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Summary: Background & aims: We investigated possible involvements of bile acids (BA) and leptin in hepatogenous insulin resistance being present in up to 90% of cirrhotic patients. Methods: Blood was analysed in 10 cirrhotic patients (8m/2f, 48 ± 10.4 yrs) and 10 controls (8m/2f, 43 ± 9.3 yrs) after oral nutrition and during 1 h of parenteral feeding. In patients, leptin was additionally analysed from mesenteric and arterial blood. Results: Cirrhosis patients showed typical signs of hepatogenous insulin resistance (hyperinsulinaemia, normoglycaemia, hyperglucagonaemia). Both fasting BA (r = .714, p = 0.047) and fasting leptin (r = .867, p = 0.001) correlated to HOMA and predicted insulin response after oral feeding (R²adj = .783, p = 0.002). But during parenteral nutrition only leptin predicted insulin response (p = 0.005). The prandial glucose response was negatively correlated to the BA increase after oral nutrition (r = −.733, p = 0.028) and to the change in leptin during parenteral nutrition (r = −.738, p = 0.037) pointing towards a nutritional route-dependent positive impact on glucose tolerance of both substances. Prandial glucagon response was correlated to BA under both feeding conditions (p < 0.05). We found no relevant intestinal release of leptin during fasting or feeding conditions. Conclusion: Our results suggest a substantial involvement of BA and leptin by improving postprandial glucose tolerance related to liver cirrhosis. [Copyright &y& Elsevier]

Profound and multifaceted effects of hormones on the development, maturation and function of the CNS are well documented. Recent developments in magnetic resonance imagining (MRI) and positron emission tomography (PET) permit detailed in vivo studies of cerebral structure and function in humans. Techniques to measure subtle differences in cerebral structure, regional brain activation, changes in blood flow and other physiological biomarkers allow us to translate experimental evidence of hormone effects obtained from animal models to humans. Here we review the imaging techniques available to support studies of hormone effects on the CNS, emphasizing the recent developments of MRI. In summarizing the major current studies we discuss the potential of these techniques for an emerging new field in endocrinology. [Copyright &y& Elsevier]

Summary: Background & aims: Circulating leptin and insulin concentrations are physiologically representing energy homeostasis. However, the artificial situation of long-term total parenteral nutrition (TPN) and its effects on serum leptin and insulin is not fully understood. Methods: We studied 42 gastroenterological patients who received TPN for 19±11 days. Serum leptin and insulin levels as well as body composition assessed by bioelectrical impedance analysis were evaluated on days 0, 7 and 14. Insulin sensitivity was estimated by calculating the QUICKI. Results: Before the start of TPN, leptin correlated positively with female gender (), BMI (), fat mass (), insulin levels () and QUICKI (). Within the first week of TPN, an increase of leptin levels was found only in patients with a body fat mass of >30% (). As these were predominantly women, their leptin levels increased likewise (). In regression analysis, fat mass (), female gender (), insulin levels (), and i.v. glucose supply rates () were independently associated to leptin levels. Conclusions: TPN—especially glucose—induces a neurohumoral response as shown here for leptin and insulin that is mainly depending on the fat mass. Better understanding of this regulatory mechanism during artificial nutrition could offer a new approach to improve its therapeutic effects. [Copyright &y& Elsevier]

Summary: Pro-opiomelanocortin (POMC) expressing neurons mediate the regulation of orexigenic drive by peripheral hormones such as leptin, cholecystokinin, ghrelin, and insulin. Most research effort has focused on α-melanocyte-stimulating hormone (α-MSH) as the predominant POMC-derived neuropeptide in the central regulation of human energy balance and body weight. Here we report a missense mutation within the coding region of the POMC-derived peptide β-MSH (Y5C-β-MSH) and its association with early-onset human obesity. In vitro and in vivo data as well as postmortem human brain studies indicate that the POMC-derived neuropeptide β-MSH plays a critical role in the hypothalamic control of body weight in humans. [Copyright &y& Elsevier]

Background: Leptin has been considered a starvation hormone, but its role in malnourished patients is unknown. Objective: We aimed to characterize the role of leptin in metabolic adaptation in women with anorexia nervosa (AN). Design: In a cross-sectional study, 57 women with AN [mean (±SD) body mass index (kg/m2) on admission: 15.2 ± 1.5] were compared with 49 healthy, normal-weight women (mean body mass index: 22.3 ± 2.3). Nineteen patients were reinvestigated during weight gain 43 and 84 d after baseline. We measured serum concentrations of leptin, soluble leptin receptor, insulin, ghrelin, and thyroid hormones [thyrotropin, triiodothyronine (T3), and thyroxine]; fat mass (FM) and fat-free mass (FFM); resting energy expenditure (REE); energy intake; and eating behavior. Results: Compared with values in the control women, leptin, T3, REE, FM, and FFM were lower in the women with AN, but the leptin secretion rate was not significantly different. Leptin correlated with FM (r = 0.83, P < 0.001), T3 (r = 0.68, P < 0.001), respiratory quotient (r = -0.47, P < 0.001), and REE (r = 0.58, P < 0.001). The association with REE weakened after adjustment for FFM and disappeared after further adjustment for T3. Hunger and appetite had positive, whereas satiety and restraint had negative, associations with leptin. During weight gain (9.0 ± 3.3 kg in 84 d), serum leptin and the leptin secretion rate increased. Changes in leptin secretion were associated with energy intake and REE. The initial changes in the leptin secretion rate (ie, the difference between baseline and 43 d) were negatively associated with changes in body weight from 43 to 84 d. Conclusions: Leptin contributes to metabolic adaptation in women with AN. The leptin responseis associated with weight gain. [ABSTRACT FROM AUTHOR]

Summary: Leptin, a potent anorectic, 16-kDa, adipose tissue-derived protein, predominantly acts in hypothalamic nuclei, signaling obesity and modulating ingestive behavior. To reach this brain area, leptin, probably has to cross the blood-brain barrier (BBB). In some cases of obesity, enhanced leptin levels in the blood do not result in anorectic effects, probably due to an altered leptin transport across the BBB. Therefore, we investigated the BBB in lean and diet-induced obese Lewis rats. To obtain information about the presence of microvessels with barrier dysfunction we examined three brain areas (hypothalamus, cortex, hippocampus) using a monoclonal antibody which detects intact microvessels of the BBB (anti-endothelial barrier antigen, anti-EBA). The results showed a significantly reduced EBA staining in the brain sections of the obese animals, except the hippocampus, compared to the control group. In a second step we injected I125-labeled leptin intravenously (i.v.) in permanent i.v.-cannulated, unrestrained Lewis rats (lean and obese). We measured the radioactivity in the cerebrospinal fluid after puncture of the cisterna magna, in the blood and brain tissue 90min after injection. The leptin content in the cerebrospinal fluid and brain was not reduced in obese compared to lean rats, thus showing a similar transport capacity of the BBB in both experimental groups. Therefore, the results of the in vivo investigations do not indicate an impairment of the BBB in diet-induced obesity, despite the immunohistological findings. Further functional and morphological studies are necessary to evaluate the specific role of other organs and distinct forms of leptin (free and protein-bound) in the pathogenesis of diet-induced obesity. [Copyright &y& Elsevier]

Abstract: Background and purpose: Undifferentiated follicular and anaplastic thyroid tumours often respond poorly to radiotherapy and show increased metastatic potential. We evaluated radiation-induced effects on metastasis in thyroid carcinoma cells and tumours, mechanistically focusing on phosphatidylinositide 3-kinase (PI3K) and associated pathways. Material and methods: Migration was analysed in follicular (FTC133) and anaplastic (8505c) cells following radiotherapy (0–6 Gray) with concomitant pharmacological (GDC-0941) or genetic inhibition of PI3K. Hypoxia-inducible factor-1 (HIF-1)-activity was measured using luciferase reporter assays and was inhibited using a dominant-negative variant. Activation and subcellular localisation of target proteins were assessed via Western blot and immunofluorescence. In vivo studies used FTC133 xenografts with metastatic lung dissemination assessed ex vivo. Results: Radiation induced migration in a HIF-dependent manner in FTC133 cells but decreased migration in 8505c’s. Post-radiation HIF-activity correlated with migratory phenotype. PI3K-targeting inhibited migration under basal and irradiated conditions through inhibition of HIF-1α, Rho-GTPase expression/activity and localisation whilst having little effect on src/FAK. In vivo, radiation induced PI3K, HIF, Rho-GTPases and src but only PI3K, HIF and Rho-GTPases were inhibited by GDC-0941. Co-treatment with GDC-0941 and radiation significantly reduced metastatic dissemination versus radiotherapy alone. Conclusions: Radiation modifies metastatic characteristics of thyroid carcinoma cells, which can be successfully inhibited by targeting PI3K using GDC-0941 in vitro and in vivo. [Copyright &y& Elsevier]

Highlights • Aim: to evaluate the effects of induced subclinical mild hypothyroidism on brain network connectivity, showing global changes of brain function. • Method: 15 otherwise healthy hypothyroid subjects under long term levothyroxine substitution reduced their pretest levothyroxine dosage by 30% for 52–56 days. Before and after reduction, resting state fMRI, neuropsychological analysis and laboratory testing was performed. • Results: Desired subclinical hypothyroidism was achieved.This lead to a significant decrease in resting-state functional connectivity in the cuneus. This was caused by weaker connectivity to cerebellum and regions of default mode network. The decrease of the connectivity in the cuneus was correlated to the increase in TSH serum levels. Subjects showed a longer reaction time and less accuracy in a memory task after levothyroxine withdrawal. • Conclusion: The human brain seems especially sensitive to even short-term thyroid hormone changes. Even slight cognitive changes in subclinical hypothyroidism may be mediated by the cuneus, connected to the cerebellum, posterior and medial frontal cortex. Abstract Objective The aim of this study was to experimentally evaluate the effects of subclinical mild hypothyroidism on brain network connectivity as determined by resting state fMRI (rsfMRI) which serves as a proxy for global changes in brain function. Methods Fifteen otherwise healthy patients with complete hypothyroidism under stable, long term levothyroxine substitution volunteered for the study. They reduced their pretest levothyroxine dosage by 30% for 52–56 days. Basally and after partial levothyroxine withdrawal, rsfMRI along with a neuropsychological analysis was performed. RsfMRI was subjected to graph-theory-based analysis to investigate whole-brain intrinsic functional connectivity. Results The desired subclinical hypothyroidism was achieved in all subjects. This was associated with a significant decrease in resting-state functional connectivity specifically in the cuneus (0.05 FWE corrected at cluster level) which was mainly caused by a weaker functional connectivity to the cerebellum and regions of the default mode network, i.e. the medial prefrontal cortex, the precuneus and the bilateral angular gyri. The decrease in cuneus connectivity was correlated to the increase in TSH serum levels. A working memory task showed a slightly longer reaction time and less accuracy after partial levothyroxine withdrawal. Conclusion Even short-term partial levothyroxine partial withdrawal leads to deficits in working memory tasks and to a weaker integration of the cuneus within the default mode network. [ABSTRACT FROM AUTHOR]

Highlights • We examined how the synthetic glucocorticoid prednisolone impacts brain areas linked with hedonic eating and motor control. • The synthetic glucocorticoid prednisolone increased amygdala and insula reactivity to approach-associated food pictures. • Prednisolone-induced insula reactivity to food approach pictures was associated with greater subsequent caloric intake. • Prednisolone increased overall commission error rate and postcentral gyrus activity durng response inhibition. Abstract Short- and long-term treatment with glucocorticoids is widely used in clinical practice and frequently induces features of iatrogenic Cushing syndrome, such as abdominally centered weight gain. Despite decades of glucocorticoids usage, the mechanisms underlying these side effects are still only partly understood. One possibility is that glucocorticoids impact subcortical (hypothalamus, amygdala, insula) and cortical (orbitofrontal and cingulate cortex) brain regions involved in appetite regulation and reward processing. In the present study, we used functional magnetic resonance imaging (fMRI) to study the acute effects of a prednisolone infusion on reactivity of brain reward systems to food stimuli. Twenty healthy normal-weight men were tested in a randomized, double-blind, cross-over study. After an overnight fast and infusion of either 250 mg prednisolone or placebo (always administered between 8 and 9 A M), fMRI scans were taken while presenting food and object pictures in a Go/NoGo (GNG) task. At home, participants were asked to register what they had eaten. On the following morning they came back to the lab and had a supervised ad libitum breakfast at a standardized buffet. Food-Go in contrast to Object-Go pictures yielded increased blood oxygen level dependent (BOLD) activity in hippocampus, amygdala, orbitofrontal cortex, insula and anterior cingulate cortex. Prednisolone increased activation in the bilateral amygdala and right insula for approach-associated food pictures. The buffet test did not reveal significant differences in calorie consumption or preferences of different macronutrients. However, prednisolone-induced insula reactivity to Food-Go images was associated with greater caloric intake, both at home and in the standardized buffet. In sum, we observed a specific effect of prednisolone on the BOLD response of the amygdala and insula to approach-associated food stimuli. As these brain areas have previously been implicated in hedonic eating, the present pattern of results may reflect an increased anticipated reward value of food modulated by glucocorticoids. These effects might potentially drive increased food intake and weight gain under prolonged glucocorticoid treatment. [ABSTRACT FROM AUTHOR]

Objectives: The aims of this study were to ascertain whether women with high levels of serum total testosterone (TT) or low levels of sex hormone-binding globulin (SHBG) are more likely to develop cardiovascular disease (CVD), and to investigate potential associations between sex hormones and mortality (all-cause, as well as cause-specific) in the general population.Study Design and Main Outcome Measures: Data on 2129 women with a mean age of 49.0 years were obtained from the population-based Study of Health in Pomerania over a median follow-up of 10.9 years. Associations of baseline levels of TT, SHBG, and rostenedione (ASD), and free testosterone (fT), and of the free androgen index (FAI), with follow-up CVD morbidity, as well as all-cause and CVD mortality, were analyzed using multivariable regression modeling.Results: At baseline the prevalence rate of CVD was 17.8% (378 women) and the incidence of CVD over the follow-up was 50.9 per 1000 person-years. We detected an inverse association between SHBG and baseline CVD in age-adjusted models (relative risk per standard deviation increase: 0.83; 95% confidence interval: 0.74-0.93). We did not detect any significant associations between sex hormone concentrations and incident CVD in age- and multivariable-adjusted Poisson regression models. Furthermore, none of the sex hormones (TT, SHBG, ASD, fT, FAI) were associated with all-cause mortality.Conclusions: This population-based cohort study did not yield any consistent associations between sex hormones in women and incident CVD or mortality risk. [ABSTRACT FROM AUTHOR]

Summary Adult onset hyperthyroidism may impact on different cognitive domains, including attention and concentration, memory, perceptual function, language and executive function. Previous PET studies implicated changed functionality of limbic regions, the temporal and frontal lobes in hyperthyroidism, whereas it is unknown whether cognitive effects of hyperthyroidism may be due to changed brain connectivity. This study aimed to investigate the effect of experimentally induced short-term hyperthyroidism thyrotoxicosis on resting-state functional connectivity using functional magnetic resonance imaging. Twenty-nine healthy male right-handed subjects were examined twice, once prior and once after 8 weeks of oral administration of 250 μg levothyroxine per day. Resting-state fMRI was subjected to graph-theory based analysis methods to investigate whole-brain intrinsic functional connectivity. Despite a lack of subjective changes noticed by the subjects significant thyrotoxicosis was confirmed in all subjects. This induced a significant increase in resting-state functional connectivity specifically in the rostral temporal lobes (0.05 FDR corrected at the cluster level), which is caused by an increased connectivity to the cognitive control network. The increased connectivity between temporal poles and the cognitive control network shown here under experimental conditions supports an important function of thyroid hormones in the regulation of paralimbic structures. [ABSTRACT FROM AUTHOR]

Abstract: Obese patients have an increased incidence of systemic infections and higher morbidity and mortality rates than normal weight subjects. Ghrelin is a potent orexigenic signal from the stomach and seems to play a role in the generation and control of immune interactions. To examine a possible benefit of a single ghrelin application on acute endotoxemia, chronic intravenous (i.v.) cannulated lean and diet-induced obese male LEW rats were treated with a bolus injection of either ghrelin (10 nmol/kg) or vehicle, 10 min prior to a challenge with a sublethal bolus of endotoxin (100 μg/kg) or vehicle. Multiple blood samples were taken within a period from 24 h before the experiment up to 24 h after the endotoxin challenge to measure ghrelin and cytokine levels. Additionally, food consumption was recorded and ghrelin expression in fore- and glandular stomach was evaluated immunohistochemically. Despite higher serum ghrelin levels, the food consumption was significantly decreased in obese endotoxemic rats compared to lean littermates after ghrelin treatment. Furthermore we could show an increase of anti-inflammatory IL-10 serum levels after ghrelin treatment of normal weight endotoxemic and an opposite effect in obese animals. As the therapy of disease-associated cachexia and various immunological problems in endotoxemia is still insufficient, peptides such as ghrelin with their modulating abilities for the endocrine and the immune system are of special interest. However, the present study shows that the beneficial effects of ghrelin were attenuated in obese endotoxemic animals. These data further document the necessity to differentiate between normal weight and obese subjects in the attempt to establish ghrelin as a therapeutic target in endotoxemia. [Copyright &y& Elsevier]

Hypothyroidism is a common clinical problem during 131Iodine-therapy of thyroid cancer. In the present investigation, possible cognitive dysfunction during hypothyroid state was assessed by means of neuropsychological tests and the recording of event-related brain potentials (ERPs).Fifteen patients undergoing therapy for thyroid cancer were examined twice: (1) substituted with thyroid hormones, (2) during hypothyroid state immediately prior to treatment. Standard neuropsychological tests were applied during both sessions and subjects showed a mild-to-moderate impairment in their hypothyroid state. In addition, ERPs were recorded from 19 scalp sites while subjects performed two visual search tasks. The serial task required the effortful one-by-one scanning of several items within a visual array, while the parallel task allowed processing of all stimulus items in parallel and automatically. ERPs showed a marked amplitude decrement and delay of the P3 component known to index the speed of stimulus evaluation and the amount of available processing resources. This effect was present only for the serial search task, while no changes were seen in the parallel search task. These data show that hypothyroidism during 131Iodine-therapy is associated with clinically relevant cognitive dysfunctions, especially with effortful attention demanding tasks. [Copyright &y& Elsevier]

: ObjectiveTo investigate the serum pattern of free leptin, bound leptin, and soluble leptin receptor throughout the physiological menstrual cycle.: DesignProspective observational study.: SettingTertiary care center for gynecological endocrinology and reproductive medicine and a university research laboratory.: Patient(s)Thirty regularly cycling volunteers (age, 29 ± 4.2 years).: Intervention(s)Blood sampling was performed at different phases (early and mid follicular phase, preovulatory phase, and early and late luteal phase) of three consecutive menstrual cycles; each phase of the menstrual cycle was investigated twice.: Main outcome measure(s)Free leptin, bound leptin, soluble leptin receptor, LH, E2, P, vaginal ultrasound.: Result(s)A peak of serum free leptin levels was found in the late luteal phase followed by a significant drop in the early follicular phase and again by a continuous increase up to the next luteal peak. There were no significant alterations in serum bound leptin and soluble leptin receptor levels.: Conclusion(s)The present study shows that there are significant circacyclic fluctuations of free leptin levels with the highest concentrations in the late luteal phase and the lowest levels in the early follicular phase, which suggests that circulating free leptin is up-regulated by the C21-steroid (P). Circulating bound leptin and soluble leptin receptor are not altered by the cyclic hormone status. The significant rise of the leptin bioequivalent, free leptin, in the late luteal phase might be of importance for the luteal-follicular and the luteal-preimplantatory functional shift. [Copyright &y& Elsevier]

Free fatty acids (FFA) have been proposed to participate in the regulation of insulin release from pancreatic beta-cells (β-cells). As a rise in cytosolic free Ca2+ ([Ca2+]i) is a key event for the stimulation of insulin secretion, the effects of saturated FFA on [Ca2+]i were investigated. Palmitate was used as a reference compound and [Ca2+]i was measured in single fura-2 loaded HIT-T15 and in primary mouse β-cells. Stimulation of single β-cells with palmitate (100 μM) caused either repetitive Ca2+ transients or a plateau-like rise in [Ca2+]i. In HIT-T15 and in mouse β-cells, the number of palmitate-responsive cells, and the amplitude of the palmitate-induced Ca2+-signals were dependent on the extracellular glucose concentration. In Ca2+-free medium palmitate (100 μM) caused only 1 or 2 Ca2+ transients indicating mobilization of Ca2+ from internal stores. Withdrawal of external Ca2+, the addition of voltage-sensitive Ca2+ channel (VSCC) blockers, as well as the KATP-channel opener diazoxide (100 μM) reversibly blocked the palmitate-induced cytosolic Ca2+ responses. This demonstrates that Ca2+ influx through VSCC of the L-type coupled to membrane depolarization through closure of KATP-channels are crucial for a sustained Ca2+-signal in response to palmitate. Methyl palmoxirate (100 μM) and 2-bromopalmitate (100 μM), which both inhibit transport of acyl-CoA into the mitochondria, reversibly blocked the palmitate-induced Ca2+-signals in HIT-T15 as well as in primary mouse β-cells. By contrast, cerulenin (100 μM), an inhibitor of protein acylation, had no effect on the palmitate-induced changes in [Ca2+]i, which suggests that mitochondrial palmitate metabolism is required for eliciting the Ca2+-signals. Simultaneous measurement of [Ca2+]i and the mitochondrial membrane potential (ΔΨ) revealed palmitate-induced depolarization of ΔΨ which demonstrates that palmitate does not enhance mitochondrial ATP production. Therefore mitochondrial signals other than ATP appear to be generated from palmitate metabolism that underly the palmitate-induced Ca2+-signals in pancreatic β-cells. [Copyright &y& Elsevier]

Objectives. To detect changes in growth hormone (GH) serum levels during different penile conditions in the cavernous and systemic blood of patients with erectile dysfunction and compare them with the course of GH registered in healthy men. It has been suggested that human GH is involved in sexual maturation and plays a regulatory role in male reproductive function. Deficiency may result in fatigability, loss of sexual desire and erection, or oligospermia or azoospermia. It is assumed that the biologic effects of GH include insulin-like growth factor 1-mediated stimulation of endothelial nitric oxide formation. It has recently been demonstrated that GH serum levels in the systemic and cavernous blood of healthy men increases during developing penile erection.Methods. Thirty-five healthy adult men and 45 patients with erectile dysfunction of either organogenic or psychogenic etiology were exposed to visual and tactile erotic stimuli to elicit penile tumescence and, in the group of healthy subjects, rigidity. Whole blood was simultaneously aspirated from the corpus cavernosum and the cubital vein during the different functional conditions of the penis. Serum levels of GH were determined by means of an immunoradiometric assay.Results. In the healthy subjects, systemic GH serum levels significantly increased during penile tumescence, followed by a transient decline from tumescence to rigidity and detumescence. In the unselected patients, the mean GH levels during penile flaccidity were determined to be about sevenfold lower than those registered in the blood of the healthy men. During penile tumescence, the mean increase in the GH levels in the systemic and cavernous blood of psychogenic patients was comparable to that seen in healthy men, but, in the group of organogenic patients, this increase was found to be negligible.Conclusions. We believe our data provide strong evidence that GH may be of major importance in the maintenance of male erectile capability—probably through a stimulating effect on cyclic guanosine monophosphate generation in human cavernous smooth muscle—and that a decline in GH release may contribute to the manifestation of erectile dysfunction. [Copyright &y& Elsevier]

In β-cells activation of the cyclic AMP (cAMP)-signaling cascade stimulates c-fos mRNA expression, which involves cAMP- and Ca2+-mediated mechanisms. To delineate potential crosstalk between both pathways at the transcriptional level we simultaneously measured c-fos promoter-driven enhanced green fluorescent protein (EGFP) expression and cytosolic free calcium ([Ca2+]i) in single β-cells (HIT-T15). Forskolin stimulated a rapid rise in cellular cAMP and in [Ca2+]i through activation of voltage-sensitive Ca2+-influx and enhanced wild-type c-fos promoter-driven EGFP (pF711d2EGFP) expression about 4-fold after 6 h. The voltage-sensitive Ca2+ channel (VSCC)-blocker nifedipine, which completely blocked the forskolin-induced rise in [Ca2+]i, partially inhibited the forskolin-induced increase in pF711d2EGFP expression, while it was completely abolished in Ca2+-free medium. VSCC-dependent Ca2+-influx per se when stimulated by K+ (45 mM) increased pF711d2EGFP expression only minimally. No correlations could be delineated between the forskolin-induced amplitude of the Ca2+ signal and the expression of pF711d2EGFP at the single cell level, which may indicate that small rises in [Ca2+]i are sufficient to fully activate the Ca2+-dependent pathways required for cAMP-dependent c-fos promoter regulation. In experiments with various deletion constructs of the c-fos promoter, it could be shown that cAMP-mediated activation of the c-fos promoter involves both the cAMP-responsive element (CRE) and the serum-responsive element (SRE). While nifedipine completely abrogated the cAMP-dependent activation of c-fos transcription via the SRE, the CRE-mediated effect of cAMP on the c-fos promoter remained unaffected by nifedipine. Thus, cAMP and Ca2+ are required for full c-fos promoter activation by the cAMP-signaling pathway in β-cells. cAMP-dependent Ca2+-influx through VSCC is crucial for c-fos gene transcription via the SRE, whereas cAMP-mediated activation of the CRE demands Ca2+-influx, which is distinct from voltage-sensitive Ca2+-influx. This indicates a complex interplay between cAMP and Ca2+ in controlling c-fos gene transcription and suggests that the mode of Ca2+ entry may differentially act on signaling pathways leading to gene transcription in β-cells. [Copyright &y& Elsevier]