Analysis of neuronal compartments has revealed many state-dependent changes in geometry but establishing synapse-specific mechanisms at the nanoscale has proven elusive. We co-expressed ...channelrhodopsin2-GFP and mAPEX2 in a subset of hippocampal CA3 neurons and used trains of light to induce late-phase long-term potentiation (L-LTP) in area CA1. L-LTP was shown to be specific to the labeled axons by severing CA3 inputs, which prevented back-propagating recruitment of unlabeled axons. Membrane-associated mAPEX2 tolerated microwave-enhanced chemical fixation and drove tyramide signal amplification to deposit Alexa Fluor dyes in the light-activated axons. Subsequent post-embedding immunogold labeling resulted in outstanding ultrastructure and clear distinctions between labeled (activated), and unlabeled axons without obscuring subcellular organelles. The gold-labeled axons in potentiated slices were reconstructed through serial section electron microscopy; presynaptic vesicles and other constituents could be quantified unambiguously. The genetic specification, reliable physiology, and compatibility with established methods for ultrastructural preservation make this an ideal approach to link synapse ultrastructure and function in intact circuits.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
This study tests the hypothesis that plasminogen activator inhibitor-1 (PAI-1) contributes to aldosterone-induced renal and cardiac injury. The effects of 12-week aldosterone (2.8 μg/day)/salt (1% ...drinking water) versus vehicle/salt on renal and cardiac histology and mRNA expression were determined in wild-type (WT) and PAI-1 deficient (PAI-1(−/−)) mice. Systolic blood pressure was similar in aldosterone-infused WT and PAI-1(−/−) mice until 12 weeks, when it was significantly higher in the WT mice. At 12 weeks, urine volume, sodium excretion, and sodium/potassium ratio were similarly increased in the two aldosterone-infused groups. In contrast, urine albumin excretion was greater in aldosterone-infused WT mice (mean±s.d.: 699.0±873.0 μg/24 h) compared to vehicle-infused WT (23.6±9.0 μg/24 h, P=0.003) or aldosterone-infused PAI-1(−/−) mice (131.6±110.6 μg/24 h, P=0.007). Aldosterone increased glomerular area to a greater extent in WT (4651±577 versus 3278±488 μm2/glomerulus in vehicle-infused WT, P<0.001) than in PAI-1(−/−) mice (3713±705 μm2/glomerulus, P=0.001 versus aldosterone-infused WT), with corresponding mesangial expansion. Renal collagen content was also increased in aldosterone-infused WT versus PAI-1(−/−) mice. In WT mice, aldosterone increased renal mRNA expression of PAI-1, collagen I, collagen III, osteopontin, fibronectin, monocyte chemoattractant protein-1 (MCP-1), and F4/80 (all P<0.05), but not transforming growth factor beta (TGF-β). In PAI-1(−/−) mice, aldosterone increased renal expression of collagen I, osteopontin, fibronectin, and MCP-1, and tended to increase collagen III. Renal osteopontin expression was diminished in aldosterone-treated PAI-1(−/−) compared to aldosterone-treated WT mice (P=0.05). Aldosterone induced cardiac hypertrophy but not fibrosis in WT and PAI-1(−/−) mice. PAI-1 contributes to aldosterone-induced glomerular injury.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Little Lambda, Who Made Thee? GOTTESMAN, Max E; WEISBERG, Robert A
Microbiology and Molecular Biology Reviews,
12/2004, Volume:
68, Issue:
4
Journal Article
Aspiration in patients with acute stroke Daniels, Stephanie K.; Brailey, Kevin; Priestly, Daniel H. ...
Archives of physical medicine and rehabilitation,
1998, 1998-Jan, 1998-01-00, 19980101, Volume:
79, Issue:
1
Journal Article
Peer reviewed
Objectives: To determine the frequency and clinical predictors of aspiration within 5 days of acute stroke.
Design: Case series.
Setting: Tertiary care center.
Patients: Consecutive stroke patients (
...n = 55) with new neurologic deficit evaluated within 5 days of acute stroke.
Main Outcome Measures: Comparison of features identified on clinical swallowing and oromotor examinations and occurrence of aspiration (silent or overt) evident on videofluoroscopic swallow study (VSS).
Results: Aspiration occurred in 21 of 55 patients (38%). Whereas 7 of 21 patients (33%) aspirated overtly, 14 (67%) aspirated silently on VSS. Chi-square analyses revealed that dysphonia, dysarthria, abnormal gag reflex, abnormal volitional cough, cough after swallow, and voice change after swallow were significantly related to aspiration and were predictors of the subset of patients with silent aspiration. Logistic regression revealed that abnormal volitional cough and cough with swallow, in conjunction, predicted aspiration with 78% accuracy.
Conclusions: Silent aspiration appears to be a significant problem in acute stroke patients because silent aspiration occurred in two thirds of the patients who aspirated. The prediction of patients at risk for aspiration was significantly improved by the presence of concurrent findings of abnormal volitional cough and cough with swallow on clinical examination.
We describe a mechanism by which nascent RNA inhibits transcriptional pausing.
PutL RNA of bacteriophage HK022 suppresses transcription termination at downstream terminators and pausing within a ...nearby U-rich sequence. In vitro transcription and footprinting assays reveal that this pausing results from backtracking of RNA polymerase and that binding of nascent
putL RNA to polymerase limits backtracking by restricting re-entry of the transcript into the RNA exit channel. The restriction is local and relaxes as the transcript elongates. Our results suggest that
putL RNA binds to the surface of polymerase close to the RNA exit channel, a region that includes amino acid residues important for antitermination. Although binding is essential for antipausing and antitermination, these two activities of
put differ: antipausing is limited to the immediate vicinity of the
putL site, but antitermination is not. We propose that RNA anchoring to the elongation complex is a widespread mechanism of pause regulation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
In addition to the contractile proteins actin and myosin, contractile filaments of striated muscle contain other proteins that are important for regulating the structure and the interaction of the ...two force-generating proteins. In the thin filaments, troponin and tropomyosin form a Ca-sensitive trigger that activates normal contraction when intracellular Ca is elevated. In the thick filament, there are several myosin-binding proteins whose functions are unclear. Among these is the myosin-binding protein C (MBP-C). The cardiac isoform contains four phosphorylation sites under the control of cAMP and calmodulin-regulated kinases, whereas the skeletal isoform contains only one such site, suggesting that phosphorylation in cardiac muscle has a specific regulatory function. We isolated natural thick filaments from cardiac muscle and, using electron microscopy and optical diffraction, determined the effect of phosphorylation of MBP-C on cross bridges. The thickness of the filaments that had been treated with protein kinase A was increased where cross bridges were present. No change occurred in the central bare zone that is devoid of cross bridges. The intensity of the reflections along the 43-nm layer line, which is primarily due to the helical array of cross bridges, was increased, and the distance of the first peak reflection from the meridian along the 43-nm layer line was decreased. The results indicate that phosphorylation of MBP-C (i) extends the cross bridges from the backbone of the filament and (ii) increases their degree of order and/or alters their orientation. These changes could alter rate constants for attachment to and detachment from the thin filament and thereby modify force production in activated cardiac muscle.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
In this issue of
Molecular Cell,
Luo et al. (2008) show that S10 protein can function in the ribosome or the transcript elongation complex with minimal structural change, providing new insights into ...the roles of S10 and NusB in transcript elongation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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