Distribution of oxyntomodulin and glucagon in the gastrointestinal tract and the plasma of the rat

Oxyntomodulin (OXM), an intestinal glucagon-containing peptide extended at its C-terminal end by an octa-peptide, is one of the gut glucagon-like immunoreactants (GLI) or enteroglucagon. The distribution of OXM and glucagon was determined in the gastrointestinal tract and in the plasma of the rat. Reversed-phase HPLC, associated with RRA or RIA, performed with an N-terminally directed glucagon antiserum (GOL), was used. HPLC of intestinal extracts or plasma separated the GOL immunoreactivity into three peaks: two major peaks coeluting with a preparation of rat glicentin (peak I, partially purified from rat intestine) and porcine or rat OXM, respectively, and a smaller peak coeluting with glucagon. The behavior of the three peaks in the analytical systems matched that of glicentin, OXM, and glucagon, respectively, allowing their identification. The concentrations of OXM picomoles per g of tissue) gradually increased from the duodenum (9 +/- 1) to ileum (93 +/- 4), thereafter decreasing in cecum and colon (22 +/- 3). In the gut, OXM, glucagon, and peak I averaged 40%, 1%, and 59% of the total GLI, respectively. OXM was present in significant amounts in the pancreas (18% of GLI) and stomach (27% of GLI), two tissues in which it accounted, together with glucagon, for almost the totality of GLI. In 24 h-fasted rats, plasma concentrations of OXM, glucagon, and peak I, determined after HPLC with GOL antiserum, were 15.1 pM, 8.6 pM, and 12.3 pM, respectively. Two hours after refeeding, both OXM and peak I were significantly increased (P less than 0.05 and P less than 0.02) by a similar factor (2-fold), while glucagon remained unchanged. When the HPLC results were compared with RIA measurement of GLI (GOL antiserum) and glucagon (with a C-terminal glucagon antiserum) in plasma, enteroglucagon (GOL--C-terminal glucagon antiserum immunoreactivities) correlated well with the sum of OXM plus peak I. The combination of HPLC and RRA or RIA allows the unambiguous determination of OXM, glucagon, and glicentin (peak I) in tissues and plasma. In the rat intestine and in the plasma, OXM and glicentin appear roughly in the same ratio and seem to be the major components, if not the totality, of enteroglucagon.     

Effect of intraluminal administration of amino acids upon plasma glicentin

To see what effect intraluminal amino acids would have on glicentin secretion, we put a mixture of 10 amino acids (1 g/kg) into the duodenum of five normal, conscious piglets. Their plasma nitrogen rose, as did insulin and glucagon measured with C-terminal-specific antiserum. Plasma total immunoreactive glucagon, determined with non-specific antiserum, rose from 2753 +/- 460 pg/ml to a peak of 4434 +/- 1352 pg/ml at 30 min. Plasma glicentin, determined with R 64 antiserum, rose from a fasting level of 297 +/- 70 pmol/l to a peak of 702 +/- 167 pmol/l at 45 min. We also gave oral arginine to 6 pancreatectomized dogs to investigate why the plasma glicentin rises after amino acid ingestion. Arginine raised the plasma total immunoreactive glucagon from 1120 +/- 214 pg/ml to a peak of 2266 +/- 512 pg/ml at 45 min. We conclude that intraluminally administered amino acids enhance glicentin secretion from the gut.     

Response of plasma glicentin to intraduodenal administration of glucose in piglets

Controversial results concerning the secretion of glicentin prompted us to investigate the response of circulating glicentin to intraduodenal administration of glucose in piglets. A 20% solution of glucose (2 g/kg) was administered into the duodenum of six piglets in a fully conscious state. As blood glucose rose, plasma insulin increased to a peak of 21 +/- 4 microU/ml. Plasma glucagon, determined by C-terminal-specific antiserum, was 70 +/- 30 pg/ml at fasting and slightly increased after the glucose load. Plasma immunoreactive glucagon measured by cross-reacting glucagon antiserum increased from the baseline of 1563 +/- 260 to a peak of 4738 +/- 415 pg/ml at 120 min. Plasma glicentin determined by antiserum R 64 was 463 +/- 81 pmol/l at baseline and reached a peak level of 1081 +/- 174 pmol/l at 90 min. The percent changes of plasma glucagon from the fasting level measured by cross-reacting antiserum and glicentin were 296 and 233%, respectively. There was a significant correlation between plasma glucagon measured by cross-reacting antiserum and glicentin (r = 0.817, P less than 0.001). Chromatography of plasma obtained during glucose load revealed the heterogeneity of glicentin. It can be concluded from the present study that glicentin is clearly secreted in response to intraluminal administration of glucose.     

Control of glucagon-like immunoreactive peptide secretion from fetal rat intestinal cultures

Some of the mechanisms underlying intestinal glucagon-like immunoreactive (GLI) peptide secretion from cultured fetal rat intestinal cells were investigated using modulators of the adenylate cyclase pathway [(Bu)2cAMP, theophylline, isobutylmethylxanthine], calcium fluxes (ionomycin, A23187), and protein kinase-C (phorbol ester). All of these agents were found to stimulate GLI peptide release, to 120-230% of paired control values (P less than 0.05-0.001). (Bu)2cAMP, but not the phorbol ester, also increased the total cell content of GLI peptides over the 2-h incubation period (P less than 0.05). No synergism between any of the three pathways was detected. When the mol wt distribution of the stored and secreted GLI peptides was determined in control and (Bu)2 cAMP-stimulated samples, 68 +/- 2% of the peptide corresponded to glicentin, while the remainder eluted with the same distribution coefficient as oxyntomodulin. No 3.5K glucagon was detected in any of the extracts. GLI peptide secretion by the cells was not altered by several pancreatic glucagon secretagogues (cortisol, bombesin, and prostaglandins E1 and D2), but was stimulated by the opioid peptide beta-endorphin (1 microM; P less than 0.02). These studies have indicated that the control of secretion of fetal rat intestinal GLI peptides is complex, involving activation of any one or a combination of the three major second messenger systems. A role for the adenylate cyclase pathway in regulating GLI peptide biosynthesis is also suggested.     

Prehepatic portal hypertension in rats modifies norepinephrine metabolism in hypothalamus medulla oblongata and portal vein

The present experiments investigated the possible relationship between portal hypertension and norepinephrine metabolism in the central nervous system (hypothalamus and medulla oblongata) and the portal vein in the rat. Group I (72), portal hypertensive, and group II (70) sham-operated animals, were sacrificed day 14, and endogenous norepinephrine content, uptake and release from hypothalamus, medulla oblongata, and portal vein were investigated. In group I our results showed increases in norepinephrine storage (69%; 8.3%) and release (19.7%; 43.8%) and a diminished uptake (42.3%; 27.5%) in the hypothalamus and medulla oblongata, respectively. Portal veins showed a decreased content and uptake (62.5% and 43.5%, respectively) and increased release (25%) compared to group II rats. These results suggest a close relationship between the central nervous system and rat portal hypertension, perhaps related to modifications of central sympathetic activity.This work was supported in part by Consejo de Investigaciones Técnicas y Cientificas of Argentina (CONICET) by grants PIA 2042/90 and 0738/91-010 and by the University of Buenos Aires grant FA 031.     

Identification and localization of glucagon-related peptides in rat brain.

Immunochemical and immunocytochemical techniques have been used to identify and characterize glucagon-related peptides of the rat central nervous system. These peptides show immunoreactivity with antiglucagon sera directed towards the central portion of the hormone, but not with antisera specific for the free COOH terminus of glucagon. Highest concentrations were found in hypothalamus (6.1 +/- 1.6 ng/g wet weight) although lower amounts (approximately 2 ng/g) were found in cortex, thalamus, cerebellum, and brain stem. Gel filtration of brain extracts revealed at least two immunoreactive forms, which have molecular weights of about 12,000 and 8000. Both peptides had radioimmunoassay dilution curves parallel to the curve for glucagon and both had identical counterparts in extracts of rat intestine. Digestion of the brain and intestinal peptides with trypsin plus carboxypeptidase B released the immunoreactive COOH-terminal tryptic fragment of pancreatic glucagon from these larger forms. Immunocytochemical studies using antiglucagon serum and peroxidase-antiperoxidase staining identified glucagon-like material in neuronal cell bodie and processes in the magnocellular portion of the paraventricular nucleus, as well as in scattered cells in the supraoptic nucleus and in fibers in the median eminence. These results suggest that glucagon-containing peptides that have undergone the intestinal type of posttranslational modification are present in neuronal cells of the rat hypothalamus.     

Immunoreactive calcitonin in brain regions and pituitary of sheep

In this study we have investigated the presence of immunoreactive calcitonin in the central nervous system and pituitary of sheep. The calcitonin concentrations were determined radioimmunologically by two different antibodies. We have demonstrated calcitonin in extracts of areas of the central nervous system, whole pituitary, thyroid gland and plasma of 21 sheep. The concentrations were (ng/g wet weight, mean values +/- SE): thyroid 16.0 +/- 4.4, pituitary 2.03 +/- 0.34, reticular formation 1.64 +/- 0.25, substantia nigra 1.53 +/- 0.46, dentate nucleus 1.11 +/- 0.27, putamen 1.05 +/- 0.35, hippocampus 0.97 +/- 0.17, fornix 0.96 +/- 0.15, anterior thalamus 0.92 +/- 0.28, mammillary body 0.88 +/- 0.12, cerebellum 0.86 +/- 0.09, caudate nucleus 0.84 +/- 0.11, posterior hypothalamus 0.83 +/- 0.19, epiphysis 0.75 +/- 0.25, thalamus centralis 0.71 +/- 0.10, almond nucleus 0.69 +/- 0.16, medulla oblongata 0.67 +/- 0.15, anterior hypothalamus 0.66 +/- 0.20, precentral gyrus 0.66 +/- 0.16, globus pallidus 0.63 +/- 0.31, postcentral gyrus 0.36 +/- 0.08 and plasma (ng/ml) 0.058 +/- 0.013. Our results demonstrate that immunoreactive calcitonin is present in the central nervous system (CNS) of sheep, compatible with a neurotransmitter function for this hormone.     

Fetal rat intestinal cells in monolayer culture: a new in vitro system to study the glucagon-like immunoreactive peptides

To establish an in vitro model to investigate the glucagon-related peptides, fetal rat intestinal cells were enzymatically dispersed and placed into culture for up to 7 days. After 1 day in culture, the presence of epithelial-like cells containing glucagon-like immunoreactivity (GLI) was demonstrated using immunocytochemical techniques. The cell peptides were extracted by passage through a cartridge of octadecylsilyl silica and characterized by gel filtration and RIA. Two GLI moieties were detected with apparent mol wts of 11,000-12,000 and 5,000-6,000. The immunoreactive profile obtained for the cells in culture was identical to that of both whole fetal rat intestine and adult rat ileum. The presence of glucagon could not be demonstrated in any of the extracts. The basal levels of GLI and apparent immunoreactive glucagon (IRGa) were 1,457 +/- 381 and 198 +/- 57 pg/dish, respectively, on day 1 of culture. The GLI content of the cells, but not the IRGa, declined with time in culture for up to 5-7 days (P less than 0.03). Addition of insulin to the culture medium (10 or 100 mU/ml) did not influence the decrease in GLI content of the cells, but did inhibit the production of IRGa (P less than 0.05). Addition of 500 mg/dl glucose to the cells in the presence of 20 microU/ml insulin increased the secretion of GLI by 42 +/- 7% over 2 h (P less than 0.05). The stimulation by glucose was not seen in the absence of insulin or with higher insulin concentrations (100 microU/ml), nor did insulin alone (100 microU/ml) have any effect on the release of GLI. Thus, fetal rat intestinal cells in culture produce the GLI peptides, and secrete them in response to glucose. This system may provide a means by which the synthesis and control of secretion of the glucagon-related peptides can be investigated.     

Brain/gut peptides in fed and fasted rats

The concentrations and contents of vasoactive intestinal peptide (VIP) and cholecystokinin (CCK) in the brain and of these peptides along with secretin and glucagon-like immunoreactivity (GLI) in the gut were compared in a group of 16 5-day fasted adult Sprague-Dawley rats with the corresponding peptides in a group of 16 nonfasted littermates. The mean weight of the fasted rats at the beginning of the study was 263 +/- 10 g (+/- SEM) and was 177 +/- 7 g before killing, for a net loss of 33% of initial body weight; the 16 fed rats increased their mean weight from 225 +/- 11 to 284 +/- 12 g, for a net gain of 12%. During the 5-day fast there was no change in the weight of the cortex, hypothalamus, or brain stem. However, the weight of tissues from the gut decreased to about half the weight of the corresponding tissues in the fed animals. There was no significant change in brain VIP or CCK. VIP content in the gut was unchanged. However, because of the decrease in organ weight, its concentration almost doubled. Secretin concentrations in the gut of fasted rats did not change significantly, but organ contents fell to about half. The gut content of GLI also fell by half or more. The concentrations of CCK in methanol extracts of the duodenum and jejunum remained relatively constant, but those in acid extracts fell by 40% in the fasted animals. This represents an approximately 70% decrease in organ content of CCK. These findings are interpretable as demonstrating that during a prolonged fast neuronal CCK and VIP are well conserved, but endocrine CCK, secretin, and GLI are markedly decreased because of loss of intestinal mucosa.     

Modulation of norepinephrine release by galanin in rat medulla oblongata.

Galanin, a 29-amino acid peptide, is widely distributed in both the central and peripheral nervous systems and is colocalized with catecholamines, although its physiological significance remains to be elucidated. In the present study we investigated the regulatory mechanisms of galanin on norepinephrine release in rat medulla oblongata. In slices of medulla oblongata of Sprague-Dawley rats, galanin inhibited the stimulation-evoked [3H]norepinephrine release in a concentration-dependent manner (fractional release ratio during electrical stimulation: control 0.937 +/- 0.043, mean +/- SEM, n = 6; galanin 1 x 10(-7) M 0.501 +/- 0.037, n = 6, p less than 0.05; and galanin 1 x 10(-6) M 0.299 +/- 0.018 n = 6, p less than 0.05). Galanin potentiated inhibition of [3H]norepinephrine release by the alpha 2-agonists (UK 14,304 and clonidine). The blockade of alpha 2-adrenergic receptors by RX 781094 diminished the inhibition of norepinephrine release by galanin. Pretreatment of pertussis toxin, which interferes with the coupling of inhibitory guanosine triphosphate-binding proteins to adenylate cyclase, significantly attenuated the suppressive effects of galanin on norepinephrine release. In slices of medulla oblongata obtained from spontaneously hypertensive rats (SHR), the inhibitory effect of galanin on norepinephrine release was significantly less than in those from age-matched Wistar-Kyoto rats. These results show that galanin might inhibit the stimulation-evoked norepinephrine release in rat medulla oblongata, at least partially mediated by alpha 2-adrenergic receptors and the pertussis toxin-sensitive guanosine triphosphate-binding proteins. Moreover, less suppression of norepinephrine release by galanin in SHR suggests that galanin might be involved in the regulation of central sympathetic nervous activity in hypertension.     

Decreased hypothalamic and medullary GABA turnover in spontaneously hypertensive rats.

OBJECTIVE: The aim was to assess whether Gamma-aminobutyric acid (GABA) neurone activities in the central nervous system, especially in the hypothalamus and medulla oblangata, are altered in hypertension. METHODS: Central GABA content and turnover rate were measured in spontaneously hypertensive rats (SHR) and their normotensive Wistar Kyoto controls (WKY). GABA content was determined with high performance liquid chromatography, and in vivo GABA turnover rates were estimated by GABA accumulation after injection of amino-oxyacetic acid, a selective inhibitor of GABA degrading system. Two groups of nine week old male rats (32 SHR and 32 WKY) were used. RESULTS: GABA concentrations in cerebrospinal fluid were lower in SHR than in WKY. Since hypothalamus and medulla oblongata are the possible active sites of this system, basal GABA contents and in vivo GABA turnover rates were measured in hypothalamus and medulla oblongata. Basal GABA content in the medulla oblongata and hypothalamus was almost equal in SHR and WKY. On the other hand, GABA turnover rates were significantly lower in SHR than in WKY in both the hypothalamus and the medulla. CONCLUSIONS: Since it is known that GABA is an inhibitory neurotransmitter in the central nervous system and that it controls autonomic and cardiovascular activities, the findings suggest that the decreased hypothalamic and medullary GABAergic activities may permit sympathetic hyperactivity to contribute to the increase in blood pressure in SHR.     

Response of plasma glicentin to fat ingestion in piglets

In order to elucidate the response of plasma glicentin to fat ingestion, butter, glycerol or palmitate was administered into the duodenum of piglets in a fully conscious state and plasma glicentin and glucagon were determined. Butter instillation did not change blood glucose. Plasma triglyceride rose gradually 120 min after butter loading. Plasma insulin and glucagon measured by antiserum specific to the C-terminal slightly increased following butter administration and plasma total glucagon and glicentin increased gradually and significantly. The increments of total glucagon and glicentin were 179 and 158%, respectively. However, chromatography of porcine plasma obtained during fat loading revealed heterogeneity of glicentin-related peptides. Glycerol ingestion induced a slight rise of plasma total glucagon. Administration of palmitate revealed an increase in plasma total glucagon and glicentin. The present study clearly demonstrates the secretion of glicentin following fat ingestion, which might be caused by the hydrolysates of triglyceride, as suggested in previous dog experiments.     

The forgotten members of the glucagon family

From proglucagon, at least six final biologically active peptides are produced by tissue-specific post-translational processing. While glucagon and GLP-1 are the subject of permanent studies, the four others are usually left in the shadow, in spite of their large biological interest. The present review is devoted to oxyntomodulin and miniglucagon, not forgetting glicentin, although much less is known about it. Oxyntomodulin (OXM) and glicentin are regulators of gastric acid and hydromineral intestinal secretions. OXM is also deeply involved in the control of food intake and energy expenditure, properties that make this peptide a credible treatment of obesity if the question of administration is solved, as for any peptide. Miniglucagon, the C-terminal undecapeptide of glucagon which results from a secondary processing of original nature, displays properties antagonistic to that of the mother-hormone glucagon: (a) it inhibits glucose-, glucagon- and GLP-1-stimulated insulin release at sub-picomolar concentrations, (b) it reduces the in vivo insulin response to glucose with no change in glycemia, (c) it displays insulin-like properties at the cellular level using only a part of the pathway used by insulin, making it a good basis for developing a pharmacological workaround of insulin resistance.     

肾上腺髓质素在中枢神经系统内的分布

目的研究肾上腺髓质素（ＡｄＭ）在中枢神经系统内的分布。方法应用免疫组织化学（ＡＢＣ法）和反转录－聚合酶链式反应（ＲＴ－ＰＣＲ）方法，观察免疫活性的ＡｄＭ及其ＡｄＭｍＲＮＡ在大鼠和人脑内的分布。结果在所检查的大鼠每一个脑区，包括大脑皮层、室旁组织、下丘脑、中脑、延髓和小脑均发现有免疫活性的ＡｄＭ及ＡｄＭｍＲＮＡ存在，应用半定量ＲＴ－ＰＣＲ分析方法发现，在大鼠的室旁组织和延髓中，ＡｄＭｍＲＮＡ表达水平较高。在人脑也发现有ＡｄＭｍＲＮＡ表达。结论在大鼠和人的中枢神经系统中存在肾上腺髓质素，它可能作为一种神经递质、神经调质或神经激素发挥生物学作用。     

Alterations in catecholamine release in the central nervous system of spontaneously hypertensive rats.

The aim of the present study was to investigate alterations in catecholamine release in the central nervous system of spontaneously hypertensive rats. Slices of hypothalamus, medulla oblongata and striatum were prepared from spontaneously hypertensive rats (SHR: 9-10 weeks old) and age-matched Wistar Kyoto rats (WKY). The slices were incubated with (3H)norepinephrine (NE) or (3H)dopamine (DA), superfused with Krebs-solution in vitro, and the release of the catecholamines was compared between the two strains. The basal release of hypothalamic (3H)NE did not differ between SHR and WKY slices. However, stimulation (1 Hz)-evoked (3H)NE release was significantly greater in SHR than in WKY (percent fractional release of total tissue NE: WKY 0.494 +/- 0.019%, n = 6, SHR 0.730 +/- 0.053%, n = 6, p less than 0.05). The stimulation-evoked (3H)NE release from the medulla oblongata did not differ significantly between SHR and WKY slices. Finally stimulation-evoked release of striatal (3H)DA was significantly depressed in SHR (percent fractional release of total tissue DA: WKY 2.048 +/- 0.24%, n = 6, SHR 1.460 +/- 0.068%, n = 6, p less than 0.05). These results indicate that the release of hypothalamic NE and striatal DA are altered in SHR. It is suggested that enhanced hypothalamic noradrenergic activity and reduced striatal dopaminergic activity can increase sympathetic outflow to the periphery, which may play a role in the pathogenesis of this form of hypertension.     

Regulation of intestinal proglucagon-derived peptide secretion by intestinal regulatory peptides.

The physiological regulation of intestinal proglucagon-derived peptide secretion has not been well studied. We have therefore used a fetal rat intestinal cell culture model to investigate the control of secretion of the gut glucagon-like immunoreactive (GLI) peptides by other intestinal regulatory peptides in vitro. Secretion of the intestinal GLI peptides was found to be stimulated in a dose-dependent fashion by the intestinal endocrine peptide, gastric inhibitory peptide (at greater than or equal to 10(-10) M, P less than 0.05), and by the neurocrine peptides, gastrin-releasing peptide (at greater than or equal to 10(-12) M, P less than 0.05), and calcitonin gene-related peptide (at greater than or equal to 10(-8) M, P less than 0.05). Gastrin-releasing peptide and its amphibian equivalent, bombesin were equipotent in stimulating GLI peptide secretion. In contrast, the endocrine and neurocrine intestinal somatostatin-related peptides, somatostatin-28 and -14, inhibited release of the GLI peptides, at concentrations of 10(-10) (P less than 0.01) and 10(-8) (P less than 0.01) M, respectively, with significant differences in potency between the two peptides detected at 10(-10) M (P less than 0.05). The inhibitory effects of both somatostatin-28 and -14 could be blocked by preincubation of the cells with pertussis toxin (P less than 0.05). Dose-dependent stimulation of gut GLI peptide secretion was also detected in response to treatment of cultured cells with sodium oleate (at 10(-4) M; P less than 0.05), or with the cholinergic agonist bethanecol (at greater than or equal to 100 microM; P less than 0.05). Other endocrine [cholecystokinin, glucagon, glucagon-like peptide-1(1-37), glucagon-like peptide-1(7-37), glucagon-like peptide-2, neurotensin, and peptide YY] and neurocrine (vasoactive intestinal peptide) peptides, and the synthetic glucocorticoid, dexamethasone, were without effect on secretion of the gut GLI peptides, at doses of 10(-12) to 10(-6) M. The results of the present study therefore demonstrate that secretion of the intestinal proglucagon-derived peptides is under the regulatory control of a wide variety of intestinal endocrine and neurocrine peptides, as well as nutrients (fats) and neurotransmitters (acetylcholine).     

Glucagon-like immunoreactive peptides in a rat ileal epithelial cell line (IEC-18)

The presence of cells containing glucagon-like immunoreactive (GLI) peptides was demonstrated in a rat ileal epithelial cell line (IEC-18) by both immunofluorescence and radioimmunoassay. When cell extracts were subjected to gel filtration chromatography, the cells were found to contain 3.5 Kd glucagon in addition to significant quantities of large molecular weight GLI peptides (apparent molecular weights of 4, 6, 8 and 10 Kd) and a 9 Kd peptide with apparent glucagon immunoreactivity. This was in contrast to extracts of adult rat ileum, which contained only large molecular weight GLI peptides (apparent molecular weights of 6 and 12 Kd). Production of GLI peptides by the IEC-18 cells was stimulated by glucose (p less than 0.02) and inhibited by insulin (p less than 0.01). In conclusion, these results demonstrate that the IEC-18 cells produce both GLI peptides and glucagon, and thus support the notion that proglucagon processing is cell-specific. IEC-18 cells may therefore provide a tool for investigations of some aspects of GLI peptide and glucagon synthesis.     

Regional brain concentrations of calcitonin gene-related peptide in spontaneously hypertensive and normotensive Wistar-Kyoto rats

The regional brain and spinal cord concentrations of calcitonin gene-related peptide (CGRP) were measured in age-matched (22-23-week-old) spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats. The highest concentration of CGRP in the WKY rats was in the spinal cord (172 +/- 9 pmol/g), followed by the medulla oblongata/pons (88 +/- 5 pmol/g). The relative order of distribution in the remaining regions was: hypothalamus (12.6 +/- 0.8 pmol/g) = striatum greater than thalamus greater than midbrain = hippocampus greater than cortex (2.1 +/- 0.3 pmol/g). The concentration of CGRP in the cerebellum was at the level of the assay's sensitivity (0.5 pmol/g). The relative order of distribution in the SHR strain was essentially the same. However, in comparison with the WKY rats, the SHR had significantly lower levels of CGRP in the hippocampus (-47%), striatum (-49%) and medulla oblongata/pons (-24%), and in the spinal cord (-24%). In younger age-matched (16-17-week-old) rats, the spinal cord and medulla oblongata/pons concentrations of CGRP were also lower in SHR than in WKY rats. CGRP is a putative neurotransmitter which, when administered centrally or peripherally, has potent cardiovascular effects. The reduced levels of this peptide may be an important factor in the cardiovascular and/or behavioural abnormalities of the SHR strain.