Fiberoptic system for recording dendritic calcium signals in layer 5 neocortical pyramidal cells in freely moving rats

Calcium influx into the dendritic tufts of layer 5 neocortical pyramidal neurons modifies a number of important cellular mechanisms. It can trigger local synaptic plasticity and switch the firing properties from regular to burst firing. Due to methodological limitations, our knowledge about Ca2+ spikes in the dendritic tuft stems mostly from in vitro experiments. However, it has been speculated that regenerative Ca2+ events in the distal dendrites correlate with distinct behavioral states. Therefore it would be most desirable to be able to record these Ca2+ events in vivo, preferably in the behaving animal. Here, we present a novel approach for recording Ca2+ signals in the dendrites of populations of layer 5 pyramidal neurons in vivo, which ensures that all recorded fluorescence changes are due to intracellular Ca2+ signals in the apical dendrites. The method has two main features: 1) bolus loading of layer 5 with a membrane-permeant Ca2+ dye resulting in specific loading of pyramidal cell dendrites in the upper layers and 2) a fiberoptic cable attached to a gradient index lens and a prism reflecting light horizontally at 90 degrees to the angle of the apical dendrites. We demonstrate that the in vivo signal-to-noise ratio recorded with this relatively inexpensive and easy-to-implement fiberoptic-based device is comparable to conventional camera-based imaging systems used in vitro. In addition, the device is flexible and lightweight and can be used for recording Ca2+ signals in the distal dendritic tuft of freely behaving animals.

Sensitive and highly specific quantitative real-time PCR and ELISA for recording a potential transfer of novel DNA and Cry1Ab protein from feed into bovine milk

To address food safety concerns of the public regarding the potential transfer of recombinant DNA (cry1Ab) and protein (Cry1Ab) into the milk of cows fed genetically modified maize (MON810), a highly specific and sensitive quantitative real-time PCR (qPCR) and an ELISA were developed for monitoring suspicious presence of novel DNA and Cry1Ab protein in bovine milk. The developed assays were validated according to the assay validation criteria specified in the European Commission Decision 2002/657/EC. The detection limit and detection capability of the qPCR and ELISA were 100 copies of cry1Ab microL(-1) milk and 0.4 ng mL(-1) Cry1Ab, respectively. Recovery rates of 84.9% (DNA) and 97% (protein) and low (<15%) imprecision revealed the reliable and accurate estimations. A specific qPCR amplification and use of a specific antibody in ELISA ascertained the high specificity of the assays. Using these assays for 90 milk samples collected from cows fed either transgenic (n = 8) or non-transgenic (n = 7) rations for 6 months, neither cry1Ab nor Cry1Ab protein were detected in any analyzed sample at the assay detection limits.

How does the ascending aorta geometry change when it dissects?

OBJECTIVES The purpose of this study is to delineate changes in aortic geometry and diameter due to dissection. BACKGROUND Aortic diameter is the major criterion for elective ascending aortic replacement for dilated ascending aortas to prevent aortic dissection. However, recommendations are made on the basis of clinical experience and observation of diameters of previously dissected aortas. METHODS Six tertiary centers on 2 continents reviewed their acute aortic dissection type A databases, which contained 1,821 patients. Included were all non-Marfan patients with nonbicuspid aortic valves who had undergone computed tomography angiography <2 years before and within 12 h after aortic dissection onset. Aortic geometry before and after dissection onset were compared. RESULTS Altogether, 63 patients were included (27 spontaneous and 36 retrograde dissections, median age 68 [57; 77] years; 54% were men). In all but 1 patient, maximum ascending aortic diameter was <55 mm before aortic dissection onset. The largest increase in diameter and volume induced by the dissection were observed in the ascending aorta (40.1 [36.6; 45.3] mm vs. 52.9 [46.1; 58.6] mm, +12.8 mm; p < 0.001; 124.0 [90.8; 162.5] cm(3) vs. 171.0 [147.0; 197.0] cm(3), +47 cm(3); p < 0.001). Mean aortic arch diameter increased from 39.8 (30.5; 42.6) mm to 46.4 (42.0; 51.6) mm (+6.6 mm; p < 0.001) and descending thoracic aorta diameter from 31.2 (27.0; 33.3) mm to 34.9 (30.9; 39.5) mm (+3.7 mm; p < 0.001). Changes in thoracic aorta geometry were similar for spontaneous and retrograde etiology. CONCLUSIONS Geometry of the thoracic aorta is affected by aortic dissection, leading to an increase in diameter that is most pronounced in the ascending aorta. Both spontaneous and retrograde dissection result in similar aortic geometry changes.

Geometry and kinematics of the Roisan-Cignana Shear Zone, and the orogenic evolution of the Dent Blanche Tectonic System (Western Alps)

The Dent Blanche Tectonic System (DBTS) is a composite thrust sheet derived from the previously thinned passive Adriatic continental margin. A kilometric high-strain zone, the Roisan-Cignana Shear Zone (RCSZ) defines the major tectonic boundary within the DBTS and separates it into two subunits, the Dent Blanche s.s. nappe to the northwest and the Mont Mary nappe to the southeast. Within this shear zone, tectonic slices of Mesozoic and pre-Alpine meta-sediments became amalgamated with continental basement rocks of the Adriatic margin. The occurrence of high pressure assemblages along the contact between these tectonic slices indicates that the amalgamation occurred prior to or during the subduction process, at an early stage of the Alpine orogenic cycle. Detailed mapping, petrographic and structural analysis show that the Roisan-Cignana Shear Zone results from several superimposed Alpine structural and metamorphic stages. Subduction of the continental fragments is recorded by blueschist-facies deformation, whereas the Alpine collision is reflected by a greenschist facies overprint associated with the development of large-scale open folds. The postnappe evolution comprises the development of low-angle brittle faults, followed by large-scale folding (Vanzone phase) and finally brittle extensional faults. The RCSZ shows that fragments of continental crust had been torn off the passive continental margin prior to continental collision, thus recording the entire history of the orogenic cycle. The role of preceding Permo-Triassic lithospheric thinning, Jurassic rifting, and ablative subduction processes in controlling the removal of crustal fragments from the reactivated passive continental margin is discussed. Results of this study constrain the temporal sequence of the tectono-metamorphic processes involved in the assembly of the DBTS, but they also show limits on the interpretation. In particular it remains difficult to judge to what extent precollisional rifting at the Adriatic continental margin preconditioned the efficiency of convergent processes, i.e. accretion, subduction, and orogenic exhumation.