Voluntary exercise is motivated by ghrelin, possibly related to the central reward circuit.

We previously reported that voluntary exercise contributed to the amelioration of abnormal feeding behavior with a concomitant restoration of ghrelin production in a rat model of obesity, suggesting a possible relationship between exercise and appetite-regulating hormones. Ghrelin is known to be involved in the brain reward circuits via dopamine neurons related to motivational properties. We investigated the relevance of ghrelin as an initiator of voluntary exercise as well as feeding behavior. The plasma ghrelin concentration fluctuates throughout the day with its peak at the beginning of the dark period in the wild-type (WT) mice with voluntary exercise. Although predominant increases in wheel running activity were observed accordant to the peak of plasma ghrelin concentration in the WT mice, those were severely attenuated in the ghrelin-knockout (GKO) mice under either ad libitum or time-restricted feeding. A single injection of ghrelin receptor agonist brought about and reproduced a marked enhancement of wheel running activity, in contrast to no effect by the continuous administration of the same drug. Brain dopamine levels (DAs) were enhanced after food consumption in the WT mice under voluntary exercise. Although the acceleration of DAs were apparently blunted in the GKO mice, they were dramatically revived after the administration of ghrelin receptor agonist, suggesting the relevance of ghrelin in the reward circuit under voluntary exercise. These findings emphasize that the surge of ghrelin plays a crucial role in the formation of motivation for the initiation of voluntary exercise possibly related to the central dopamine system.

Beneficial effects of metformin on energy metabolism and visceral fat volume through a possible mechanism of fatty acid oxidation in human subjects and rats.

OBJECTIVE: Metformin is known to have a beneficial effect on body weight and body composition, although the precise mechanism has not been elucidated yet. The aim of this study is to investigate the effects of metformin on energy metabolism and anthropometric factors in both human subjects and rats. METHODS: In human studies, metformin (1500mg/day) was administered to 23 healthy subjects and 18 patients with type 2 diabetes for 2 weeks. Metabolic parameters and energy metabolism were measured during a meal tolerance test in the morning before and after the treatment of metformin. In animal studies, 13 weeks old SD rats were fed 25-26 g of standard chow only during 12-hours dark phase with either treated by metformin (2.5mg/ml in drinking water) or not for 2 weeks, and metabolic parameters, anthropometric factors and energy metabolism together with expressions related to fat oxidation and adaptive thermogenesis were measured either in fasting or post-prandial state at 15 weeks old. RESULTS: Post-prandial plasma lactate concentration was significantly increased after the metformin treatment in both healthy subjects and diabetic patients. Although energy expenditure (EE) did not change, baseline respiratory quotient (RQ) was significantly decreased and post-prandial RQ was significantly increased vice versa following the metformin treatment in both groups. By the administration of metformin to SD rats for 2 weeks, plasma levels of lactate and pyruvate were significantly increased in both fasting and post-prandial states. RQ during a fasting state was significantly decreased in metformin-treated rats compared to controls with no effect on EE. Metformin treatment brought about a significant reduction of visceral fat mass compared to controls accompanied by an up-regulation of fat oxidation-related enzyme in the liver, UCP-1 in the brown adipose tissue and UCP-3 in the skeletal muscle. CONCLUSION: From the results obtained, beneficial effects of metformin on visceral fat reduction has been demonstrated probably through a mechanism for a potential shift of fuel resource into fat oxidation and an upregulation of adaptive thermogenesis independent of an anorexigenic effect of this drug.

Experimental validation of talc pleurodesis for carcinomatous pleuritis in an animal model.

OBJECTIVE: This study was performed to evaluate the change of pleural cavity by talc pleurodesis for carcinomatous pleuritis in an animal model. METHODS: Models of malignant pleuritis were produced by an intra-venous injection of lung adenocarcinoma cells to male BALB/c nude mice at 6 weeks of age. Two weeks after the injection, either talc or saline was injected into the left thoracic cavity and the mice were further observed for 4 weeks. Six weeks after the injection, they were killed and the occurrence of lung cancer, amount of pleural effusion, and histological change of the pleural cavity were examined and compared between the two groups with or without talc administration. RESULTS: Talc administration caused a significant reduction in pleural effusion with no increase of pleural adhesion. Talc administration resulted in marked pleural thickening compared to saline treatment. The vascular architecture and its area did not differ between the two groups. CONCLUSION: Talc pleurodesis to reduce pleural effusion is valid for carcinomatous pleuritis, potentially through an acceleration of pleural thickening.

Voluntary exercise contributed to an amelioration of abnormal feeding behavior, locomotor activity and ghrelin production concomitantly with a weight reduction in high fat diet-induced obese rats.

In the present study, effects of voluntary exercise in an obese animal model were investigated in relation to the rhythm of daily activity and ghrelin production. Male Sprague-Dawley rats were fed either a high fat diet (HFD) or a chow diet (CD) from four to 16 weeks old. They were further subdivided into either an exercise group (HFD-Ex, CD-Ex) with a running wheel for three days of every other week or sedentary group (HFD-Se, CD-Se). At 16 weeks old, marked increases in body weight and visceral fat were observed in the HFD-Se group, together with disrupted rhythms of feeding and locomotor activity. The induction of voluntary exercise brought about an effective reduction of weight and fat, and ameliorated abnormal rhythms of activity and feeding in the HFD-Ex rats. Wheel counts as voluntary exercise was greater in HFD-Ex rats than those in CD-Ex rats. The HFD-obese had exhibited a deterioration of ghrelin production, which was restored by the induction of voluntary exercise. These findings demonstrated that abnormal rhythms of feeding and locomotor activity in HFD-obese rats were restored by infrequent voluntary exercise with a concomitant amelioration of the ghrelin production and weight reduction. Because ghrelin is related to food anticipatory activity, it is plausible that ghrelin participates in the circadian rhythm of daily activity including eating behavior. A beneficial effect of voluntary exercise has now been confirmed in terms of the amelioration of the daily rhythms in eating behavior and physical activity in an animal model of obesity.

Changes in the nitric oxide-soluble guanylate cyclase system and natriuretic peptide receptor system in placentas of pregnant Dahl salt-sensitive rats.

AIM: Diminished vasodilator activity during pregnancy, which augments vascular responses to vasoconstrictors, is one reason for the onset of pre-eclampsia and superimposed pre-eclampsia. It is known that Dahl salt-sensitive (Dahl-S) rats develop salt-sensitive hypertension like African-Americans. The present study attempted to assess the changes and the interactions of the NOS-NO-sGC-cGMP and NP-NPR-cGMP systems in the hypertensive placenta using Dahl-S rats as an animal model of superimposed pre-eclampsia. MATERIAL AND METHODS: Pregnant Dahl-S rats were fed a high-salt diet to induce the development of hypertension and fetal growth restriction. Using these rats, we investigated the regulation of these two vasodilatation systems, including the kinetics of cyclic guanosine monophosphate (cGMP), soluble guanylate cyclase (sGC), endothelial nitric oxide synthase (NOS), cytokine-inducible NOS, natriuretic peptides (NP) (atrial NP, brain NP and C-type NP), and NP receptors (NPR) (NPR-A, NPR-B, NPR-C). RESULTS: Dahl-S rats fed a high-salt diet exhibited hypertension, fetal growth restriction and thickening of the walls in decidual vessels. The placental cGMP level in the rats fed the high-salt diet was significantly decreased compared with that in controls. The expression levels of endothelial NOS and cytokine-inducible NOS mRNA increased significantly, while that of sGCα2-sunbnit declined significantly. Messenger RNA levels of NPR-C, a clearance-type receptor of NP, declined significantly, whereas those of NP and their functional receptors NPR-A and NPR-B were unchanged. CONCLUSIONS: As Dahl-S rats with excess salt-loading during pregnancy exhibited pathological changes similar to those observed in female humans with pre-eclampsia/superimposed pre-eclampsia, this rat could be useful as an animal model of superimposed pre-eclampsia. In the placentas of hypertensive Dahl-S rats, vasodilatation seemed to be disturbed by the deregulation of both the NO-sGC-cGMP and NP-NPR-cGMP systems.

Changes in Subcellular Distribution of n-Octanoyl or n-Decanoyl Ghrelin in Ghrelin-Producing Cells.

BACKGROUND: The enzyme ghrelin O-acyltransferase (GOAT) catalyzes the acylation of ghrelin. The molecular form of GOAT, together with its reaction in vitro, has been reported previously. However, the subcellular processes governing the acylation of ghrelin remain to be elucidated. METHODS: Double immunoelectron microscopy was used to examine changes in the relative proportions of secretory granules containing n-octanoyl ghrelin (C8-ghrelin) or n-decanoyl ghrelin (C10-ghrelin) in ghrelin-producing cells of mouse stomachs. The dynamics of C8-type (possessing C8-ghrelin exclusively), C10-type (possessing C10-ghrelin only), and mixed-type secretory granules (possessing both C8- and C10-ghrelin) were investigated after fasting for 48 h or after 2 weeks feeding with chow containing glyceryl-tri-octanoate (C8-MCT) or glyceryl-tri-decanoate (C10-MCT). The dynamics of C8- or C10-ghrelin-immunoreactivity (ir-C8- or ir-C10-ghrelin) within the mixed-type granules were also investigated. RESULTS: Immunoelectron microscopic analysis revealed the co-existence of C8- and C10-ghrelin within the same secretory granules (mixed-type) in ghrelin-producing cells. Compared to control mice fed standard chow, the ratio of C10-type secretory granules increased significantly after ingestion of C10-MCT, whereas that of C8-type granules declined significantly under the same treatment. After ingestion of C8-MCT, the proportion of C8-type secretory granules increased significantly. Within the mixed-type granules the ratio of ir-C10-ghrelin increased significantly and that of ir-C8-ghrelin decreased significantly upon fasting. CONCLUSION: These findings confirmed that C10-ghrelin, another acyl-form of active ghrelin, is stored within the same secretory granules as C8-ghrelin, and suggested that the types of medium-chain acyl-molecules surrounding and available to the ghrelin-GOAT system may affect the physiological processes of ghrelin acylation.

Blood pressure variability activates cardiac mineralocorticoid receptor and induces cardiac remodeling in hypertensive rats.

BACKGROUND: Hypertensive patients with large blood pressure variability (BPV) have aggravated target organ damage. Because the aldosterone/mineralocorticoid receptor (MR) system is a possible mechanism of hypertensive organ damage, we investigated in spontaneously hypertensive rats (SHRs) whether a specific MR blocker, eplerenone, would prevent BPV-induced aggravation of hypertensive cardiac remodeling. METHODS AND RESULTS: A rat model of a combination of hypertension and large BPV was created by performing bilateral sinoaortic denervation (SAD) in SHRs. SAD increased BPV without changing mean BP. SAD induced perivascular macrophage infiltration and aggravated myocardial fibrosis and cardiac hypertrophy, resulting in LV systolic dysfunction. Immunohistostaining revealed SAD-induced translocation of MRs into the nuclei (ie, MR activation) of the intramyocardial arterial medial cells and cardiac myocytes. SAD increased phosphorylation of p21-activated kinase1 (PAK1), a regulator of MR nuclear translocation. Chronic administration of a subdepressor dose of eplerenone prevented MR translocation, macrophage infiltration, myocardial fibrosis, cardiac hypertrophy, and LV dysfunction, while not affecting BPV. Circulating levels of aldosterone and cortisol were not changed by SAD. CONCLUSIONS: Eplerenone inhibited the aggravation of cardiac inflammation and hypertensive cardiac remodeling, and thereby prevented progression of LV dysfunction in SHRs with large BPV. This suggests that the PAK1-MR pathway plays a role in cardiac inflammation and remodeling induced by large BPV superimposed on hypertension, independent of circulating aldosterone.

Ghrelin acylation by ingestion of medium-chain fatty acids.

We found in a primary study that ingestion of medium-chain fatty acids (MCFAs) or medium-chain triacylglycerols (MCTs) increased the stomach contents of acyl ghrelin, and we further showed that the carbon-chain length of the acyl groups that modified the nascent ghrelin peptides corresponded to that of the ingested MCFAs or MCTs. These findings clearly demonstrated that the ingested MCFAs are directly used for the acyl-modification of ghrelin. Before the discovery of ghrelin-O-acyltransferase (GOAT), our in vivo study suggested that the putative GOAT preferred MCTs (composed of C6:0 to C10:0 FFAs) to either short- or long-chain triglycerides. In another study, we suggested that MCFAs or MCTs might represent a potential therapeutic modality for the clinical manipulation of energy metabolism through the modulation of ghrelin activity. After the discovery of GOAT, many studies have been done on the acylation of ghrelin using MCFAs, MCTs, or their derivatives; however, results and interpretations have been inconsistent, largely due to the differences in experimental conditions. This chapter describes detailed methods for the analysis of ghrelin acylation in vivo to facilitate future research in this field.

A-type natriuretic peptide level in angiotensin II type 1a receptor knockout mice.

A-type (atrial) natriuretic peptide (ANP) levels in the heart and plasma were examined by immunohistochemistry, electron microscopy, and radioimmunoassay (RIA) in angiotensin II type 1a receptor knockout (Agtr1a KO) mice. Additionally, the ANP mRNA level in the heart was measured using a real-time polymerase chain reaction (PCR) assay. The blood pressure in Agtr1a KO mice was significantly lower than that in wild-type (WT) mice. The number of ANP granules and ANP immunoreactivity in the auricular cardiocytes were significantly lower in Agtr1a KO mice than in WT mice. Ultrastructurally, the ventricular cardiocytes in Agtr1a KO mice occasionally had ANP-like granules, which were not present in WT mice. The plasma, auricular, and ventricular ANP and plasma cyclic guanosine monophosphate (cGMP) concentrations were significantly higher in Agtr1a KO mice than in WT mice. The ANP mRNA levels of the auricular and ventricular cardiocytes in the Agtr1a KO mice were almost twice as large as those in WT mice. The present data suggest that a notable increase in the ANP biosynthesis and release in the heart of Agtr1a KO mice may account for the reduction in blood pressure together with the lack of an AGTR1A receptor in this model.

Disrupted regulation of ghrelin production under antihypertensive treatment in spontaneously hypertensive rats.

BACKGROUND: Ghrelin is an acylated peptide hormone mainly secreted from the stomach. When administrated externally it modulates vascular tone mainly through the regulation of autonomic nerve activity. However, the effects of blood pressure (BP) on the production and secretion of ghrelin remain to be clarified. METHODS AND RESULTS: We examined the stomach and plasma levels of ghrelin in spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats after a 4-week-intervention with antihypertensive agents (candesartan-cilexetil [ARB], doxazosin [DZN], metoprolol [MP], reserpine [RES]) to clarify the influence of BP on the secretion of ghrelin. The effect of these agents on ghrelin production and secretion were examined by comparing vehicle-treated controls (WKY-Intact, SHR-Intact). Treatment with the 4 antihypertensive drugs all yielded a significant decline in systolic BP in both SHR and WKY. Under these conditions, significantly lower levels of stomach and plasma ghrelin were detected in WKY treated with ARB (P<0.05), DZN (P<0.05), MP (P<0.05) and RES (P<0.05) compared with WKY-Intact, whereas no significant change in the ghrelin levels in the stomach and plasma were detected in SHR under the same treatments. CONCLUSIONS: The findings imply that the production and secretion of ghrelin are controlled by the ambient vascular tone and vice versa in normotensive WKY. This inter-relationship between ghrelin and BP seems to be disrupted in SHR.

Increased production of active ghrelin is relevant to hyperphagia in nonobese spontaneously diabetic Torii rats.

An abnormal eating behavior is often associated with diabetes mellitus in individuals. In the present study, we investigated the mechanisms underlying the relationship among uncontrolled diabetes, food intake, and the production of ghrelin, an orexigenic hormone, in spontaneous diabetic Torii (SDT) rats. Male SDT rats and age-matched control Sprague-Dawley (SD) rats were housed from 8 to 38 weeks of age. Body weight and daily food intake were measured weekly, whereas blood and whole stomach samples were obtained at the age of 8, 25, and 38 weeks in both SDT and SD rats. The SDT rats at both 25 and 38 weeks of age demonstrated significantly lower body weights despite almost doubled food consumption compared with the SD rats of the same age. The SDT rats showed overt hyperglycemia at 25 and 38 weeks of age with concomitant hypoinsulinemia. The plasma active ghrelin levels and the ratio to total ghrelin levels of SDT rats at 38 weeks of age were significantly higher than those of SD rats of the same age. Stomach ghrelin and ghrelin O-acyltransferase messenger RNA expression levels were higher in SDT rats than in SD rats after the induction of diabetes, with a concomitant decrease of stomach ghrelin-immunopositive cell numbers in SDT rats at 38 weeks of age. The SDT rats with uncontrolled hyperglycemia show hyperphagia with a concomitant increase of plasma active ghrelin concentration. This report is the first to clarify the relevance of ghrelin to hyperphagia in diabetic state over an extended period.

Different A-type natriuretic peptide level in five strains of mice.

Atrial (A-type) natriuretic peptide (ANP) is vasodilative hormone involved in the regulation of blood pressure and volume homeostasis. In this study, we examined the differences of the auricular and plasma ANP distribution by immunohistochemistry, ultrastructural morphometry, and radioimmunoassay in five strains of mice. The ANP-immunoreactivities of the auricle were most intense in ICR, and moderate in C57BL/6J and C3H/HeN, and weakest in BALB/cA and DBA/2Cr. The number of ANP-granules was greatest in ICR followed by C57BL, C3H or BALB/c, and DBA/2 mice, in this order. The auricular ANP content was highest in ICR, but the plasma ANP concentration was comparable in all strains. The present study demonstrates that there are differences in the ANP circulating system between five strains.

Exaggerated blood pressure variability superimposed on hypertension aggravates cardiac remodeling in rats via angiotensin II system-mediated chronic inflammation.

Hypertensive patients with large blood pressure variability (BPV) have aggravated end-organ damage. However, the pathogenesis remains unknown. We investigated whether exaggerated BPV aggravates hypertensive cardiac remodeling and function by activating inflammation and angiotensin II-mediated mechanisms. A model of exaggerated BPV superimposed on chronic hypertension was created by performing bilateral sinoaortic denervation (SAD) in spontaneously hypertensive rats (SHRs). SAD increased BPV to a similar extent in Wistar Kyoto rats and SHRs without significant changes in mean blood pressure. SAD aggravated left ventricular and myocyte hypertrophy and myocardial fibrosis to a greater extent and impaired left ventricular systolic function in SHRs. SAD induced monocyte chemoattractant protein-1, transforming growth factor-beta, and angiotensinogen mRNA upregulations and macrophage infiltration of the heart in SHRs. The effects of SAD on cardiac remodeling and inflammation were much smaller in Wistar Kyoto rats compared with SHRs. Circulating levels of norepinephrine, the active form of renin, and inflammatory cytokines were not affected by SAD in Wistar Kyoto rats and SHRs. A subdepressor dose of candesartan abolished the SAD-induced left ventricular/myocyte hypertrophy, myocardial fibrosis, macrophage infiltration, and inductions of monocyte chemoattractant protein-1, transforming growth factor-beta, and angiotensinogen and subsequently prevented systolic dysfunction in SHRs with SAD. These findings suggest that exaggerated BPV induces chronic myocardial inflammation and thereby aggravates cardiac remodeling and systolic function in hypertensive hearts. The cardiac angiotensin II system may play a role in the pathogenesis of cardiac remodeling and dysfunction induced by a combination of hypertension and exaggerated BPV.

Regional distribution and the dynamics of n-decanoyl ghrelin, another acyl-form of ghrelin, upon fasting in rodents.

n-Decanoyl ghrelin (D-ghrelin), a member of ghrelin-derived peptides, is found in plasma and the stomach; however, there have so far been no studies describing its dynamics. A D-ghrelin-specific radioimmunoassay was established to examine the tissue distribution and the kinetics of D-ghrelin in mice. The effect of D-ghrelin on food intake was also examined and compared to n-octanoyl ghrelin (O-ghrelin). D-ghrelin was detected throughout the gastrointestinal tissue and plasma with highest level in the stomach. An immunofluorescent study revealed the co-localization of D- and O-ghrelin in the same stomach cells. Upon fasting, the levels of D-ghrelin in the stomach and plasma significantly increased, while that of O-ghrelin in the stomach declined. D-ghrelin increased the 2 h food consumption in mice as O-ghrelin does. These findings indicate that D-ghrelin is mainly produced in the stomach to work in concert with O-ghrelin. The different kinetics of D- and O-ghrelin in the stomach upon fasting implies the possibility of D-ghrelin-specific bioregulation.

Changes in A-type natriuretic peptide and its receptors induced by a neutral endopeptidase inhibitor in a rat model of sepsis.

PURPOSE: Elevated plasma A-type natriuretic peptide (ANP) levels in sepsis cause fluid transfer into extravascular spaces. We investigated the changes in ANP concentrations and natriuretic peptide receptor (NPR) expression induced by thiorphan, a neutral endopeptidase (NEP) inhibitor, in a rat model of sepsis. METHODS: Fifteen male rats were divided into three groups: a control group (n = 5), a lipopolysaccharide (LPS) group (n = 5), and an LPS-thiorphan group (n = 5). We measured ANP concentrations in the plasma and lung, and NPR mRNA expression in the lung 4 h after administering LPS, and compared the values with those in the control group. RESULTS: Plasma and lung ANP levels in the LPS group were significantly higher than those in the control group (P < 0.05), but were significantly decreased by thiorphan administration (P < 0.05). NPR-A mRNA levels did not differ significantly among the groups. NPR-C mRNA levels in the LPS-thiorphan group were significantly higher than those in the other groups (P < 0.05). CONCLUSIONS: Elevated ANP levels were decreased by thiorphan administration, which increased NPR-C mRNA levels in the lung. Thus, thiorphan might be effective for reducing elevated ANP levels in sepsis.

Immunohistochemical localization of ghrelin in rodent kidneys.

Ghrelin is a novel peptide hormone, originally identified in the rat and human stomach that plays various important roles. In the present study, we report the intra-renal localization of ghrelin in laboratory rodents. Kidneys from 3 month-old mice, rats and hamsters of both sexes were analyzed by immunohistochemistry. Positive signals were clearly observed in the epithelium of the distal tubules, whereas other segments of the nephron or interstitial cells, including juxtaglomerular cells, showed negative reactions. Pre-embedding immunoelectron microscopy revealed positive signals exclusively on the basolateral membrane in the distal tubular cells and in the collecting ducts. In addition, prepro-ghrelin gene expression was assessed by RT-PCR, and the expected 329-bp prepro-ghrelin mRNA was clearly detected in the kidney. On Western blot analysis, although a specific band for ghrelin (3 kDa) was not detected in the kidney, the expected band for prepro-ghrelin (13 kDa) was clearly detected in both the stomach and the kidney. This paper clarified the intra-renal localization of ghrelin.

Developmental changes in the pattern of ghrelin's acyl modification and the levels of acyl-modified ghrelins in murine stomach.

Ghrelin is an acylated peptide hormone secreted primarily from endocrine cells in the stomach. The major active form of ghrelin is a 28-amino acid peptide with an n-octanoyl modification at Ser(3) (n-octanoyl ghrelin), which is essential for its activity. In addition to n-octanoyl ghrelin, other forms of ghrelin peptide exist, including des-acyl ghrelin, which lacks an acyl modification, and other minor acylated ghrelin species, such as n-decanoyl ghrelin, whose Ser(3) residue is modified by n-decanoic acid. Multiple reports have identified various physiological functions of ghrelin. However, until now, there have been no reports that explore the process of ghrelin acyl modification, and only a few studies have compared the levels of des-acyl, n-octanoyl, and/or other minor populations of acylated ghrelin peptides. In this study we report that the amount of n-octanoyl ghrelin in murine stomachs increases gradually during the suckling period to a maximal level at 3 wk of age and falls sharply after the initiation of weaning. However, the concentration (picomoles per milligram of wet weight tissue) of total ghrelin, which includes des-acyl and all acylated forms of ghrelin peptides with intact C termini in murine stomach, remains unchanged across this suckling-weaning transition. Prematurely weaned mice exhibited a significant decrease in the amount of n-octanoyl or n-decanoyl ghrelin in the stomach. Orally ingested glyceryl trioctanoate, a medium-chain triacylglyceride rich in milk lipids, significantly increased the level of n-octanoyl-modified ghrelin in murine stomach. Fluctuations in the proportion of this biologically active, acyl-modified ghrelin could contribute to or be influenced by the change in energy metabolism during the suckling-weaning transition.

Characterization and species differences in gastric ghrelin cells from mice, rats and hamsters.

Ghrelin is a newly identified gastric peptide hormone that has various important functions, including growth-hormone release and appetite stimulation. Ghrelin-immunoreactive cells (ghrelin cells) are characterized by X-type endocrine cells in the rat stomach. In the present study, we analysed ghrelin cells in fundi of stomach from ICR mice and Syrian hamsters immunohistochemically, immunoelectron microscopically and morphometrically, and compared the results with those from Wistar rats. Immunohistochemistry revealed that ghrelin cells were sparsely distributed in the proper gastric glands in all species. The number of ghrelin cells per unit area in hamsters was significantly lower than that in rats. Immunoelectron microscopy detected ghrelin immunolabelling in granules in the X-type endocrine cells. However, the diameter of granules in the hamsters was significantly smaller than that in the mice and rats. Gastric ghrelin contents were determined by radioimmunoassay, and levels in the hamsters were significantly lower than those in mice and rats. The results from mice were identical to those from rats. In conclusion, gastric ghrelin cells in mice and hamsters are characterized by X-type endocrine cells, as has been observed in rats. However, the data indicated that gastric ghrelin production was lower in hamster than in mouse or rat.

A-type natriuretic peptide level in hypertensive transgenic mice.

A-type (atrial) natriuretic peptide (ANP) levels in heart and plasma were examined by immunohistochemistry, electron microscopy, and radioimmunoassay (RIA) in hypertensive transgenic mice (Tsukuba hypertensive mice; THM). Additionally, the ANP mRNA level in the heart was measured using real-time polymerase chain reaction (PCR) assay. The blood pressure and the ratio of heart weight to body weight in THM was significantly higher than those in the control mice (C57BL/6J). The number of ANP-granules and ANP immunoreactivity in the auricular cardiocytes were significantly lower in THM than in the control. Ultrastructurally, the ventricular cardiocytes in the THM occasionally had ANP-like granules, which were not present in the controls. Using RIA, the plasma, auricular, and ventricular ANP concentrations were significantly higher in THM than in the control, but there was no significant difference in plasma cyclic guanosine monophosphate (GMP) concentration between THM and the control. The ANP mRNA levels of the auricular and ventricular cardiocytes in the THM were siginificantly higher than those in the controls. The present study suggested that the ANP release system of the auricular cardiocytes in these transgenic mice is different from normal (control mice).

Effectiveness of angiotensin-converting enzyme inhibitor or angiotensin II receptor blocker on atrial natriuretic peptide.

The aim of this study was to evaluate the effectiveness of an angiotensin-converting enzyne inhibitor (ACEI, quinapril) or angiotensin II receptor blocker (ARB, candesartan) on atrial natriuretic peptide (ANP) activity in rats with hypertension induced by nitric oxide (NO) inhibition. ACEI and ARB have a number of pharmacologic effects, including blood pressure reduction, myocardial preservation, and an unknown effect in the circulation. The changes in ANP in NO inhibitor-induced hypertensive rats were evaluated in order to elucidate the interaction between ANP and NO in the regulation of blood pressure. Thirty-six rats were divided into 4 groups and administered the experimental agents for 8 weeks: group CONTROL was given regular food (n=9), group N(G)-nitro-L-arginine (L-NNA) was administered L-NNA (25 mg. kg(-1). day(-1), n=9), group ACEI was administered L-NNA and quinapril (10 mg. kg(-1). day(-1), n=9), and group ARB was administered L-NNA and candesartan (10 mg. kg(-1). day(-1), n=9). Blood pressure, plasma ANP, atrial ANP, ANP mRNA, and ANP granules were measured. A significant elevation in blood pressure was observed in group L-NNA. However, there were no increases in plasma ANP (L-NNA: 138.8+/-64.4, CONTROL: 86.7+/-36.4), ANP mRNA (L-NNA: 2.2+/-1.0, CONTROL: 1.7+/-0.5) or ANP granules (L-NNA: 61.1+/-10.2, CONTROL: 64.5+/-8.5). No increase in blood pressure was seen in groups ACEI and ARB. However, plasma ANP (ACEI: 1,392.3+/-1,034.4, ARB: 1,142.8+/-667.3), ANP mRNA (ACEI: 52.8+/-29.1, ARB: 42.9+/-21.2), and ANP granules (ACEI: 122.5+/-23.4, ARB: 136.3+/-33.2) increased significantly. NO inhibitor-induced hypertension caused no changes in ANP concentrations. However, the ACEI and ARB had a direct effect on the induction of ANP secretion. The findings suggest that ANP secretion is directly effected by ACEI and ARB, which seems to play a key role in lowering blood pressure, relieving heart failure symptoms, and preserving the myocardium.