Three-dimensional synaptic ultrastructure in the dentate gyrus and hippocampal area CA3 in the Ts65Dn mouse model of Down syndrome

Down syndrome (DS) results from trisomy of human chromosome 21. Ts65Dn mice are an established model for DS and show several phenotypes similar to those in people with DS. However, there is little data on the structural plasticity of synapses in the trisynaptic pathway in the hippocampus. Here we investigate 3D ultrastructure of synapses in the hippocampus of age-matched control (2N) and Ts65Dn male mice. Serial ultrathin sections and 3D reconstructions characterize synapses in the middle molecular layer (MML) of dentate gyrus and in thorny excrescences (TEs) in proximal portions of apical dendrites of CA3 pyramidal neurons. 3D analysis of synapses shows phenotypes that distinguish Ts65Dn from 2N mice. For the MML, synapse density was reduced by 15% in Ts65Dn vs. 2N mice (P < 0.05). Comparative 3D analyses demonstrate a significant decrease in the number of thorns per TE in CA3 in Ts65Dn vs. 2N mice (by ≈45%, P = 0.01). Individual thorn volume was 3 times smaller in Ts65Dn vs. 2N mice (P = 0.02). A significant decrease in the number of thorn projections per TE in Ts65Dn vs. 2N mice was accompanied by a decrease of filopodium-like protrusions on the surface of TEs (P = 0.02). However, the volume of postsynaptic densities in CA3 Ts65Dn and 2N mice was unchanged (P = 0.78). Our findings suggest that the high degree of plasticity of CA3 thorns may be connected with their filopodial origin. Alterations of 3D synaptic structure in Ts65Dn mice may further contribute to the diminished plasticity in DS.     

Pharmacy student perceptions of adverse event reporting

Objective. To assess US pharmacy students’ knowledge and perceptions of adverse event reporting.Methods. To gauge pharmacy students'' impressions of adverse event reporting, a 10-question survey instrument was administered that addressed student perceptions of the reporting procedures of the Food and Drug Administration (FDA) and pharmaceutical manufacturers, as well as student understanding of the Health Insurance Portability and Accountability Act (HIPAA) and its relationship to adverse event reporting.Results. Two hundred twenty-eight pharmacy students responded to the survey. The majority of respondents believed that the FDA is more likely than a pharmaceutical company to take action regarding an adverse event. There were misconceptions relating to the way adverse event reports are handled and the influence of HIPAA regulations on reporting.Conclusions. Communication between the FDA and pharmaceutical manufacturers regarding adverse event reports is not well understood by pharmacy students. Education about adverse event reporting should evolve so that by the time pharmacy students become practitioners, they are well acquainted with the relevance and importance of adverse event reporting.     

Leucettines, a class of potent inhibitors of cdc2-like kinases and dual specificity, tyrosine phosphorylation regulated kinases derived from the marine sponge leucettamine B: modulation of alternative pre-RNA splicing

We here report on the synthesis, optimization, and biological characterization of leucettines, a family of kinase inhibitors derived from the marine sponge leucettamine B. Stepwise synthesis of analogues starting from the natural structure, guided by activity testing on eight purified kinases, led to highly potent inhibitors of CLKs and DYRKs, two families of kinases involved in alternative pre-mRNA splicing and Alzheimer's disease/Down syndrome. Leucettine L41 was cocrystallized with CLK3. It interacts with key residues located within the ATP-binding pocket of the kinase. Leucettine L41 inhibits the phosphorylation of serine/arginine-rich proteins (SRp), a family of proteins regulating pre-RNA splicing. Indeed leucettine L41 was demonstrated to modulate alternative pre-mRNA splicing, in a cell-based reporting system. Leucettines should be further explored as pharmacological tools to study and modulate pre-RNA splicing. Leucettines may also be investigated as potential therapeutic drugs in Alzheimer's disease (AD) and in diseases involving abnormal pre-mRNA splicing.     

Photosensitizer Radachlorin®: Skin cancer PDT phase II clinical trials

"Radachlorin"(?), also known in the EU as Bremachlorin, a composition of 3 chlorophyll a derivatives in an aqueous solution, was introduced into the Russian Pharmacopoeia. Its GMP (Good Manufacturing Practice) facility based manufacturing method was patented. Laboratory experiments and clinical phase I were performed. Protocols were designed for PDT of basal cell carcinoma of the skin to result in GCP (Good Clinical Practice)-conformed randomized phase II clinical studies. "Radachlorin"(?) solution for intravenous infusions 0.35% 10mL in the doses of 0.5-0.6 and 1.0-1.2mg/kg and a gel for topical application 0.1% 25g in the dose of 0.1g/cm(2) were photoactivated by 2.5W 662nm semiconductor laser "LAKHTA-MILON(?)" (St. Petersburg, Russia) in light doses of 200, 300 (solution), 400, 600, 800 (gel) J/cm(2). Safety study showed no side effects and a good tolerability of "Radachlorin"(?) by patients. There was no normal skin/subdermal tissue damage after both laser and sun light exposure. The main part (98%) of the drug was excreted or metabolized in the first 48h. Drug administration at a dose of 1.0-1.2mg/kg and irradiation at 3h with 662±3nm light at a dose of 300J/cm(2) (solution) and 4 PDT sessions at an interval of 1 week with 3h gel exposure, followed by 400J/cm(2) light exposure (gel) were found to be the optimal treatment regimes. Having successfully passed clinical trials, "Radachlorin"(?) achieved marketing authorization in Russia in 2009 and a conditional approval in South Korea in 2008. It is a candidate for phase III clinical trials in the EC and may be commercialized as a prospective second-generation photosensitizer.     

PI3Kα inhibitors that inhibit metastasis

Previous genetic analyses have suggested that mutations of the genes encoding PI3Kα facilitate invasion and metastasis but have less effect on primary tumor growth. These findings have major implications for therapeutics but have not been factored into pre-clinical drug development designs. Here we show that the inhibition of PI3Kα by newly designed small molecule inhibitors prevented metastasis formation in mice but had much less effect on the growth of subcutaneous xenografts or primary intra-abdominal tumors. These data support the idea that PI3Kα plays an important role in the metastatic process and suggest a more informed strategy for selecting drugs worthy of further development for clinical application.     

Synapsin- and actin-dependent frequency enhancement in mouse hippocampal mossy fiber synapses

The synapsin proteins have different roles in excitatory and inhibitory synaptic terminals. We demonstrate a differential role between types of excitatory terminals. Structural and functional aspects of the hippocampal mossy fiber (MF) synapses were studied in wild-type (WT) mice and in synapsin double-knockout mice (DKO). A severe reduction in the number of synaptic vesicles situated more than 100 nm away from the presynaptic membrane active zone was found in the synapsin DKO animals. The ultrastructural level gave concomitant reduction in F-actin immunoreactivity observed at the periactive endocytic zone of the MF terminals. Frequency facilitation was normal in synapsin DKO mice at low firing rates (approximately 0.1 Hz) but was impaired at firing rates within the physiological range (approximately 2 Hz). Synapses made by associational/commissural fibers showed comparatively small frequency facilitation at the same frequencies. Synapsin-dependent facilitation in MF synapses of WT mice was attenuated by blocking F-actin polymerization with cytochalasin B in hippocampal slices. Synapsin III, selectively seen in MF synapses, is enriched specifically in the area adjacent to the synaptic cleft. This may underlie the ability of synapsin III to promote synaptic depression, contributing to the reduced frequency facilitation observed in the absence of synapsins I and II.     

Systemic hypertension in two patients with ASL deficiency: A result of nitric oxide deficiency?

Argininosuccinic aciduria (ASA) is an inborn error of ureagenesis which if untreated leads to hyperammonemia, accumulation of argininosuccinic acid and arginine depletion. The presence of high blood pressure in patients with ASA has been reported so far as transient in one newborn. We describe the first two patients, one child and one young adult, with ASA and persistent systemic hypertension. Extensive evaluation of both patients excluded secondary causes of systemic hypertension. The intriguing link between nitric oxide (NO) production and hypertension lead us to hypothesize that the deficiency of endogenously synthesized arginine caused by ASL deficiency is responsible for the increased blood pressure.