The hypothalamo-hypophysial system of the lamprey,Lampetra fluviatilis L.

The distribution of monoaminergic structures was studied in the proximal neurosecretory contact region and neurohypophysis of the lamprey by light and electron microscopic radioautography. Only weak radioautographic reactions were found in the proximal neurosecretory contact region 1 h after injection of 3H-dopamine. High-resolution radioautography revealed some labeled neurosecretory terminals mainly in contact with the basement membrane of the connective tissue layer separating the proximal neurosecretory contact region from the hypophysial pars distalis. The number of silver grains as well as the number of neurosecretory terminals marked by the presence of labeled dopamine was much higher in the neurohypophysis of the same species. In the latter, labeled neurosecretory terminals were found in contact with the connective tissue layer containing blood vessels of the general circulation. Some neurosecretory terminals make synaptoid contacts with tanycyte perikarya and their basal processes. According to their ultrastructure and the size of their granules, the labeled neurosecretory terminals are identical with the B type terminals described in both neurohemal regions (transmission electron microscopy). No labeled neurosecretory terminals were observed in the proximal neurosecretory contact region and the neurohypophysis of lampreys treated with the serotonin precursor, 3H-5-hydroxytryptophan.     

Nuclear structures in the late oogenesis of Rana temporaria. III. Electron microscopic studies]

The ultrastructural organization of the vitellogenic oocyte nucleus (stage VI, according to Duryee, 1950) was studied in normal and in vitro hormone-stimulated maturing oocytes of Rana temporaria. At this stage, numerous nucleoli are gathered around the knot of highly contracted chromosomes (the karyosphere) thus making the karyosphere capsule. Light microscope observations reveal three zones in the capsule: a central fibrous zone separating the chromosomes from the nucleoli, a middle zone, consisting of numerous nucleoli and a distinct fibrous componen; in addition a fibrous zone on the capsule periphery is seen. The nucleoli are fibrillar, bear no proribosomal granules and do not synthesize RNA. This period is characterized by an intensive fragmentation and segregation of the nucleolar material. Numerous micronucleoli and nuclear bodies occur in the nucleus. The nucleoli are normally compound and irregular in shape to become spherical in hormone-stimulated maturing oocytes. In the central fibrous zone of the capsule, separating the chromosomes from the nucleoli, some peculiar abundant accumulations of annuli were detected lacking the membranes component. Annuli are linked with the fibrous material and are regularily packed making peculiar pseudomembranes (PMM). The chromatin is connected with PMM directly. In the middle zone of the capsule, accumulations of PMM are also seen, though less abundant and less regularly packed; along with annuli, membranous areas of various size and form are met in PMM. PMM are connected with the micronucleoli with filaments 20 nm thick. In the peripheral zone of the capsule, a variety of membranous structures is detected: intranuclear annuli lamellae, component of the capsule consists of different membranous and pseudomembranous (with annuli) structures. A question of the contribution of the chromatin material in the formation of the fibrous capsular component (PMM and membranous structures) is discussed.     

On the synthesis of viral ribonucleic acids and ribonucleoproteins in the submitochondrial system completely free of interfering cytoplasmic contaminations

Summary The synthesis of virus-specific macromolecules was studied in the reconstituted system containing inner membrane-matrix fraction from rat liver mitochondria and infectious RNA of Venezuelian equine encephalomyelitis (VEE) virus. In a series of preliminary experiments it was shown that isolated submitochondrial fraction was completely free of interfering cytoplasmic contaminations and particularly, of cytoplasmic 80S ribosomes. VEE RNA when added to submitochondrial system caused significant stimulation of RNA and protein synthesis. These processes were resistant to actinomycin D which inhibited profoundly the synthesis of proper mitochondrial macromolecules. The stimulating effect of VEE RNA in experiments with submitochondrial system was about three times higher than that with intact mitochondria. The stimulation of¹⁴C-amino acid incorporation increased as a function of incubation time; a certain lag-period being observed. The newly formed virus-specific RNA's and ribonucleoproteins were identified with the aid of sedimentation analysis. In particular, radioactive RNA's with sedimentation coefficients 40S and 26-18S were isolated from the incubated system. These RNA's are similar respectively to VEE genome RNA and doublestranded VEE replicative RNA. In double labelling experiments with³H-uridine and¹⁴Camino acids it was shown that VEE RNA induced synthesis of ribonucleoproteins containing newly formed RNA and protein. These RNP possessed sedimentation coefficients 60-80S, 140S and 300S in sucrose gradient and buoyant densities 1.32 and 1.50 g/cm³ in cesium chloride gradients. These properties of ribonucleoproteins synthesized de novo in submitochondrial system are close to those of RNP intermediates of VEE virus reproduction in the infected cells. We concluded that viral RNA could program virus-specific synthesis in the submitochondrial system under conditions that eliminated the contribution of cytoplasmic ribosomes.     

Hen egg white lysozyme fibrillation: a deep‐UV resonance Raman spectroscopic study

Amyloid fibrils are associated with numerous degenerative diseases. The molecular mechanism of the structural transformation of native protein to the highly ordered cross‐β structure, the key feature of amyloid fibrils, is under active investigation. Conventional biophysical methods have limited application in addressing the problem because of the heterogeneous nature of the system. In this study, we demonstrated that deep‐UV resonance Raman (DUVRR) spectroscopy in combination with circular dichroism (CD) and intrinsic tryptophan fluorescence allowed for quantitative characterization of protein structural evolution at all stages of hen egg white lysozyme fibrillation in vitro. DUVRR spectroscopy was found to be complimentary to the far‐UV CD because it is (i) more sensitive to β ‐sheet than to α ‐helix, and (ii) capable of characterizing quantitatively inhomogeneous and highly light‐scattering samples. In addition, phenylalanine, a natural DUVRR spectroscopic biomarker of protein structural rearrangements, exhibited substantial changes in the Raman cross section of the 1000‐cm^–1 band at various stages of fibrillation. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Molecular Co-occupancy Identifies Transcription Factor Binding Cooperativity In Vivo

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Caffeine has a dual influence on NMDA receptor–mediated glutamatergic transmission at the hippocampus

Caffeine, a stimulant largely consumed around the world, is a non-selective adenosine receptor antagonist, and therefore caffeine actions at synapses usually, but not always, mirror those of adenosine. Importantly, different adenosine receptors with opposing regulatory actions co-exist at synapses. Through both inhibitory and excitatory high-affinity receptors (A₁R and A₂R, respectively), adenosine affects NMDA receptor (NMDAR) function at the hippocampus, but surprisingly, there is a lack of knowledge on the effects of caffeine upon this ionotropic glutamatergic receptor deeply involved in both positive (plasticity) and negative (excitotoxicity) synaptic actions. We thus aimed to elucidate the effects of caffeine upon NMDAR-mediated excitatory post-synaptic currents (NMDAR-EPSCs), and its implications upon neuronal Ca²⁺ homeostasis. We found that caffeine (30–200 μM) facilitates NMDAR-EPSCs on pyramidal CA1 neurons from Balbc/ByJ male mice, an action mimicked, as well as occluded, by 1,3-dipropyl-cyclopentylxantine (DPCPX, 50 nM), thus likely mediated by blockade of inhibitory A₁Rs. This action of caffeine cannot be attributed to a pre-synaptic facilitation of transmission because caffeine even increased paired-pulse facilitation of NMDA-EPSCs, indicative of an inhibition of neurotransmitter release. Adenosine A_2ARs are involved in this likely pre-synaptic action since the effect of caffeine was mimicked by the A_2AR antagonist, SCH58261 (50 nM). Furthermore, caffeine increased the frequency of Ca²⁺ transients in neuronal cell culture, an action mimicked by the A₁R antagonist, DPCPX, and prevented by NMDAR blockade with AP5 (50 μM). Altogether, these results show for the first time an influence of caffeine on NMDA receptor activity at the hippocampus, with impact in neuronal Ca²⁺ homeostasis.

     

Gadolinium-containing carbon nanomaterials for magnetic resonance imaging: Trends and challenges

Gadolinium-containing carbon nanomaterials are a new class of contrast agent for magnetic resonance imaging. They are characterized by a superior proton relaxivity to any current commercial gadolinium contrast agent and offer the possibility to design multifunctional contrasts. Intense efforts have been made to develop these nanomaterials because of their potential for better results than the available gadolinium contrast agents. The aim of the present work is to provide a review of the advances in research on gadolinium-containing carbon nanomaterials and their advantages over conventional gadolinium contrast agents. Due to their enhanced proton relaxivity, they can provide a reliable imaging contrast for cells, tissues or organs with much smaller doses than currently used in clinical practice, thus leading to reduced toxicity (as shown by cytotoxicity and biodistribution studies). Their active targeting capability allows for improved MRI of molecular or cellular targets, overcoming the limited labelling capability of available contrast agents (restricted to physiological irregularities during pathological conditions). Their potential of multifunctionality encompasses multimodal imaging and the combination of imaging and therapy.