Mitochondrial CB₁ receptors regulate neuronal energy metabolism

The mammalian brain is one of the organs with the highest energy demands, and mitochondria are key determinants of its functions. Here we show that the type-1 cannabinoid receptor (CB(1)) is present at the membranes of mouse neuronal mitochondria (mtCB(1)), where it directly controls cellular respiration and energy production. Through activation of mtCB(1) receptors, exogenous cannabinoids and in situ endocannabinoids decreased cyclic AMP concentration, protein kinase A activity, complex I enzymatic activity and respiration in neuronal mitochondria. In addition, intracellular CB(1) receptors and mitochondrial mechanisms contributed to endocannabinoid-dependent depolarization-induced suppression of inhibition in the hippocampus. Thus, mtCB(1) receptors directly modulate neuronal energy metabolism, revealing a new mechanism of action of G protein-coupled receptor signaling in the brain.

Effects of colostrum versus formula feeding on hepatic glucocorticoid and α1- and β2-adrenergic receptors in neonatal calves and their effect on glucose and lipid metabolism

Neonatal energy metabolism in calves has to adapt to extrauterine life and depends on colostrum feeding. The adrenergic and glucocorticoid systems are involved in postnatal maturation of pathways related to energy metabolism and calves show elevated plasma concentrations of cortisol and catecholamines during perinatal life. We tested the hypothesis that hepatic glucocorticoid receptors (GR) and α₁- and β₂-adrenergic receptors (AR) in neonatal calves are involved in adaptation of postnatal energy metabolism and that respective binding capacities depend on colostrum feeding. Calves were fed colostrum (CF; n=7) or a milk-based formula (FF; n=7) with similar nutrient content up to d 4 of life. Blood samples were taken daily before feeding and 2h after feeding on d 4 of life to measure metabolites and hormones related to energy metabolism in blood plasma. Liver tissue was obtained 2 h after feeding on d 4 to measure hepatic fat content and binding capacity of AR and GR. Maximal binding capacity and binding affinity were calculated by saturation binding assays using [(3)H]-prazosin and [(3)H]-CGP-12177 for determination of α₁- and β₂-AR and [(3)H]-dexamethasone for determination of GR in liver. Additional liver samples were taken to measure mRNA abundance of AR and GR, and of key enzymes related to hepatic glucose and lipid metabolism. Plasma concentrations of albumin, triacylglycerides, insulin-like growth factor I, leptin, and thyroid hormones changed until d 4 and all these variables except leptin and thyroid hormones responded to feed intake on d 4. Diet effects were determined for albumin, insulin-like growth factor I, leptin, and thyroid hormones. Binding capacity for GR was greater and for α₁-AR tended to be greater in CF than in FF calves. Binding affinities were in the same range for each receptor type. Gene expression of α₁-AR (ADRA1) tended to be lower in CF than FF calves. Binding capacity of GR was related to parameters of glucose and lipid metabolism, whereas β₂-AR binding capacity was negatively associated with glucose metabolism. In conclusion, our results indicate a dependence of GR and α₁-AR on milk feeding immediately after birth and point to an involvement of hepatic GR and AR in postnatal adaptation of glucose and lipid metabolism in calves.

The chemokine receptors ACKR2 and CCR2 reciprocally regulate lymphatic vessel density

Macrophages regulate lymphatic vasculature development; however, the molecular mechanisms regulating their recruitment to developing, and adult, lymphatic vascular sites are not known. Here, we report that resting mice deficient for the inflammatory chemokine-scavenging receptor, ACKR2, display increased lymphatic vessel density in a range of tissues under resting and regenerating conditions. This appears not to alter dendritic cell migration to draining lymph nodes but is associated with enhanced fluid drainage from peripheral tissues and thus with a hypotensive phenotype. Examination of embryonic skin revealed that this lymphatic vessel density phenotype is developmentally established. Further studies indicated that macrophages and the inflammatory CC-chemokine CCL2, which is scavenged by ACKR2, are associated with this phenotype. Accordingly, mice deficient for the CCL2 signalling receptor, CCR2, displayed a reciprocal phenotype of reduced lymphatic vessel density. Further examination revealed that proximity of pro-lymphangiogenic macrophages to developing lymphatic vessel surfaces is increased in ACKR2-deficient mice and reduced in CCR2-deficient mice. Therefore, these receptors regulate vessel density by reciprocally modulating pro-lymphangiogenic macrophage recruitment, and proximity, to developing, resting and regenerating lymphatic vessels.

Glucose-dependent insulinotropic polypeptide receptors in most gastroenteropancreatic and bronchial neuroendocrine tumors

Gastrointestinal peptide hormone receptors overexpressed in neuroendocrine tumors (NET), such as somatostatin or glucagon-like peptide-1 (GLP-1) receptors, are used for in vivo tumor targeting. Unfortunately, not all NET express these receptors sufficiently.

Transfection of drug-specific T-cell receptors into hybridoma cells: tools to monitor drug interaction with T-cell receptors and evaluate cross-reactivity to related compounds

In the context of drug hypersensitivity, our group has recently proposed a new model based on the structural features of drugs (pharmacological interaction with immune receptors; p-i concept) to explain their recognition by T cells. According to this concept, even chemically inert drugs can stimulate T cells because certain drugs interact in a direct way with T-cell receptors (TCR) and possibly major histocompatibility complex molecules without the need for metabolism and covalent binding to a carrier. In this study, we investigated whether mouse T-cell hybridomas transfected with drug-specific human TCR can be used as an alternative to drug-specific T-cell clones (TCC). Indeed, they behaved like TCC and, in accordance with the p-i concept, the TCR recognize their specific drugs in a direct, processing-independent, and dose-dependent way. The presence of antigen-presenting cells was a prerequisite for interleukin-2 production by the TCR-transfected cells. The analysis of cross-reactivity confirmed the fine specificity of the TCR and also showed that TCR transfectants might provide a tool to evaluate the potential of new drugs to cause hypersensitivity due to cross-reactivity. Recombining the alpha- and beta-chains of sulfanilamide- and quinolone-specific TCR abrogated drug reactivity, suggesting that both original alpha- and beta-chains were involved in drug binding. The TCR-transfected hybridoma system showed that the recognition of two important classes of drugs (sulfanilamides and quinolones) by TCR occurred according to the p-i concept and provides an interesting tool to study drug-TCR interactions and their biological consequences and to evaluate the cross-reactivity potential of new drugs of the same class.

Effects of dexamethasone on mRNA abundance of nuclear receptors and hepatic nuclear receptor target genes in neonatal calves

Hepatic nuclear receptors (NR), particularly constitutive androstane receptor (CAR) and pregnane X receptor (PXR), are involved in the coordinated transcriptional control of genes that encode proteins involved in the metabolism and detoxification of xeno- and endobiotics. A broad spectrum of metabolic processes are mediated by NR acting in concert with ligands such as glucocorticoids. This study examined the role of dexamethasone on hepatic mRNA expression of CAR, PXR and several NR target genes. Twenty-eight male calves were allotted to one of four treatment groups in a 2 x 2 arrangement of treatments: feed source (colostrum or milk-based formula) and glucocorticoid administration (twice daily intramuscular dexamethasone). Liver biopsies were obtained at 5 days of age. Real-time reverse transcription polymerase chain reaction was used to quantify mRNA abundances. No effects of feed source on mRNA abundances were observed. For the NR examined, mRNA abundance of both CAR and PXR in dexamethasone-treated calves was lower (p < 0.05) by 39% and 40%, respectively, than in control calves. Abundance of NR target genes exhibited a mixed response. SULT1A1 mRNA abundance was 39% higher (p < 0.05) in dexamethasone-treated calves compared with control calves. mRNA abundance of CYP2C8 tended also to be higher (+44%; p = 0.053) after dexamethasone treatment. No significant treatment effects (p > 0.10) were observed for mRNA abundances of CYP3A4, CYP2E1, SULT2A1, UGT1A1 or cytochrome P450 reductase (CPR). In conclusion, an enhanced glucocorticoid status, induced by pharmacological amounts of dexamethasone, had differential and in part unexpected effects on NR and NR target systems in 5-day-old calves. Part of the unexpected responses may be due the immaturity of NR and NR receptor target systems.

Synergy-inducing chemokines enhance CCR2 ligand activities on monocytes

The migration of monocytes to sites of inflammation is largely determined by their response to chemokines. Although the chemokine specificities and expression patterns of chemokine receptors are well defined, it is still a matter of debate how cells integrate the messages provided by different chemokines that are concomitantly produced in physiological or pathological situations in vivo. We present evidence for one regulatory mechanism of human monocyte trafficking. Monocytes can integrate stimuli provided by inflammatory chemokines in the presence of homeostatic chemokines. In particular, migration and cell responses could occur at much lower concentrations of the CCR2 agonists, in the presence of chemokines (CCL19 and CCL21) that per se do not act on monocytes. Binding studies on CCR2(+) cells showed that CCL19 and CCL21 do not compete with the CCR2 agonist CCL2. Furthermore, the presence of CCL19 or CCL21 could influence the degradation of CCL2 and CCL7 on cells expressing the decoy receptor D6. These findings disclose a new scenario to further comprehend the complexity of chemokine-based monocyte trafficking in a vast variety of human inflammatory disorders.

The role of purinergic signaling in the liver and in transplantation: effects of extracellular nucleotides on hepatic graft vascular injury, rejection and metabolism

Extracellular nucleotides (e.g. ATP, UTP, ADP) are released by activated endothelium, leukocytes and platelets within the injured vasculature and bind specific cell-surface type-2 purinergic (P2) receptors. This process drives vascular inflammation and thrombosis within grafted organs. Importantly, there are also vascular ectonucleotidases i.e. ectoenzymes that hydrolyze extracellular nucleotides in the blood to generate nucleosides (viz. adenosine). Endothelial cell NTPDase1/CD39 has been shown to critically modulate levels of circulating nucleotides. This process tends to limit the activation of platelet and leukocyte expressed P2 receptors and also generates adenosine to reverse inflammatory events. This vascular protective CD39 activity is rapidly inhibited by oxidative reactions, such as is observed with liver ischemia reperfusion injury. In this review, we chiefly address the impact of these signaling cascades following liver transplantation. Interestingly, the hepatic vasculature, hepatocytes and all non-parenchymal cell types express several components co-ordinating the purinergic signaling response. With hepatic and vascular dysfunction, we note heightened P2- expression and alterations in ectonucleotidase expression and function that may predispose to progression of disease. In addition to documented impacts upon the vasculature during engraftment, extracellular nucleotides also have direct influences upon liver function and bile flow (both under physiological and pathological states). We have recently shown that alterations in purinergic signaling mediated by altered CD39 expression have major impacts upon hepatic metabolism, repair mechanisms, regeneration and associated immune responses. Future clinical applications in transplantation might involve new therapeutic modalities using soluble recombinant forms of CD39, altering expression of this ectonucleotidase by drugs and/or using small molecules to inhibit deleterious P2-mediated signaling while augmenting beneficial adenosine-mediated effects within the transplanted liver.

Arginine vasopressin in vasodilatory shock: effects on metabolism and beyond

PURPOSE OF REVIEW: To describe the effects of arginine vasopressin other than its vasoconstrictive and antidiuretic potential in vasodilatory shock. RECENT FINDINGS: Arginine vasopressin influences substrate metabolism by stimulation of hepatic glucose release, gluconeogenesis, ureogenesis and fatty acid esterification. Although arginine vasopressin is a secretagogue of different hormones, only prolactin increases during arginine vasopressin therapy. Plasmatic and cellular coagulation are affected by arginine vasopressin, resulting in thrombocyte aggregation. Therefore, platelet count typically decreases following arginine vasopressin infusion in critically ill patients. In addition, arginine vasopressin reduces bile flow and may increase bilirubin concentrations. Despite its potential to decrease serum sodium, no change in electrolytes was observed in critically ill patients receiving arginine vasopressin. Although arginine vasopressin is an endogenous antipyretic, body temperature is not decreased by central venous arginine vasopressin infusion. In addition, arginine vasopressin modulates immune function through V1 receptors. Compared with norepinephrine, arginine vasopressin may have protective effects on endothelial function. Net arginine vasopressin effects on gastrointestinal motility seem to be inhibitory and are dose dependent. SUMMARY: Except for its antidiuretic and vasoconstrictive actions, the effects of arginine vasopressin in patients with vasodilatory shock have so far only been partially examined. Potential influences of arginine vasopressin on metabolism and immune, liver and mitochondrial function remain to be assessed in future studies.

Dynamics of lipoprotein metabolism in adult growth hormone deficiency

Dyslipidaemia is often associated with adult growth hormone (GH) deficiency. Reduced removal of very-low-density lipoprotein (VLDL) apolipoprotein B-100 (apo B-100) can, in part, explain the "unfavourable" lipid profile of these patients. By modifying VLDL composition and through its action on low-density lipoprotein (LDL) receptors, GH may improve the lipid profile by increasing direct hepatic uptake of VLDL apo B-100, thereby decreasing conversion to LDL. Although GH stimulates VLDL apo B-100 secretion, this is exceeded by its effects in upregulating LDL receptors and modifying VLDL composition. We hypothesize that the improved lipid profile, in particular the decrease in cholesterol-rich VLDL particles, may contribute to a possible antiatherogenic action of GH. GH appears to have an important role in hepatic apo B-100 metabolism. However, we are just at the beginning of understanding the underlying mechanism. Further studies are required to investigate the effect of GH on other lipoprotein classes, in particular VLDL subfractions, intermediate-density lipoprotein, LDL and high-density lipoprotein. The key question, however, remains as to whether GH replacement therapy can reduce cardiovascular mortality. Long-term studies with sufficient numbers of patients are required to answer this question.

Posttranslational modifications of cardiac ryanodine receptors: Ca2+ signaling and EC-coupling

In cardiac muscle, a number of posttranslational protein modifications can alter the function of the Ca(2+) release channel of the sarcoplasmic reticulum (SR), also known as the ryanodine receptor (RyR). During every heartbeat RyRs are activated by the Ca(2+)-induced Ca(2+) release mechanism and contribute a large fraction of the Ca(2+) required for contraction. Some of the posttranslational modifications of the RyR are known to affect its gating and Ca(2+) sensitivity. Presently, research in a number of laboratories is focused on RyR phosphorylation, both by PKA and CaMKII, or on RyR modifications caused by reactive oxygen and nitrogen species (ROS/RNS). Both classes of posttranslational modifications are thought to play important roles in the physiological regulation of channel activity, but are also known to provoke abnormal alterations during various diseases. Only recently it was realized that several types of posttranslational modifications are tightly connected and form synergistic (or antagonistic) feed-back loops resulting in additive and potentially detrimental downstream effects. This review summarizes recent findings on such posttranslational modifications, attempts to bridge molecular with cellular findings, and opens a perspective for future work trying to understand the ramifications of crosstalk in these multiple signaling pathways. Clarifying these complex interactions will be important in the development of novel therapeutic approaches, since this may form the foundation for the implementation of multi-pronged treatment regimes in the future. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Cardiac Pathways of Differentiation, Metabolism and Contraction.