Dopamine and alcohol relapse: D1 and D2 antagonists increase relapse rates in animal studies and in clinical trials

A considerable number of animal studies on the effects of dopaminergic agents on alcohol intake behavior have been performed. Acute alcohol administration in rats induces dopamine release in the caudate nucleus and in the nucleus accumbens, an effect related among others to reinforcement. It has been repeatedly suggested that D1 and D2 receptor activation mediates reward. As alcohol consumption and dopaminergic transmission seem to have a close relationship, all kinds of dopaminergic agents may be regarded as putative therapeutics for preventing relapse. In a prospective European double-blind multicenter clinical trial, comparing the D1, D2, D3 antagonist flupenthixol and placebo in 281 chronic alcohol-dependent patients (27.4% women), the application of the Lesch typology made an outcome differentiation possible. It could be shown in which patients flupenthixol administration was followed by a significantly higher relapse rate and in which patient groups no differences were found when compared to placebo.     

Effects of sodium intake on cardiovascular variables in humans during posture changes and ambulatory conditions

The hypothesis was tested that cardiac output (CO) and stroke volume (SV) are increased by a moderate physiological elevation in sodium intake with a more pronounced effect in the ambulatory upright seated than supine position. Fourteen healthy males were investigated during ambulatory and controlled laboratory conditions at the end of two consecutive 5-day periods with sodium intakes of 70 (low) and 250 (high) mmol/24 h or vice versa, respectively. Comparing high and low sodium intake, plasma volume and plasma protein concentrations were 9 and 8% higher in the seated and the supine position, respectively. When seated during laboratory conditions, CO was 5.3 +/- 0.2 l/min on the high sodium intake vs. 4.8 +/- 0.2 l/min on the low (P < 0.05), and SV was 81 +/- 3 vs. 68 +/- 3 ml (P < 0.05). In the supine position, SV was 107 +/- 3 ml on the high vs. 99 +/- 3 ml (P < 0.05) on the low sodium intake, while CO remained unchanged. The difference in CO and SV induced by the change in sodium intake was significantly higher in the seated than in the supine position (P < 0.05). During upright ambulatory conditions, CO was 5.9 +/- 0.2 l/min during the high and 5.2 +/- 0.2 l/min during the low sodium intake (P < 0.05), and SV was 84 +/- 3 and 69 +/- 3 ml (P < 0.05), respectively. Mean arterial pressure was unchanged by the variations in sodium intake. In conclusion, increments in sodium intake within the normal physiological range increase CO and SV and more so in the seated vs. the supine position. These changes are readily detectable during upright, ambulatory conditions. The results indicate that the higher SV and CO could constitute an arterial baroreflex stimulus for the augmented renal sodium excretion.     

Control of mitotic exit in budding yeast. In vitro regulation of Tem1 GTPase by Bub2 and Bfa1

The elimination of mitotic kinase activity at the end of mitosis is essential for progression to the next stage of the eukaryotic cell cycle. In budding yeast, this process is controlled by a regulatory cascade called the mitotic exit network. Extensive genetic data indicate that mitotic exit network activity is determined by a GTP-binding protein, Tem1, and its putative regulators, Bub2, Bfa1, and Lte1. Here we describe the purification and in vitro activities of Tem1, Bub2, and Bfa1. We describe the nucleotide binding properties of Tem1 and characterize its intrinsic GTPase activity. The combination of Bfa1 and Bub2 acts as a two-component GTPase-activating protein for Tem1. In the absence of Bub2, Bfa1 inhibits the GTPase and GTP exchange activities of Tem1. This inhibition is elicited by either the N- or C-terminal regions of Bfa1, which also retain some ability to co-activate GTPase activity in the presence of Bub2. Although the C-terminal region of Bfa1 binds to Bub2, no interaction of the N-terminal half of Bfa1 with Bub2 was detected despite their combined GAP activity. Therefore, we propose that Bfa1 acts both as an adaptor to connect Bub2 and Tem1 and as an allosteric effector that facilitates this interaction.     

Structural requirements for mutational lutropin/choriogonadotropin receptor activation

Different activation mechanisms of glycoprotein hormone receptors, which are members of the G protein-coupled receptor superfamily, have been proposed. For example, the large ectodomain of glycoprotein hormone receptors may function as an inverse agonist keeping the transmembrane domain in an inactive conformation. To provide support for this hypothesis, we have generated different lutropin/choriogonadotropin receptor (LHR) constructs lacking the ectodomain. Although some ectodomain-deficient LHR constructs were targeted to the cell surface, cAMP levels remained unchanged under basal conditions and agonist application but could be increased by a mutation within the transmembrane domain 6 (D578H). Taking advantage of a constitutive activating mutation (S277N) located in the extracellular domain, we showed that the intact leucine-rich repeat-containing ectodomain is essential for constitutive activation of the LHR by mutation of the hinge region. Our findings support an activation scenario in which agonist binding or mutational alterations expose a structure within the ectodomain, which then activates the transmembrane core.     

The mitochondrial presequence translocase: an essential role of Tim50 in directing preproteins to the import channel

Mitochondrial proteins with N-terminal targeting signals are transported across the inner membrane via the presequence translocase, which consists of membrane-integrated channel proteins and the matrix Hsp70 import motor. It has not been known how preproteins are directed to the import channel. We have identified the essential protein Tim50, which exposes its major domain to the intermembrane space. Tim50 interacts with preproteins in transit and directs them to the channel protein Tim23. Inactivation of Tim50 strongly inhibits the import of preproteins with a classical matrix-targeting signal, while preproteins carrying an additional inner membrane-sorting signal do not strictly depend on Tim50. Thus, Tim50 is crucial for guiding the precursors of matrix proteins to their insertion site in the inner membrane.     

Multiple nicotinic acetylcholine receptor α-subunits are expressed in the arterial system of the rat

Neuronal nicotinic acetylcholine receptors (nAChRs) are hetero- and homopentamers built up by nine different alpha-subunits and three different beta-subunits. The subtype composition within the receptor determines ligand specificity, affinity and cation permeability. In this study we focused on the distribution of the ligand binding alpha-subunits in the rat arterial system by means of RT-PCR and immunohistochemistry. Subtypes alpha3, alpha5, alpha7 and alpha10 were found to be expressed by endothelial cells, suggesting that they are equipped both with calcium-preferring (alpha7 homopentamers) and monovalent cation-preferring (heteropentamers containing alpha3- and alpha5-subunits) nAChR channels. All alpha-subtypes except alpha9 were expressed by vascular smooth muscle cells with a highly specific distribution pattern along the vascular tree. While every alpha-subunit except alpha9 was detected in the thoracic aorta, intrapulmonary arterial branches contained only alpha7 immunoreactivity, and other vascular beds held intermediate positions with respect to the extent of alpha-subunit expression. Current knowledge does not allow to correlate these distribution patterns to specific functions, but it can be anticipated that at least some components of nAChR-mediated signalling in the arterial wall are highly specific for individual arteries.     

CPMK2, an SLT2-homologous mitogen-activated protein (MAP) kinase,is essential for pathogenesis of Claviceps purpurea on rye: evidence for a second conserved pathogenesis-related MAP kinase cascade in phytopathogenic fungi

Cpmk2, encoding a mitogen-activated protein (MAP) kinase from the ascomycete Claviceps purpurea, is an orthologue of SLT2 from Saccharomyces cerevisiae, the first isolated from a biotrophic, non-appressorium-forming pathogen. Deletion mutants obtained by a gene replacement approach show impaired vegetative properties (no conidiation) and a significantly reduced virulence, although they retain a limited ability to colonize the host tissue. Increased sensitivity to protoplasting enzymes indicates that the cell wall structure of the mutants may be altered. As the phenotypes of these mutants are similar to those observed in strains of the rice pathogen, Magnaporthe grisea, that have been deprived of their MAP kinase gene mps1, the ability of cpmk2 to complement the defects of delta mps1 was investigated. Interestingly, the C. purpurea gene, under the control of its own promoter, was able to complement the M. grisea mutant phenotype: transformants were able to sporulate and form infection hyphae on onion epidermis and were fully pathogenic on barley leaves. This indicates that, despite the differences in infection strategies, which include host and organ specificity, mode of penetration and colonization of host tissue, CPMK2/MPS1 defines a second MAP kinase cascade (after the Fus3p/PMK1 cascade) essential for fungal pathogenicity.