D-AKAP2, a novel protein kinase A anchoring protein with a putative RGS domain

Subcellular localization directed by specific A kinase anchoring proteins (AKAPs) is a mechanism for compartmentalization of cAMP-dependent protein kinase (PKA). Using a two-hybrid screen, a novel AKAP was isolated. Because it interacts with both the type I and type II regulatory subunits, it was defined as a dual specific AKAP or D-AKAP1. Here we report the cloning and characterization of another novel cDNA isolated from that screen. This new member of the D-AKAP family, D-AKAP2, also binds both types of regulatory subunits. A message of 5 kb pairs was detected for D-AKAP2 in all embryonic stages and in all adult tissues tested. In brain, skeletal muscle, kidney, and testis, a 10-kb mRNA was identified. In testis, several small mRNAs were observed. Therefore, D-AKAP2 represents a novel family of proteins. cDNA cloning from a mouse testis library identified the full length D-AKAP2. It is composed of 372 amino acids which includes the R binding fragment, residues 333-372, at its C-terminus. Based on coprecipitation assays, the R binding domain interacts with the N-terminal dimerization domain of RIalpha and RIIalpha. A putative RGS domain was identified near the N-terminal region of D-AKAP2. The presence of this domain raises the intriguing possibility that D-AKAP2 may interact with a Galpha protein thus providing a link between the signaling machinery at the plasma membrane and the downstream kinase.     

The apolipoprotein E allele epsilon 4 is overrepresented in patients with the Lewy body variant of Alzheimer's disease

We determined apolipoprotein E (ApoE) genotypes in 122 autopsied demented patients. The frequency of the ApoE epsilon 4 allele was 39.6% in Alzheimer's disease (AD), 29.0% in the Lewy body variant of AD (LBV), and 6.25% in diffuse Lewy body disease. For AD and LBV patients, the epsilon 4 frequency was significantly higher than that reported in nondemented controls (10 to 15%). Therefore, LBV and AD share ApoE epsilon 4 as a genetic risk factor, providing further evidence that these conditions overlap.     

Metabotropic glutamate receptors are involved in long-term potentiation in isolated slices of rat medial frontal cortex

The prelimbic region of medial frontal cortex in the rat receives a direct input from the hippocampus and this functional connection is essential for aspects of spatial memory. Activity-dependent changes in the effectiveness of synaptic transmission in the medial frontal cortex, namely long-term potentiation (LTP) and long-term depression (LTD) can persist for tens of minutes or hours and may be the basis of learning and memory storage. Glutamatergic activation of ionotropic receptors is required to induce both LTP and LTD. We now present evidence of the involvement of metabotropic glutamate receptors in LTP in isolated slices of frontal cortex. Repetitive bursts of stimulation at theta frequencies (TBS) were applied to layer II, and monosynaptic EPSPs were monitored in layer V neurons of the prelimbic area. TBS was found to be more effective at inducing LTP than tetanic stimulation at 100 Hz and produced LTP that lasted >30 min in 8 out of 14 neurons. Tetanic stimulation at 100 Hz in the presence of the N-methyl--aspartate (NMDA)-antagonist 2-amino-5-phosphonopentanoate (AP5) was reported to be a reliable method of inducing LTD in prelimbic cortex (). However we found that this protocol did not facilitate the induction of LTD. The role of metabotropic glutamate receptors (mGluR) in LTP was assessed by using the selective, broad-spectrum antagonist (R, S)-alpha-methyl-4- carboxyphenylglycine (MCPG). This drug significantly reduced the incidence of LTP after TBS to only 1 of 14 neurons (P < 0.02, chi2 test). The pooled responses to TBS in MCPG showed significantly reduced potentiation [(P < 0.02, analysis of variance (ANOVA)]. The broad-spectrum mGluR agonist (1S, 3R)-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD) and the selective group I agonist S-3 hydroxyphenylglycine(S-3HPG) both produced membrane depolarization, an increase in number of spikes evoked by depolarizing current pulses, and a reduction in the afterhyperpolarization. Similar effects were produced by these agonists even when synaptic transmission was blocked by use of the gamma-aminobutyric acid-B (GABAB) receptor agonist, 200 microM baclofen, which suggests that group I mGluRs are present on layer V neurons. We conclude that mGluRs participate in the production of LTP in prelimbic cortex, and that this excitatory effect could be mediated by the postsynaptic group I mGluRs.     

Influence of age and health on immune functions and trace elements

Apparently healthy elderly donors were screened according to a simple protocol that included clinical examination and the determination of hematological and biochemical values. This screening was performed to detect subclinical alterations which might interfere with immune responses and trace element status. The elderly were divided into two groups. The first group consisted of 22 (age 76 +/- 1 years) positively selected elderly (PSE), i.e. healthy subjects with no hematological and laboratory alterations, the second one comprised 13 (age 75 +/- 1 years) negatively selected elderly (NSE). Data were then compared with those obtained from 40 (age 35 +/- 2 years) healthy young controls. In both groups of elderly donors, plasma zinc levels were normal, while plasma copper concentrations were increased. Intracellular values of zinc and copper in mono- and polymorphonuclear cells from both groups of elderly were within reference limits. After in vitro activation, granulocyte chemiluminescence activity was impaired only in NSE. A decrement in the number of circulating CD3 lymphocytes and an increase in CD8d, CD57 cells were found in PSE, while NSE showed an increased number of CD3,DR cells and CD8d, CD57, CD8b,CD57 and CD16,CD56 positive cells. Our results indicate that only plasma copper levels were affected by age, whereas subclinical alterations in hematological or biochemical values appear to impair immune responses in the elderly.     

Common migraine ("migraine without aura"): localization of the initial pain of attack

The importance of maximal versus submaximal exercise testing and the significance of heart failure on the prognostic value of exercise-provoked ST-segment depression > or = 0.1 mV was studied in 143 patients recovering from acute myocardial infarction. Patients were exercise tested prior to discharge and follow up lasted for up to 18 months (mean 17 months). End-point was first major event (i.e. first non-fatal reinfarction or death). A symptom-limited exercise test was superior to a heart-rate-limited test in detecting ST-segment depressions (27% vs. 20%: P < 0.5), and patients with ST-segment depression at lower heart rates did not have an increased risk of subsequent events compared with patients with ST-segment depression at higher heart rates (14% vs. 27%; NS). Heart failure surpassed ST-segment depression as a risk predictor (34% vs. 18%). Based on a meta-analysis including 13 studies (1987 patients) exercise-provoked ST-segment depression possessed an increased risk of subsequent major events (P < 0.0001; risk ratio = 1.90; 95% confidence limits 1.43,2.51). Thus, ST-segment depression provoked by a symptom-limited test selects patients with an increased risk of subsequent major events. In patients with a history of heart failure exercise-provoked ST-segment depression is of limited value.     

Predation of livestock ticks by chickens as a tick-control method in a resource-poor urban environment

The increasing prevalence of streptococci as causes of potentially fatal nosocomial bacteremia requires that antimicrobial agents used for empiric therapy in hospitalized patients include both pneumococci and viridans group streptococci as well as beta-hemolytic streptococci in their activity profile. In this study, the in vitro activity of cefepime, a new fourth-generation cephalosporin, was compared with other cephalosporins versus 197 nosocomial blood stream isolates of streptococci (20 Streptococcus pneumoniae, 104 viridans group, and 73 beta-hemolytic) isolated from patients at more than 30 medial centers from 1995 to 1997. Additional agents tested included penicillin, erythromycin, and vancomycin. Overall, cefepime inhibited 83% of the isolates at concentrations < or = 0.5 microgram/mL and 100% at < or = 8 micrograms/mL. By comparison, ceftazidime inhibited 35 and 88% of isolates at the same concentrations. Cefepime was approximately eightfold more potent than ceftazidime against S. pneumoniae, viridans group streptococci, and beta-hemolytic streptococci. Among the 42 isolates with penicillin MICs > 0.12 microgram/mL, 100% were inhibited by cefepime and only 48% by ceftazidime at < or = 8 micrograms/mL. The rank order of activity for all six agents against the 197 isolates was vancomycin > ceftriaxone > cefepime > penicillin > erythromycin > ceftazidime. Based on the results of the present study, cefepime and ceftriaxone were the superior cephalosporins in potency and spectrum for empiric coverage of patients at risk for streptococcal blood stream infections.     

The use of quantitative bacteriologic assessment of bone

A new sub-class of binding protein-dependent transporter with specificity for a broad range of polar amino acids has been identified by sequence comparison, in Rhizobium leguminosarum, Rhodobacter capsulatus, Escherichia coli and Pseudomonas fluorescens. Southern blotting and PCR analysis has shown that transporters from this new sub-class are widely distributed in Gram-negative bacteria, including, in addition to the above, Citrobacter freundii, Erwinia carotovorum and Rhizobium meliloti. ABC transporters of polar amino acids can be divided into two groups: those with narrow solute specificity and the newly identified sub-class with broad solute specificity. The binding and inner membrane proteins from transporters with a broad solute specificity are larger by approximately 30% than those with a narrow solute specificity. Multiple alignment of the inner membrane proteins from all sequenced polar amino acid transporters indicates there is an N-terminal conserved region that may be involved in solute specificity. A conserved arginine or lysine at residue 30 of this region is changed to glutamate in arginine transporters. Residue 53 also has a strong correlation with the charge on the transported solute, with basic amino acid transporters replacing an aliphatic amino acid at this position with a negatively charged amino acid. The general amino acid permease from R. leguminosarum, which will transport aliphatic as well as basic and acidic amino acids, juxtaposes two prolines at residues 52 and 53 of the N-terminal conserved region.