to the start gradient as an injection solvent. Furthermore, the systematic investigation underlined the high impact of the container material, demonstrating polypropylene as the most favorable material. A conjoint injection solvent optimum was found to determine both peptides simultaneously by the conduction of a sweet-spot analysis. The experimental design substantially reduced nonspecific peptide adsorption and enabled the simultaneous and selective determination of endogenous substance P and hemokinin-1 plasma levels.
Asunto(s)
Cromatografía Liquida/métodos , Péptidos/química , Sustancia P/aislamiento & purificación , Taquicininas/aislamiento & purificación , Espectrometría de Masas en Tándem/métodos , Adsorción , Cromatografía Liquida/instrumentación , Proyectos de Investigación , Sustancia P/análisis , Espectrometría de Masas en Tándem/instrumentaciónRESUMEN
Neurocysticercosis (NCC), a helminth infection of the brain, is a major cause of seizures. The mediators responsible for seizures in NCC are unknown, and their management remains controversial. Substance P (SP) is a neuropeptide produced by neurons, endothelial cells and immunocytes. The current studies examined the hypothesis that SP mediates seizures in NCC. We demonstrated by immunostaining that 5 of 5 brain biopsies from NCC patients contained substance P (SP)-positive (+) cells adjacent to but not distant from degenerating worms; no SP+ cells were detected in uninfected brains. In a rodent model of NCC, seizures were induced after intrahippocampal injection of SP alone or after injection of extracts of cysticercosis granuloma obtained from infected wild type (WT), but not from infected SP precursor-deficient mice. Seizure activity correlated with SP levels within WT granuloma extracts and was prevented by intrahippocampal pre-injection of SP receptor antagonist. Furthermore, extracts of granulomas from WT mice caused seizures when injected into the hippocampus of WT mice, but not when injected into SP receptor (NK1R) deficient mice. These findings indicate that SP causes seizures in NCC, and, suggests that seizures in NCC in humans may be prevented and/or treated with SP-receptor antagonists.
Asunto(s)
Encefalopatías/complicaciones , Granuloma/parasitología , Neurocisticercosis/complicaciones , Convulsiones/etiología , Sustancia P/metabolismo , Animales , Encéfalo/patología , Encefalopatías/parasitología , Encefalopatías/patología , Modelos Animales de Enfermedad , Femenino , Eliminación de Gen , Granuloma/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Neurocisticercosis/parasitología , Neurocisticercosis/patología , Antagonistas del Receptor de Neuroquinina-1/uso terapéutico , Ratas , Ratas Sprague-Dawley , Receptores de Neuroquinina-1/genética , Receptores de Neuroquinina-1/fisiología , Convulsiones/tratamiento farmacológico , Convulsiones/parasitología , Convulsiones/prevención & control , Sustancia P/análisis , Sustancia P/antagonistas & inhibidores , Sustancia P/aislamiento & purificación , Taenia/fisiologíaRESUMEN
The neurokinin-1 receptor plays a profound role in inflammatory processes and is involved in immune cell differentiation, cytokine release, and mast cell activation. Due to their similar peptide structures, the neurokinin-1 receptor does not discriminate between the endogenous ligands substance P (SP) and human hemokinin-1 (hHK-1), which both demonstrate biological receptor affinity. In addition, due to cross-reactivity, the current bioanalytical method of choice-immunoassays-also displays limitations in differentiating between these peptides. Thus, a recently developed mass spectrometric assay was utilized for the selective quantification of SP and hHK-1 in various biofluids and tissue. By applying the sample processing protocols developed, SP was quantified in porcine brain tissue (4.49 ± 0.53 nM), human saliva (113.3 ± 67.0 pM), and human seminal fluid (0.52 ± 0.15 nM) by mass spectrometric analysis. As previously reported, neither SP nor hHK-1 could be detected in human plasma by mass spectrometry. Comparison with analysis using a commercial immunoassay of the same plasma sample revealed SP like-immunoreactivity concentrations of 37.1-178.0 pM. The previously reported carboxylic acid of SP, whose identity was confirmed by high-resolution mass spectrometric analysis, did not show cross-reactivity in the applied immunoassay and did not contribute to SP-like immunoreactivity results. Subsequent compound discovery of the immunocaptured substance indicated the presence of a precursor of SP as possible cross-reactor in human plasma samples. The found cross-reactivity might be the cause for the high variance of SP plasma levels in former determinations.
Asunto(s)
Inflamación/genética , Receptores de Neuroquinina-1/aislamiento & purificación , Sustancia P/aislamiento & purificación , Taquicininas/aislamiento & purificación , Animales , Líquidos Corporales/química , Encéfalo/metabolismo , Humanos , Inflamación/metabolismo , Inflamación/patología , Espectrometría de Masas , Péptidos/química , Péptidos/aislamiento & purificación , Receptores de Neuroquinina-1/química , Receptores de Neuroquinina-1/genética , Saliva/química , Semen/química , Sustancia P/química , Sustancia P/genética , Porcinos , Taquicininas/química , Taquicininas/genéticaRESUMEN
Peripheral neuropathy is induced by multiple doses of oxaliplatin and interferes with the clinical utility of the drug in patients with colorectal cancer. In this study, we sought to determine whether cell loss or selective neuronal damage was the basis for the peripheral neuropathy caused by oxaliplatin. Adult female rats were given 1.85 mg/kg oxaliplatin twice per week for 8 weeks. Nerve conduction and L5 dorsal root ganglia (DRG) were studied 1 week after the completion of all treatment. No mortality occurred during oxaliplatin treatment, but the rate of body weight gain was reduced compared to age-matched vehicle-treated controls. Oxaliplatin slowed conduction velocity and delayed conduction times in peripheral sensory nerves, without affecting central or motor nerve conduction. In L5 DRG, total numbers of neurons were unchanged by oxaliplatin, but there were significant reductions in neuronal size distribution, ganglion volume, average cell size and the relative frequency of large cells. In addition, the relative frequency of small DRG cells was increased by oxaliplatin. Oxaliplatin significantly altered the size distribution and average cell body area of the predominantly large parvalbumin-immunoreactive DRG neurons without affecting the frequency of parvalbumin staining. On the contrary, neither the staining frequency nor the size distribution of the predominantly small substance P-immunoreactive DRG neurons was changed by oxaliplatin. In conclusion, oxaliplatin causes selective atrophy of a subpopulation of DRG neurons with predominantly large parvalbumin-expressing cells without inducing neuronal loss. Because DRG cell body size and axonal conduction velocity are positively correlated, neuronal atrophy may be the morphological basis for the development of decreased sensory nerve conduction velocity that characterizes oxaliplatin-induced peripheral neuropathy.
Asunto(s)
Antineoplásicos/toxicidad , Ganglios Espinales/efectos de los fármacos , Conducción Nerviosa/efectos de los fármacos , Compuestos Organoplatinos/toxicidad , Enfermedades del Sistema Nervioso Periférico/inducido químicamente , Animales , Femenino , Oxaliplatino , Parvalbúminas/aislamiento & purificación , Enfermedades del Sistema Nervioso Periférico/patología , Ratas , Ratas Wistar , Sustancia P/aislamiento & purificaciónRESUMEN
The aim of this study was to compare immunoreactivities for substance P with other enteric neuropeptides and GAP-43, a general marker for enteric nerves, in normal human colon and in different stages of ulcerative colitis. Tissue samples from normal colon and regions of ulcerative colitis colon were obtained at surgery and immunostained for substance P, vasoactive intestinal polypeptide (VIP), somatostatin, calcitonin gene-related peptide (CGRP), enkephalin, galanin, GAP-43, and neuron-specific enolase (NSE). Visual examination and semiquantitative analysis revealed a clear increase in the immunoreactivity for substance P in ulcerative colitis, whereas no differences were observed in the distribution of the other peptides. Therefore, quantitative analysis was performed only for substance P immunoreactivity in the lamina propria, circular muscle layer, and myenteric ganglia. In the lamina propria, the score of total intensity of substance P immunoreactivity was 0.55 +/- 0.15 (mean +/- SEM) in normal colon, 1.30 +/- 0.35 (p = 0.087) in least affected colon, and 2.22 +/- 0.28 (p < 0.001) in moderately affected colon, whereas no significant differences were observed in immunoreactivities for GAP-43. Similar results were obtained for the mean substance P- or GAP-43-immunoreactive area. In the circular muscle layer, the number, density, total intensity, and perimeter of substance P- and GAP-43-immunoreactive fibers were essentially similar in normal colon, and in mild or moderately affected colon. We conclude that ulcerative colitis does not change the density of gut innervation as a whole. However, the density of substance P-containing nerves is specifically increased, probably due to increased peptide synthesis leading to better visibility of the fibers.
Asunto(s)
Colitis Ulcerosa/patología , Colon/patología , Sistema Nervioso Entérico/patología , Proteína GAP-43/aislamiento & purificación , Sustancia P/aislamiento & purificación , Adulto , Anciano , Anciano de 80 o más Años , Colon/inervación , Ganglios Autónomos/patología , Humanos , Inmunohistoquímica , Persona de Mediana Edad , Plexo Mientérico/patología , Distribución TisularRESUMEN
1. The effect of an acid extract of the carp intestinal bulb (ECI) on guinea-pig ileum longitudinal smooth muscle (GPLM) and carp intestinal bulb longitudinal smooth muscle (CIBLM) was examined. 2. ECI caused a concentration-dependent contraction of GPLM and CIBLM. This ECI-induced response was reduced by atropine to 30-40% of the control, indicating that part of the contracting activity of ECI is attributable to acetylcholine. The atropine-resistant contracting activity of ECI was not mediated by histamine, 5-hydroxytryptamine, ATP, ADP, angiotensin II, neurotensin, vasoactive intestinal peptide or an opioid peptide. 3. The active material mediating the atropine-resistant contracting activity is probably a peptide, because the contraction in response to ECI was abolished on incubation with pepsin or alpha-chymotrypsin. 4. [D-Pro2, D-Trp7,9]-substance P, [D-Pro4, D-Trp7,9]-substance P (4-11) decreased the atropine-resistant contracting activity of ECI as did desensitization induced by substance P. 5. On a Sephadex G 25 column, the active material was eluted as one peak. The active fractions were pooled and then applied to another Sephadex G25 column to compare the Ve/Vo value for the active material with those for peptides of known molecular weights. The molecular weight of the active material was estimated to be 1200-1700 (1410 +/- 70, n = 6). 6. The results indicate the presence of a substance P-like peptide in the carp intestinal bulb.
Asunto(s)
Carpas/fisiología , Cyprinidae/fisiología , Intestinos/fisiología , Sustancia P/aislamiento & purificación , Extractos de Tejidos/fisiología , Acetilcolina/farmacología , Adenosina Trifosfato/farmacología , Angiotensina II/farmacología , Animales , Cromatografía en Gel , Quimotripsina/farmacología , Femenino , Histamina/farmacología , Técnicas In Vitro , Masculino , Contracción Muscular/efectos de los fármacos , Músculo Liso/efectos de los fármacos , Neurotensina/farmacología , Pepsina A/farmacología , Serotonina/farmacología , Sustancia P/farmacología , Extractos de Tejidos/análisisRESUMEN
An active gelatinase has been purified from the conditioned medium of granulation tissue culture formed by carrageenin injection in rats. The purified gelatinase gave a single band corresponding to a M(r) of 57 kDa on both sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and SDS-gelatin PAGE. The granulation tissue-derived gelatinase selectively cleaved the Gln6-Phe7 bond of substance P (SP) with a Km of 0.17 mM and a Vmax of 0.027 nmol SP7-11/min/micrograms protein, resulting in the generation of biologically inactive fragments, SP1-6 and SP7-11. Our data suggest that the gelatinase produced by granulation tissue participates in the inactivation of SP in the inflammatory site.
Asunto(s)
Tejido de Granulación/enzimología , Inhibidores de Proteasas/metabolismo , Sustancia P/antagonistas & inhibidores , Animales , Captopril/farmacología , Técnicas de Cultivo , Electroforesis en Gel de Poliacrilamida , Gelatina/metabolismo , Hidrólisis , Cinética , Masculino , Fragmentos de Péptidos/metabolismo , Ratas , Ratas Wistar , Sustancia P/análogos & derivados , Sustancia P/aislamiento & purificación , Sustancia P/metabolismo , Tiorfan/farmacologíaRESUMEN
The neuron morphology and distribution of four putative transmitters were investigated in the myenteric plexus of frog (Rana esculenta) midgut. The gross morphology was revealed by NADH-diaphorase histochemistry, and the shape of the neurons by silver impregnation. Nerve cells had heterogeneous distribution: they either formed ganglia or placed as solitary neurons in the duodenum, while in the rest of the midgut only solitary neurons were observed. Three morphologically distinct cell types were revealed by silver impregnation: mainly type I and type II neurons cells were seen in the duodenum, while the rest of the intestine contained type II and III cells. Catecholamine fluorescence was revealed in nerve fibres in the duodenum, while few small nerve cells were observed in the small intestinal region. Acetylcholinesterase histochemistry showed strongly reactive nerve cells that were associated with the main fibre bundles in the duodenum. Only longitudinally oriented fibres and occasionally stained neurons were seen in the small intestine. Substance P immunocytochemistry revealed an extensive plexus, which contained a moderate number of stained perikarya in the full length of the midgut. Gamma-aminobutyric acid showed non-uniform distribution in the two parts of the midgut: a stronger and more regular fibre staining was found in the duodenum then in the rest of the intestine. Ultrastructural observations demonstrated that intrinsic neurons received synaptic inputs from the profiles contained agranular vesicles, while "P"-type profiles established close contacts with neurons. Both profile types formed close contacts with the smooth muscle cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Intestino Delgado/anatomía & histología , Plexo Mientérico/química , Neuronas/química , Rana esculenta/anatomía & histología , Acetilcolinesterasa/aislamiento & purificación , Animales , Biomarcadores , Catecolaminas/aislamiento & purificación , Dihidrolipoamida Deshidrogenasa/aislamiento & purificación , Femenino , Histocitoquímica , Intestino Delgado/química , Intestino Delgado/inervación , Masculino , Plexo Mientérico/anatomía & histología , Plexo Mientérico/ultraestructura , Neuronas/citología , Neuronas/ultraestructura , Tinción con Nitrato de Plata , Sustancia P/aislamiento & purificación , Ácido gamma-Aminobutírico/aislamiento & purificaciónRESUMEN
The occurrence and distribution of the neurohormone substance P in the urogenital tract suggest a role for substance P in regulation of blood flow, smooth-muscle activity, and sensation. The distribution and pharmacologic effects of substance P in the brain suggest an involvement of substance P in regulating the release of gonadotropins and prolactin from the pituitary and in the gonadal-hypothalamic feedback. As substance P is also present in endocrine-like cells of the diffuse endocrine system in the urogenital tract, it may serve as a tumor marker for carcinoid and related tumors, particularly in the ovary and the uterine cervix. This review is limited to the distribution and effects of substance P in the female genital tract and its relevance in female reproduction.
Asunto(s)
Genitales Femeninos/fisiología , Sustancia P/fisiología , Animales , Gatos , Glándulas Endocrinas/embriología , Trompas Uterinas/análisis , Trompas Uterinas/fisiología , Femenino , Genitales Femeninos/análisis , Cobayas , Humanos , Sistema Hipotálamo-Hipofisario/fisiología , Trabajo de Parto , Ovario/análisis , Sistema Hipófiso-Suprarrenal/fisiología , Embarazo , Prurito Vulvar/fisiopatología , Ratas , Sustancia P/aislamiento & purificación , Útero/análisis , Útero/fisiología , Vagina/análisisRESUMEN
Substance P-like immunoreactivity (SP-LI) as measured by RIA was found to be present in a variety of submammalian species and invertebrates. We analyzed this SP-LI in extracts from submammalian species by high performance liquid chromatography. The following species were investigated for the presence of SP-LI (RIA) which was further characterized by subsequent HPLC (investigated areas in parentheses): Hagfish (brain plus spinal cord), (brain, intestine, skin), frog (brain, intestine), turtle (brain, intestine), lizard (brain, intestine, skin) and mouse (spinal cord). RIA alone was performed in extracts from branchiostoma and cricket. The concentrations of SP-LI in brain, spinal cord and intestine of different submammalian species except branchiostoma brain and intestine and turtle brain, were in a similar range (2.1-5.3 fmol/mg in the brain, 0.2-2.0 fmol/mg in the spinal cord, 0.3-4.2 fmol/mg in the intestine). In the turtle brain, extremely high SP-LI concentrations (210 fmol/mg) were found, whereas brain and intestine of branchiostoma contained very little SP-LI (0.1 fmol/mg). In the skin of different species, SP-LI concentrations varied from 0.04 fmol/mg (trout) to 2.0 fmol/mg (lizard). In the cricket, high SP-LI concentrations were found in the cerebral ganglion (15 fmol/mg protein) and in the subesophageal ganglion (27 fmol/mg protein). HPLC analysis of extracts showed that all tissues investigated contained a substance which co-eluted with synthetic SP, and in most tissues a peak was present which co-eluted with SP sulfoxide. Only in mouse spinal cord, trout brain and hagfish brain were these the only peaks.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Insectos/análisis , Sustancia P/aislamiento & purificación , Urocordados/fisiología , Vertebrados/fisiología , Secuencia de Aminoácidos , Animales , Evolución Biológica , Cromatografía Líquida de Alta Presión , Radioinmunoensayo , Sustancia P/inmunologíaRESUMEN
The concentrations of tachykinins in rat spinal cord and dorsal root ganglia (DRGs) were measured using a combination of high performance liquid chromatography (HPLC) and radioimmunoassays (RIAs). Substance P-like immunoreactivity (SPLI) was found to be significantly higher than either substance K-like immunoreactivity (SKLI) or neuromedin K-like immunoreactivity (NMKLI) in both tissues. In the spinal cord, the concentration of SKLI was comparable to that of NMKLI. In DRGs, NMKLI is present at concentrations much lower than those of SKLI or SPLI. In addition to immunoreactive components co-eluting with the three mammalian tachykinins SP, SK and NMK, analyses using reverse-phase HPLC revealed an immunoreactive peak co-eluting with the C-terminal octapeptide of SK (SK3-10), and a yet to be identified peak eluting before SK. This study also demonstrates the use of a novel and highly specific RIA for NMK to measure NMKLI without the need of reverse-phase HPLC.
Asunto(s)
Ganglios Espinales/análisis , Médula Espinal/análisis , Taquicininas/aislamiento & purificación , Animales , Cromatografía Líquida de Alta Presión/métodos , Reacciones Cruzadas , Femenino , Masculino , Radioinmunoensayo/métodos , Ratas , Ratas Endogámicas , Sustancia P/aislamiento & purificación , Taquicininas/inmunologíaRESUMEN
The medial septal nucleus provides one of the major afferents to the hippocampal formation. The two major types of neurons present in the medial septum are cholinergic and GABAergic, but other types of neurons are also present. A small population of substance P-containing neurons is present along the border between the medial and lateral septum, but it is unclear whether these project to the hippocampus. The present study, by employing both anterograde and retrograde tracing techniques, combined with immunocytochemistry for substance P, provides direct morphological evidence for a substance P projection from the lateral region of medial septum to a portion of CA2/3a, which is restricted to the mid-septotemporal portion of the hippocampus.
Asunto(s)
Hipocampo/anatomía & histología , Hipotálamo Medio/anatomía & histología , Sustancia P/aislamiento & purificación , Animales , Hipotálamo Medio/química , Inmunohistoquímica , Neuronas/química , Neuronas/citología , Ratas , Ratas EndogámicasRESUMEN
Peripheral injury produces long term changes in peptide content in dorsal root ganglion (DRG) cells that contribute to the inflammatory process in the periphery and neuronal plasticity in the spinal cord. We report here the proportion of colonic afferents labeled for calcitonin gene-related peptide (CGRP), substance P (SP) or somatostatin (Som) in the T13-L2 and L6-S2 DRG and changes in the percentage of SP or CGRP labeled afferents 6, 24, and 72 h following induction of experimental colitis. Following injection of fluorogold (FG) into the descending colon, significantly more FG labeled DRG cells were observed in the T13-L2 than L6-S2 DRG. In noninflamed rats, in both spinal regions, 60-70% of the colonic afferents that were labeled with FG were double labeled for SP. Similar results were obtained when double labeling for CGRP. Only 20-30% of the FG labeled afferents were double labeled for Som. Following experimental colitis induced by intracolonic zymosan, there was a significant decrease in the percentage of cells double labeled for SP in the T13-L2 and L6-S2 DRG at 6, 24, and 72 h. The percentage of CGRP double labeled cells was decreased in the T13-L2 DRG at all time points, but only at 24 h in the L6-S2 DRG. The cell bodies of CGRP labeled colonic afferents were significantly larger than SP or Som in control rats. Inflammation did not affect the mean size of the double labeled cells. These results suggest that colonic inflammation increases SP and CGRP release in the spinal cord and the colon that is manifest as a decrease in peptide content in the cell bodies of the colonic afferents during the first 72 h following injury.
Asunto(s)
Dolor Abdominal/etiología , Colitis/inducido químicamente , Ganglios Espinales/química , Neuropéptidos/aislamiento & purificación , Aferentes Viscerales/química , Animales , Péptido Relacionado con Gen de Calcitonina/aislamiento & purificación , Colon/inervación , Región Lumbosacra , Masculino , Ratas , Ratas Sprague-Dawley , Sustancia P/aislamiento & purificación , TóraxRESUMEN
The feasibility of extracting neuropeptides from rat knee joints for quantitation by radioimmunoassay was tested. The investigation, based on 25 adult Lewis rats, focused on substance P, calcitonin gene-related peptide, neuropeptide Y, and vasoactive intestinal polypeptide. The relative recovery of the peptides in different extraction media was assessed Both knee joints including the articulating epiphysis were dissected and cut into small pieces. The series was divided into five subgroups, 10 joints in each, for extraction in five different media: 1) 1 M acetic acid in 4% EDTA, 2) 2 M acetic acid in 4% EDTA, 3) neutral water in 4% EDTA, 4) 2 M acetic acid in 4% EDTA and 95% alcohol, and 5) 2 M acetic acid without EDTA. Measureable concentrations of the four neuropeptides were reproducibly assessed by RIA. Although all extraction media provided measurable concentrations, 2 M acetic acid in 4% EDTA was found to give the highest overall yield of the four neuropeptides analyzed. Reverse-phase HPLC confirmed that the immunoreactivities assessed by RIA corresponded to the four neuropeptides of interest. Experimental and clinical evidence suggest a neurogenic involvement in the pathophysiology of inflammatory joint disease, e.g., rheumatoid arthritis. The extraction procedure described offers a means of determining neuropeptide concentrations in joint tissue under normal and pathologic conditions by RIA.
Asunto(s)
Articulación de la Rodilla/química , Neuropéptidos/aislamiento & purificación , Radioinmunoensayo/métodos , Animales , Artritis Experimental/metabolismo , Péptido Relacionado con Gen de Calcitonina/inmunología , Péptido Relacionado con Gen de Calcitonina/aislamiento & purificación , Cromatografía Líquida de Alta Presión , Femenino , Masculino , Neuropéptido Y/inmunología , Neuropéptido Y/aislamiento & purificación , Neuropéptidos/inmunología , Ratas , Sustancia P/inmunología , Sustancia P/aislamiento & purificación , Extractos de Tejidos , Péptido Intestinal Vasoactivo/inmunología , Péptido Intestinal Vasoactivo/aislamiento & purificaciónRESUMEN
The distribution of calcitonin gene-related peptide (CGRP), substance P/tachykinin (SP/TK), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY) and gastrin-releasing peptide (GRP) immunreactivities (IR) in the rat pancreas was investigated using radioimmunoassay and immunohistochemistry. CGRP, NPY and VIP tissue contents are much higher than GRP and SP/TK concentrations. Peptide-containing nerves are distributed to both the exocrine and endocrine pancreas. However, differences exist in terms of density and targets of innervation for each peptidergic system. In the acini and through the stroma, fibers IR for CGRP, NPY and VIP are greater than GRP- and SP/TK-containing processes. The vasculature is supplied by a prominent NPY, CGRP and, to a lesser extent, SP/TK innervation. VIP-IR is found occasionally, and GRP-IR is never detected, in fibers associated with blood vessels. Around ducts, CGRP- and NPY-positive neurites are greater than SP/TK- greater than or equal to VIP-IR fibers, whereas GRP-containing nerves are not visualized. In the islets, the density of peptidergic nerves is: VIP-, GRP- greater than or equal to CGRP-IR greater than NPY or SP/TK. In intrapancreatic ganglia. VIP- and, to a lesser extent, NPY-IRs are found in numerous neuronal cell bodies and in nerve fibers; GRP-IR is present in numerous nerve processes and in few cell bodies; CGRP- and SP/TK-IRs are detected only in fibers wrapping around unlabeled ganglion cells. The majority of CGRP-IR fibers contain SP/TK-IR. The existence of differential patterns of peptidergic nerves suggests that peptides exert their effects on pancreatic functions via different pathways.
Asunto(s)
Islotes Pancreáticos/inervación , Tejido Nervioso/química , Neuropéptidos/aislamiento & purificación , Páncreas/inervación , Animales , Péptido Relacionado con Gen de Calcitonina/aislamiento & purificación , Ganglios/química , Péptido Liberador de Gastrina , Inmunohistoquímica , Masculino , Neuropéptido Y/aislamiento & purificación , Péptidos/aislamiento & purificación , Radioinmunoensayo , Ratas , Ratas Endogámicas , Sustancia P/aislamiento & purificación , Péptido Intestinal Vasoactivo/aislamiento & purificaciónRESUMEN
The medial nucleus of the amygdala (Me), bed nucleus of the stria terminalis (BNST), and medial preoptic area (MPOA) regulate copulation in the male hamster. The present study identified neuropeptide Y-immunoreactive (NPY-IR) neurons in the BNST and Me with the greatest concentration in the posteromedial and posteriordorsal subdivisions of these nuclei, respectively. NPY-IR filters are found in all three nuclei with dense plexi of NPY-IR varicosities in the most medial subdivisions. Substance P neurons are also densely concentrated in the posterior BNST and Me; however, no neurons contained both peptides. Thus, NPY and substance P neurons comprise two distinct populations within the BNST and Me of the hamster.
Asunto(s)
Sistema Límbico/química , Neuronas/química , Neuropéptido Y/aislamiento & purificación , Sustancia P/aislamiento & purificación , Amígdala del Cerebelo/química , Animales , Copulación/fisiología , Cricetinae , Inmunohistoquímica , Masculino , Mesocricetus , Área Preóptica/química , Núcleos Talámicos/química , Distribución TisularRESUMEN
In this study, Met-enkephalin (Met-enk), substance P (SP) and tyrosine hydroxylase (TH) immunostaining was assessed in caudate nucleus biopsies from 15 Parkinson's disease patients who were treated surgically. According to the combination of changes in Met-enk, SP and TH immunostaining, several subgroups of parkinsonian patients were disclosed. Group I: Patients showing low SP and normal Met-enk immunostaining, and variably reduced TH immunoreactivity. Group II: both SP and Met-enk immunostaining were apparently of normal intensity in these PD patients, but they showed the greatest decrease in TH labeling. Group III: PD patients that showed normal SP, very low Met-enk and variably reduced TH immunostaining. Low Met-enk immunostaining tended to correlate with the severity of the disease as judged by higher Unified Parkinson's disease Rating Scale and gait scores. These results suggest that different neurochemical phenotypes may exist among Parkinson's disease patients. Peptidergic deficits should be taken into account for therapeutic intervention.
Asunto(s)
Núcleo Caudado/química , Encefalina Metionina/aislamiento & purificación , Enfermedad de Parkinson/clasificación , Sustancia P/aislamiento & purificación , Anciano , Análisis de Varianza , Biopsia , Núcleo Caudado/anatomía & histología , Núcleo Caudado/enzimología , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Destreza Motora , Estadísticas no Paramétricas , Tirosina 3-Monooxigenasa/aislamiento & purificaciónRESUMEN
The coexistence of neurokinin A (NKA) with substance P (SP) and serotonin (5-HT) in ventral medullary neurons of the parapyramidal region and nucleus raphe pallidus of the rat was studied using multiple immunofluorescence labeling. Nearly all of the NKA-immunoreactive (IR) cells in the parapyramidal region and raphe pallidus were SP-IR nd 5-HT-IR, whereas about 70% of the SP-IR neurons and about 60% of the 5-HT-IR neurons contained NKA-IR. There were no apparent differences in the patterns of coexistence between parapyramidal and raphe pallidus neurons. NKA-IR neurons, which colocalized SP-IR and 5-HT-IR, were studied for projections to the lumbar and thoracic spinal cord by use of retrograde transport of fluorescent tracer. Whereas about 50% of the retrogradely labeled neurons of the parapyramidal region and raphe pallidus contained NKA-IR, nearly all of the NKA-IR neurons projected to the thoracic and lumbar spinal cord. In addition, some NKA-IR neurons in the ventral medulla were retrogradely labeled with tracer from localized injections into the thoracic intermediolateral cell column. In summary, this study demonstrated that NKA-IR is colocalized with SP-IR in bulbospinal serotonergic neurons of the parapyramidal region and raphe pallidus, which are known to regulate sensory, motor, and autonomic activities of the spinal cord.
Asunto(s)
Sistema Nervioso Central/química , Neuroquinina A/aislamiento & purificación , Neuronas/química , Serotonina/aislamiento & purificación , Sustancia P/aislamiento & purificación , Animales , Sistema Nervioso Central/anatomía & histología , Inmunohistoquímica , Masculino , Bulbo Raquídeo/química , Neuroquinina A/inmunología , Tractos Piramidales/química , Núcleos del Rafe/química , Ratas , Ratas Sprague-Dawley , Serotonina/inmunología , Sustancia P/inmunologíaRESUMEN
The central nucleus of the amygdala, bed nucleus of the stria terminalis, and central gray are important components of the neural circuitry responsible for autonomic and behavioral responses to threatening or stressful stimuli. Neurons of the amygdala and bed nucleus of the stria terminalis that project to the midbrain central gray were tested for the presence of peptide immunoreactivity. To accomplish this aim, a combined immunohistochemical and retrograde tracing technique was used. Maximal retrograde labeling was observed in the amygdala and bed nucleus of the stria terminalis after injections of retrograde tracer into the caudal ventrolateral midbrain central gray. The majority of the retrogradely labeled neurons in the amygdala were located in the medial central nucleus, although many neurons were also observed in the lateral subdivision of the central nucleus. Most of the retrogradely labeled neurons in the BST were located in the ventral and posterior lateral subdivisions, although cells were also observed in most other subdivisions. Retrogradely labeled neurotensin, corticotropin releasing factor (CRF), and somatostatin neurons were mainly observed in the lateral central nucleus and the dorsal lateral BST. Retrogradely labeled substance P-immunoreactive cells were found in the medial central nucleus and the posterior and ventral lateral BST. Enkephalin-immunoreactive retrogradely labeled cells were not observed in the amygdala or bed nucleus of the stria terminalis. A few cells in the hypothalamus (paraventricular and lateral hypothalamic nuclei) that project to the central gray also contained CRF and neurotensin immunoreactivity. The results suggest the amygdala and the bed nucleus of the stria terminalis are a major forebrain source of CRF, neurotensin, somatostatin, and substance P terminals in the midbrain central gray.
Asunto(s)
Amígdala del Cerebelo/anatomía & histología , Sustancia Gris Periacueductal/anatomía & histología , Prosencéfalo/anatomía & histología , Tálamo/anatomía & histología , Animales , Hormona Liberadora de Corticotropina/aislamiento & purificación , Encefalinas/aislamiento & purificación , Técnica del Anticuerpo Fluorescente , Inmunohistoquímica/métodos , Masculino , Neuronas/química , Neuropéptidos/aislamiento & purificación , Neurotensina/aislamiento & purificación , Ratas , Somatostatina/aislamiento & purificación , Sustancia P/aislamiento & purificaciónRESUMEN
Gamma(glutamyl5)spermine derivative of substance P (Spm-SP) was synthesized in vitro in the presence of purified guinea pig liver transglutaminase and Ca2+. The spermine adduct of the neuropeptide was purified by HPLC on a reversed-phase column and characterized by fast atom bombardment mass spectrometry. The biological activities of Spm-SP were tested by assaying, in comparison with substance P, its ability to induce both the contractions of smooth muscle in vitro and the edema formation in vivo. Spm-SP was shown not to elicit contractile responses in the isolated rat stomach strip and duodenum and not to antagonize the spasmogenic effect evoked by the native neuropeptide. Furthermore, Spm-SP was unable, when administered into rats by plantar injection, either to provoke an acute inflammatory response in the hind limb or to antagonize the edema formation induced by a concurrent administration of substance P. These results indicate that the introduction of a large size hydrophilic moiety at the glutamine5 level negatively affects the ability of the neuropeptide to bind to its receptor(s), thus supporting the view that the hydrophobic middle portion of substance P plays a key role in receptor recognition.