1. Universidad San Pablo-CEU

The effect of the histamine H3 receptor antagonist thioperamide on morphine reinforcement and physical dependence was studied by using the conditioned place preference paradigm and naloxone-precipitated withdrawal in the rat. Coadministration of low doses of thioperamide (0.2 and 2 mg/kg) with morphine did not produce consistent changes of the place conditioning effect of the opioid; however, the highest dose of thioperamide tested (10 mg/kg) completely blocked morphine preference. This effect did not seem to result from thioperamide-induced dysphoria since this drug did not elicit place aversion per se. Thioperamide (10 mg/kg) failed to alter the signs of opiate withdrawal when acutely injected in morphine-dependent rats 1 h before naloxone administration. These results show that thioperamide could decrease the positive reinforcing properties of morphine, an effect that could be related to an activation of the central histaminergic function. On the other hand, the results obtained do not support a prominent role for histamine H3 receptors in the expression of opiate withdrawal.

Cholecystokinin (CCK) is a postprandial hormone that elicits a satiating effect and regulates feeding behaviour. CCK has been shown to enhance the effect of leptin in several experimental paradigms. The goal of this work was to characterize the effect of endogenous CCK on plasma leptin content by using CCK receptor antagonists. Therefore, we administered SR-27,897, a selective CCK1 receptor antagonist, and L-365,260, a selective CCK2 receptor antagonist, to fed and food-deprived rats and determined plasma leptin concentration by enzyme immunoassay. Plasma insulin and glucose concentration as well as food intake were also determined. Under our conditions, SR-27,897 increased plasma concentration of leptin both in fed and food-deprived rats. It also increased food intake as well as plasma concentration of insulin in fed animals. L-365,260 increased plasma leptin concentration only in fed rats. In animals receiving exogenous leptin, CCK-8 increased the ratio between the concentration of leptin in cerebrospinal fluid and plasma. These results show that CCK receptor antagonists increases plasma concentration of leptin and suggest that endogenous CCK may facilitate the uptake of plasma leptin to the cerebrospinal fluid.

Peripheral cholecystokinin (CCK) elicits satiety by acting on hypothalamic nuclei. The anoretic effect of CCK is mediated by the vagus nerve and involves brainstem areas receiving vagal inputs, such as the nucleus tractus solitarius (NTS) and the area postrema (AP). This work aims to analyze, by measuring c-Fos expression, the effect of selective CCK receptor agonists on brain areas involved in food-intake/satiety process. We observed that SR-146,131, a CCK1R agonist, increased c-Fos expression in NTS and AP as well as in some hypothalamic nuclei. CCK-4, a CCK2R agonist which does not cross the blood-brain barrier (BBB), only was effective in the hypothalamus. Our data show that the activation of the brainstem is not a requisite to obtain a hypothalamic effect of peripheral CCK and suggest that CCK-4 may indirectly stimulate hypothalamic areas endowed with BBB, without previous activation of neither NTS nor AP.

Activation of the hypothalamic–pituitary–adrenal axis by fasting seems to involve cholecystokinin (CCK) receptors. This work aims to characterize the role of endogenous CCK in the activity of the paraventricular nucleus (PVN) of the hypothalamus during food withdrawal. We investigated, by c-Fos immunohistochemistry, the effect of CCK1 and CCK2 receptor antagonists (SR-27,897 and L-365,260, respectively) on c-Fos levels expression induced by food deprivation. Under our conditions, the number of cells expressing c-Fos was reduced both by SR-27,897 and L-365,260 in food-deprived rats. To investigate the importance of glucose availability, we studied the effect of CCK receptor antagonists on c-Fos synthesis induced by the glucose antimetabolite 2-deoxyglucose. In these animals, only SR-27,897 decreased c-Fos expression in the PVN. Our results indicate that the effect of CCK antagonists is mainly perceptible when glucose availability decreases, and suggest that CCK-ergic inputs could drive the activity of the PVN under fasting/low glucose conditions.

The objective of this work was to characterize the adaptation of cardiac metabolism to a lipid overload in a model of diet-induced obesity (DIO) in mice. After 8 wk dietary treatment, mice receiving a high-fat diet exhibited an increase in the amount of adipose tissue, accompanied by a surge in plasma leptin concentration (from 5.4-16.0 ng/ml). This was associated with: 1) an induction of uncoupling protein-2 (120%), 2) an increase in the phosphorylated form of AMP-activated protein kinase (120%), and 3) a reduction in lactate concentration and lactate dehydrogenase activity in myocardial tissue (40%). Because DIO induces leptin resistance, we analyzed leptin receptor functionality by measuring phospho-signal transducer and activator of transcription 3 in response to acute leptin (1 mg/kg). We observed that leptin receptor signaling remained unaltered within the heart but was fully impaired within the hypothalamus. Taken together, these data show that during DIO development, there is a metabolic shift in the heart aimed at increasing fatty acid oxidation to the detriment of carbohydrates. This effect seems to be leptin-dependent, suggesting that the increased adiposity observed during the onset of obesity might contribute to impairing ectopic lipidic deposition in the heart

Regulation of body weight (BW) results from the interplay between different hormonal systems acting at central and peripheral level. This study aims at characterizing the involvement of cholecystokinin (CCK) in BW and energy balance regulation. We have characterized, in free-feeding rats, the effect of CCK-8 on 1) food intake, BW, and adiposity; 2) skeletal muscle metabolism; 3) leptin signaling pathway within the arcuate nucleus of the hypothalamus; and 4) the permeability of brain barriers to leptin. We demonstrate here that CCK-8 acutely decreases BW by a mechanism partially dependent on central leptin pathways, based on the following results: 1) the effect of CCK was less intense in rats lacking functional leptin receptors (Zucker fa/fa), 2) CCK-8 facilitated the uptake of leptin from peripheral circulation to cerebrospinal fluid (CSF), 3) the concentration of leptin in CSF of rats receiving CCK was more elevated in those animals showing higher loss of BW, and 4) CCK activated leptin signaling pathways within the hypothalamus as well as phosphorylation of AMP-activated protein kinase in skeletal muscle. We also suggest that gain of BW may be linked to individual susceptibility to the effect of CCK, because we observed that in animals treated with this hormone, the increase of BW negatively correlated with leptin concentration within the CSF. Our data show that CCK has a negative impact on energy balance and suggest that CCK facilitates the access of leptin to hypothalamic areas, thus allowing leptin to act on hypothalamic targets involved in BW control.

Cholecystokinin (CCK) and leptin act coordinately in the brain to regulate food intake and energy balance. Recently we have reported that CCK enhances the permeability of brain barriers to leptin and we have proposed that CCK enhances energy expenditure in rats by activating in the hypothalamus the janus kinase/signal transducer and activator of transcription (JAK/STAT) signalling pathway, which is coupled to leptin receptors. Because plasma leptin concentration follows a circadian rhythm (plasma leptin concentration rise maximal values during the night, after rats start eating), we have hypothesized that the interaction between leptin and CCK should be more intense in animals receiving CCK during the night, i.e., during periods of positive energy balance. In order to further characterize the physiological relevance of the interplay between leptin and CCK we have compared the effect of diurnal vs. nocturnal administration of the C-terminal octapeptide of CCK (CCK-8) on (i) body weight and food intake, and (ii) STAT3 activation, by analyzing phosphorylated STAT3 (pSTAT3) immunostaining within the arcuate nucleus of the hypothalamus. Our results show that CCK decreases body weight and food intake only after p.m. administration. Accordingly pSTAT3 immunostaining within the hypothalamus was more intense in p.m. than in a.m.-treated animals. These data suggest that the effect of CCK on leptin pathways follows a circadian rhythm linked to the energy balance status and gives further support to the interaction between leptin and CCK.

The aim of this study was to determine whether alterations in periadventitial adipose tissue and its anti-contractile effect precede hypertension development. We used 4-week-old male Wistar Kyoto (WKY) and spontaneously hypertensive rats (SHR), which were pre-hypertensive. Vascular function was studied in the perfused mesenteric bed (MB, 1.5 mL/ min). MB weight was lower in SHR (8.0±0.3 mg/g body weight) than in WKY (9.0±0.3 mg/g body weight) rats. Concentration-response curves to KCI (6 to 75 mmol/L) and to acetylcholine (10-9 to 10-5 mol/L) were similar between groups. Contractile responses to serotonin (10-9 to 10-5 mol/L) were significantly higher in SHR compared to WKY. 4-Aminopyridine (4-AP, 2 mmol/L), a blocker of Kv channels, induced a similar increase in perfusion pressure in both strains. However, 4-AP (2 mmol/L) significantly increased the contractile response to serotonin (10-9 to 10-5 mol/L) only in WKY. The anti-contractile effect of fat was confirmed by a comparison of (+) fat and (-) fat mesenteric arteries, which revealed that 4-AP significantly enhanced contractions only in (+) fat rings from WKY. These results show that alterations in visceral periadventitial fat mass and function in SHR precede hypertension, suggesting a constitutive mechanism independent of age and the hypertensive state.

Background and purpose: Leptin regulates energy expenditure and body weight by acting both on the hypothalamus and on peripheral targets. Central actions of leptin are enhanced by cholecystokinin (CCK). The interaction between leptin and CCK makes physiological sense, as rats lacking CCK 1 receptors are resistant to peripheral leptin but not to leptin directly infused into the brain. We have recently reported that CCK enhances leptin effects by increasing the entry of leptin into the CNS. The aim of this work was to further characterize the effect of CCK (10 μg kg -1) on leptin kinetics as well as the CCK receptor subtype involved in the interaction between CCK and leptin. Experimental approach: Experiments were carried out both in free-feeding and in fasted rats receiving a single dose of leptin (100 μg kg -1; i.p.). Parameters analysed over the next 6 h were plasma and cerebrospinal fluid concentrations of leptin. Key results: We observed that CCK-8 depressed the increase in plasma leptin that followed the i.p. injection and simultaneously increased leptin concentration in the cerebrospinal fluid from 92±25 to 230±24 pg mL -1 (P<0.05). The effect of CCK-8 was totally prevented by the CCK 1 receptor antagonist, SR-27,897 (0.3 mg kg -1, s.c.), but not by the CCK 2 receptor antagonist, L-365,260 (1 mg kg -1). Conclusions and implications: These results show that CCK plays a role in regulating the access of leptin to the brain and suggest that CCK analogues, acting on CCK 1 receptors, might be useful drugs in improving leptin actions within the brain. © 2008 Nature Publishing Group All rights reserved

Background and purpose: Overfeeding increases adipose tissue mass and leptin production and up-regulates the renin-angiotensin system in adipose tissue in rodents. Here, we determined the effect of chronic treatment with the renin inhibitor, aliskiren, in a model of diet-induced obesity in mice, on: (i) body weight, adipose tissue weight and plasma leptin; (ii) food intake and caloric efficiency; and (iii) angiotensin II (Ang II) in adipose tissue. Experimental approach: Four-week-old C57BL/6J mice (n = 40) received aliskiren (50 mg·kg -1·day -1; 6 weeks) by means of a subcutaneous osmotic Alzet minipump. Animals were given either a low-fat (10% kcal from fat) or a high-fat diet (45% kcal from fat) during this period. Food-intake and body-weight variation were monitored during treatment. Key results: In addition to a decrease of plasma renin activity, aliskiren reduced body-weight gain, adipose pads and plasma leptin concentration, independent of the diet. In adipose tissue, local concentrations of Ang II were also reduced by aliskiren. Conclusions and implications: Aliskiren limited the gain of adiposity in young mice. This effect was not due to changes in food intake or caloric efficiency and might be related to a down-regulation of the local renin-angiotensin system in adipose tissue. These effects were accompanied by reduced plasma leptin levels. As Ang II favours differentiation of adipocytes, it is possible that the decreased adipose tissue was linked to changes in adipocyte size and number