Bombesin-like Immunoreactivity Research Articles

Distribution and molecular forms of immunoreactive bombesin in the ovine median eminence.

The distribution and molecular forms of bombesin-like immunoreactivity (BLI) were determined in the ovine median eminence using a new, C-terminally directed antiserum. BLI was confined to neurons of the external zone of the median eminence, near small blood vessels, many of which also were immunoreactive for CRF. Each median eminence contained about 10 pmol of BLI (533 pmol/g tissue). Gel filtration and reverse-phase high pressure liquid chromatography (HPLC) demonstrated the existence of two molecular forms of BLI, which co-eluted with porcine gastrin releasing peptide (GRP)1-27 and GRP18-27 in a molar ratio of 1:2. The presence of BLI in the ovine median eminence and its co-localization in some neurons with CRF, suggest a possible role for peptides of the bombesin family in the regulation of pituitary function.

Bombesin-like immunoreactivity in human seminal fluid

Bombesin-like immunoreactivity in human seminal fluid

Bombesin-like immunoreactivity in nervous and epithelial elements of earthworm ( [formula omitted

Bombesin-like immunoreactivity in nervous and epithelial elements of earthworm ( [formula omitted

Distribution of bombesin-like immunoreactivity in the rat brain and spinal cord

Distribution of bombesin-like immunoreactivity in the rat brain and spinal cord

Bombesin microinfusion into the paraventricular nucleus suppresses gastric acid secretion in the rat

Bombesin is a particularly potent inhibitor of gastric acid secretion when injected intracisternally in the rat. Because bombesin-like immunoreactivity is found in several forebrain regions implicated in gut regulation, the ability of bombesin to affect gastric secretion was tested in these areas by direct microinfusion. Bombesin significantly and dose-relatedly suppressed gastric acid secretion when it was infused into the hypothalamic paraventricular nucleus. Bombesin microinfusion into the ventromedial or lateral hypothalamic areas, or the caudate-putamen, had no significant effect. A further experiment using glass micropipets showed that back-diffusion of bombesin along the cannula track to a distant site of action was unlikely to account for the results obtained, and provided further evidence that the active site is limited to the paraventricular nucleus and possibly the ventralmost nucleus reuniens. The results suggest that the bombesin receptors and immunoreactive terminals previously identified in this region may be involved in the central regulation of gastric secretion.

Differential control of bombesin-like immunoreactivity (BLI), gastrin- and somatostatin (SLI) secretion from the isolated perfused rat stomach during vagal stimulation

Differential control of bombesin-like immunoreactivity (BLI), gastrin- and somatostatin (SLI) secretion from the isolated perfused rat stomach during vagal stimulation

Projections of bombesin-like immunoreactive fibers from the rat stomach to the celiac ganglion revealed by a double-labeling technique

Gastrofugal bombesin (BOM)-like immunoreactive (BOMI) structures in the rat were studied by immunocytochemistry combined with retrograde labeling. Transection of the mesenteric nerve peripheral to the celiac ganglion resulted in the complete disappearance of BOMI nerve terminals, whereas transection of the splanchnic nerves did not alter the immunoreactivity. Injection of biotinylated wheat germ agglutinin into the celiac ganglion labeled several neurons in the myenteric ganglion of the stomach. Simultaneous staining with antiserum against BOM showed that some of them are BOMI-positive. These findings demonstrate that BOMI neurons in the myenteric ganglion of the rat stomach project to the celiac ganglion.

Effects of neuromedin B and neuromedin C on exocrine and endocrine rat pancreas

Neuromedin B and neuromedin C are novel decapeptides that have recently been isolated from porcine spinal cord and canine intestinal mucosa and show striking sequence homology with bombesin and gastrin-releasing peptide (GRP-27) at the carboxyl-terminal region. The effects of synthetic neuromedin B and C on exocrine pancreatic function and insulin release have been compared with bombesin and GRP-27 in isolated pancreatic acini and isolated perfused pancreas in rat. Neuromedin B and C as well as bombesin and GRP-27 were able to cause stimulation of amylase release. The relative efficacy of neuromedin B, C, bombesin, and GRP-27 was the same as that of cholecystokinin octapeptide (CCK-8). Bombesin and GRP-27 were equipotent, and both were approximately 15-fold less potent than CCK-8. Neuromedin C was approximately 2-fold more potent, whereas neuromedin B as approximately 10-fold less potent than bombesin and GRP-27. All of these peptides stimulated insulin release that was limited to the first 3 min of a 20-min perfusion. However, GRP-27 and its related peptides were weak stimulants of insulin release compared with their abilities to stimulate exocrine pancreatic secretion. Bombesin and neuromedin B id not stimulate insulin release at doses stimulating pancreatic exocrine secretion. Neuromedin B was also approximately 10-fold less potent than neuromedin C, bombesin, and GRP-27 in eliciting insulin secretion. Because bombesin-like immunoreactivity is found to be present in nerves in the pancreas, neuromedin B and C may be neurotransmitters or neuromodulators and exert a direct local neurocrine action on enzyme secretion by acinar cells and insulin secretion by the islets.

Distribution of gastrin-releasing peptide (GRP)-like immunoreactivity in the rat.

Rat tissues were examined for gastrin-releasing peptide (GRP)-like immunoreactivity, using carboxy-terminus-specific antibody made against GRP (19-27), which is thought to be the biologically active site and to be common among species. The distribution of GRP-like immunoreactivity in the rat was similar to that of bombesin-like immunoreactivity. Sephadex G 50 chromatography revealed two peaks of GRP-like reactivity in the rat stomach. Immunohistochemical studies using the antiserum to GRP (19-27) revealed numerous nerve fibers in the mucosa of the rat stomach. In the antral mucosa, GRP-containing nerves were present mainly around the base of the antral glands. Some GRP-containing nerves were in contact with gastrin-containing cells and somatostatin-containing cells. GRP-containing nerve fibers were numerous in the middle portion of the oxyntic gland, where somatostatin-containing cells were also detected. None of the endocrine cells stained positively with anti-GRP serum. These results support the hypothesis that GRP is a neurotransmitter in the stomach and that the peptide plays a physiological role in the gastrointestinal tract.

Distribution of bombesin-like peptides in the alimentary canal of several vertebrate species

The objective of this study was to quantitate and characterize the variants of bombesin-like immunoreactivity in the alimentary canal of the rat, rabbit, hawk, owl, dog, monkey and human. Bombesin-like immunoreactivity was found throughout the entire gastrointestinal tract of all species studied. In the rat, the highest concentration of bombesin-like immunoreactivity was found in the colon. Gel chromatography showed that bombesin-like immunoreactivity corresponded to gastrin-releasing peptide (GRP-27) and GRP-10. In the dog, the greatest concentration of bombesin-like immunoreactivity was observed in the mucosal layer of the fundus, whereas the concentration of bombesin-like immunoreactivity in the muscle layer of the dog did not vary significantly from region to region. Gel chromatography showed that bombesin-like immunoreactivity in the dog corresponded to GRP-27, bombesin, GRP-10, and a smaller fragment. In the human, the concentration of bombesin-like immunoreactivity did not vary significantly from region to region in the mucosal and muscular layers. Gel chromatography of human fundal mucosa showed that bombesin-like immunoreactivity peaks occur in the regions of GRP-27, bombesin and GRP-10. These findings substantiate the observation that bombesin-like peptides play a variety of roles in the regulation of gut function.

Characterization of three new variant type cell lines derived from small cell carcinoma of the lung

Three new, well growing cell lines (GLC-1, GLC-2, and GLC-3) have been established from small cell lung carcinoma (SCLC) and characterized. A subclone (GLC-1-M13) markedly different from its parent line GLC-1 was also isolated and characterized. Cytogenetic analysis of the cell lines revealed deletions in the short arm of chromosome 3 as a most consistent chromosomal aberration. The deleted region was not identical in all metaphases, 3p(21-23) being the shortest region of overlap. Despite their SCLC origin GLC-1, GLC-2, and GLC-3 do not show pronounced SCLC differentiation features. Neurosecretory granula were very rare (GLC-1) or completely absent (GLC-2 and GLC-3), whereas the SCLC-related enzyme and hormone markers L-3,4-dihydroxyphenylalanine decarboxylase, neuron-specific enolase, creatine kinase BB, and bombesin-like immunoreactivity were variably expressed. Although the subclone GLC-1-M13 was derived from the poorly differentiated GLC-1, it behaved according to the above criteria as a differentiated "classic" SCLC cell line. When assessed with specific monoclonal antibodies the different cell lines appeared to express different subsets of intermediate filament proteins, indicative for different stages and directions of differentiation: "undifferentiated" (GLC-1 and GLC-2); "neural tissue related" (GLC-2); "simple epithelium" related (GLC-1-M13); and a combination of simple and squamous epithelium related (GLC-3). We conclude that GLC-1, GLC-2, and GLC-3 represent dedifferentiated forms of SCLC, related to the recently described "variant" type of SCLC, whereas the clonal derivate GLC-1-M13 behaves like a differentiated "classic" SCLC cell line.

THE DISTRIBUTION OF VASOACTIVE INTESTINAL POLYPEPTIDE‐LIKE, SUBSTANCE P‐LIKE AND BOMBESIN‐LIKE IMMUNOREACTIVITY IN THE DIGESTIVE SYSTEM OF THE SHEEP

Peptides which are possibly related to the non-cholinergic, non-adrenergic division of the autonomic nervous system have been identified by immunofluorescence in the digestive system of mature sheep. Vasoactive intestinal polypeptide-, substance P-, and bombesin-like immunoreactivity were localized in neural elements throughout the ovine gastrointestinal tract (g.i.t.). Vasoactive intestinal polypeptide-like immunoreactivity (VIP-l.i.) was demonstrable in the submaxillary, parotid and the sublingual salivary glands close to small blood vessels and the acini. VIP-l.i. was also demonstrable in the upper oesophagus in connective tissue near small blood vessels. In the forestomachs, abomasum, and small and large intestines reactive fibres were present in the mucosa, submucosa, smooth muscle layers and the plexuses. The plexuses also contained reactive nerve cell bodies. VIP-reactive fibres were found in the pancreas, the gall bladder and the common bile and pancreatic duct but were not found in the intestinal mesentery, portal vein, and liver tissue. Substance P-like immunoreactivity (SP-l.i.) was demonstrable in nerve fibres in all the layers of the g.i.t. and in nerve cell bodies in the gut plexuses. The pancreas and the gall bladder also contained a few scattered fibres. Additionally, SP-l.i. was present in open-type endocrine cells throughout the mucosa of the small and large intestines but no SP-l.i. was found in the salivary glands or the oesophagus. Bombesin-like immunoreactivity (B-l.i.) was associated with nerve fibres and was demonstrable in the mucosa and myenteric plexuses throughout the g.i.t. B-l.i. in the smooth muscle appeared to be restricted to nerve fibres in the forestomachs, the abomasum, and the upper small intestine. No B-l.i. was found in the salivary glands, oesophagus, liver tissue, pancreas, gall bladder or intestinal mesentery.

Establishment and identification of small cell lung cancer cell lines having classic and variant features

Using a chemically defined medium containing hydrocortisone, insulin, transferrin, 17 beta-estradiol and selenium, with or without serum supplementation (2.5% v/v), continuous cell lines can be established from 72% of all fresh biopsy specimens of small cell lung cancer (SCLC) containing tumor cells. No differences were observed in the rate of establishing cell lines from newly diagnosed untreated patients, or from patients who have relapsed from prior therapy, or from a variety of different organ sites. Biochemical characterization of 50 SCLC cell lines for the expression of L-dopa decarboxylase; bombesin-like immunoreactivity; neuron-specific enolase, and the brain isozyme of creatine kinase, revealed that SCLC cell lines can be subdivided into two distinct classes: classic SCLC cell lines (35 lines), which express elevated levels of all four biomarkers; and variant SCLC cell lines (15 lines) which have undetectable levels of L-dopa-decarboxylase and bombesin-like immunoreactivity, but continue to express neuron-specific enolase and the brain isozyme of creatine kinase. The presence of the latter two markers distinguishes variant lines fron non-SCLC cell lines. In addition, four distinct classes were identified morphologically. The biomedical differences among established SCLC cell lines may account for the differences in response rates to cytotoxic therapy observed in newly diagnosed SCLC patients. A prospective study of biomarker characterization of SCLC tumors will determine if clinical differences exist between classic and variant SCLC tumors.

Association of changes in bombesin immunoreactive neuroendocrine cells in lungs of newborn infants with persistent fetal circulation and brainstem damage due to birth asphyxia.

The pulmonary neuroendocrine (NE) cells, from 16 term infants dying at 1-4 days of age from birth asphyxia, were immuno stained for bombesin-like immunoreactivity by the immunoperoxidase method. The distribution and frequency of bombesin-immunoreactive NE cells were quantified morphometrically and correlated with the presence or absence of brainstem function and persistent fetal circulation (PFC). In infants with loss of brainstem function, the frequency of bombesin immunoreactive NE cells was significantly increased compared to infants with intact brainstem function, i.e. meconium aspiration with PFC. Infants with brainstem injury, with one exception, failed to develop PFC. Pathological changes in the tegmentum of the brainstem, i.e. containing the respiratory center, correlated in nine of 10 cases with loss of brainstem function. These data suggest an inverse relationship between brainstem function, release of bombesin-like peptide from the pulmonary NE cells and the functional state of the pulmonary vasculature. Intact brainstem function appears to be essential for both the release of bombesin-like peptide from the NE cells and for pulmonary vasoconstriction leading to PFC; absence of brainstem function is, on the other hand, associated with failure to release bombesin-like peptide and loss of PFC type reactivity in the pulmonary vasculature. However, it appears unlikely that bombesin itself is a direct mediator of pulmonary vasoconstriction.

Changes in the phenotype of human small cell lung cancer cell lines after transfection and expression of the c-myc proto-oncogene.

Small cell lung cancer growing in cell culture possesses biologic properties that allow classification into two categories: classic and variant. Compared with classic small cell lung cancer cell lines, variant lines have altered large cell morphology, shorter doubling times, higher cloning efficiencies in soft agarose, and very low levels of L dopa decarboxylase production and bombesin-like immunoreactivity. C-myc is amplified and expressed in some small cell lung cancer cell lines and all c-myc amplified lines studied to date display the variant phenotype. To investigate if c-myc amplification and expression is responsible for the variant phenotype, a normal human c-myc gene was transfected into a cloned classic small cell lung cancer cell line not amplified for or expressing detectable c-myc messenger RNA (mRNA). Clones were isolated with one to six copies of c-myc stably integrated into DNA that expressed c-myc mRNA. In addition, one clone with an integrated neo gene but a deleted c-myc gene was isolated and in this case c-myc was not expressed. C-myc expression in transfected clones was associated with altered large cell morphology, a shorter doubling time, and increased cloning efficiency, but no difference in L dopa decarboxylase levels and bombesin-like immunoreactivity. We conclude increased c-myc expression observed here in transfected clones correlates with some of the phenotypic properties distinguishing c-myc amplified variants from unamplified classic small cell lung cancer lines.

Modulatory glucose effect on bombesin-like immunoreactivity and gastrin secretion from isolated perfused rat stomach.

Previous studies have demonstrated mostly inhibitory effects of elevated plasma glucose levels on gastric exo- and endocrine as well as motor functions. Because increased plasma glucose levels reduce vagal activity via the central nervous system, it remains unclear if glucose exerts a direct effect on gastric functions. Therefore, our study was designed to determine the effect of acute changes in glucose concentrations on the release of gastrin and bombesin-like immunoreactivity (BLI) from the isolated perfused rat stomach. Acute elevations of perfusate glucose from 100 to 200 mg/dl or from 100 to 300 mg/dl augmented BLI secretion significantly without affecting gastrin release. During an acute decrease from 200 to 30 mg/dl, the secretion of both peptides remained unchanged. When acetylcholine was administered to stimulate BLI and gastrin secretion, the elevation of perfusate glucose to 200 mg/dl and the decrease to 30 mg/dl attenuated BLI secretion, whereas gastrin secretion remained unchanged compared with the control experiments at 100 mg/dl glucose. On the other hand, the perfusion of vasoactive intestinal peptide (VIP) and Leu-enkephalin had no effect on BLI and gastrin secretion during 100 mg/dl glucose perfusion, but both peptides elicited a significant stimulatory effect on BLI secretion during a perfusate glucose concentration of 200 mg/dl without affecting gastrin secretion. In conclusion, our study demonstrates first that an acute increase of glucose augments basal BLI secretion. Second, cholinergically induced BLI secretion is attenuated by hypo- and hyperglycemia. Third, hyperglycemia augments BLI secretion in response to the neuropeptides VIP and Leu-enkephalin. Fourth, basal and stimulated gastrin secretion remains unchanged during acute alterations of perfusate glucose levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Modulatory glucose effect on bombesin-like immunoreactivity and gastrin secretion from isolated perfused rat stomach

Modulatory glucose effect on bombesin-like immunoreactivity and gastrin secretion from isolated perfused rat stomach

Bombesin inhibits growth hormone response to insulin-induced hypoglycemia in humans

Intravenous administration of bombesin (5ng/kg/min× 2.5h) significantly reduced growth hormone (GH) response to insulin-induced (0.15 U/kg, i.v.) hypoglycemia in 8 male volunteers without affecting its basal plasma levels. These data, together with the presence of bombesin-like immunoreactivity in the human brain, suggest that — as in experimental animals — this neurogastrointestinal peptide may be of physiological significance in the control of GH secretion also in man.

Effect of insulin on secretion of bombesin-like immunoreactivity and gastrin from the isolated rat stomach in response to acetylcholine, VIP and leucine-enkephalin

Bombesin-like immunoreactivity (BLI), a putative peptidergic neurotransmitter of the gastrointestinal intrinsic nervous system is released from the isolated perfused rat stomach in response to the classical neurotransmitter acetylcholine and in response to other putative peptidergic neurotransmitters such as vasoactive intestinal peptide (VIP), peptide histidine isoleucine (PHI) or growth hormone releasing factor (GRF). The secretion of BLI is modulated not only by gastric factors such as the intragastric pH but also by changes of perfusate glucose concentrations indicating that alterations of carbohydrate metabolism might have an effect on gastric neuroendocrine regulation. Since previous studies have shown that insulin, the major regulatory hormone of glucose metabolism, reduces gastric somatostatin and glucagon secretion it was of interest to determine the effect of insulin on gastric BLI and gastrin secretion. The experiments were performed in the isolated perfused rat stomach model. The addition of porcine insulin to the perfusate at concentrations of 50 and 100 μU/ml had no effect on basal BLI and gastrin secretion. The infusion of acetylcholine (2 × 10 −6M and 4 × 10 −6M) elicited a stimulation of BLI and gastrin secretion which was not altered by the addition of insulin (100 μU/ml). On the other hand, significant effects of insulin were observed during administration of the two putative peptidergic neuro-transmitters VIP and leu-enkephalin. The infusion of VIP at 10 −11M and 10 −8M had no effect on BLI and gastrin secretion in the absence of insulin, however, with the addition of insulin (100 μU/ml) the higher dose of VIP (10 −8M) elicited a significant stimulation of BLI secretion while both doses of VIP (10 −11M and 10 −8M) significantly increased gastrin release. Simlar to VIP the infusion of leu-enkephalin at doses of 10 −9M and 10 −6M had no effect on BLI and gastrin secretion in the absence of insulin. When insulin was added to the perfusate both doses of leu-enkephalin elicited a significant stimulation of BLI secretion while gastrin remained unchanged. The addition of the specific opiate receptor antagonist naloxone (10 −5M) did not block the effect of leu-enkephalin in the presence of insulin. In addition the effect of naloxone was also examined during cholinergic stimulation. The addition of naloxone (10 −5M) during the infusion of acetylcholine abolished the stimulatory effect on BLI secretion in the absence of insulin, whereas in the presence of insulin naloxone did not alter cholinergically-induced BLI secretion. In conclusion the present data demonstrate that in the rat stomach insulin has no effect on cholinergically-induced BLI and gastrin secretion but it augments BLI secretion in response to VIP and leu-enkephalin and gastrin release in response to VIP. This suggests that insulin has a modulatory effect on the action and release of putative peptidergic neurotransmitters of the rat peripheral autonomic nervous system. Furthermore, endogenous opioids apparently mediate the cholinergic stimulation of BLI secretion and insulin antagonizes the effect of the opiate-receptor antagonist naloxone via as yet unknown mechanisms.

Effect of vasoactive intestinal peptide, peptide histidine isoleucine and growth hormone-releasing factor-40 on bombesin-like immunoreactivity, somatostatin and gastrin release from the perfused rat stomach

Bombesin-like immunoreactivity (BLI) has been demonstrated in neurons of the gastrointestinal tract and gastric BLI secretion can be demonstrated in response to the classical neurotransmitter acetylcholine. Since structurally related peptides VIP, PHI and GRF have to be considered as peptidergic neurotransmitters it was of interest to determine their effect on gastric BLI secretion. Additionally, somatostatin (SLI) and gastrin secretion was examined. The isolated stomach of overnight fasted rats was perfused with Krebs-Ringer buffer via the celiac artery and the effluent was collected via the portal vein. The gastric lumen was perfused with isotonic saline at pH7 or pH2. All four peptides were tested at a dose of 10 −11 M and 10 −8 M at both pH levels and in addition the effect of VIP and PHI was examined at 10 −14 M and 10 −12 M during luminal pH2. At luminal pH7 VIP and PHI stimulated SLI release at 10 −8 M but had no effect on BLI or gastrin secretion. rGRF and hpGRF were both ineffective on SLI and gastrin release while rGRF inhibited and hpGRF stimulated BLI secretion. This effect was not dose related. At luminal pH2 all four peptides stimulated BLI secretion. Stimulation by PHI was already observed at a dose of 10 −14 M while VIP elicited a stimulatory effect at 10 −12 M. PHI at the two lowest concentrations of 10 −14 and 10 −12 M elicited a stimualtion of SLI and gastrin release while the same doses of VIP and the higher doses of all four peptides had no effect on SLI and gastrin secretion at an acidic intraluminal pH. The present data demonstrate that at a low intragastric pH BLI secretion is stimulated predominantly by putative peptidergic neurotransmitters such as PHI and VIP and possibly GRF in contrast to the previously reported cholinergic stimulation at neutral pH. The effect of low dose PHI on gastrin secretion suggests that this peptide might participate in the previously observed vagally-induced gastrin secretion during low intragastric pH. On the other hand, VIP and PHI might contribute to postprandial stimulation of somatostatin release during meal-induced increase of the intragastric pH.