Effects of gastric inhibitory polypeptide (GIP) immunoneutralization on mouse motor coordination and memory.

Gastric inhibitory polypeptide (GIP) is a regulatory peptide expressed in the mammalian upper small intestine, and both GIP and its receptor (GIPR) are expressed in the cortex and hippocampus regions of the brain as well. While learning and memory deficits have been observed in GIPR-/- mice, the effects of peripheral GIP immunoneutralization on motor-coordination, learning, and memory have not been examined. In the present study, adult GIPR-/- mice (KO) and age-matched wild-type C57BL/6 J mice (WT) received weekly vehicle PBS injections for 12 weeks, while a third group of wild-type mice were injected weekly for 12 weeks with 30 mg/kg body weight humanized GIP-mAb (AB) to assess the possibility of long-term effects of peripheral GIP antagonism on rodent memory and behavior. All mice groups then underwent a battery of tests that evaluated motor behavior, body coordination, and memory. Performance deficits in several memory studies after 12 weeks of treatment were demonstrated in KO, but not in AB or WT mice. Body coordination performance showed no significant differences among the 3 groups. A similar short-term study (3 injections over 9 days) was also conducted and the results were similar to those from the long-term study. Thus, short-term and long-term peripheral GIP antagonism by GIP-mAb did not appear to affect learning and memory in mice, consistent with the notion that the GIP-mAb does not cross the blood brain barrier. Furthermore, our studies indicate that GIP signaling in the brain appears to involve local neurocrine pathways.

GI inflammation Increases Sodium-Glucose Cotransporter Sglt1.

A correlation between gastrointestinal (GI) inflammation and gut hormones has reported that inflammatory stimuli including bacterial endotoxins, lipopolysaccharides (LPS), TNFα, IL-1ß, and IL-6 induces high levels of incretin hormone leading to glucose dysregulation. Although incretin hormones are immediately secreted in response to environmental stimuli, such as nutrients, cytokines, and LPS, but studies of glucose-induced incretin secretion in an inflamed state are limited. We hypothesized that GI inflammatory conditions induce over-stimulated incretin secretion via an increase of glucose-sensing receptors. To confirm our hypothesis, we observed the alteration of glucose-induced incretin secretion and glucose-sensing receptors in a GI inflammatory mouse model, and we treated a conditioned media (Mϕ 30%) containing inflammatory cytokines in intestinal epithelium cells and enteroendocrine L-like NCI-H716 cells. In GI-inflamed mice, we observed that over-stimulated incretin secretion and insulin release in response to glucose and sodium glucose cotransporter (Sglt1) was increased. Incubation with Mϕ 30% increases Sglt1 and induces glucose-induced GLP-1 secretion with increasing intracellular calcium influx. Phloridzin, an sglt1 inhibitor, inhibits glucose-induced GLP-1 secretion, ERK activation, and calcium influx. These findings suggest that the abnormalities of incretin secretion leading to metabolic disturbances in GI inflammatory disease by an increase of Sglt1.

Gastric inhibitory polypeptide immunoneutralization attenuates development of obesity in mice.

Previous reports have suggested that the abrogation of gastric inhibitory polypeptide (GIP) signaling could be exploited to prevent and treat obesity and obesity-related disorders in humans. This study was designed to determine whether immunoneutralization of GIP, using a newly developed specific monoclonal antibody (mAb), would prevent the development of obesity. Specific mAb directed against the carboxy terminus of mouse GIP was identified, and its effects on the insulin response to oral and to intraperitoneal (ip) glucose and on weight gain were evaluated. Administration of mAb (30 mg/kg body wt, BW) to mice attenuated the insulin response to oral glucose by 70% and completely eliminated the response to ip glucose coadministered with human GIP. Nine-week-old C57BL/6 mice injected with GIP mAbs (60 mg·kg BW(-1)·wk(-1)) for 17 wk gained 46.5% less weight than control mice fed an identical high-fat diet (P < 0.001). No significant differences in the quantity of food consumed were detected between the two treatment groups. Furthermore, magnetic resonance imaging demonstrated that subcutaneous, omental, and hepatic fat were 1.97-, 3.46-, and 2.15-fold, respectively, lower in mAb-treated animals than in controls. Moreover, serum insulin, leptin, total cholesterol (TC), low-density lipoprotein (LDL), and triglycerides were significantly reduced, whereas the high-density lipoprotein (HDL)/TC ratio was 1.25-fold higher in treated animals than in controls. These studies support the hypothesis that a reduction in GIP signaling using a GIP-neutralizing mAb might provide a useful method for the treatment and prevention of obesity and related disorders.

Distribution of K and L cells in the feline intestinal tract.

Glucose-dependent insulinotropic peptide (GIP), glucagon-like peptide (GLP)-1 and GLP-2 are hormones secreted from specialized K cells (GIP) and L cells (GLP-1, GLP-2) in the intestinal mucosa. These hormones play major roles in health and disease by modulating insulin secretion, satiety, and multiple intestinal functions. The aim of this study was to describe the distribution of K cells and L cells in the intestines of healthy cats. Samples of duodenum, mid-jejunum, ileum, cecum, and colon were collected from 5 cats that were euthanized for reasons unrelated to this study and had no gross or histologic evidence of gastrointestinal disease. Samples stained with rabbit-anti-porcine GIP, mouse-anti-(all mammals) GLP-1, or rabbit-anti-(all mammals) GLP-2 antibodies were used to determine the number of cells in 15 randomly selected 400× microscopic fields. In contrast to other mammals (eg, dogs) in which K cells are not present in the ileum and aborally, GIP-expressing cells are abundant throughout the intestines in cats (>6/high-power field in the ileum). Cells expressing GLP-1 or GLP-2 were most abundant in the ileum (>9/high-power field) as in other mammals, but, although GLP-1-expressing cells were abundant throughout the intestines, GLP-2-expressing cells were rarely found in the duodenum. In conclusion, the distribution of GIP-secreting K cells in cats is different from the distribution of K cells that is described in other mammals. The difference in distribution of GLP-2- and GLP-1-expressing cells suggests that more than 1 distinct population of L cells is present in cats.

Evaluation of the long-term effects of gastric inhibitory polypeptide-ovalbumin conjugates on insulin resistance, metabolic dysfunction, energy balance and cognition in high-fat-fed mice.

The effects of active immunisation with gastric inhibitory polypeptide (GIP) or (proline3)GIP-ovalbumin conjugates on insulin resistance, metabolic dysfunction, energy expenditure and cognition were examined in high-fat-fed mice. Normal mice were injected (subcutaneously) once every 14 d for 98 d with GIP-ovalbumin conjugates, with transfer to a high-fat diet on day 21. Active immunisation resulted in GIP antibody generation and significantly (P < 0·01 to P < 0·001) reduced circulating non-fasting plasma insulin concentrations compared to high-fat control mice from day 70 onwards. The glycaemic responses to intraperitoneal glucose or nutrient ingestion were significantly improved in all treated mice, with corresponding stimulated plasma insulin levels depressed compared to high-fat controls. These changes were associated with substantially (P < 0·001) improved glucose-lowering responses to exogenous insulin and decreases of muscle and fat TAG, pancreatic insulin, circulating total and LDL-cholesterol levels (P < 0·01 to P < 0·001). Treatment with GIP-ovalbumin conjugates was not associated with alterations in energy expenditure, indirect calorimetry or aspects of cognitive function. The observed changes were almost identical in GIP and (Pro3)GIP immunised mice and were independent of any effects on food intake or body weight. Further tests established that coupling of GIP peptides to ovalbumin abolished any intrinsic insulin-releasing or glucose-lowering activity. These results suggest that induction of GIP-neutralising antibodies with GIP-ovalbumin conjugates is an effective means of countering the metabolic abnormalities induced by high-fat feeding and does not adversely have an impact on a marker of cognition function or energy expenditure.

Dual-monoclonal, sandwich immunoassay specific for glucose-dependent insulinotropic peptide1-42, the active form of the incretin hormone.

BACKGROUND: Glucose-dependent insulinotropic peptide (GIP) is an incretin peptide secreted by intestinal K cells that stimulates insulin secretion in a glucose-dependent manner. It is secreted as an active, intact 42-amino acid peptide GIP(1-42), which is rapidly degraded by dipeptidyl peptidase 4 to GIP(3-42), which is inactive. There is currently no described monoclonal antibody-based sandwich immunoassay to quantify concentrations of GIP(1-42), the active form of the peptide. METHODS: To create a sandwich ELISA for GIP(1-42), we generated a monoclonal antibody specific for the intact N-terminus of the peptide, which was further optimized to increase its affinity. We used this antibody as a conjugate antibody in a sandwich ELISA and paired it with an anti-total GIP capture monoclonal antibody to create a dual monoclonal sandwich ELISA for GIP(1-42). RESULTS: The sandwich ELISA was highly specific for GIP(1-42) and did not recognize GIP(3-42). The ELISA demonstrated a broad dynamic range and a lower limit of quantification of 5 ng/L. Using the ELISA, we were able to show that GIP(1-42) concentrations in healthy volunteers increased dramatically in the postprandial state compared to the fasting state. GIP(1-42) values were correlated with total GIP values overall; however, there was substantial interindividual variation. CONCLUSIONS: The use of an N-terminal-specific monoclonal antibody in a sandwich ELISA format provides a robust and convenient method for measuring concentrations of GIP(1-42), the active form of the incretin hormone. This ELISA should help to improve our understanding of the role of GIP(1-42) in regulating glucose-dependent insulin secretion.

The small intestine of the adult New Hampshire chicken: an immunohistochemical study.

The presence and distribution of glucose-dependent insulinotropic polypeptide or gastric inhibitory polypeptide (GIP), gastric-releasing peptide (GRP) and glucagon immunoreactivity were studied in the small intestine of the New Hampshire chicken using immunohistochemistry. This is the first report of the presence of GIP-immunoreactive (ir) cells in avian small intestine. GIP, GRP and glucagon immunoreactivity was localized in the epithelium of the villi and crypts of the duodenum, jejunum and ileum. In particular, both in the duodenum and in the jejunum immunoreactive endocrine cells to GIP, GRP and glucagon were observed. In the ileum, we noticed GIP-ir and glucagon-ir cells. GRP-ir was found in nerve fibres of all three segments of the small intestine. The distribution of these bioactive agents in the intestinal tract of the chicken suggests that GIP and glucagon may play a role in the enteropancreatic axis in which intestinal peptides modulate pancreas secretion.

Active immunization with glucose-dependent insulinotropic polypeptide vaccine influences brain function and behaviour in rats.

Glucose-dependent insulinotropic polypeptide (GIP) is involved in the aetiology of obesity induced by overnutrition, and blocking GIP activity may be valuable to anti-obesity treatment. However, GIP and GIP receptor are closely related to various brain functions which have caused very little data to be published concerning this cerebral functionality after blocking GIP activity. Here, we showed that active vaccination of mature rats with GIP immunoconjugates [GIP-keyhole limpet haemocyanin (KLH)] was associated with changes in body weight. Furthermore, we also observed significant changes in brain function and behaviour. Data indicated that GIP-KLH-immunized rats showed decreased spontaneous activity in the open field test, decreased cerebral glucose utilization assessed by 18F-fluorodeoxyglucose-positron emission tomography/computed tomography (PET/CT), and increased apoptosis and proliferation of hippocampal granule cells marked by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) or proliferating cell nuclear antigen method. In conclusion, we have shown that vaccine-induced antibodies inhibited GIP activity in vivo and led to significant changes in brain function and behaviour, which underscore the need to address any potential problems GIP-targeted immunotherapy may involve in further research.

Active immunization against (Pro(3))GIP improves metabolic status in high-fat-fed mice.

AIM: Ablation of gastric inhibitory polypeptide (GIP) receptor signalling can prevent many of the metabolic abnormalities associated with dietary-induced obesity-diabetes. The present study was designed to assess the ability of active immunization against (Pro(3))GIP to counter metabolic dysfunction associated with diet-induced obesity in high-fat-fed mice. METHODS: Normal male Swiss NIH mice were injected (s.c.) once every 14 days for 98 days with complexed (Pro(3))GIP peptide, with transfer to a high-fat diet on day 21. RESULTS: Active immunization against (Pro(3))GIP resulted in circulating GIP antibody production and significantly (p < 0.05 p < 0.01) reduced circulating blood glucose concentrations compared to high-fat control mice from day 84 onwards. Glucose levels were not significantly different from lean controls. The glycaemic response to i.p. glucose was correspondingly improved (p < 0.01) in (Pro(3))GIP-immunized mice. Furthermore, circulating and glucose-stimulated plasma insulin levels were significantly (p < 0.01 to p < 0.001) depressed compared to high-fat control mice. Liver triglyceride, pancreatic insulin and circulating LDL-cholesterol levels were also significantly reduced in (Pro(3))GIP-immunized mice. These changes were independent of any effects on food intake or body weight. The glucose-lowering effect of native GIP was annulled in (Pro(3))GIP-immunized mice consistent with the induction of biologically effective GIP-specific neutralizing antibodies. CONCLUSION: These results suggest that immunoneutralization of GIP represents an effective means of countering the disruption of metabolic processes induced by high-fat feeding.

Differential processing of pro-glucose-dependent insulinotropic polypeptide in gut.

Glucose-dependent insulinotropic polypeptide (GIP) is a hormone released from enteroendocrine K cells in response to meals. Posttranslational processing of the precursor protein pro-GIP at residue 65 by proprotein convertase subtilisin/kexin type 1 (PC1/3) in gut K cells gives rise to the established 42-amino-acid form of GIP (GIP(1-42)). However, the pro-GIP peptide sequence contains a consensus cleavage site for PC2 at residues 52-55 and we identified PC2 immunoreactivity in a subset of K cells, suggesting the potential existence of a COOH-terminal truncated GIP isoform, GIP(1-30). Indeed a subset of mouse and human K cells display GIP immunoreactivity with GIP antibodies directed to the mid portion of the peptide, but not with a COOH-terminal-directed GIP antibody, indicative of the presence of a truncated form of GIP. This population of cells represents approximately 5-15% of the total GIP-immunoreactive cells in mice, depending on the region of intestine, and is virtually absent in mice lacking PC2. Amidated GIP(1-30) and GIP(1-42) have comparable potency at stimulating somatostatin release in the perfused mouse stomach. Therefore, GIP(1-30) represents a naturally occurring, biologically active form of GIP.

Immunoassays for the incretin hormones GIP and GLP-1.

The measurement of the incretin hormones, glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), using immunologically based assays is made difficult by the fact that the processing of the precursor molecules gives rise to a number of different peptides which cross-react with antisera raised against the two hormones. For GLP-1, the picture is further complicated because of the necessity to differentiate between the intestinal and pancreatic proglucagon products. Finally, once secreted, both incretins are rapidly degraded by the enzyme dipeptidyl peptidase-4 (DPP-4) to generate metabolites which have lost their insulinotropic activities. These metabolites are the major circulating forms of the incretins, accounting for 60-80% of total immunoreactive GLP-1 and GIP in the peripheral plasma, while concentrations of the intact forms can be very low and, in some cases, barely detectable. The use of highly specific assays using well-characterised antisera and careful sample handling is therefore required for a reliable determination of incretin hormone concentrations.

Evidence for beneficial effects of compromised gastric inhibitory polypeptide action in obesity-related diabetes and possible therapeutic implications.

Gastric inhibitory polypeptide (GIP) is a physiological gut peptide secreted from the intestinal K-cells with well documented insulin-releasing actions. However, the GIP receptor is widely distributed in peripheral organs, including the pancreas, gut, adipose tissue, heart, adrenal cortex and brain, suggesting that it may have other functions. The presence of functional GIP receptors on adipocytes and the key role played by GIP in lipid metabolism and fat deposition suggest a possible beneficial effect of compromised GIP action in obesity and insulin resistance. Several key observations in animal models of obesity-related diabetes with chemically or genetically mediated biological GIP deficiency support this concept. Thus, obese diabetic animals with compromised GIP action due to peptide-based GIP receptor antagonists, small molecular weight GIP receptor antagonists, vaccination against GIP, genetic knockout of GIP receptor or targeted K-cell destruction are protected against obesity and associated metabolic disturbances. In addition, by causing preferential oxidation of fat, blockade of GIP signalling clears triacylglycerol deposits from liver and muscle, thereby restoring mechanisms for suppression of hepatic glucose output and improving insulin sensitivity. Emerging evidence also suggests that rapid cure of diabetes in grossly obese patients undergoing bypass surgery is mediated, in part, by surgical removal of GIP-secreting K-cells in the upper small intestine.

Active immunisation against gastric inhibitory polypeptide (GIP) improves blood glucose control in an animal model of obesity-diabetes.

Recent research suggests that long-term ablation of gastric inhibitory polypeptide (GIP) receptor signalling can reverse or prevent many of the metabolic abnormalities associated with dietary and genetically induced obesity-diabetes. The present study was designed to assess the sub-chronic effects of passive or active immunisation against GIP in ob/ob mice. Initial acute administration of GIP antibody together with oral glucose in ob/ob mice significantly increased the glycaemic excursion compared to controls (p<0.05). This was associated with a significant reduction (p<0.05) in the overall glucose-mediated insulin response. However, sub-chronic passive GIP immunisation was not associated with any changes in body weight, food intake or metabolic control. In contrast, active immunisation against GIP for 56 days in young ob/ob mice resulted in significantly (p<0.05) reduced circulating plasma glucose concentrations on day 56 compared to controls. There was a tendency for decreased circulating insulin in GIP immunised mice. The glycaemic response to intraperitoneal glucose was correspondingly improved (p<0.05) in mice immunised against GIP. Glucose-stimulated insulin levels were not significantly different from controls. Furthermore, insulin sensitivity was similar in mice immunised against GIP and respective controls. Overall, the results reveal that active, as opposed to passive, immunisation against GIP improves blood glucose control ob/ob mice.

Vaccination against GIP for the treatment of obesity.

BACKGROUND: According to the WHO, more than 1 billion people worldwide are overweight and at risk of developing chronic illnesses, including cardiovascular disease, type 2 diabetes, hypertension and stroke. Current therapies show limited efficacy and are often associated with unpleasant side-effect profiles, hence there is a medical need for new therapeutic interventions in the field of obesity. Gastric inhibitory peptide (GIP, also known as glucose-dependent insulinotropic polypeptide) has recently been postulated to link over-nutrition with obesity. In fact GIP receptor-deficient mice (GIPR(-/-)) were shown to be completely protected from diet-induced obesity. Thus, disrupting GIP signaling represents a promising novel therapeutic strategy for the treatment of obesity. METHODOLOGY/PRINCIPAL FINDINGS: In order to block GIP signaling we chose an active vaccination approach using GIP peptides covalently attached to virus-like particles (VLP-GIP). Vaccination of mice with VLP-GIP induced high titers of specific antibodies and efficiently reduced body weight gain in animals fed a high fat diet. The reduction in body weight gain could be attributed to reduced accumulation of fat. Moreover, increased weight loss was observed in obese mice vaccinated with VLP-GIP. Importantly, despite the incretin action of GIP, VLP-GIP-treated mice did not show signs of glucose intolerance. CONCLUSIONS/SIGNIFICANCE: This study shows that vaccination against GIP was safe and effective. Thus active vaccination may represent a novel, long-lasting treatment for obesity. However further preclinical safety/toxicology studies will be required before the therapeutic concept can be addressed in humans.

Liver transplantation restores endocrine cells in patients with familial amyloidotic polyneuropathy.

BACKGROUND: The aim of this study was to investigate familial amyloidotic polyneuropathy, Portuguese type patients' endocrine cell content in the stomach and duodenum before and after liver transplantation, and to relate the findings to the patients' gastrointestinal disturbances. METHODS: Ten liver-transplanted familial amyloidotic polyneuropathy, Portuguese type patients and 10 healthy controls were seen. Endocrine cells were identified by immunohistochemistry and quantified with computerized image analysis. The activity of the cells was appraised by measurements of the cell secretory index and nuclear area. Clinical symptoms were obtained from the patients' medical records. RESULTS: After transplantation, a significant increase of several endocrine cell types were noted, and the pretransplant depletion of several types of endocrine cells disappeared. For no type of endocrine cell was any difference compared with controls noted after transplantation. There was no significant decrease of the amount of amyloid in the biopsies after liver transplantation. The patients' symptoms remained generally unchanged after transplantation, although a substantial time lapse between pretransplant evaluation and transplantation was present. CONCLUSIONS: Liver transplantation restores the endocrine cells in the upper part of the gastrointestinal tract. The restoration was not correlated with an improvement of the patients' symptoms. No decrease of the amyloid deposits was noted.

The reactivity of anti-porcine gastric inhibitory polypeptide (GIP) rabbit serum R65 with synthetic human and porcine GIP.

Porcine gastric inhibitory peptide (GIP) has been used extensively as a standard and as [125I]-GIP in radio-immunoassays (RIAs) to determine immunoreactive GIP (IR GIP) in human plasma even though porcine and human GIP have slightly different amino acid sequences. A rabbit anti-porcine GIP serum (R65) which has been widely used in the RIA for IR GIP reacts with a part of the GIP molecule which is identical in human and porcine GIP. In order to validate the use of R65 the IR GIP contents of synthetic human and porcine GIP were measured, using natural porcine GIP as standard, and found to be 40% and 35% on a weight basis respectively. These materials were considered unsuitable for use as RIA standards. The IR GIP in samples of human plasma was then assayed with 125I-labelled synthetic human or porcine GIP using natural porcine GIP as standard. The values measured for IR GIP did not depend on the labelled peptide used (p = 0.84, n = 81) and were linearly correlated (r = 0.98, p less than 0.001, n = 81). It is concluded that the value of IR GIP measured in human plasma using R65 and standards of porcine GIP is linearly proportional to the level of human GIP in plasma.

Effect of exogenous or endogenous gastric inhibitory polypeptide (GIP) on plasma triglyceride responses in rats.

We examined the effects of exogenous and endogenous GIP on plasma triglyceride levels in rats, pretreated with a fat-enriched diet, during intraduodenal infusion of a lipid test meal (Lipomul, 8 ml/h). Following the fat load the plasma triglyceride levels increased nearly linearly from a fasting value of 0.621 +/- 0.031 mmol/l to 3.32 +/- 0.403 mmol/l at 150 min. Simultaneously, the plasma GIP levels rose from 47.1 +/- 5.1 at fasting to a peak value of 268.4 +/- 32.2 pmol/l at 120 min. When porcine GIP was infused intravenously during the fat load, the plasma triglyceride increments were significantly smaller (control 1.64 +/- 0.264 mmol/l versus 0.949 +/- 0.114 mmol/l during GIP infusion at 60 min; p less than 0.002). GIP infusion in the absence of the fat load did not change fasting triglyceride levels. The effect of endogenous GIP was investigated by neutralization of GIP by injection of GIP antiserum (0.3 ml). Rats pretreated with the antiserum exhibited a significantly greater triglyceride increment late in the time course of the fat load. These data demonstrate that exogenous and endogenous GIP are able to lower the plasma triglyceride response to a fat load. Both, inhibition of fat absorption or stimulation of triglyceride uptake by peripheral tissues may be responsible for the GIP effects. The gut peptide GIP seems to represent an important hormonal regulator of postprandial triglyceride response.

Autoimmunity in diarrhoeal disease.

Evidence for autoimmunity in diarrhoeal disease is reviewed. Firstly, coeliac disease (CD) is considered. The incidence of tissue-reactive autoantibodies in both adults and children with CD (68% and 65%, respectively) is higher than the incidence of these autoantibodies in controls (6% in normal adults, and 14% and 9% in disease controls drawn respectively from adult and child populations). The R1 antireticulin antibody, when present, was found to disappear after several weeks on a gluten-free diet, but in contrast, other autoantibodies persisted. Secondly, a case is argued for a new disease category, namely "autoimmune enteropathy." Seven cases are reviewed in which patients presented with protracted diarrhoea, a small intestinal enteropathy which failed to heal during periods of total parenteral nutrition, and evidence of a predisposition to autoimmunity (namely, the presence of high titre autoantibodies including one specific for gut epithelium, and/or the presence of associated diseases regarded to be autoimmune). Thirdly, evidence for autoimmunity in inflammatory bowel disease is reviewed and includes discussion of serum goblet cell antibodies and of circulating T cells which participate in antibody-dependent cellular cytotoxicity in vitro using colonic epithelial cells as targets. Finally, an unusual child is described who presented with chronic diarrhoea and a flat small intestinal mucosa, who responded to gluten withdrawal but who later relapsed spontaneously during a strict gluten-free diet. Her mucosa healed only after a period of total parenteral nutrition and treatment with oral steroids. This child's enteropathy was also associated with thyrotoxicosis and a microscopic colitis.

Peptides of the APUD system in amphibian skins.

Methanol extracted skins from 84 species of amphibia were screened, measuring by RIAs: gastrin-CCK, VIP, calcitonin, GIP, PP and motilin. G-CCK-like immunoreactivity was found in 97.6%; VIP-like immunoreactivity in 41%; CT-like immunoreactivity in 34%; GIP-like immunoreactivity in 10%; PP-like immunoreactivity in 40% and MT-like immunoreactivity in 60% of the samples. The use of a sequence-specific radioimmunoassay and of gel-chromatography confirmed the caerulein-CCK-8-like nature of the immunoreactive material. Detected amounts of the other peptides (VIP, CT, GIP, PP, MT) were too low for bioassay or chromatographic studies, thus leaving the question open if they are due to some kind of unspecific interferences or, most likely, to species-specificity differences of the used antisera.